Anaerobic Infections Part 3

Diseases from Neurotoxic Clostridia

Tetanus All of the clinical features of tetanus are caused by a potent neurotoxin—tetanospasmin—produced by C. tetani. The toxin travels to the spinal cord and suppresses the inhibitory neurotransmitter GABA in the neuromuscular junction, resulting in severe muscle spasms.

Tetanus is now rare in industrialized countries because of widespread immunization. In the United States, fewer than 50 cases are reported annually, mostly in inadequately immunized older adults.32 In developing countries, tetanus is still a major problem, causing the deaths of an estimated one million persons annually, half of whom are newborns. C. tetani is commonly found in the soil, in the intestines of domestic animals, and occasionally in human feces. The organism is commonly introduced by a laceration or puncture wound that is usually sustained outdoors. Tetanus may also occur in association with pregnancy (postpartum and postabortion tetanus), injection of illicit narcotics, surgery (postoperative tetanus), burns, vaccination, intramuscular injections, chronic skin ulcers, dog bites, and umbilical stump infection in newborns (neonatal tetanus). In 10% to 20% of patients with tetanus, there is no history of injury or evidence of an infected lesion.

The disease is characterized by generalized rigidity and intermittent, intense muscle spasms. The incubation period ranges from 1 to 55 days, but onset of symptoms occurs within 14 days after the initial injury in over 80% of patients. The usual presenting symptoms are restlessness; pain caused by muscle spasm; and stiffness of the back, neck, thighs, and abdomen. When muscle spasms occur, the characteristic clinical features are determined by the relative strengths of the opposing muscles: the greater strength of the masseter over the opposing digastricus and mylohyoid results in trismus; the greater strength of the extensor groups over the flexors in the lower extremities produces characteristic extension at the hips and knees; and the greater strength of the biceps results in flexion of the forearms. This combination of flexion of the upper extremities and extension of the lower extremities is termed opisthotonos [see Figure 4].

Figure 4 Spasms in a wounded soldier with tetanus are illustrated in this drawing by Scottish surgeon and anatomist Sir Charles Bell in his topic The Anatomy and Philosophy of Expression, published in 1832. The classic signs of tetanus—risus sardonicus, trismus, and opisthotonos—are shown.

Difficulty in opening the mouth (trismus, or lockjaw) is the first symptom in more than 50% of patients. Dysphagia, caused by spasm of pharyngeal muscles, may be an early symptom. Deep tendon reflexes are hyperactive, but the plantar responses are flexor. As the process progresses, violent spasms of the paraspinal, abdominal, and limb musculature occur, but the patient remains conscious. Trismus and stiffness of the facial muscles produce risus sardonicus, a characteristic sneering expression. Sudden stimuli (e.g., bright light or noise) can precipitate tonic seizure accompanied by diaphragmatic, intercostal, glottal, or laryngeal spasm; such spasms can result in hypoxia and respiratory arrest. Fever may be caused by the marked muscular rigidity and spasms alone. Severe sympathetic hyperactivity, evidenced by labile hypertension or hypotension, tachypnea, tachycardia, arrhythmias, profuse sweating, and marked intermittent vasoconstriction, may occur singly or in varying combinations. After the manifestations of tetanus have peaked, they persist at the same level for about a week and then gradually diminish over several weeks. Residual stiffness may persist for several more weeks. In patients who have survived moderate to severe tetanus, respiratory assistance has been required for 2 to 4 weeks; the average hospital stay has been 5 to 8 weeks. The major complications of tetanus are respiratory arrest secondary to tetanic spasms, pneumonia secondary to aspiration, pulmonary emboli, cardiac problems related to sympathetic overactivity or to cardiomyopathy, and fractures of thoracic vertebrae caused by violent spasms.

Botulism About 100 cases of botulism are reported in the United States each year, but a surge in cases associated with black tar heroin and the threat of international bioterrorism have renewed interest in a disease that was first recognized in the early 17th century.

As with tetanus, all the clinical manifestations of botulism are caused by a potent neurotoxin produced by C. botulinum. The organism has been isolated from soil everywhere in the world. The spores are very hardy, resisting dryness and extremes of temperature. Spores that are introduced into food, wounds, or the human intestinal tract can germinate and elaborate botulinum toxin. Although seven antigenically distinct forms of the toxin (serotypes A through G) have been identified, just three types (A, B, and E) account for nearly all human disease. Botulism occurs in three main forms: food-borne botulism, wound botulism, and infant botulism.

