Category: Bacterial Infectious Diseases (Including Chlamydia, Mycoplasma, and Legionella Infections)

Bacterial Infectious Diseases (Including Chlamydia, Mycoplasma, and Legionella Infections)

  • Nontuberculous Mycobacteria

    There are other mycobacteria besides Mycobacterium tuberculosis, some of which are frequently pathogenic for humans and some of which rarely cause human infection. The nontuberculous mycobacteria were originally called “atypical mycobacteria.” The first useful classification was that of Runyon, who subdivided the nontuberculous mycobacteria into four groups, depending on growth speed and colony characteristics (Table 14-2). These groups have some clinical value as rough guides to the type of organism present while awaiting more definitive identification and speciation. It is desirable to place the organisms in the correct species, since some members (species) of any of the Runyon groups may not be pathogenic very often or may differ in degree of pathogenicity. Mycobacterium intracellulare or Mycobacterium avium-intracellulare complex (formerly known as the Battey Mycobacterium), a member of Runyon group III, and Mycobacterium kansasii, from Runyon group I, together cause the majority of significant nontuberculous mycobacterial infections in about equal proportions. These produce a disease similar to pulmonary tuberculosis (although often milder or more indolent) that is much more frequent in adults. M. avium-intracellulare infections are very frequent in persons with acquired immunodeficiency syndrome (AIDS) or AIDS-related conditions. The mycobacterial infection frequently becomes bacteremic or disseminated due to compromise of the immune system by HIV-1 causing AIDS. Runyon group II organisms are more frequent in children and clinically tend to cause cervical lymphadenopathy. Diagnosis of the nontuberculous mycobacteria is essentially the same as for M. tuberculosis. Skin tests (old tuberculin or PPD) for M. tuberculosis will also cross-react with the nontuberculous mycobacteria. In general, the nontuberculous mycobacteria tend to produce less reaction to standard tuberculin skin tests than M. tuberculosis. In fact, several studies claim that the majority of positive intermediate-strength tuberculin skin test results that produce a reaction of less than 10 mm diameter are due to nontuberculous mycobacterial infection rather than TB. Skin test antigens are available for each of the nontuberculous mycobacterial groups, although some reports challenge the specificity of these preparations. The main clinical importance of these nontuberculous organisms is resistance that many have toward one or more of the standard antituberculous chemotherapeutic agents.

    Classification of the atypical mycobacteria

    Table 14-2 Classification of the atypical mycobacteria

    Several reports have linked one of the non-MTB organisms, M. paratuberculosis, to Crohn’s disease (regional ileitis).

  • Tuberculosis and Mycobacterial Disease

    Tuberculosis is caused by Mycobacterium tuberculosis (MTB), a rod-shaped bacterium that requires special media for culture and that has the peculiarity of “acid-fastness” (resistance to decolorization by strong acidic decolorizing chemicals such as acid alcohol after being stained by certain stains such as carbol fuchsin). Tuberculosis is still very important and common despite advances in drug therapy. It has been reported that about 25% of persons exposed to MTB will become infected; and of those infected, about 10% will develop clinical disease (range, 5%-40%). The disease usually begins in the chest due to inhalation of airborne infectious material. This material is carried to some localized area of the lung alveoli and provokes a host response of granulomatous inflammation around the material (the “Ghon complex”). It is thought that in many cases there is also a silent hematogenous spread of the organisms. In most cases the host is able to contain and eventually destroy the organisms in the chest and those reaching other locations. Those in the lungs seem better able to survive than those deposited elsewhere. In some cases the organisms remain dormant, and the infection can be reactivated at a later date; in some cases the initial infection spreads; and in some cases reinfection takes place. If infection in the lungs progresses to clinical disease, the most important symptoms are cough, fever, and hemoptysis. (The most important diseases to rule out are lung carcinoma and bronchiectasis.) The kidney is involved in a small percentage of advanced cases, with the main symptom being hematuria (Chapter 12). A small number of patients develop widespread extrapulmonary disease, known as miliary tuberculosis. The laboratory findings in tuberculosis depend to some extent on the stage and severity of the disease.

    Chest x-ray films

    Chest x-ray films often provide the first suggestion of tuberculosis and are a valuable parameter of severity, activity, and response to therapy. Depending on the situation, there are a variety of possible roentgenographic findings. These may include one or more of the following:

    1. Enlargement of hilar lymph nodes.
    2. Localized pulmonary infiltrates. These occur characteristically in an upper apical location or, less commonly, in the superior segment of the lower lobes. Cavitation of lesions may occur.
    3. Miliary spread (small punctate lesions widely distributed). This pattern is not common and may be missed on routine chest x-ray films.
    4. Unilateral pleural effusion. The most common causes are tuberculosis, carcinoma, and congestive heart failure. Tuberculosis has been reported to cause 60%-80% of so-called idiopathic pleural effusions, although this percentage varies greatly depending on the patient’s geographic location and other factors.

    Sputum smear

    Sputum smears provide a rapid presumptive diagnosis in pulmonary tuberculosis. The smear is usually stained by one of the acid-fast (acid-fast bacillus, or AFB) procedures (usually the Ziehleelsen or Kinyoun methods). Fluorescent Auramine-o staining methods are available, faster, and somewhat more sensitive. Smears require about 5 Ч 103 organisms/ml of specimen for microscopic detection. The more advanced the infection, the more likely it is to yield a positive smear. Therefore, the rate of positive findings is low in early, minimal, or healing tuberculosis. Also, the smear may be normal in a substantial minority of advanced cases. Culture is more reliable for detection of tuberculosis and also is necessary for confirmation of the diagnosis, for differentiation of MTB from the “atypical” mycobacteria, and for sensitivity studies of antituberculous drugs. According to the literature, false negative smears (smear negative but culture positive) have been reported in an average of 50% of cases (literature range, 16%-70%). Some of these false negative results may be due to laboratory technique problems and differences in smear staining methods. A high centrifugation speed when concentrating the specimen is said to increase the yield of positive smears. False positive smears (positive smear but negative culture) have been reported, averaging about 1%-5% of positive smears (literature range, 0.5%-55%). Some of these were apparently due to contamination of water used in the smear-staining procedure by saprophytic mycobacteria. Control slides are necessary to prevent this. Some authorities believe that only 1-2 acid-fast organisms/300 oil immersion fields should be considered negative (although indicative of need for further specimens). Smears may sometimes be positive for up to 2 months when cultures are negative if the patient is on antituberculous drug therapy (this would not be considered a genuine false positive, since the drugs inhibit mycobacterial growth or the organisms may be nonviable). After 2 months, persistence of positive smears raises the question of treatment failure. Temporary persistence of positive smears with negative cultures is more likely to occur in severe cavitary disease (in one series, this occurred in 20% of cases). Sputum specimens should be collected (for culture and smear of the concentrated specimen) once daily for at least 3 days. If the smear is definitively positive, further smears are not necessary. Also, a definitively positive smear means high probability that culture of the specimens already collected will obtain positive results, and it is not necessary to collect more than three specimens or to proceed to more complicated diagnostic procedures. If smears are negative, one must consider the possibility that the culture may also be negative, and conventional cultures on standard solid media average 20 days to produce growth from MTB smear-positive specimens and about 27 days from MTB smear-negative specimens (overall range, 2–8 weeks).

    Culture

    Sputum culture is preferred for pulmonary tuberculosis (gastric aspiration may be done if adequate sputum specimens cannot be obtained); urine culture is preferred for renal involvement; and bone marrow culture is preferred in miliary tuberculosis. Reports indicate that an early morning specimen, either of sputum or urine, produces almost as many positive results as a 24-hour specimen and has much less problem with contamination. Special mycobacteria culture media are needed. The necessity for adequate sputum culture specimens, regardless of the concentrated smear findings, should be reemphasized. Several reports indicate that aerosol techniques produce a significantly greater yield of positive cultures than ordinary sputum collection. The aerosol mixture irritates the bronchial tree and stimulates sputum production. At any rate, it is necessary to get a “deep cough” specimen; saliva alone, although not completely useless, is much less likely to reveal infection and is much more likely to be contaminated. If sputum cultures are negative or if the patient is unable to produce an adequate sputum sample, gastric aspiration may be used. Gastric contents are suitable only for culture; nontuberculous acid-fast organisms may be found normally in the stomach and cannot be distinguished from M. tuberculosis on AFB smear. If renal tuberculosis is suspected, urine culture should be done (Chapter 12). However, renal tuberculosis is uncommon; and even with urine specimens obtained on 3 consecutive days, only about 30% of cases are positive.

    Cultures should be grown in high carbon dioxide atmosphere, since this is reported to increase the number of positive cultures by at least 10%. Inoculation on several varieties of media increases the number of positive results by 5%-10%. The 4% sodium hydroxide traditionally used to digest and decontaminate sputum before concentration also kills some mycobacteria. Use of weaker digestion agents increases culture yield, but troublesome overgrowth by other bacteria may also increase.

    Culture should be done on all tissue specimens when tuberculosis is suspected. Acid-fast stains on tissue slides reveal tuberculosis organisms in only 30%-40% of cases that are positive by culture. Several newer methods such as BACTEC (which uses liquid media and a machine that detects metabolic products of bacterial growth) have been able to decrease detection time for MTB smear-positive specimens to 8 days and time for MTB smearegative specimens to 14 days (overall range, 1-3 weeks). The system is about 93% sensitive compared to conventional multimedia culture. Once culture growth occurs, the organism must be identified. Conventional methods require biochemical and growth tests to be performed that may take 3-6 weeks to complete. The BACTEC system has a nucleic acid phosphate method that can identify MTB (only) in 3-5 days. Commercial DNA probes are available that can identify MTB and certain non-MTB mycobacteria in 1 day. Gas-liquid chromatography and high-performance liquid chromatography have also been used. Antibiotic sensitivity studies are recommended when a mycobacterial organism is isolated, since multiresistant MTB is increasing in various areas and non-MTB mycobacteria have often been multiresistant. Conventional culture methods take 21 days; BACTEC takes about 5 days.

    Data on sputum culture sensitivity using conventional AFB media is difficult to find since culture is usually considered the gold standard of AFB detection. Sputum culture appear to average about 75% sensitivity (range, 69%-82%). Sensitivity using BACTEC averages about 85% (range, 72%-95%).

    Nucleic acid probe

    Nucleic acid (DNA) probe methods are now becoming available that permit direct nonsmear detection of mycobacteria in clinical specimens. The first such test available (Gen-Probe, Inc.) is reported in two studies to be 83%-85% sensitive compared with culture using sputum specimens. However, it has been reported that antituberculous drugs can interfere with the probe; this study found sensitivity in nontreated patients to be over 90% (when comparing probe to culture, culture only detects about 75%-80% of cases). Ten percent of specimens were positive by probe but negative by culture, which may represent additional true positives that could not be confirmed. A DNA probe is also available specifically for M. tuberculosis. Same-day results can be obtained. Disadvantages of this first-generation method are need for considerable technologist time and certain special equipment. Compared with culture, the general Mycobacterium screen probe will not differentiate M. tuberculosis from other mycobacteria, whereas the specific M. tuberculosis probe will not detect the other mycobacteria. Neither probe would provide therapeutic drug sensitivity information. The major drawback regarding its use as replacement for the acid-fast smear is the relatively high cost of the probe method, very high if only one specimen at a time is processed. DNA probes with PCR amplification have been reported (e.g., Roche Diagnostics) that are said to have a sensitivity of 3-30 organisms/ml (compared to at least 5 Ч 103 organisms/ml required for a positive acid-fast smear). Nevertheless, one study involving 7 prestigious worldwide reference laboratories who were sent sputum or saliva specimens to which various quantities of BCG (M. Bovis) mycobacteria were added, showed false positive PCR rates of 3%-77%. In specimens containing 102 organisms, sensitivity ranged from 0%-55%; in specimens containing 103 organisms, 2%-90%; and in specimens containing 101 organisms, 20%-98%.

    Skin test (Mantoux test)

    This test is performed with an intradermal injection of purified protein derivative (PPD) or old tuberculin (Table 14-1). A positive result is represented by an area of induration having a specified diameter by 48 hours. The diameter used to be 10 mm but was redefined in 1990 to require different diameters depending on the person’s risk group (see box). In addition, a distinction was made between “reaction” (diameter or width of induration without record of previous test result) and “conversion” (increase in reaction width within 2 years from last previous reaction width). For all persons younger than 35 years of age whose previous reaction was negative, an increase in PPD induration of 10 mm or more in diameter within a period of 2 years would be considered a conversion and presumptive suspicion for occult tuberculosis (TB), whereas the change would have to be at least 15 mm for persons 35 years of age or more (that is, for nonrisk persons above or below age 35 who have had a PPD within 2 years, conversion criteria would replace reaction size criteria).

    Table 14-1 Comparison of tuberculosis skin tests*
    Comparison of tuberculosis skin tests


    A positive skin test is a manifestation of hypersensitivity to the tubercle bacillus. This reaction usually develops about 6 weeks after infection, although it may take several months. A positive reaction means previous contact and infection with TB; the positive reaction does not itself indicate whether the disease is currently active or inactive. However, in children under 3 years of age it usually means active TB infection. Apparently, once positive, the reaction persists for many years or for life, although there is evidence that a significant number of persons revert to negative reactions if the infection is completely cured early enough. In a few cases of infection the test never becomes positive. The Mantoux test may revert to negative or fail to become positive in the following circumstances:

    1. In about 20% of seriously ill patients, due to malnutrition (severe protein deficiency).
    2. In newborns and occasionally in old age.
    3. In some persons with viral infections, or within 1 month after receiving live virus vaccination.
    4. In 50% or more of patients with miliary tuberculosis.
    5. In a high percentage of patients with overwhelming pulmonary tuberculosis.
    6. In a considerable number of patients who are on steroid therapy or immunosuppressive therapy.
    7. In many persons who also have sarcoidosis or Hodgkin’s disease.
    8. In some persons with chronic lymphocytic leukemia or malignant lymphoma.
    9. In some patients with chronic renal failure or severe illness of various types.
    10. In some persons with old infection (“waning” of reactivity).
    11. When there is artifact due to improper skin test technique (e.g, subcutaneous rather than intradermal injection).

    In cachectic patients and those with protein malnutrition, treatment with an adequate protein diet can restore Mantoux test reactivity to most patients after about 2 weeks. In patients after age 50 with M. tuberculosis infection, especially those with old previous infection, the PPD skin test sometimes may slowly decrease in reactivity and eventually become negative. (How often this occurs is controversial; the best estimate seems to be 8%-10%, but studies range from 0.1%-21%, possibly influenced by the elapsed time period since infection and time intervals between testing.) If another skin test is performed, the new skin test itself stimulates body reaction and may restore reactivity (“booster phenomenon”). This phenomenon could simulate a new infection if previous infection were not known, since the next time a skin test is performed the physician would see only conversion of a negative to a positive reaction. Restoration of skin reactivity can take place in only 1 week, so that retesting 1 week after the first negative reaction can usually show whether or not there is potential for the booster reaction. (The 1-week interval would in most cases not be long enough for true conversion in persons with their first infection.) Repeated skin tests will not cause a nonexposed person to develop a positive reaction. Some investigators recommend skin testing with antigens used to demonstrate the presence of skin test anergy (e.g., Candida or Trichophyton antigen) if the Mantoux test is repeatedly negative in a person with substantial suspicion of mycobacterial infection.

    The standard procedure for skin testing is to begin with an intermediate strength PPD (or the equivalent). If the person has serious infection, some clinics recommend starting with a first-strength dose to avoid necrosis at the injection site. A significant minority of patients with tuberculosis (9%-17%) fail to react to intermediate strength PPD; a second-strength dose is then indicated.

    Miliary tuberculosis

    Miliary TB is clinically active TB that is widely disseminated in the body by hematogenous spread. Clinical symptoms are often nonspecific, such as fever, weakness, and malaise. There frequently is an associated condition, such as alcoholism, intravenous (IV) drug abuse, or malignancy, that decreases immunologic defenses. About 20% have a negative tuberculin skin test reaction. About 35% do not show a miliary pattern on chest x-ray film. If routine clinical and culture methods fail, biopsy of bone marrow or liver may be useful. Liver biopsy has a fairly good positive yield (up to 80%), considering that a needle biopsy specimen is such a tiny random sample of a huge organ. However, it is usually difficult to demonstrate acid-fast organisms on liver biopsy even when tubercles are found, and without organisms the diagnosis is not absolutely certain. Bone marrow aspiration is probably the best procedure in such cases. Bone marrow yields much better results for mycobacterial culture than for demonstration of tubercles. Routine marrow (Wright-stained) smears are worthless for histologic diagnosis in TB. Aspirated material may be allowed to clot in the syringe, then formalin-fixed and processed as a regular biopsy specimen for histologic study. Before clotting, some of the aspirate is inoculated into a suitable TB culture medium. It should be emphasized that bone marrow aspiration or liver biopsy is not indicated in pulmonary tuberculosis (since this disease is relatively localized), only in miliary TB.

    PPD Reaction Size Considered “Positive” (Intracutaneous 5 TU* Mantoux Test at 48 Hours)
    5 MM OR MORE
    Human immunodeficiency virus
    (HIV) infection or risk factors for HIV
    Close recent contact with active TB case
    Persons with chest x-ray consistent with healed TB
    10 MM OR MORE
    Foreign-born persons from countries with high TB prevalence in Asia, Africa, and Latin America
    Intravenous (IV) drug users
    Medically underserved low-income population groups (including Native Americans, Hispanics, and African Americans)
    Residents of long-term care facilities (nursing homes, mental institutions)
    Medical conditions that increase risk for TB (silicosis, gastrectomy, undernourished, diabetes mellitus, high-dose corticosteroids or immunosuppression RX, leukemia or lymphoma, other malignancies
    Employees of long-term care facilities, schools, child-care facilities, health care facilities
    15 MM OR MORE
    All others not listed above
    * TU, tuberculin units.

    Renal tuberculosis

    Renal TB is almost always bilateral and presumably results from nonhealing infection produced during the transient bacteremia of the lung primary stage. There is usually a latent period of many years before clinical infection becomes evident. The estimated incidence of eventual active renal infection is about 2%-5%, but this probably represents incidence in high-risk groups. About 25% (range, 20%-75%) of patients are said to have a normal chest x-ray film. About 14% (range, 12%-15%) of patients are reported to have a negative PPD skin test result. Even the intravenous pyelogram results (IVP) are normal in about 25% (range, 14%-39%) of cases. Most patients do not have systemic symptoms, such as fever. The erythrocyte sedimentation rate was elevated in 23% of patients in one report. Only 20%-56% have urinary tract symptoms. Gross hematuria is the classic finding in renal TB but is present in only about 20% of patients. Pyuria (with negative urine culture) and microscopic hematuria are more frequent, occurring in about 65%-85% of cases. Some patients have a positive urine culture with some ordinary pathogen in addition to renal TB. Urine culture for TB was mentioned previously; 30%-80% of patients have positive cultures when three 24-hour or early morning specimens are collected (true culture sensitivity is probably less than 80%, since the diagnosis could easily be missed with negative cultures).

  • Anaerobic Bacteria

    The three major sources of anaerobic organisms are the dental and mouth area, the lower intestinal tract (ileum and colon), and the female external genital tract (vagina and vulva area). Anaerobes comprise about 95% of the bacterial flora of the colon and outnumber aerobes in the mouth and vagina. From the mouth the organisms can reach the lungs and brain; from the vagina they may involve the remainder of the female genital tract; diseases of the lower intestinal tract may release anaerobes into the abdominal cavity; and all of these locations may be the source of organisms found in the bloodstream. In addition, anaerobic bacteria are frequently associated with chronic infection, such as bronchiectasis, abscess, or chronic osteomyelitis. Infections with anaerobes frequently are mixed infections that also contain aerobes. The types of infection most frequently associated with anaerobes are listed in the box below.

    Types of Infection Most Frequently Associated With Anaerobes (Alone or as a Mixed Infection)
    Dental or mouth area infection
    Chronic sinus infection
    Bite wounds
    Bronchiectasis and aspiration pneumonia
    Gynecologic intraabdominal and extra abdominal infection
    Abscess of any area other than skin; including brain, abdominal and thoracic cavities, and any organ
    Infections associated with the intestinal tract, especially the colon (diverticulitis, appendicitis, bowel perforation, etc.)
    Deep tissue infection or necrosis
    Biliary tract infection
    Chronic osteomyelitis

    Anaerobic infections usually center on three groups of organisms: Clostridia species, Bacteroides species, and the anaerobic streptococci.

    Clostridia

    These gram-positive anaerobic rods include several important organisms. Clostridium perfringens (Clostridium welchii) is the usual cause of gas gangrene. It is a normal inhabitant of the GI tract and reportedly can be isolated from the skin in about 20% of patients and from the vagina and female genitalia in about 5%. Therefore, just as with S. aureus, a culture report of C. perfringens isolated from an external wound does not necessarily mean that the organism is producing clinical infection. When isolated from abdominal or biliary tract infections, C. perfringens most often is part of a polymicrobial infection and, although serious, is not quite as alarming as isolation from a deep tissue wound. Clostridium perfringens occasionally is a cause of “food poisoning” (discussed later).

    Clostridium tetani causes tetanus. The organism can be found in the human GI tract but is more common in animals. Spores are widely distributed in soil. Clinical disease is produced by release of bacterial exotoxin after local infection in a manner analogous to diphtheria. Puncture-type wounds are notorious for high risk of C. tetani infection. The incubation time is 3-14 days in most patients, a few cases being reported as early as 24 hours after exposure. Cultures for C. tetani are said to be positive in less than 50% of patients. Clostridium difficile is the most frequent proven cause of antibiotic-associated diarrhea. In the 1950s and early 1960s, broad-spectrum oral antibiotics were frequently used, and S. aureus was thought to be the major cause of antibiotic-associated enteritis. Beginning in the late 1960s parenteral antibiotics became much more common in hospitals, and C. difficile was eventually proven to be the major etiology. Clostridium difficile enteritis usually occurs during or after therapy with antibiotics, although some patients have not received antibiotics. One report indicated a possible association with diarrhea induced by cancer chemotherapeutic agents. The condition may consist only of varying degrees of diarrhea, may progress to inflammation of portions of the colon mucosa, and in the more severe form (known as pseudomembraneouscolitis) there is inflammation and partial destruction of varying areas of the colon mucosa, with formation of a pseudomembrane of fibrin, necrotic cells, and segmented neutrophils on the surface of the remnants of the affected mucosa. Some patients develop intestinal perforation and sepsis.

    Diagnosis of C. difficile colitis is not always easy. Only about 25%-30% (range, 20%-33%) of diarrhea occurring with antibiotic therapy is due to C. difficile, with most of the remainder not associated with currently known infectious agents. Sigmoidoscopy or colonoscopy can be done to document pseudomembrane formation, but this procedure runs the risk of perforation; and even in pseudomembraneous enterocolitis (PMC), pseudomembranes can be demonstrated in only about 50% (range, 40%-80%) of patients. The patient stools in PMC typically contain WBCs and often contain red blood cells, but gross blood is uncommon. However, WBCs on Gram stain actually are present in only about 45%-50% of cases. For several years, culture was the usual means of diagnosis. Today, this is not often done. C. difficile can be cultured, with reliable results, only on special media, so diagnosis usually is made through detection of C. difficile cytotoxin. Stool specimens must be frozen and sent to a reference laboratory with dry ice. The specimen container must be sealed tightly since the carbon dioxide from the dry ice can inactivate the toxin. Another problem is the fact that C. difficile can be found in the stool of some clinically healthy persons, including 30%-40% (range, 11%-63%) of clinically healthy neonates, 10%-15% of children, 3% (range, 0%-10%) of adults, 20% (range, 11%-36%) of hospitalized persons without diarrhea and not taking antibiotics, and about 30% (range, 20%-46%) of patients taking antibiotics but without diarrhea. Even in patients with endoscopy-proven PMC, stool culture is positive for C. difficile in only about 90% (range, 75%-95%) of cases. Most laboratories today rely more on tests to detect C. difficile toxin, which has been shown to correlate much better with C. difficile clinical infection than does stool culture. There are several C. difficile toxins, of which the best characterized are called toxins A and B. Toxin B is a cytotoxin that must be detected by a tissue culture system with an incubation period of 48 hours. In endoscopy-proven PMC, toxin B sensitivity is reported to be about 90% (range, 64%-100%). Results may be positive in some patients with positive C. difficile cultures who are clinically normal, especially in infants. An enzyme immunoassay kit is commercially available that detects both toxin B and toxin A (Cytoclone A+B). Sensitivity reported to date is about 85%-90% (range, 76%-99%). Toxin A is an enterotoxin that originally was assayed with a biologic system, the rabbit ileal loop test. Several EIA tests are commercially available for toxin A, with reported sensitivity of about 85% (range, 65%-97%). There is a simple latex agglutination kit available (CDT or Culturette CDT) that can provide same-day results. This kit was originally thought to detect toxin A but subsequently was found to be detecting an enzyme (glutamate dehydrogenase) from C. difficile. The test detects non-toxin-producing as well as toxin-producing C. difficile and cross-reacts with Clostridium sporogenes and a few strains of other clostridia. Nevertheless, in various studies this test frequently gave results equivalent to those of the cytotoxicity test (about 75%-90%; range, 38%-97% positive in patients with proven PMC, with roughly 90% specificity). Another company has a similar test (Meritec-CD). In one comparison between CDT and Meritec-CD, Meritec-CD showed about 10% less sensitivity.