Food-borne botulism results from the ingestion of home-processed foods that are improperly cooked or refrigerated. The vegetative organisms produce the toxin, which is heat labile and can be destroyed by boiling for 10 minutes or by heating to 80° C for 30 minutes. However, if contaminated food is not heated sufficiently, the toxin resists gastric acid and intestinal trypsin and is absorbed from the intestinal tract. About half the cases of food-borne botulism are caused by type A toxin; the rest are divided evenly between types B and E.

Wound botulism typically follows severe trauma, such as a crush injury involving an extremity. However, heroin use has become the leading predisposing factor in the United States; most cases have occurred in California after subcutaneous injection of black tar heroin from Mexico. About 80% of wound botulism is caused by type A toxin; most of the rest is caused by type B toxin.

Infant botulism is caused by the ingestion of C. botulinum spores rather than preformed toxin. The spores then germinate, colonizing the intestinal tract with toxin-producing organisms. Although the source of the spores eludes detection in most cases, honey has been implicated in about 15% of cases. Toxin type A and type B each accounts for about half the cases of infant botulism.

The symptoms of food-borne botulism usually begin 18 to 36 hours after ingestion of the toxin; the incubation period for wound botulism is typically longer. Although food-borne botulism may be heralded by gastrointestinal symptoms such as nausea, vomiting, abdominal cramps, and diarrhea (earlier stage) or constipation (later stage), neurologic manifestations soon predominate. In wound botulism, gastrointestinal symptoms are absent and the wound may appear surprisingly benign. Cranial nerve symptoms such as blurred vision and diplopia are usually the earliest neurologic complaints, followed by dysphagia, dysarthria, and dry mouth. Symmetrical motor paralysis ensues, characteristically progressing in a descending fashion that begins with the arms and then involves the respiratory muscles and lower body. Autonomic dysfunction can produce constipation, urinary retention, and orthostatic hypotension. Sensory deficits are absent, and mentation is normal. Respiratory arrest occurs in severe cases; mechanical ventilation and respiratory support may be required for weeks to months before full recovery. Infant botulism presents as lethargy, constipation, poor feeding, and floppiness, typically in the second month of life.

Newly recognized anaerobes

Molecular tools such as 16S ribosomal DNA sequencing are continuing to identify new genera and species of clinically relevant anaerobic bacteria.34,35 For example, the anaerobic gram-negative bacillus Bilaphila wadsworthia is now known to be an important pathogen and is frequently isolated from gangrenous ap-pendicitis.36 Newly described anaerobic cocci, gram-positive non-spore-forming rods, and clostridia have also been isolated from various infections.35,37

Diagnosis of Anaerobic Infections

Clinical clues to anaerobic infections

Apart from actinomycosis and clostridial myonecrosis, infections involving obligate anaerobes are generally indistinguishable from infections caused by other pathogens. Clinical manifestations are largely determined by the organ system involved and by the extent and chronicity of the infection. The two most helpful clinical clues are the presence of local tissue ischemia or necrosis and the proximity of infection to mucosal surfaces where obligate anaerobes normally reside. A putrid, foul-smelling discharge is virtually diagnostic of infection involving anaerobes, although the absence of foul odor does not rule out this possibility. Similarly, crepitus or black discoloration of affected tissue is only suggestive evidence.

Microbiologic diagnosis of anaerobic infections

A well-performed Gram stain of appropriately collected clinical material is a very useful diagnostic tool. Anaerobic infections are typically polymicrobial, and the characteristic cellular morphology of certain anaerobic pathogens may be recognized by an accomplished microscopist [see Figure 1]. The finding of so-called sterile pus by conventional culture methods in the face of a positive Gram stain should be considered presumptive evidence of an anaerobic infection. In the final analysis, however, the accurate diagnosis of anaerobic infection depends on the ability of the laboratory to isolate these fastidious organisms from clinical material likely to yield meaningful bacteriologic data.