    Clostridium botulinus produces botulism. Botulism is a severe food poisoning due to ingestion of preformed botulinal endotoxin (a powerful neurotoxin) contained in the contaminated food, rather than actual infection of the patient by the organism. C. botulinum spores are widespread, but they can germinate only in anaerobic conditions at a fairly high pH (>4.6). These conditions are met in canned foods that have not been sufficiently heated during the canning process. Therefore, C. botulinum is usually associated with canned food, especially home canned. Vegetables are the most frequently involved home-canned food, but any variety of canned food may become contaminated. Fortunately, the disease is not common. Although the endotoxin is preformed, symptoms most often appear 12-36 hours after ingestion (in adult patients) and commonly include nausea, vomiting, and abdominal cramps (about 50% of patients), constipation (75%), cranial nerve signs of dysphagia, dysarthria, and dry mouth (80%-90%), upper or lower extremity weakness (70%), and diplopia or other eye abnormalities (90%). There is no fever or diarrhea. Differential diagnosis in adults includes Guillain-Barrй syndrome, drug reaction (especially phenothiazines), myasthenia gravis, cerebrovascular accident, chemical poisoning (mercury, arsenic, etc.), diphtheria, and tick bite paralysis (Landry’s ascending paralysis). Botulism may also occur in infants, usually between ages 3 and 26 weeks (median age 10 weeks). The classic syndrome is onset of constipation followed by 4-5 days of progressive feeding difficulty, ptosis, muscle hypotonia, and possibly respiratory distress. Mildly affected infants may have varying degrees of “failure to thrive” with feeding difficulty and mild muscle weakness, whereas severely affected infants may have severe respiratory distress. Some cases of sudden infant death syndrome (SIDS) have been ascribed to infant botulism, whereas some infants and older children (investigated during studies initiated by botulism cases) were found to harbor C. botulinum organisms without symptoms. Honey was incriminated in some cases as the source of infection but was not used in the majority of cases.

    Standard laboratory tests, including those on the CSF, are usually normal in botulism. Electromyography (EMG) may be helpful in differentiating adult botulism from certain other neurologic diseases, but the typical EMG findings are neither specific nor always present. The diagnosis is confirmed by stool culture and demonstration of C. botulinum toxin, neither of which can be done in the ordinary laboratory. In addition, food suspected of contamination should always be tested. Patient vomitus and gastric contents have also been tested. The basic specimens in adult botulism are stool (25 gm or more) and serum (at least 5 ml). For diagnosis of infant botulism a stool specimen is required; but infant serum rarely contains C. botulinum toxin and therefore infant serum specimens are not necessary. The specimens should be kept refrigerated, since the toxin is heat labile, and sent to a reference laboratory with a coolant (for short distances) or dry ice (for more than 1-day transit). There are several C. botulinum serotypes, but most cases are caused by types A and B, with type A predominating.

    Bacteroides

    Bacteroides are the most frequent organisms found in anaerobic infections. They comprise several species of gramegative rods normally found in the mouth, the intestine, and the female genital tract. Isolation of these organisms often raises the question of their significance or pathogenicity. It seems well established that bacteroides occasionally cause serious infection, which frequently results in abscess or gangrene. The most commonly associated clinical situations include septic abortion, aspiration pneumonia, focal lesions of the GI tract (e.g., carcinoma or appendicitis), and pelvic abscess in the female.

    Anaerobic streptococci

    Anaerobic streptococci are frequently associated with Bacteroides infection but may themselves produce disease. They are normally present in the mouth and GI tract. Septic abortion and superinfection of lesions in the perirectal area seem to be the most commonly associated factors. Anaerobic streptococci are also part of the fusiform bacteria-spirochetal synergistic disease known as Vincent’s angina.

    Laboratory diagnosis of anaerobic infections

    Anaerobic specimens cannot be processed by the laboratory as easily as those from aerobic infections. Anaerobic bacteria in general do not grow as readily as common aerobic bacteria, and prereduced media or other special media are often needed. Achieving and maintaining anaerobic culture conditions are not a simple matter. However, by far the greatest problem is the specimen received by the laboratory. If the specimen is not properly maintained in an anaerobic environment en route to the laboratory, the best and most elaborate facilities will be of little help. For abscesses or any fluid material, the preferred collection technique is to aspirate the material with a sterile syringe and needle, then expel the residual air from the needle by pushing the syringe plunger slightly, and then immediately cap the needle point with a sterile cork or other solid sterile material. The syringe must be transported immediately to the laboratory. If there is no liquid content, a swab that forms part of one of the special commercially available anaerobic transport systems should be used. Directions for creating the anaerobic environment should be followed exactly. A less satisfactory procedure is to inoculate a tube of thioglycollate medium immediately after the specimen is obtained and immediately recap the tube.

  • Gram-Positive Rods

    Listeria monocytogenes

    Listeria monocytogenes is a short gram-positive rod. Although most gram-positive rods of medi- cal importance are anaerobes, Listeria is an aerobic (rather than anaerobic) organism. It is found widely in soil, sewage, and various animals, as well as in the feces of about 15% (range, 2%-60%) of asymptomatic adults. Infection in children and adults is most often from contaminated food (salads, soft cheese and blue-veined cheeses but not hard cheese or cottage cheese, or unpasteurized milk). Listeria survives at refrigerator temperatures and can contaminate refrigerators so that leftovers, “cold cuts,” or cold processed-meat products would have to be adequately heated before serving. In newborns, infection may occur from the mother’s vagina. Most cases are found in neonates, in pregnancy, the elderly, and immunocompromised persons. Infection is especially associated with renal transplants, leukemia or lymphoma, and pregnancy. The most common infection at nearly all ages is meningitis (about 55%-80% of Listeria cases). Next most common (about 25% of cases) is bacteremia (which is the type of Listeria infection found in pregnancy). Listeria occasionally has been reported to cause stillbirth and may produce septicemia in newborns (especially in premature infants). Culture of the mother’s lochia has been suggested as an aid in diagnosis, as well as blood cultures from the infant. On Gram stain the organism may be mistaken for diphtheroids, streptococci, or improperly stained H. influenzae. Listeria may cause meningitis in early infancy, usually after the first week of life. In Listeria meningitis, CSF Gram stain is said to be negative in about one half of the patients and misinterpreted in some of the others, most often being mistaken for contaminant “diphtheroids.” About one third of patients have predominance of lymphocytes or mononuclears rather than neutrophils. CSF glucose is said to be normal in 50% or more of patients. CSF culture is the most reliable method of diagnosis. DNA probe methods have recently been described.

  • Other Gram-Negative Organisms

    Pseudomonas

    These gramegative rods are not classified with the Enterobacteriaceae, although they may be found normally in the GI tract. The most important is Pseudomonas aeruginosa. Pseudomonas is less common in community-acquired infection than with most of the major Enterobacteriaceae but becomes more frequent in hospital-acquired infections (about 10% of nosocomial infections). The conditions for which it is best known are urinary tract infection, septicemia, pneumonia (especially in patients with cystic fibrosis), extensive burn infection, malignant otitis externa, and infection associated with ocular contact lenses or physical therapy whirlpool tubs. P. aeruginosa is also known for its resistance to many antibiotics and for its high mortality rate.

    In the environment, Pseudomonas organisms live in water or on moist surfaces. In human disease, Pseudomonas has many attributes similar to “opportunistic fungi” in that infection is rare in healthy persons; and even in hospitalized persons, colonization is more common than infection. Infection is most often superimposed on serious underlying diseases such as hematologic or nonhematologic malignancy (especially during chemotherapy), severe burns, wounds, and foreign bodies (tracheostomy, catheter), or in immunocompromised patients. P. aeruginosa is resistant to many of the standard antibiotics and therefore may become a secondary invader after antibiotic therapy for the original infection. This is most frequent in urinary tract infections. Pseudomonas may be found normally on the skin (as well as in the GI tract) and thus is a very important and frequent problem in severe burns. Pseudomonas septicemia is increasing as a complication or a terminal event in patients with malignancy or immunocompromised state. In some tertiary medical centers, Pseudomonas is reported involved in as many as 20% (range, 6%-25%) of bacteremic episodes.

    Calymmatobacterium granulomatis (Donovania granulomatis)

    Granuloma inguinale is a venereal disease caused by a gramegative rod bacterium that has some antigenic similarity to the Klebsiella group. Infection is transmitted by sexual contact. After incubation, an elevated irregular flattened granulomatous lesion develops, usually in or around the medial aspect of the inguinal area or on the labia. The organism is difficult to culture and requires special media, so culture is not usually done. Diagnosis is accomplished by demonstration of the organisms in the form of characteristic Donovan bodies, found in the cytoplasm of histiocytes. The best technique is to take a punch biopsy of the lesion, crush the fresh tissue between two glass slides, and make several smears with the crushed tissue. These smears are air-dried and stained with Wright’s stain. (It is possible to process a biopsy specimen in the routine manner and do special stains on tissue histologic sections, but this is not nearly as effective.) Granuloma inguinale is sometimes confused with lymphogranuloma venereum, a totally different disease, because of the similarity in names and because both are venereal diseases.

    Haemophilus (hemophilus)

    This genus is one of three in the family Pasteurellaceae (the other two are Pasteurella and Actinobacillus). The genus Haemophilus contains several species, most of which are normal nasopharyngeal inhabitants, that on Gram stain are very small gramegative rods (“coccobacilli”). The most important species is Haemophilus influenzae, which is the most common etiology of meningitis between the ages of 2 months and 5 years (about 70% of cases). H. influenzae occasionally produces a serious type of laryngitis (croup) in children known as acute epiglottitis and occasionally is associated with childhood otitis media, sinusitis, meningitis, bacteremia, and pneumonia. H. influenzae infection in adults is far less common than in early childhood, but the same types of diseases may occur. H. influenzae exists in both capsulated and noncapsulated forms. In early childhood, 90%-95% of cases of meningitis or bacteremia are caused by capsulated type B. Non-B encapsulated serotypes and nonencapsulated forms increase in frequency in localized pediatric infection and in adult infection. Currently, most typing of Haemophilus organisms is done with antibodies to the capsular antigens of H. influenzae. “Nontypable” H. influenzae means a noncapsulated strain and cannot be ignored, on this basis alone, as a cause for serious disease. Organisms in the genus Haemophilus require one or both substances called X and V factors. Therefore, the traditional H. influenzae culture plate contains blood agar (supplying the X factor) on which is a small area previously inoculated with S. aureus (“staph streak”), which supplies V factor (“satellite test”). However, other satisfactory culture and identification systems are now available. In addition to culture, latex agglutination tests are available to detect H. influenzae antigen in spinal fluid. These will be discussed in more detail in the chapter on CSF tests. In very limited studies involving patients with infections other than CSF, latex agglutination test results on urine were positive in 92%-100% of cases. Haemophilus influenzae is considered normal flora of the nasopharynx (3%-5% of children over age 6 months) and also sputum (which acquires these organisms as it passes through the oropharynx). Heavy growth or predominance of a single “normal flora” organism increases the suspicion that it may be pathogenic.

    Other members of the Haemophilus genus that deserve special mention are H. aegyptius (Koch-Weeks bacillus), which produces purulent conjunctivitis; and H. ducreyi, which is the etiologic agent of the venereal disease chancroid. Haemophilus aphrophilus is being reported more often in recent years as a pathogen.

    Pasteurella

    The most important organism in this genus is Pasteurella multocida. This is a small gramegative bacillus found normally in the mouth of many (but not all) cats and dogs. Dog bites and either cat bites or cat scratches inoculate the organism, leading to cellulitis and occasionally to osteomyelitis or infection of prosthetic devices. The organism grows on ordinary laboratory media. Isolation and identification is made much easier if the laboratory knows that the specimen is from a dog- or cat-inflicted wound.

    Bordatella

    For a long time these organisms were included in the genus Haemophilus. The most important of this group is Bordatella pertussis, the etiologic agent of pertussis (“whooping cough”). The disease is spread by aerosol droplets and is highly contagious, with attack rates in family members of patients ranging from 50% to over 90%. In one study about two thirds of those infected were asymptomatic. The incubation period averages 7 days (range, 6-20 days). In children there are three clinical stages. The catarrhal stage lasts 1-2 weeks (range, 1-3 weeks), associated with symptoms similar to a mild or moderate viral upper respiratory infection. The paroxysmal stage lasts 2-4 weeks (sometimes more), with paroxysms of coughing alternating with relatively asymptomatic periods being the main symptoms. The convalescent stage usually lasts 1-2 weeks but may persist up to 6 months. The most frequent complication is pneumonia (15% of cases in one study); this is responsible for over 90% of deaths. In adults, clinical symptoms most often consist of chronic cough. In one study, 20%-25% of adults with chronic cough had serological evidence of pertussis.

    Laboratory findings consist mostly of leukocytosis (64% of cases in one study) that averages 21,000/mm3 (21 Ч 109/L) but can reach 70,000/mm3(70 x 109/L). There typically is an absolute lymphocytosis; since young children normally have more lymphocytes than neutrophils on peripheral smear, the percentage of lymphocytes is most often within the age-adjusted reference range for young children. There typically is a significant percentage of small lymphocytes with a clefted or distorted nucleus (12%-56% of the lymphocytes). Most of these are T-lymphocytes. In one study lymphocytosis persisted for more than 1 week in 70% of cases and for more than 2 weeks in 14% of cases. In young children, one study found an abnormal chest x-ray in 42% of patients.

    Options for diagnosis include posterior nasopharyngeal culture, direct tests for antigen on nasopharyngeal swabs, and serologic tests. Material for culture or antigen tests should be obtained from the pharynx at the posterior end of the nose, using a calcium alginate or Dacron-tipped flexible wire swab. Cotton-tipped swabs decrease chances of a positive culture. A positive culture is most likely (80%-90% of cases) during the first and second weeks of the catarrhal stage; during the later part of the third week, and the beginning of the paroxysmal (fourth week) stage, culture positivity falls off rapidly. After the fifth week only 15%-20% of cases are culture-positive. The original (and still useful) culture medium was Bordet-Gengou; however, the newer Regan-Lowe media can increase culture yield by 20% or more. Reported rates of culture diagnosis vary considerably (30%-60%; range, 20%-90%), depending on the stage of disease culture was obtained and the details of culture technique. Culture takes 4-7 days. As noted previously, recovery results over 50% are most likely to be obtained in the first 3 weeks except during epidemics. Direct fluorescent antibody (DFA) tests on posterior nasopharyngeal specimens are less sensitive than culture in the early nontreated stage but may be more sensitive later in clinical illness or after antibiotic therapy. In general, DFA is 50%-90% as sensitive as culture (range, 11%-100% depending on circumstances). False positive rates of 7%-40% have been reported (although some of these may have been true but nonconfirmed positives). Serologic tests are available but difficult to obtain. The most useful clinically are IgG antibodies against pertussis toxin or filamentous hemagglutinins (FHA). Acute and convalescent specimens give best results. Sensitivity of FHA-IgG tests is reported to be 75%-80% (range, 54%-100%).

    Campylobacter

    There are several species within this genus, of which the most important is Campylobacter fetus. This is a gramegative curved or spiral organism that originally was classified as a Vibrio. There are three subspecies (ssp.) of which two may infect humans. One of these is Campylobacter fetus ssp. fetus (formerly, ssp. intestinalis), which causes abortion in sheep and cattle and which can rarely infect humans. It produces disseminated disease without diarrhea, presenting as newborn or infant meningitis and childhood or adult septicemia. It also can produce thrombophlebitis. Patients usually are immunocompromised or have some debilitating disease such as alcoholism, cancer, or renal failure. Diagnosis is made by blood culture in septicemia and by spinal fluid culture in meningitis. Ordinary culture media can be used. The other Campylobacter organism, C. fetus ssp. jejuni, is much more common and infects cattle, dogs, and birds, in addition to humans. Poultry, especially turkeys, have been responsible for some epidemics. C. fetus ssp. jejuni produces enteric infection with diarrhea, which in some areas is responsible for as many cases as Salmonella and Shigella combined. Overall, both children and adults are affected in about equal proportion, although there is variance in different reports. Published studies indicate that C. fetus ssp. jejuni can be isolated from approximately 4%-8% of patients with diarrhea (literature range, 3%-32%).

    Typical symptoms include a prodromal period of 12-24 hours with fever, headache, abdominal pain, and malaise, followed by diarrhea with crampy abdominal pain. Fever often disappears after onset of diarrhea. The diarrhea is sometimes grossly bloody, typically lasts 2-3 days, and usually is self-limited. In some patients it is more prolonged. The severity of illness varies considerably.

    Laboratory tests. Helpful laboratory studies include tests for fecal blood and Gram stain of the feces for WBCs. About two thirds of patients demonstrate gross or occult stool blood (literature range, 60%-90%), and about the same number have segmented neutrophils in the stool. About two thirds of patients have peripheral blood leukocytosis. Fecal culture is the mainstay of diagnosis (blood cultures are usually negative in ssp. jejuni infection). However, routine stool culture will be negative, because the organism is microaerophilic and must be incubated 48-72 hours on special media in special gas mixtures. It grows best at 42°C but will grow at 37°C. Routine laboratories can perform the cultures, but it takes special effort and special techniques. The organism is extremely sensitive to drying. If specimens are sent to an outside laboratory, they should be inoculated into special transport media (not ordinary media) and sent with ice if more than 1 day’s travel is necessary. Gram stain of the feces can demonstrate Campylobacter in about 50% of cases (range, 43%-65%). The organism is gram negative and has different shapes; most typically curved, S-shaped, and “gull winged.” A few reports indicate that 1% aqueous basic fuchsin stain is more sensitive than Gram stain.

    Aeromonas

    The Aeromonas organism, a gramegative rod, is frequently found in water (usually nonchlorinated although chlorinated water is not always safe) and in soil. It infects fish and amphibians as well as humans. Sources of human infection are water; uncooked or poorly cooked meat, poultry, shellfish, and fish; and raw milk. Clinical infection in humans is not common but has been reported in those with traveler’s diarrhea, immunosuppressed patients, and sporadic cases. Asymptomatic infection has been reported in 0.1%-27% of persons examined. Diarrhea and abdominal cramps are the most common symptoms. Persons of all ages can become infected; acute self-limited diarrhea is more common in infants and young children and chronic infection in adults. Occasional extraintestinal disease (osteomyelitis, urinary tract infection, septicemia, and others) has been reported. Diagnosis is most often made by stool culture; enrichment media followed by special selective media is said to give best overall results, although the organisms can grow on standard culture media.

    Helicobacter

    Helicobacter pylori (formerly Campylobacter pylori or pyloridis) is an S-shaped (curved, or “gull-winged” like Campylobacter), small gramegative bacillus found in the mucus covering the gastric mucosa, with the organisms located next to the gastric lining cells of the surface and gastric pits. H. pylori is associated with acute and chronic gastritis in the gastric antrum and is present in about 90% (range, 70%-100%) of patients with duodenal ulcer, in 70%-75% (range, 40%-90%) of patients with gastric ulcer, about 50% (range, 30%-75%) of patients with nonulcer dyspepsia, and about 20%-25% of patients with gastric cancer. Significant H. pylori antibody levels can be detected in about 20%-25% of clinically normal U.S. and European adults (range, 5%-75% depending on age). This suggests that H. pylori infection is often subclinical. Indeed, in patients with clinical infection, biopsy of normal-appearing gastric mucosal areas often contains demonstrable H. pylori organisms. Chronic or recurrent duodenal ulcer is very highly associated with both excess acid production and H. pylori infection. However, one exception is duodenal ulcer due to the Zollinger-Ellison syndrome, in which incidence of H. pylori infection is zero.

    Incidence of H. pylori is age-related; in one study 5% or less of adults age 25-45 had antibodies to H. pylori; 20% of adults age 45-55, 50% of adults 55-65, and 75% of adults over age 65 had the antibodies. African Americans are reported to have about twice the incidence seen in Europeans. Besides antral gastritis, H. pylori is associated to a lesser degree with patchy gastritis in the body of the stomach.

    The traditional gold standard for diagnosis has been culture of gastric mucosal biopsy specimens. This should be placed in 2-3 ml of sterile isotonic saline, plated on enriched media such as chocolate agar or selective media such as Skirrow’s, and incubated 5-7 days at 37°C. However, based on comparison with results of other tests, culture only detects about 75% (range, 50%-95%) of cases. Giemsa stain of gastric mucosa biopsy tissue is reported to detect about 90% of cases (range, 88%-93%), although in my experience it has been less. Warthin-Starry silver stain has a little better sensitivity than Giemsa stain and Gram stain has somewhat less (about 75%; range, 69%-86%). H. pylori has considerable ability to metabolize urea by means of the enzyme urease that forms the basis for several other tests. The urea breath test using radioactive carbon-13 or carbon-14 incorporated into a urea test dose is administered orally and expired air is then tested for radioactivity 20-120 minutes later. This is probably the best test of all those available, with sensitivity in most reports over 95% (range, 90%-100%) and specificity over 95%. In fact, at present it is probably a better gold standard than culture. However, the test is available only in a relatively few large medical centers, is expensive, requires special equipment, and uses long-lived isotopes that are becoming serious disposal problems. Other tests are based on fresh gastric mucosal biopsy tissue placed in a urea-containing medium with some indicator system to signal metabolism of the urea. The best-studied of these is the CLO test; it is reported to have a sensitivity of about 90% (range, 86%-96%).

    Some immunoassays for antibody to H. pylori have become commercially available, mostly in EIA format, which detects IgG, IgA, or IgM anti- body alone or total antibody. In experimental H. pylori infection, an IgM elevation first begins at about 3 weeks, becoming nondetectable about 8 weeks later. IgA and IgG levels rise at about 8-9 weeks after infection, with the IgG level remaining elevated a year or more. The height of elevation does not correlate well with severity of infection. IgM and IgA levels often fall in about 4 weeks after successful treatment, whereas IgG levels are not affected. Most of the antibody assays evaluated in the literature to date have been homemade (which often give better results than commercially available kits); these have reported sensitivity of 80%-100%. Several new commercial kits claim sensitivity of 95% or more. However, some of these sensitivity claims are less because the tests against which the commercial EIA was compared had sensitivity themselves less than 100%. Also, more evaluations must be done to get adequate information about each EIA kit. In general, most current interest seems to be toward IgG antibody. However, this antibody has the disadvantage that it often remains elevated for a year or more after adequate therapy.

    Brucella

    Three members of the Brucella genus produce an uncommon febrile disease known as “brucellosis,” which sometimes must be considered in the differential diagnosis of fever of unknown origin. The Brucella organism is a gramegative coccobacillus with three main species. One species infects cattle, the second one infects goats, and the third is found in swine. Classic brucellosis was most often transmitted to humans by infected milk or milk products. However, persons most likely to contact the infection today in the United States are workers in the meat-processing industry, especially those working with swine. Veterinarians and dairy farmers are also at risk. Clinical symptoms include fever, chills, and myalgia. About 25% of patients develop single-joint arthralgia. Lymph nodes are usually not enlarged. Splenomegaly occurs in about 20% of cases. Some patients develop pneumonia. WBC counts are usually normal or decreased. Blood culture is positive in 30%-50% of cases, but the organisms need added carbon dioxide and grow very slowly on ordinary culture media, so that under ordinary circumstances the brucellae may either not grow at all or the culture bottle is discarded as negative before growth is evident. Special media are available that will assist growth. A slide agglutination test is the most frequent method for diagnosis. Test results greater than 1:80 are suspicious for brucellosis. After recovery, antibody titer elevation lasts 1-2 years.

    Francisella

    Formerly included in the genus Pasteurella, the genus Francisella contains one medically important species, Francisella tularensis, which is the causative agent of tularemia. F. tularensis is a very small gramegative aerobic coccobacillus that requires special media for adequate growth. The major reservoir for human infection in the United States is wild animals, predominantly wild rabbits, and in some cases deerflies or ticks. Most persons who contract the disease are those who handle raw wild rabbit meat. Tularemia may exist in several clinical forms, but the most common symptoms are painful single-area lymphadenopathy (ulceroglandular form, comprising 75%-80% of cases) and prolonged low-grade fever. Much less common is a septicemic form (5%-15% of cases). Pneumonia may occur in 15% or more of either the ulceroglandular or the septicemic forms, especially the septicemic type. Rabbit-associated infection is usually associated with a small ulcerated skin lesion at the place of entry on the hand and also with axillary adenopathy, whereas in tick-associated infection the ulcer is located on the trunk or lower extremities and the lymph nodes affected are most commonly in the inguinal area. About 15% of affected persons lack either the ulcer or both the ulcer and the adenopathy. WBC counts are normal in at least 50% of patients and slightly or mildly elevated in the remainder. The differential cell count is usually normal. Inconsistent elevation of one or more liver function test results is present in about one half of the patients.

    Tularemia must occasionally be considered in the differential diagnosis of fever of unknown origin. The organisms will not grow on standard laboratory culture media. Special media can be obtained and culture performed from lymph node aspiration, but this is rarely done because of an unusually high rate of laboratory technologist infection from the specimens. The standard diagnostic test is a slide agglutination procedure. Titers are said to be negative during the first week of illness and begin to rise at some time during the second week, become maximal in 4-8 weeks, and remain elevated for several years. A fourfold rising titer is necessary to prove acute infection. There is considerable disagreement regarding a single titer level that could be considered presumptive evidence of infection; 1:160 seems to be the least controversial value.

    Vibrios

    Vibrios are gramegative small curved rods. The most important species is Vibrio cholerae, which produces cholera by means of an enterotoxin. The V. cholerae species is subdivided into many serotypes, of which only one, the 01 strain, is associated with epidemic cholera in the United States. A different strain is found in Latin America than in the United States. About 75% of cholera organism infections are asymptomatic, about 20% produce mild to moderate diarrhea, and about 5% produce the severe watery diarrhea ordinarily associated with the name “cholera” that can lead to death from loss of fluid and electrolytes. Humans usually become infected by drinking contaminated water or eating shellfish from contaminated water. Diagnosis has traditionally been made by stool culture. The organism is very sensitive to drying, and if swabs are used, they must be placed in transport media as soon as the specimen is obtained. Also culture has been reported to miss 10%-20% of cases.

    Recently, two stool tests have been developed by the same manufacturer using a monoclonal antibody called COLTA that is claimed to be specific for V. Cholerae 01 strain. CholeraScreen is a coagglutination slide test and CholeraSmart is a tube agglutination test. Both are said to be 95%-100% sensitive compared to culture. Only a few evaluations have been published to date.