Specimen Collection and Transport

One of the major handicaps in the recovery of anaerobic bacteria is improper specimen collection and transport. Care must be taken to avoid specimens that may be contaminated by commensal flora of mucocutaneous surfaces where anaerobes normally reside (e.g., throat swabs, expectorated sputum, voided urine, bronchoscopic and nasotracheal aspirates, vaginal secretions, feces, colostomy effluent, or superficial wound swabs). Blood and other body fluids that are normally sterile and asepti-cally obtained should be routinely cultured for anaerobic bacteria. Other clinical materials likely to yield meaningful bacterio-logic data for anaerobic infections include specimens from tissue biopsy or curettage or from deep wounds during surgery.

Proper specimen transport to preclude aeration is critical for microbiologic confirmation of an anaerobic infection. Many fastidious organisms are extremely oxygen sensitive and cannot withstand even a brief moment of exposure to air. Furthermore, in mixed infections, the presence of facultative organisms that grow faster than anaerobes frequently precludes recovery of the latter. Several commercially available systems for anaerobic transport of clinical specimens have been evaluated and have been shown to provide excellent recovery of fastidious anaer-obes.38 If commercial anaerobic transport vials are not available, specimens should be collected with a sterile needle and syringe. Air in the syringe is carefully expelled. The needle is capped to minimize aeration, and the specimen should be promptly delivered to the clinical laboratory. If swabs are to be used, they should be prepared, stored, and transported in gas-filled containers under anaerobic conditions. Immediate processing of specimens by the laboratory also improves recovery, but in practice, this is often not feasible.

Radiologic and imaging studies

Noninvasive tests such as computed tomography, magnetic resonance imaging, ultrasonography, and gallium or indium scanning are most useful for localization of suppurative infections in the central nervous system and in intra-abdominal and pelvic organs. The sensitivity and specificity of these tests in the detection of abscess and the differentiation from tumor, hemato-ma, and other noninflammatory space-occupying lesions in various sites remain to be determined by careful prospective study. In general, it may be said that a positive scan is highly suggestive, particularly when supported by the clinical picture; however, a negative scan is much less useful in ruling out infection.

Specific anaerobic infections

Actinomycosis

The diagnosis of actinomycosis depends on identification of the organism by smear or culture and by characteristic histo-pathology from tissue biopsy. The identification of sulfur granules establishes the diagnosis of actinomycosis [see Figure 3]; however, sulfur granules may constitute no more than 1% of total tissues in a given lesion and, hence, are easily missed by routine tissue staining. Similar granules may be seen with other microorganisms, notably Nocardia brasiliensis and Streptomyces madurae (both of which can cause mycetoma), as well as S. aureus (a cause of botryomycosis). However, these other granules do not have peripheral clubs, which appear to be specific to Actino-myces species. Not all Actinomyces species form sulfur granules (e.g., A. odontolyticus does not), and a peripheral fringe of clubs may be absent in certain instances, such as in a tonsillar crypt infection or in pelvic actinomycosis associated with an IUD. Additionally, Actinomyces species can be morphologically differentiated from Nocardia; moreover, Nocardia is acid fast in modified acid-fast stains, whereas Actinomyces is not.

Clostridial Myonecrosis versus Crepitant Cellulitis

Surgical exploration is necessary to distinguish myonecrosis from anaerobic cellulitis. In gas gangrene, the involved muscle looks cooked and lacks contractility, whereas in clostridial cel-lulitis, the muscle is visibly healthy. A presumptive diagnosis is based on a typical Gram stain of the wound drainage or aspirate that reveals many clostridia but few leukocytes. The presence of gas in subcutaneous tissue is not pathognomonic of clostridial infection. Patients with diabetes mellitus are particularly prone to crepitant cellulitis caused by enteric bacteria or Bacteroides species. Perineal phlegmons, which result from extension of perirec-tal abscesses caused by mixed anaerobic and facultative organisms, may also involve subcutaneous gas formation. Crepitus from trapped air after traumatic injury can usually be distinguished from anaerobic cellulitis or myonecrosis by the fact that the former does not spread.