  • Enteric Bacilli (Enterogacteriacae)

    Enteric bacilli form a large family of gramegative rods (see Table 37-6). As their name implies, most are found primarily in the intestinal tract. These include species such as Salmonella, Shigella, Escherichia coli, Enterobacter, Klebsiella, Proteus, and several others (see Table 37-6). Many are normal inhabitants and cause disease only if they escape to other locations or if certain pathogenic types overgrow; others are introduced from contaminated food or water. Salmonellae and shigellae are not normal gastrointestinal inhabitants and always indicate a source of infection from the environment.

    Salmonella

    These organisms cause several clinical syndromes. Typhoid fever is produced by Salmonella typhi (Salmonella typhosa). The classic symptoms are a rising fever during the first week, a plateau at 103°F-104°F (39.4°C-40.0°C) for the second week, then a slow fall during the third week, plus GI symptoms and splenomegaly. Despite fever, the pulse rate tends to be slow (bradycardia). This picture is often not present in its entirety. Diarrhea occurs in 30%-60% of patients. There typically is mild leukopenia with lymphocytosis and monocytosis. However, in one series only 10% of patients had leukopenia and 7% had a WBC count more than 15,000/mm3.

    Laboratory diagnosis. During the first and second weeks of illness, blood cultures are the best means of diagnosis; thereafter, the incidence of positive specimens declines rapidly. During the latter part of the second week to the early part of the fourth week, stool cultures are the most valuable source of diagnosis. However, stool cultures may occasionally be positive in the first week; in carriers, positive stool cultures may persist for long periods. (About 3%-5% of typhoid patients become carriers—persons with chronic subclinical infection.) Urine cultures may be done during the third and fourth weeks but are not very effective. Even with maximum yield, blood cultures miss at least 20% of cases, stool cultures miss at least 25%, and urine cultures miss at least 75%. Repeated cultures increase the chance of diagnosis. Besides cultures, serologic tests may be performed. There are three major antigens in S. typhi: the H (flagellar), O (somatic), and Vi (capsule or envelope) antigens. Antibody titers against these antigens constitute the Widal test. Most authorities agree that of the three antibodies, only that against the O antigen is meaningful for diagnosis. Vaccination causes a marked increase in the anti-H antibodies; the level of anti-O antibodies rises to a lesser degree and returns much more quickly to normal. The Widal test (anti-O) antibodies begin to appear 7-10 days after onset of illness. The highest percentage of positive test results is reported to be in the third and fourth weeks. As with any serologic test, a fourfold (change of at least two dilution levels) rising titer is more significant than a single determination. There has been considerable controversy over the usefulness of the Widal test in the diagnosis of Salmonella infections. It seems to have definite but limited usefulness. Drawbacks to the Widal test include the following: (1) antibodies do not develop early in the illness and may be suppressed by antibiotic therapy; (2) antibody behavioris often variable and often does not correlate with the severity of the clinical picture; (3) an appreciable number of cases (15% or more) do not have a significantly elevated anti-O titer, especially if only one determination is done. Only about 50% (one study obtained only 22%) display a fourfold rise in titer. In some cases, therapy may suppress the response. A normal Widal titer is 0-1:40.

    To summarize, in typhoid fever, blood cultures during the first and second weeks and stool cultures during the second, third, and fourth weeks are the diagnostic tests of choice. The Widal test may occasionally be helpful. Negative diagnostic test results do not exclude the diagnosis.

    Paratyphoid fever (enteric fever) is produced by salmonellae other than S. typhi; the clinical picture is similar to typhoid fever but milder. Salmonella typhimurium and Salmonella enteritidis (formerly Salmonella paratyphi) are usually the most common causes in the United States. Diagnosis is similar to that for typhoid fever.

    In the United States, Salmonella gastroenteritis is more frequent than typhoid fever or enteric fever. The gastroenteritis syndrome has a short incubation, features abdominal pain, nausea, and diarrhea, and is most commonly produced by S. typhimurium. There is usually leukocytosis with a minimal increase in neutrophils, in contrast to the leukopenia in textbook cases of typhoid fever. Blood cultures are said to be negative; stool cultures are usually positive.

    Other salmonella diseases. Septicemia may occasionally be found, and salmonellae may rarely cause focal infection in various organs, resulting in pneumonia, meningitis, and endocarditis. Salmonella osteomyelitis has been associated with sickle cell anemia. Salmonella bacteremia is relatively frequent in patients with systemic lupus erythematosis. Salmonella infections, including bacteremia, are also more frequent in the acquired immunodeficiency syndrome (AIDS) and in patients with leukemia or lymphoma.

    Sources of salmonella infection. Salmonella typhi is found only in humans, and infection is transmitted through fecal contamination of food and water. Other salmonellae infect poultry and animals. Nontyphoid salmonellosis is most commonly acquired from poultry and eggs, which in turn are most frequently associated with S. enteritidis infection. It is necessary to cook an egg thoroughly to avoid S. enteritidis infection, even when the eggshell has no evidence of being cracked. However, Centers for Disease Control (CDC) reports that one third of egg-related S. enteritidis infections took place when eggs had been cooked at recommended time and temperature. Only when cooking causes all of the egg yolk to become solid will the egg become safe. Pasteurized eggs are another option. Contaminated meat and nonpasteurized or powdered milk have been an occasional problem. Reptiles are reported to carry Salmonella species in 36%-84% of those cultured.

    Due to DNA hybridization and other research work, the genus Salmonella is being reorganized to include certain organisms, such as Arizona, that previously were not considered salmonellae. The organisms previously considered Salmonella species are now serotypes (serovars) of the subspecies choleraesuis from the species Salmonella choleraesuis. The other organisms, such as Arizona, form part of other subspecies of S. choleraesuis.

    Shigella

    The shigellae are also important sources of intestinal infection as well as the salmonellae. Shigella organisms cause so-called bacillary dysentery. Shigellae usually remain localized to the colon and do not enter the peripheral blood; blood cultures are therefore negative, in contrast to blood cultures in early Salmonella infection. Stool culture is the main diagnostic test. Besides salmonellae and shigellae, certain other bacteria (e.g., Yersinia enterocolitica, Campylobacter jejuni, Clostridium difficile, and enteropathogenic E. coli) may cause diarrhea from GI infection; these are discussed elsewhere.

    Enterobacter and Klebsiella

    These bacteria are normal GI tract inhabitants. Nomenclature has been particularly confusing in relation to these organisms. Enterobacter was formerly called Aerobacter. Enterobacter and Klebsiella react differently to certain test media but are similar enough that previous classifications included both in the same group. According to previous custom, if infection by these organisms was in the lungs it was called Klebsiella; if it was in the urinary tract, it was called Aerobacter. Current classification does not make this distinction. The species of Klebsiella called Klebsiella pneumoniae is the organism that produces so-called Friedlьnder’s pneumonia, a resistant necrotizing pneumonia that often cavitates and that is characteristically found in alcoholics and debilitated patients. Klebsiella pneumoniae also is an important cause (about 10%) of both community-acquired and hospital-acquired urinary tract infection. The species of Enterobacter called Enterobacter aerogenes is an important agent (about 5%) in nosocomial urinary tract infections, is often resistant to therapy, and occasionally produces septicemia.

    Present classification differentiates Enterobacter from Klebsiella. Sources of confusion include the family name Enterobacteriaceae, which is similar to the name of one component genus, Enterobacter. Enterobacteriaceae is a group of several tribes, each of which contains a genus or genera. One tribe, Klebsiellae, has a similar name to one of its three component genera, Klebsiella, and also includes the genus Enterobacter. A further source of difficulty is that the predominant species of Klebsiella is K. pneumoniae, which, despite its name, is found more frequently in the urinary tract than in the lungs.

    Escherichia

    Escherichia coli is the most common cause of community-acquired (about 70%) and of hospital-acquired urinary tract infection (about 50%). E. coli is likewise the most common cause of gramegative bacteremia or septicemia, both of community-acquired or hospital-acquired origin. The primary site of infection leading to bloodstream entry is most often the urinary tract, as happens with the majority of the gramegative rod bacteria. This is more frequent with urinary tract obstruction. E. coli is one of the most common causes of severe infection in the newborn, especially meningitis. E. coli causes periodic outbreaks of epidemic diarrhea in newborns and infants. E. coli has been incriminated as an important cause of traveler’s diarrhea, which may affect U.S. tourists in other countries. E. coli is a normal inhabitant of the colon, so a stool culture growing E. coli does not prove that the organism is the causative agent of diarrhea.

    In the past it was thought that only certain strains of E. coli were responsible for diarrhea, and that serotyping (using antisera against the so-called enteropathogenic strains) was helpful in deciding whether or not E. coli was responsible for diarrhea by demonstrating or excluding the presence of the enteropathogenic subspecies (see Table 37-9). However, so many exceptions have been noted that serotyping is no longer advocated except as an epidemiologic tool to show that patients in an epidemic are infected by the same E. coli strain. Nevertheless, there is a condition caused by a specific strain of E. coli (0157:H7) called hemorrhagic colitis, manifested by severe diarrhea with abdominal pain and bloody stools. In the United States this organism predominately exists by colonizing cattle (10%-20% prevalence rate). Diarrhea in humans is caused by production of a verotoxin. Although bloody diarrhea is the organism’s trademark, about 30% of cases are said to have diarrhea without blood. Stools usually do not contain many WBCs. About 10%-30% of patients need hospitalization and 2%-7% develop the hemolytic-uremic syndrome (hemolytic anemia with red blood cell [RBC] fragmentation, thrombocytopenia, and renal failure). Diagnosis can be made by using selective media for E. coli 0157-H7 strain such as sorbitol-MacConkey agar (SMA), by fluorescent antibody stains applied to stool smears, or by verotoxin assay. In one study, verotoxin assay was 20% more sensitive than culture.

    Proteus

    This is a group of gramegative rods that assumes most importance as the cause of urinary tract infection in hospitalized patients (about 5%) but occasionally causes community-acquired cystitis and some cases of hospital-acquired bacteremia and other infections.

    Yersinia

    The genus Yersinia, which includes several organisms formerly located in the genus Pasteurella, contains three important species: Yersinia pestis, Yersinia pseudotuberculosis, and Yersinia enterocolitica. Yersiniapestis is the cause of plague, which was transmitted historically by rat fleas, followed in some cases by nasopharyngeal droplet dissemination from infected humans. Today, plague is not a serious menace in Western nations, although a few cases in rodents (including prairie dogs) are reported each year from the American West and Southwest, from which source the disease is occasionally transmitted to humans. There are three clinical forms: septicemic, bubonic, and pneumonic. The septicemic type is responsible for 5%-10% of cases and is associated with a bacteremic form of febrile illness that progresses to septicemia and shock. Diagnosis is made by blood culture. When lymph nodes are primarily involved, the condition is known as bubonic plague. This constitutes the great majority of plague cases. However, septicemia or pneumonia may develop. Only one group of lymph nodes is enlarged in the majority of cases.

    Diagnosis is made by blood culture (positive in about 80% of cases) or lymph node aspiration and culture (positive in about 80% of cases). Pneumonic plague has been uncommon in recent years and is rapidly fatal. Blood culture and sputum culture provide the diagnosis. Y. pestis grows on ordinary laboratory culture media.

    Yersinia pseudotuberculosis is found in many wild and domestic animals and in various domestic fowl. Human infection is uncommon, or at least, is uncommonly recognized, and most often consists of mesenteric adenitis. There usually is abdominal pain, fever, and leukocytosis, a clinical picture that simulates acute appendicitis. Diagnosis is made by culture of affected lymph nodes (lymph nodes are likely to be placed in chemical fixative for microscopic examination, which makes culture impossible unless the surgeon orders a culture to be performed and directs personnel not to fix the specimen).

    Yersinia enterocolitica is by far the most common pathogen of these three Yersinia species. Y. enterocolitica most often produces acute enteritis, with clinical features including headache, fever, malaise, crampy abdominal pain, and nonbloody diarrhea. The organism also can produce mesenteric adenitis and, rarely, intraabdominal abscess or septicemia. Enteritis is more common in children and mesenteric adenitis in adolescents and adults. Diagnosis is made by stool culture in patients with enteritis and by lymph node culture in those with mesenteric adenitis. Culture is less reliable for Y. enterocolitica than for the other yersiniae, because the organism grows somewhat slowly and forms tiny pinpoint colonies on ordinary media used for gramegative rods, which can easily be overlooked among normal stool organism colonies. The organism grows much better at room temperature (25°C) than at the usual incubation temperature of 37°C. Some investigators believe that “cold enrichment” is necessary (special media incubated at 4°C for several weeks). Therefore Y. enterocolitica will usually be missed on routine culture unless the laboratory takes special precautions.

    “Opportunistic” enterobacteriaceae

    There are a considerable number of species located in various genera of the Enterobacteriaceae that are less common pathogens. It was originally thought that these organisms were nonpathogenic or rarely pathogenic and that their importance was only in the necessity for differentiation from more pathogenic enteric organisms such as Salmonella. Now, however, it is recognized that these organisms produce disease, most often urinary tract infections but also septicemia and pulmonary infections, although the frequency of infection for any individual species is not high. Most of these organisms have been renamed and reclassified since initial studies were done. Some of the more important organisms include Arizona, Providentia (formerly Proteus inconstans and Proteus rettgeri), Citrobacter (formerly Bethesda group and Escherichia freundii), and Serratia. Infections caused by these organisms are often associated with decreased host resistance and with long-term bladder catheterization. Serratia is usually considered the most dangerous of these organisms. About 90% of Serratia infections involve the urinary tract, the great majority following urinary tract surgery or cystoscopy or associated with indwelling catheters. As many as 50% of Serratia urinary tract infections are asymptomatic. Bacteremia develops in about 10%. A few patients develop Serratia pneumonia, although respiratory tract colonization is much more frequent than pneumonia. The most common (but not the only) means of Serratia spread from patient to patient is the hands of hospital personnel.

  • Other Venereal Diseases

    Other venereal diseases include lymphogranuloma venereum (caused by a subspecies of C. trachomatis different from the one that causes nongonococcal urethritis), syphilis, granuloma inguinale, trichomoniasis, chancroid, herpesvirus type 2, molluscum contagiosum, and condyloma acuminatum. Most of these will be discussed elsewhere.

    Gardnerella vaginalis

    This organism (also called Corynebacterium vaginalis and Haemophilus vaginalis) is a small bacillus or coccobacillus that gives variable results on Gram stain, most often gram negative, but sometimes gram positive. The organism has been implicated as the cause of most cases of “nonspecific vaginitis” (i.e., vaginitis not due to Candida albicans or Trichomonas). Clinical infection in women occurs only when estrogen levels are normal or increased, so postmenopausal women usually are not involved. Gardnerella vaginalis can also be cultured from the urethra of many male sexual partners of infected women, so the disease is postulated to be at least potentially sexually transmissible.

    Clinically, there is a vaginal discharge that typically is malodorous (although this odor was present in only two thirds of cases in one report). Other than the discharge, there usually are few or no symptoms in the female and none in the male.

    Diagnosis is usually made either from aspiration of vaginal discharge or from swab specimens of the vaginal wall (avoiding the cervix). The vaginal discharge specimens have a pH greater than 4.5, with 90% between 5.0 and 5.5 (normal pH is <4.5 except during menstruation, when the presence of blood raises the pH to 5.0 or more). Trichomonas parasitic infection may also produce a vaginal discharge with a pH greater than 4.5, whereas the pH in Candida infection usually is less than 4.5. Therefore, a discharge with a pH less than 4.5 is strong evidence against Trichomonas or Gardnerella infection, whereas a discharge with a pH greater than 5.0 suggests infection by these organisms. Addition of 10% sodium hydroxide to the specimen from G. vaginalis infection typically results in a fishy odor. A wet mount or Gram stain of discharge or swab material from the vaginal wall (avoiding the cervix) demonstrates clue cells in about 90% of G. vaginalis infections (literature ranges in the few studies available are 76%-98% for wet mount and 82%-100% for Gram stain). Clue cells are squamous epithelial cells with a granular appearance caused by adherence of many tiny G. vaginalis gramegative bacteria. Wet mount may sometimes be difficult to interpret due to degeneration of the squamous epithelial cells or because of only partial coverage of the cell by the Gardnerella organisms. Wet mount may occasionally produce a false positive result, and Gram stain is usually easier to interpret and generally more accurate; but one investigator found it to be somewhat more liable to false positive errors by mistaking diphtheroids for G. vaginalis. Papanicolaou cytology stain can also be used. The organism can be cultured on special media or on the same Thayer-Martin medium with increased carbon dioxide that is used for the diagnosis of gonorrhea. However, isolation of Gardnerella is not diagnostic of vaginitis, since Gardnerella can be cultured from the vagina in 42%-50% of clinically normal women. In addition, there is substantial evidence that anaerobic bacteria are associated with symptomatic infection by G. vaginalis.

    Several studies suggest that Gardnerella is responsible for urinary tract infection in some pregnant patients and some patients with chronic renal disease. Since Gardnerella does not grow on culture media routinely used for urine, urine culture in these patients would be negative. However, Gardnerella has been isolated with equal frequency in similar patients without clinical evidence of urinary tract infection.

  • Gram-Negative Diplococci

    Gramegative diplococci include several Neisseria species, such as meningococci and gonococci; and Branhamella catarrhalis. Branhamella catarrhalis (formerly Neisseria catarrhalis) is a member of the genus Moraxella but closely resembles neisseriae in many respects, including microscopic appearance. B. catarrhalis, as well as some of the neisseriae, is normally found in the upper respiratory tract. Although usually nonpathogenic, it occasionally can produce pneumonia (especially in patients with chronic lung disease) and very uncommonly has caused bacteremia.

    Meningococci

    Meningococcal infection is still the most common type of meningitis, although the incidence varies with age (Chapter 19). Meningococci are present in the nasopharynx of approximately 5%-15% (range, 0.9%-40%) of clinically normal persons, and it takes special subculturing to differentiate these from nonpathogenic Neisseria species that are normal common inhabitants of the nasopharynx. In rare instances one of the (usually) nonpathogenic Neisseria species may be associated with serious disease, such as septicemia or meningitis.

    Gonococci

    Neisseria gonorrhoeae causes gonorrheal urethritis and initiates the majority of cases of acute salpingitis, so-called pelvic inflammatory disease (PID). Gonorrhea is not symptomatic in about 75%-80% (range, 60%-83%) of infected females and in about 10%-15% (range, 5%-42%) of infected males. Currently, about 2% of gonococcal strains are resistant to penicillin (range, 0%-6%, depending on the location in the United States; rates of 10%-30% have been reported in parts of the Far East and Africa). A presumptive diagnosis of gonorrhea can often be made by a Gram-stained smear of discharge from the male urethra. Gonococci appear as bean-shaped gramegative intracellular diplococci, located within the cytoplasm of polymorphonuclear neutrophils. Extracellular organisms are not considered reliable for diagnosis. Acinetobacter calcoaceticus (formerly Mima polymorpha) and Moravella osloensis look somewhat like gonococci in urethral gram stain; these bacteria are usually extracellular but can be intracellular. S. aureus sometimes may simulate Diplococcus, although it is rounder than typical gonococci organisms and is gram positive rather than gram negative. Finally, rare urethral infections by Neisseria meningitidis have been reported. Nevertheless, consensus is that diagnosis of gonorrhea can be made from a male urethral smear with reasonable confidence if there are definite gramegative diplococci having characteristic Neisseria morphology within neutrophils. In males with a urethral exudate, a smear is usually all that is required. The patient can wait while the smear is processed, and if the smear is negative or equivocal, culture can be done. In females, an endocervical Gram-stained smear is positive in only 50% of gonorrhea cases (literature range, 20%-70%). Cultures should therefore be done, both to supplement the smear as a screening technique and to provide definitive diagnosis. In females, the endocervical canal is the single best site for culture, detecting about 82%-92% of cases that would be uncovered by culture from multiple sites. About 30%-50% of patients have positive rectal cultures (swab cultures using an anoscope, taking care to avoid contamination with feces). Rectal culture adds an additional 5%-10% of cases to endocervical culture. Cultures repeated on specimens obtained 1 week later will uncover an additional 5%-10% of cases. In males, a rectal culture is positive much less frequently except in male homosexuals, with one study finding approximately 30% of cases positive only by rectal culture. Pharyngeal gonorrhea is usually asymptomatic, self-limited (10-12 weeks), and visible in less than 50% of cases. Gonococcal penicillin resistance occurs in 2% (0%-6%) of cases; some are resistant to certain other antibiotics.

    Certain technical points deserve mention. Gonococci should be cultured on special media, such as Thayer-Martin or NYC, rather than on the traditional less selective media like chocolate agar, and need a high carbon dioxide atmosphere for adequate growth. Speculum instruments should not be lubricated with material that could inhibit gonococcal growth (which includes most commercial lubricants). Specimens should be inoculated immediately into special transport media. There are several commercial transport systems available specifically for gonococci, most of them based on modified Thayer-Martin medium and incorporating some type of carbon dioxide source. Some authors state that the medium on which the specimen is to be inoculated should be warmed at least to room temperature before use, since gonococci may not grow if the medium is cold. However, others obtained similar results with cold or room temperature media. To make matters more confusing, some investigators report that vancomycin used in gonococcal selective media to prevent bacterial contaminant overgrowth may actually inhibit gonococcal growth if the gonococcal strain is sensitive to vancomycin (about 5%-10% of strains, range, 4%-30%). Due to the 24- to 48-hour delay in obtaining culture results and lack of adequate Gram stain sensitivity in female infection, additional rapid tests to detect gonococcal infection have been introduced. One of these is Gonozyme, an immunologic enzyme-linked immunosorbent assay (ELISA) procedure that takes 3 hours to perform. Sensitivity (compared to culture) in males is reported to be about 97% (range, 93%-100%, similar to Gram stain results) and about 93% (range, 74%-100%) in females. Disadvantages are that Gram stain is faster and less expensive in males, the test cannot be used for rectal or pharyngeal specimens due to cross-reaction with other Neisseria species, there is an inability to determine if a positive result is a penicillin-resistant infection, and there are a significant number of false negative results in females. At present, the test is not widely used. Gonorrhea precedes the majority of cases of acute and chronic salpingitis (PID). However, gonococci can be isolated from endocervical culture in only about 40%-50% of patients with proved PID (literature range, 5%-80%). Chlamydia trachomatis apparently is also an important initiator of PID— or at least associated with it in some way—with acute infection with Chlamydia being present in a substantial minority of patients, either alone or concurrently with gonorrhea (combined infection of the endocervix is reported in 25%-50% of cases). The organisms most frequently recovered from infected fallopian tubes or from tuboovarian abscesses include gonococci, group D streptococci, anaerobes such as Bacteroides or anaerobic streptococci, and gramegative rods. Infection is often polymicrobial.

    Nongonococcal urethritis and the acute urethral syndrome. In men, nongonococcal urethritis reportedly constitutes about 40% of urethritis cases, and some believe that it is more frequent than gonorrhea. The most common symptom is a urethral discharge. Chlamydia trachomatis is the most commonly reported organism, identified in 30%-50% of male nongonococcal urethritis patients. Chlamydia has also been found in some female patients with nongonococcal cervicitis, and Chlamydia may coexist with gonococci in other patients. Ureaplasma urealyticum (formerly called T mycoplasma) is frequently cultured in male nongonococcal urethritis, but its relationship to disease is not proved. Reiter’s syndrome might also be mentioned as an occasional cause of nongonococcal urethritis. Chlamydia organisms have been found in some cases of Reiter’s syndrome. In females, nongonococcal urethritis is usually called the acute urethral syndrome. Symptoms are most commonly acute dysuria and frequency, similar to those of acute cystitis. In females with these symptoms, about 50% have acute cystitis, and about 25%-30% (range, 15%-40%) are due to the acute urethral syndrome. Some of the remainder are due to vaginitis. Differentiation between acute cystitis and the acute urethral syndrome is made through urine culture. A urine culture with no growth or quantitative growth less than 100,000/mm3 suggests acute urethral syndrome. Diagnosis of urethral infection usually requires the following steps:

    1. Symptoms of urethritis (urethral discharge or dysuria).
    2. Objective evidence of urethritis. In men, a urethral Gram-stained smear demonstrating more than four segmented neutrophils per oil immersion field or (alternatively) 10 or more leukocytes per high-power field (some require 15 rather than 10) in the centrifuged sediment from the first portion (first 10-30 ml collected separately) of a clean-catch (midstream) voided urine specimen is required. In women, a urine culture without growth or growing an organism but quantitatively less than 100,000 (105) organisms/ml is required.
    3. Exclusion of gonococcal etiology (urethral smear or culture in men, urethral culture in women). Culture or direct identification (e.g., fluorescent antibody or nucleic acid probe methods) would be needed to detect Chlamydia or Mycoplasma organisms.

  • Gram-Positive Cocci

    Streptococci

    Streptococci are gram-positive cocci that, on Gram stain, typically occur in chains. Streptococci are subclassified in several ways. The three most useful classifications are by bacterial oxygen requirements, by colony appearance on blood agar, and by specific carbohydrate from the organism. Depending on clinical oxygen environment associated with disease, streptococci may be considered aerobic, microaerophilic, or anaerobic. Most streptococci are aerobic. The microaerophilic organisms sometimes cause a chronic resistant type of skin ulcer and occasionally are isolated in deep wound infections. The anaerobic streptococci are discussed later.

    Streptococci are divided into three types by the appearance of the hemolysis that the streptococcal colonies produce on sheep blood agar—alpha, beta, and gamma. Alpha (hemolytic) streptococci are characterized by incomplete hemolysis surrounding the colony; this area usually has a greenish appearance on sheep blood agar, and streptococci producing green hemolysis are often called “viridans.” The beta hemolytic organisms produce a complete, clear (colorless) hemolysis. Gamma streptococci do not produce hemolysis on sheep blood agar. These differences have clinical value. Alpha streptococci of the viridans subgroup are one of the most frequent causes of subacute bacterial endocarditis. Beta streptococci are the causative agent of several different types of infection and syndromes, as will be discussed later, and account for the great majority of disease associated with streptococci with the exception of subacute bacterial endocarditis. Gamma streptococci are of lesser importance, but some of the organisms known as enterococci belong to this category.