C. difficile-Associated Diarrhea

The diagnosis of CDAD is established by demonstrating the toxins of C. difficile in stool specimens by immunoassays. Toxins A and B can be detected using specific antibodies. In approximately 5% to 20% of patients, more than one stool specimen is required to detect C. difficile toxin. Consequently, when enzyme-linked immunosorbent assay results are negative but clinical suspicion is high, tests should be repeated using the tissue culture cytotoxicity assay. Some clinical laboratories utilize screening tests that detect the presence of C. difficile in fecal specimens, either by culture or by detecting the presence of glutamate dehy-drogenase, a metabolic enzyme expressed at high levels by all strains of C. difficile, both toxigenic and nontoxigenic.39 Although these rapid screening tests may be cost-effective in some instances where large volumes of fecal specimens are processed, they are more suitable for excluding rather than establishing the diagnosis of CDAD because of their high negative (approximately 98%) but low positive (approximately 60%) predictive values.

Tetanus

The diagnosis of fully developed tetanus presents little difficulty. Acute strychnine poisoning is the only disease that resembles tetanus. A greater problem in the differential diagnosis occurs earlier in the course of the illness, when trismus is the principal manifestation. Trismus may occur in patients with intraoral disease, especially dental or jaw infections, and is occasionally seen in patients with trichinosis. Hepatic encephalopathy is sometimes associated with prominent muscle stiffness and rigidity. However, the associated liver disease is usually obvious. Furthermore, a sudden stimulus, such as jarring a bed rail, is likely to cause spasms in a patient with tetanus but not in a patient with hepatic encephalopathy. Trismus may also develop as an acute reaction to phenothiazines (the so-called grimacing syndrome). Unlike trismus from tetanus, the masseter muscle spasm in this drug reaction is painful and intermittent, and to some degree it can be overcome voluntarily. This drug reaction is readily reversed with intravenous diphenhydramine.

Botulism

Because botulism is uncommon, the diagnosis may not be entertained despite characteristic clinical findings. Clustering of cases in a family or community and a history of eating home-canned or spoiled foods may be important clues. The differential diagnosis includes Guillain-Barre syndrome, Eaton-Lambert syndrome, myasthenia gravis, cerebrovascular accidents, tick paralysis, and chemical intoxication. In patients with botulism, results of complete blood counts, blood chemistries, CNS imaging studies, and cerebrospinal fluid analysis are all normal. Rapid repetitive electromyography, however, is highly suggestive of botulism if it demonstrates a pattern of facilitation. A positive diagnosis can be established by demonstrating botulinum toxin in serum or stool specimens; the toxin may also be detected in food samples.

Management of Anaerobic Infections

Successful treatment of anaerobic infections requires rational antibiotic selection in conjunction with judicial surgical resection and drainage. The choice of antibiotics should be guided by culture results and antibiotic susceptibility data.

Several methods for antimicrobial susceptibility testing of obligate anaerobes have been validated by the National Committee for Clinical Laboratory Standards.40 Both agar dilution and mi-crobroth testing methods are appropriate, whereas the E-test, which utilizes a predefined gradient of antibiotic concentrations on a plastic strip, offers a more expensive but practical and fairly accurate alternative for susceptibility testing of individual anaerobic isolates. In light of the growing concern of emerging antibiotic resistance among anaerobic bacteria, the need for more regular susceptibility testing of clinical isolates of anaerobic bacteria has become evident.41 Susceptibility testing is particularly important in clinical settings where there has been a suboptimal response to empirical antibiotic regimens. Certainly, antimicrobial susceptibility testing should be routinely performed on organisms that are frequently resistant to antibiotics commonly used as empirical therapy, such as members of the B. fragilis group; pigmented Prevotella, including P. bivia and P. disiens; and certain Fusobacterium species. In the absence of specific culture or susceptibility data, initial antibiotic therapy must be chosen empirically and directed against the pathogens most likely to be present in a particular clinical setting [see Table 5], in accordance with predicted in vitro susceptibility patterns [see Table 6].

Predicted antimicrobial susceptibility

Although penicillin G has been considered the agent of choice for a number of mixed infections at various sites above the diaphragm (particularly oropulmonary and head and neck infections), p-lactamase production and treatment failure have been increasingly reported.42 p-lactamase production is increasingly recognized in oral isolates of P. intermedia, F. nucleatum, and Pep-tostreptococcus micros.42,43 Among the cephalosporins, only cefox-itin, cefotetan, and ceftizoxime have an enhanced antianaerobic spectrum. These agents appear to have comparable activity against B. fragilis, with resistance rates ranging from 10% to 20%; none are as active as clindamycin or metronidazole. Among the penems, imipenem-cilastatin, meropenem, and erzapenem are the most broadly active.44 The monobactam aztreonam is inactive against anaerobes, as well as gram-positive aerobes.