    The third classification is that of Lancefield, who discovered antibodies produced to a somatic carbohydrate of streptococcal organisms. Streptococci can be divided into groups according to the particular carbohydrate they possess on the basis of organism reaction to the different Lancefield antibodies (antisera). These groups are given a capital letter name ranging from A to G (in some classifications, even letters further in the alphabet but excluding E). Lancefield grouping does not depend on the presence of hemolysis or the type of hemolysis; for example, streptococci of group A are all beta hemolytic but streptococci of group D can be either alpha or beta hemolytic or even gamma nonhemolytic. Lancefield grouping cannot be done on some streptococci that are not beta hemolytic with the exception of those from groups B and D. Lancefield group A organisms, as mentioned earlier, are always beta hemolytic, and colonies are definitively identified with group-specific antiserum. This requires culture of a specimen, isolation of several colonies of a beta hemolytic organism resembling Streptococcus by colony appearance or Gram stain, and testing a colony with the Lancefield antisera. However, Lancefield antisera are relatively expensive, and since group A organisms seem to have an unusually marked susceptibility to the antibiotic bacitracin, Lancefield grouping is very frequently replaced by a less expensive identification method based on demonstrating inhibition of growth around a disk impregnated with a standardized concentration of bacitracin in an agar culture containing a pure growth of the organism (the organism is first identified as a Streptococcus by Gram stain or biochemical tests). However, this method is only presumptive rather than definitive, since about 5% (range, 4%-10%) of hemolytic group A streptococci are not inhibited by bacitracin (these organisms would be incorrectly excluded from group A), whereas 8%-22% of beta hemolytic streptococci from groups other than A (e.g., B, C, D, and G) have been reported to be sensitive to bacitracin (and therefore would be incorrectly assigned to group A.) Also, the bacitracin method takes 2 and sometimes even 3 days—1 day to culture the organism, 1 day to perform the disk susceptibility test, and sometimes another day to separate the organism colony from other bacterial colonies before the bacitracin test if the original culture grows several different organisms.

    Group A organisms may be further separated into subgroups (strains) by use of special antisera against surface antigens (M antigens). Strain typing is mostly useful in epidemiologic work, such as investigating outbreaks of acute glomerulonephritis, and is not helpful in most clinical situations. Group A streptococci produce certain enzymes, such as streptolysin-O, which can be detected by serologic tests (antistreptolysin-O titer, Chapter 23). Antibodies against these enzymes do not appear until 7-10 days after onset of infection, so they are usually not helpful in diagnosing acute streptococcal infection. Their major usefulness is in the diagnosis of acute rheumatic fever and acute glomerulonephritis.

    Rapid immunologic identification tests from many manufacturers have now become commercially available for group A streptococci that directly test for the organisms in swab specimens without culture. Usually the organism is extracted from the swab chemically or enzymatically and then tested with antiserum against group A antigen. The rapid tests can be performed in about 10-15 minutes (range, 5-60 minutes, not counting setup time of perhaps 5-15 minutes). Compared to throat culture, average overall sensitivity of the new direct methods is about 87%-95% (range 61%-100%, including different results on the same manufacturer’s kits by different investigators). Reported sensitivity is about 5%-10% higher if comparison culture specimens contain at least 10 organism colonies than if the culture density is less than 10 colonies. There is debate in the literature whether throat cultures with density of less than 10 colonies represent clinically significant infection. Some consider growth less than 10 colonies to represent a carrier state, but others disagree. Another point to remember is that rapid test sensitivity quoted in reports is not true clinical sensitivity, since rapid test sensitivity is usually less than culture sensitivity and culture sensitivity itself is rarely more than 95%. (It is probably less, since various changes in usual laboratory culture technical methodology for throat cultures have each been reported to increase culture sensitivity about 5%-10%, sometimes even more. In fact, one study on tonsillectomy patients found only 82% of preoperative throat cultures were positive when operative tonsillar tissue culture obtained group A streptococci.)

    Lancefield group A streptococci are also known as Streptococcus pyogenes. Certain strains are associated with specific diseases, such as acute glomerulonephritis and acute rheumatic fever. Group A beta streptococci also produce various infections without any stain specificity. The most common is acute pharyngitis. Wound infections and localized skin cellulitis are relatively frequent. Other diseases that are much less common, although famous historically, include scarlet fever, erysipelas (vesicular cellulitis), and puerperal fever. There are even a few reports describing patients with necrotizing fasciitis or a condition resembling staphylococcal toxic shock syndrome.

    Group A beta hemolytic streptococci may be isolated from throat or nasopharyngeal cultures in 15%-20% (range, 11%-60%) of clinically normal children. Nevertheless, for clinical purposes they are not considered normal inhabitants of the throat or nasopharynx since the usual current culture or serologic methods, which are done in the acute stage of infection, cannot reliably differentiate between carrier state and true pathogen, and prompt therapy for group A streptococcal nasopharyngeal infection is thought to decrease the possibility of acute rheumatic fever or acute glomerulonephritis. The accuracy of group A streptococcal isolation from throat cultures can be enhanced in several ways. Anaerobic rather than aerobic incubation has been reported to increase sensitivity about 15% (range, 0%-35%). Obtaining specimens on two swabs instead of one is reported to increase yield approximately 10%. Use of special differential media (with certain antibiotics added) helps to suppress growth of other organisms (e.g., hemolytic Haemophilus species) that may simulate beta hemolytic streptococcal colonies. However, not all of these enhancement techniques produce the same reported additive effect if they are combined. Although group A streptococci are almost always sensitive to penicillin on in vitro antibiotic sensitivity tests, 10%-30% of adequately treated patients continue to harbor the organisms and are considered carriers.

    Lancefield group B streptococci (also known as Streptococcus agalactiae) are one of the most common causes of neonatal septicemia and meningitis. Escherichiacoli is the most frequent etiology of neonatal meningitis, but it has been reported that about one third of cases are due to group B streptococci. Neonatal group B streptococcal infection may occur in two clinical forms: early onset (before age 7-10 days, usually within 48 hours of birth), and late onset (after age 7-10 days). Septic neonates may have respiratory distress that mimics the noninfectious respiratory distress syndrome. The source of Streptococcus infection is the maternal genital tract, with 20%-30% (literature range, 4.6%-40%) of mothers and a similar percentage of all women of child-bearing age being culture-positive. Vaginal and perirectal colonization are more likely than cervical or urinary tract colonization. Colonization exists in all three trimesters in approximately the same percentage of women. About two thirds of women colonized in the first trimester of pregnancy are still culture-positive at delivery.

    Group B streptococcus (GBS) causes about 20% of postpartum endometritis and 10%-20% of postpartum bacteremia. Caesarian section in a colonized mother has a higher risk of endometrial infection. GBS maternal colonization can also result in maternal urinary tract infection or bacteremia.

    About 50% of infants born to mothers with culture-proven GBS become colonized (range, 40%-73%), with the bacteria usually acquired during delivery. About 1%-2% of colonized infants develop clinical GBS infection. GBS infection accounts for 30%-50% of neonatal serious infections. Principal risk factors for neonatal clinical GBS infection are prematurity and premature rupture of the membranes (also a twin with infection, maternal urinary tract infection or bacteremia, multiple births, or African American race). About two thirds of infected (not only colonized) neonates develop early onset disease and about one third develop late onset disease.

    Diagnosis is most often made by maternal culture. The highest yield is obtained by one or more swabs from multiple sites in the distal third of the vagina plus the perineum (perirectal) area placed into selective liquid culture medium (usually Todd-Hewitt broth). Use of cervical cultures and solid culture media results in a considerably lower percentage of positive cultures. Rapid immunologic tests are available, similar in methodology to those for Group A streptococcal throat tests. However, current kit sensitivity for vaginal-rectal GBS overall is only about 40%-60% (range, 20%-88%), with the highest rates occurring in more heavily colonized patients. There is not a dependable cor- relation between degree of colonization and infant infection rate, although heavy colonization is more likely to result in neonatal infection.

    There is controversy whether all mothers should be screened for GBS colonization before delivery (usually at about 26 weeks’ gestation), at delivery only, at delivery only if risk factors are present, or no screening at all with all infants being watched closely for the first hours after birth. There is also controversy whether those who are culture-positive early in pregnancy should be treated immediately, whether those positive at delivery should be treated, or if no cultures are done, whether prophylactic antibiotics should be given to patients with risk factors or to all patients during delivery. In general, positive cultures obtained early in pregnancy have a 70%-80% chance of a positive culture at delivery, whereas a negative culture obtained early in pregnancy is much less reliable in predicting status at delivery. Intradelivery parenteral antibiotics (usually penicillin or ampicillin) beginning before delivery, are the most commonly recommended method of prophylaxis. The two most frequent recommendations for this prophylaxis are a positive culture before delivery or presence of maternal risk factors. One recent report advocates intradelivery treatment for all mothers as the most cost-effective procedure (based on review of the literature).

    GBS may also produce adult infections in nonpregnant persons (although this is not frequent), with postpartum endometritis, infection after urinary tract or gynecologic operations, pneumonia, and soft tissue infections being the most common types. Many affected adults have some underlying disease or predisposing cause. GBS also produces mastitis in cows.

    Culture is the major diagnostic test for GBS. Besides culture, rapid latex agglutination slide tests for group B streptococcal antigen in body fluids are now available from several manufacturers. Although not many studies are published, reports indicate that these tests when performed on concentrated urine specimens detect 90%-100% of cases in culture-positive neonatal group B streptococcal bacteremia. Serum or unconcentrated urine is generally less successful.

    Lancefield group C streptococci are primarily animal pathogens. However, these organisms may be found in the nasopharynx (1.5%-11%), vagina, and gastrointestinal (GI) tract of clinically normal persons. The most common strain in humans is Streptococcus anginosus. Group C streptococci occasionally produce human disease, most commonly (but not limited to) pharyngitis and meningitis (about 6% of cases; range, 3%-26%).

    Lancefield group D streptococci contain several species, of which Streptococcus faecalis (one of the enterococci) and Streptococcus bovis (one of the nonenterococci) are the most important. Group D streptococci are responsible for approximately 20% (range, 8%-21%) of infectious endocarditis, typically that subgroup formerly called subacute bacterial endocarditis (SBE), and about 10% of urinary tract infections, as well as constituting the third most common cause of biliary tract infections. Group D streptococci are also associated with mixed wound infections, intraabdominal infections and a wide variety of other conditions, although there is some controversy about their relative pathogenicity in these circumstances. The majority of serious group D infections are due to enterococci. Enterococci are certain species of group D streptococci that are found normally in the human intestinal tract and that have certain special laboratory characteristics. These include resistance to heating and growth in certain media such as bile-esculin and 6.5% sodium chloride. Some species of enterococci produce alpha hemo- lysis, some produce beta hemolysis, and some are (gamma) nonhemolytic. Besides their role in infectious endocarditis (10%-12% of cases, range, 6%-20%), enterococci assume importance because they frequently are involved in other nosocomial (hospital-acquired) infections, particularly urinary tract infections (about 10%, range 6%-16%) and because they are usually partially resistant or resistant to penicillin. Bacteremia due to nonenterococcal S. bovis has been very frequently associated with either benign or malignant colon neoplasms (about two thirds of cases, range 17%-95%). Recently it has been proposed that the enterococci should be removed from the streptococci and placed into a new genus called Enterococcus. The new genus would include Enterococcus faecalis, Enterococcus faecium, Enterococcus durans, and several others. Streptococcus group D would include S. bovis and Streptococcus equinus.

    Lancefield group F streptococci (Streptococcus anginosus also called milleri) form tiny colonies. Infection is uncommon, but they have been reported to cause bacteremia, dental abscesses, and abscesses in other body tissues, usually superimposed on preexisting abnormality.

    Lancefield group G streptococci are among the normal flora in the nasopharynx (up to 23% of persons), skin, vagina, and GI tract. The most common serious infection is bacteremia, being isolated in 3.5% of all patients with bacteremia in one study. Many of these patients had serious underlying disease, especially cancer (21%-65% of patients in this study with group G bacteremia). Other infections are uncommon, but pharyngitis and arthritis are among the more important. Other Lancefield groups have occasionally been reported to produce human infection.

    Viridans streptococci. The viridans group has been defined in several ways. The most common definition is streptococci that lack Lancefield antigens and produce alpha hemolysis on sheep blood agar. However, other investigators state that some viridans organisms are nonhemolytic, and still others say that some species can react with various Lancefield group antisera, although not with group B or D. Viridans streptococci are the most common cause of infectious endocarditis (that subgroup formerly known as SBE), being isolated in approximately 35%-40% of cases (range 30%-52%). They may also cause urinary tract infection, pneumonia, and wound infection. They are normal inhabitants of the mouth and may have a role in producing dental caries.

    Streptococcus pneumoniae. This organism is clinically known as pneumococcus and occasionally is called Diplococcus pneumoniae in the literature. Although S. pneumoniae is a member of the Streptococcus genus, most nonmicrobiologists usually think of pneumonococcus as a separate entity from the other streptococci. Pneumococci are gram-positive diplococci that can be found in the throat of 30%-70% of apparently normal persons. They are still the most common cause of bacterial pneumonia, usually comprising at least 50% (range, 26%-78%) of community-acquired cases. They also produce many cases of middle ear infection in children and are an important cause of meningitis in older children, adolescents, and adults, most commonly in debilitated persons. Pneumococcal bacteremia develops in about 20% (range, 15%-25%) of patients with pneumococcal pneumonia, and pneumococcus is found in about 18% of all bacteremias, with the incidence being even higher in children. Splenectomy (or sickle cell anemia hyposplenism) is one well-known predisposing cause. The great majority of pneumococcal strains are very sensitive to penicillin, although about 4%-5% (range, 0%-20%, depending on the geographical area) of isolates are now relatively resistant and 1%-2% (range, 0%-6%) are resistant. Relative resistance is important in blood and cavity fluid, and especially in cerebrospinal fluid (CSF).

    Pneumococci usually produce alpha (green) incomplete hemolysis on blood agar and thus mimic viridans streptococci. Morphologic differentiation from streptococci may be difficult, especially in cultures, since streptococci may appear singly or in pairs (instead of chains), and pneumococci often do not have the typical lancet shape or grouping in pairs. In the laboratory, differentiation is readily made because of the special sensitivity of pneumococci to a compound known as optochin. A disk impregnated with optochin is placed on the culture plate; inhibition of an alpha hemolytic coccus denotes pneumococci.

    Besides diagnosis by culture, rapid slide latex agglutination tests for pneumococcal antigen are now available (developed for testing of CSF in patients with meningitis). Unfortunately, in the few reports available, detection of antigen in serum or unconcentrated urine has been less than 40%. Sensitivity in concentrated urine may be considerably better, but too little data are available.

    Staphylococci

    Staphylococci are gram-positive cocci that typically occur in clusters. Originally they were divided into three groups, depending on colony characteristics on blood agar: Staphylococcus albus (Staphylococcus epidermidis), with white colonies; Staphylococcus aureus, with yellow colonies; and Staphylococcus citreus, with pale green colonies. In that classification, S. aureus was by far the most important; it was generally hemolytic, and the pathogenic species were coagulase positive. The newer classification recognizes that coagulase activity (the ability to coagulate plasma) is a better indication of pathogenicity than colony color, since a significant number of coagulase-positive organisms are not yellow on blood agar. Therefore, all coagulase-positive staphylococci are now called S. aureus. Many microbiology laboratories still issue reports such as “S. aureus coagulase positive”; it is not necessary to include the coagulase result because S. aureus by definition is coagulase positive. Detection of heat-stable anti-DNA (“thermonuclease”), another enzyme produced by S. aureus, is generally considered to be the best confirmatory test for S. aureus should this be necessary. This test is nearly as sensitive and specific as coagulase but is more expensive and time consuming.

    At this point, a few words should be said regarding coagulase tests. S. aureus produces two types of coagulase, known as bound coagulase (“clumping factor”) and free coagulase. The standard procedure is known as the tube coagulase test, based on the ability of free coagulase to clot plasma. This test requires 4 hours’ incubation and sometimes as long as 24 hours to confirm nonreactive results or weak positive reactions. Bound coagulase alone was originally used in a slide coagulase test that required only 1-2 minutes but produced about 20% false negative results. More recently, rapid 15- to 60-second slide tests based on reagents that react with either bound coagulase, protein A (another substance produced by S. aureus), or both have been introduced. Evaluations have shown sensitivities of about 98%-99% (range, 94%-100%) for these tests. However, false positive results have been reported in a few percent of most such tests, although the majority of evaluations of each test have not reported false positive results. Also, at least one investigator has reported at least 1% (and sometimes more) false negative results with each test when used with methicillin-resistant S. aureus. The family Micrococcaceae contains several genera, including Staphylococcus and Micrococcus. Both of these genera are composed of organisms that are identical on Gram stain and have certain biochemical similarities. As noted above, those that are coagulase positive are placed in the Staphylococcus genus and designated S. aureus. Gram-positive cocci that resemble S. aureus morphologically but that are coagulase negative have not yet been subjected to a uniform method of reporting. Some laboratories call all of them “S. epidermidis” or “coagulaseegative staphylococci.” Others differentiate organisms from the genus Staphylococcus and the genus Micrococcus on the basis of certain biochemical tests such as the ability of staphylococci to produce acid from glucose anaerobically. In general, non-aureus staphylococci, although much less pathogenic than S. aureus, are more pathogenic than micrococci. Some of the larger laboratories differentiate species of non-aureus staphylococci using certain tests. The species most commonly associated with human disease are S. epidermidis and Staphylococcus saprophyticus.

    Staphylococci, as well as the enteric gramegative rod organisms and some of the Clostridia gram-positive anaerobes (to be discussed later), are normal inhabitants of certain body areas. The habitat of staphylococci is the skin. Therefore, a diagnosis of staphylococcal infection should not be made solely on the basis of S. aureus isolation from an external wound; there should be evidence that S. aureus is actually causing clinical disease. Besides the skin, about one half of all adults out- side the hospital carry S. aureus in the nasopharynx; this reportedly increases to 70%-80% if cultures are performed repeatedly on the same population over a long period. More than 50% of hospitalized persons have positive nasopharyngeal cultures. Exactly what factors induce these commensal organisms to cause clinical disease is not completely understood.

    Staphylococcus aureus. Staphylococcus aureus is typically associated with purulent inflammation and characteristically produces abscesses. The most common site of infection is the skin, most frequently confined to minor lesions, such as pustules or possibly small carbuncles, but occasionally producing widespread impetigo in children and infants. The most frequent type of serious staphylococcal disease (other than childhood impetigo) is wound infection or infection associated with hospital diagnostic or therapeutic procedures.

    In a small but important number of cases, S. aureus produces certain specific infections. Staphylococcal pneumonia may occur, especially in debilitated persons, or following a viral pneumonia. Meningitis and septicemia are also occasionally found, again, more commonly in debilitated persons or those with decreased resistance—often without any apparent portal of entry. S. aureus produces about 20%-25% (range, 9%-33%) of infectious endocarditis (especially that subgroup that used to be called acute rather than subacute). S. aureus causes a type of food poisoning different from that of the usual infectious agent; symptoms result from ingestion of bacterial toxins rather than from actual enteric infection by living organisms.

    S. aureus frequently produces an enzyme known as beta-lactamase which makes the organism resistant to certain antibiotics that contain a beta-lactam ring structure such as penicillin G and ampicillin. S. aureus resistant to methicillin (MRSA) is a particularly difficult problem and will be discussed later in the section on antibiotic sensitivity testing. The nationwide incidence of MRSA (as a percentage of all S. aureus isolates) is about 4%; for individual hospitals, the range is 0%-60%. Staphylococcus epidermidis in many areas of the country is frequently resistant to a considerable number of antibiotics, including methicillin.

    Toxic shock syndrome. The toxic shock syndrome is a disease strongly linked to S. aureus infection. The syndrome has been seen in various age groups in both sexes but occurs predominantly in females of childbearing age. There is an association with use of vaginal tampons, but a considerable number of cases have occurred when tampons were not used. Many (but not all) cases occur during the menstrual period. At least 15% of cases are not related to menses or tampon use but in- stead are preceded by surgical wound infection or by infections of other types. Clinical criteria for the syndrome include temperature of 102°F (38.9°C) or greater, erythematous sunburnlike or macular rash with eventual skin peeling or desquamation (especially on the palms and soles), hypotension or shock, clinical or laboratory involvement of at least four organ systems, and no definite evidence of bacteremia.

    Involvement of organ systems is evidenced by a combination of symptoms and laboratory data. The classic syndrome includes at least four of the following organ systems (the approximate incidence of laboratory abnormality reported in one study is listed after each test):

    1. Gastrointestinal: Vomiting and watery diarrhea.
    2. Central nervous system: Headache, confusion, and disorientation. Stiff neck may occur. CSF tests are usually normal.
    3. Liver: Aspartate aminotransferase (SGOT, 75%) or alanine aminotransferase (SGPT, 50%) more than twice the upper limit of the reference range. Total bilirubin may also be increased (70%).
    4. Kidney: Blood urea nitrogen (BUN, 68%) or serum creatinine (90%) levels more than twice the upper limit of the reference range. There may be increased numbers of urinary white blood cells (WBCs) (73%) with negative urine cultures. Oliguria may occur.
    5. Mucous membranes: Conjunctivitis.
    6. Hematologic: Hemoglobin level is usually normal (77%). The WBC count may be normal (52%) or increased (48%). Neutrophil percentage is usually increased (86%), and frequently there are increased band forms. Dohle bodies are often present. Schistocytes are present in some patients (48%). The platelet count is less than 100,000/mm3 in 42%, the activated partial thromboplastin time is elevated in 46%, and fibrin split products are frequently elevated; but the fibrinogen level is normal in 86% and the prothrombin time is normal in 93%.
    7. Muscular: Myalgias; creatine phosphokinase level is more than twice the upper limit of the reference range (63%).

    It is not clear how many of the laboratory abnormalities are due to the disease itself or are secondary to hypotension with tissue hypoxia.

    Staphylococcus aureus has been cultured from the cervix or vagina in approximately 75% of toxic shock patients (literature range 67%-92%) as opposed to approximately 7% (literature range, 0%-15%) of normal persons. Some series include culture data from the nasopharynx in addition to the cervix-vaginal region. This is more difficult to interpret since about 20% of normal persons carry S. aureus in their nasopharynx. Blood cultures are usually sterile. Toxic shock S. aureus is said to be usually nonhemolytic phage type 1, able to produce toxic shock toxin-1 (TSST-1) exotoxin, sensitive to erythromycin but resistant to penicillin. Currently, there is no readily available test to differentiate toxic shock from other entities with similar clinical or laboratory manifestations. It is possible to culture the organism and send an isolate to some reference laboratory able to test for production of TSST-1, but this would take several days.

    Staphylococcus epidermidis. Staphylococcus epidermidis is found in more abundance on the skin than S. aureus and is a relatively frequent contaminant in cultures from the skin area. It may also be a contaminant in blood cultures or other cultures when the material for culture is obtained by needle puncture of the skin. However, S. epidermidis may produce disease. Infections most frequently associated with S. epidermidis are related to indwelling catheters, vascular grafts, or joint prosthetic devices; bacteremia in patients who are immunosuppressed; and infection after eye surgery (endophthalmitis). It is the causal agent of infectious endocarditis in 2%-3% of all endocarditis cases (range, 1%-8%), more often in persons with an artificial heart valve (where it may comprise up to 40% of prosthetic valve endocarditis). S. epidermidis is the most frequent cause of bacterial colonization or infection associated with indwelling vascula catheters and those used for hyperalimentation, where it is reported to cause about 20% of bacteremias and 40% of septicemias. Other catheter-related problem areas include CSF shunts (causing about 50% of infections; range, 44%-70%) and peritoneal dialysis catheters (about 50% of infections). S. epidermidis is also reported to cause about 40% (range, 25%-70%) of orthopedic joint prosthesis infections. Since S. epidermidis is a common skin-dwelling organism, it is a frequent culture contaminant. When it is obtained from a blood culture, there is always a question regarding its significance. Some rules of thumb from the literature that would tend to suggest contamination are isolation from only one of a series of cultures, isolation from only one of two cultures drawn close together in time, and isolation from only one bottle of an aerobic and anaerobic two-bottle culture set inoculated from the same syringe. However, none of these conclusively proves that the organism is a contaminant. Whereas S. aureus is differentiated from other staphylococci by a positive coagulase test result, many laboratories do not perform tests to identify the exact species of coagulaseegative staphylococci but report them all as S. epidermidis. However, not all of these organisms are S. epidermidis. Also, at least one of the coagulaseegative nonepidermidis staphylococci, S. saprophyticus, has been implicated as the causal agent in about 20% of urinary tract infections occurring in young women. On the other hand, S. epidermidis is not considered to be a urinary tract pathogen in young women, at least by some investigators. Therefore, it might be better to use the term “coagulaseegative Staphylococcus” in culture reports if the laboratory does not speciate these organisms. S. epidermidis is often resistant to many antibiotics, with 33%-66% reported to be resistant to methicillin.

  • Laboratory Classification of Bacteria

    The most useful laboratory classification of bacteria involves a threefold distinction: the Gram stain characteristics (gram positive or gram negative), morphology (coccus or bacillus), and oxygen requirements for growth (aerobic or anaerobic). Species exist that are morphologic exceptions, such as spirochetes; others are intermediate in oxygen requirements; still others are identified by other techniques, such as the acid-fast stain. Reaction to Gram stain has long been correlated with bacterial sensitivity to certain classes of antibiotics. A classic example is the susceptibility of most gram-positive organisms to penicillin. Morphology, when used in conjunction with this primary reaction, greatly simplifies identification of large bacterial groups, and oxygen growth requirements narrow the possibilities still further. The interrelationship of these characteristics also helps to control laboratory error. For example, if cocci seem to be gram negative instead of gram positive, a laboratory recheck of the decolorization step in the Gram procedure is called for since nearly all cocci are gram positive. If the staining technique is verified, the possibility of a small bacillus (Coccobacillus) or a Diplococcus must be considered.