All strains of B. fragilis produce -lactamases and are resistant to penicillin, but extended-spectrum penicillins in combination with p-lactamase inhibitors (e.g., ampicillin-sulbactam, ticarcillin-clavulanate, and piperacillin-tazobactam) are active against most strains. Increasing resistance of B. fragilis to cefoxitin and clin-damycin has also been reported, and they are no longer the agents of choice in intra-abdominal infections.45 Cefotetan, cefti-zoxime, piperacillin-tazobactam, imipenem, and meropenem remain active. Antibiotic susceptibilities of the non-fragilis species of the B. fragilis group are more variable than those of B. fragilis. Only metronidazole, imipenem, and chloramphenicol are predictably active against nearly all isolates.

Erythromycin and ketolides are relatively inactive against Fu-sobacterium species and most B. fragilis strains. Similarly, the first-or second-generation quinolones (e.g., norfloxacin, ciprofloxacin, enoxacin, ofloxacin, and levofloxacin) are relatively inactive as single-agent therapy for anaerobic or mixed infections. However, the third-generation quinolones moxifloxacin and gatifloxa-cin have good in vitro activity against most anaerobes, including B. fragilis,47 whereas gemifloxacin is less active.48

Table 6 Predicted in Vitro Susceptibility of Clinically Important Anaerobes to Major Classes of Antimicrobial Agents

	Above Diaphragm		Above or Below Diaphragm		Below Diaphragm
Antibiotic	Fusobacterium Species	Porphyromonas and Prevotella Species	Peptostreptococcus Species	Actinomyces Species	Bacteroides fragilis Group	Clostridium Species
Penicillin	S	S-R	S	S	R	S*
Ampicillin-sulbactam+	S	S	S	S	S	S
Piperacillin, ticarcillin	S	S	S	S	S-R	S
Piperacillin-tazobactam	S	S	S	S	S	S
Imipenem, meropenem	S	S	S	S	S	S
Cefazolin	S	S-R	S	S-R	R	S
Cefoxitin	S	S-R	S	S-R	S	S-R
Cefotetan	S	S	S	S-R	S-R	S
Ceftizoxime	S	S	S	S-R	S	S-R
Cefoperazone, cefotaxime	S	S	S	S-R	S-R	S
Ceftriaxone, ceftazidime	S	S-R	S	S-R	S-R	S
Clindamycin	S*	S	S*	S	S*	S-R
Macrolides	R	S-R	S	S-R	R	S
Metronidazole	S	S	S-R	S	S	S
Ciprofloxacin, levofloxacin	R	R	R	R	R	R
Moxifloxacin, gatifloxacin, gemifloxacin	S	S	S	S	S	S
Tetracycline	S	S-R	S-R	S	S-R	S-R

S—> 80% of strains sensitive S-R—30%-80% of strains sensitive R—< 30% of strains sensitive

^Emerging resistance noted.

^Similar combinations currently available, including amoxicillin-clavulanate and ticarcillin-clavulanate, are comparably active.

Metronidazole has excellent activity against B. fragilis, Fu-sobacterium species, and Clostridium perfringens. Peptostreptococcus and Bacteroides species other than B. fragilis are only moderately sensitive, whereas nonsporulating gram-positive bacilli are relatively resistant. Metronidazole lacks activity against aerobic bacteria and should not be used as a single agent for empirical therapy, because most infections involving anaerobic bacteria are in fact mixed infections. On the other hand, metronidazole is the only agent with consistent bactericidal activity against B. fragilis. Metronidazole crosses the blood-brain barrier well, so it is particularly useful for treating anaerobic brain abscess or infective endocarditis.

Tetracycline and its analogues can no longer be recommended for the empirical treatment of anaerobic infections because of the substantial resistance acquired by B. fragilis and virtually all classes of other anaerobic bacteria. Tetracycline remains useful in the treatment of actinomycosis, however. Trimethoprim-sul-famethoxazole has only limited activity against anaerobic bacteria. Vancomycin is effective against some gram-positive anaerobes (particularly C. difficile), but it has no activity against gram-negative anaerobes. Aminoglycosides are uniformly inactive against obligate anaerobes.