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The genus Bacillus includes many species of aerobic or facultative, spore-forming, Gram- positive rods. With the exception of one species, B anthracis, they are low-virulence saprophytes widespread in air, soil, water, dust, and animal products. Bacillus anthracis causes the zoonosis anthrax, a disease of animals that is occasionally transmitted to humans. The genus is made up of rod-shaped organisms that can vary from coccobacillary to rather long-chained filaments. Motile strains have peritrichous flagella. Formation of round or oval spores, which may be central, subterminal, or terminal depending on the species, is characteristic of the genus.
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Gram-positive spore-forming rods
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With Bacillus, growth is obtained with ordinary media incubated in air and is reduced or absent under anaerobic conditions. The bacteria are catalase-positive and metabolically active. The spores survive boiling for varying periods and are sufficiently resistant to heat that those of one species are used as a biologic indicator of autoclave efficiency. Spores of B anthracis survive in soil for decades.
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Aerobic conditions preferred for growth
Heat-resistant spores survive boiling
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Bacillus anthracis has a tendency to form very long chains of rods and in culture is nonmotile and nonhemolytic; colonies are characterized by a rough, uneven surface with multiple curled extensions at the edge resembling a “Medusa head.” Bacillus anthracis has a polypeptide (poly-d-γ-glutamic acid) capsule of a single antigenic type that has antiphagocytic properties similar to those of bacterial polysaccharide capsules. Bacillus anthracis endospores are extremely hardy and have been shown to survive in the environment for decades. The organism also produces a potent exotoxin complex, which consists of two enzymes, edema factor (EF) and lethal factor (LF) together with a receptor-binding protein called protective antigen (PA). When PA binds to either EF or LF it then acts as a translocase forming a pore-like site on the host cell surface. This allows the complexes to enter the cell (Figure 26–9A). Once in the cytosol multiple toxin actions are expressed including adenylate cyclase activity and host protein inactivation. Bacillus anthracis also produces multiple other proteases that digest tissue components.
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Endospores survive in nature
Polypeptide capsule is antiphagocytic
Exotoxin complex has multiple component and actions
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CLINICAL CAPSULE
Human anthrax is typically an ulcerative sore on an exposed part of the body. Constitutional symptoms are minimal, and the ulcer usually resolves without complications. If anthrax spores are inhaled, a fulminant pneumonia may lead to respiratory failure and death.
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The isolation of B anthracis, the proof of its relationship to anthrax infection, and the demonstration of immunity to the disease are among the most important events in the history of science and medicine. Robert Koch rose to fame in 1877 by growing the organism in artificial culture using pure culture techniques. He defined the stringent criteria needed to prove that the organism caused anthrax (Koch's postulates), then met them experimentally. Louis Pasteur made a convincing field demonstration at Pouilly-le-Fort to show that vaccination of sheep, goats, and cows with an attenuated strain of B anthracis prevented anthrax. He was cheered and carried on the shoulders of the grateful farmers of the district, an experience now, unhappily, largely restricted to winning football coaches.
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Pasteur produced animal vaccine with attenuated anthrax strain
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Anthrax is primarily a disease of herbivores such as horses, sheep, and cattle, who acquire it from spores of B anthracis contaminating their pastures. Humans become infected through contact with these animals or their products in a way that allows the spores to be inoculated through the skin, ingested, or inhaled. In the 1920s, more than 100 cases occurred annually in the United States among farmers, veterinarians, and meat handlers, but the control of animal anthrax in developed countries has made human cases rare. A few endemic foci persist in North America and have been the source of naturally acquired disease. Another source is animal products such as wool, hides, or bone meal fertilizer that have been imported from a country where animal anthrax is endemic.
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Infection is through injection of spores derived from herbivores into the skin
Contaminated materials are imported from countries with animal anthrax
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The real threat associated with anthrax comes from its continuing appeal to those bent on using it as an agent of biologic warfare or terrorism. The long life, stability, and low mass of the dried spores make the prospect of someone producing a “cloud of death” leading to massive pulmonary anthrax a chilling reality. A 1979 episode resulting in more than 60 anthrax deaths in the former Soviet Union is now attributed to an accidental explosion at a biologic warfare research facility that aerosolized more than 20 pounds of anthrax spores. United Nations inspection teams in Iraq uncovered facilities for the production of massive amounts of spores together with plans to create and spread infectious aerosols using missile warheads. The inhalation anthrax among postal workers after the September 11, 2001, terrorist attacks appears to have been due to the mailing of envelopes containing “weapons-grade” anthrax spores stolen from a biologic warfare research facility. Such spores had been treated to enhance their aerosolization and dissemination. The forms of anthrax are summarized in Figure 26–10.
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Use for biologic warfare is a continuing threat
Aerosols could spread pulmonary anthrax widely
Weapons-grade spores are specially treated
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When spores of B anthracis reach the rich environment of human tissues, they germinate and multiply in the vegetative state. The antiphagocytic properties of the capsule aid in survival, eventually allowing production of enough exotoxin to cause disease. The timing and relative importance of the EF, LF, and PA components is not known. The EF adenylate cyclase activity is believed to correlate with the striking edema seen at infected sites. In pulmonary anthrax the inhaled spores are taken up by alveolar macrophages but apparently do not germinate inside them at least until they drain to the mediastinum via the lymphatics. This most lethal of anthrax forms is manifest in the lung as a mediastinal process and systemically as a virulent bacteremia.
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Antiphagocytic effect of glutamic acid capsule required for virulence
Edema is produced by EF
Pulmonary focus is mediastinum
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The specific mechanisms of immunity against B anthracis are not known. Experimental evidence favors antibody directed against the toxin complex, but the relative role of the components of the toxin is not clear. The capsular glutamic acid is immunogenic, but antibody against it is not protective.
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Immune mechanisms are unknown
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ANTHRAX: CLINICAL ASPECTS
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Cutaneous anthrax usually begins 2 to 5 days after inoculation of spores into an exposed part of the body, typically the forearm or hand. The initial lesion is an erythematous papule, which may be mistaken for an insect bite. This papule usually progresses through vesicular and ulcerative stages in 7 to 10 days to form a black eschar (scab) surrounded by edema (Figure 26–9B and C). This lesion is known as the “malignant pustule,” although it is neither malignant nor a pustule. Associated systemic symptoms are usually mild, and the lesion typically heals very slowly after the eschar separates. Less commonly, the disease progresses with massive local edema, toxemia, and bacteremia.
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Initial papule evolves to malignant pustule
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Pulmonary anthrax is contracted by inhalation of spores. Historically, this has occurred when contaminated hides, hair, wool, and the like are handled in a confined space (wool-sorter disease) or after laboratory accidents. Today it is the form we would expect from the dissemination of a spore aerosol in biologic warfare. In the pulmonary syndrome, 1 to 5 days of nonspecific malaise, mild fever, and nonproductive cough lead to progressive respiratory distress and cyanosis. Spread to the bloodstream and CNS follow rapidly. Massive edema and hemorrhage are hallmark features of anthrax meningitis. Mediastinal edema was a prominent finding in the postal workers. If untreated, progression to a fatal outcome is usually very rapid once bacteremia has developed. An intestinal form of anthrax follows ingestion of contaminated food, usually meat. It is characterized by abdominal pain, ascites, and shock.
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Pulmonary anthrax is acquired by inhaling spores
Fever and cough progress to cyanosis and death
Hemorrhagic mediastinitis and meningitis
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Culture of skin lesions, sputum, blood, and CSF are the primary means of anthrax diagnosis. Given some suspicion on epidemiologic grounds, Gram stains of sputum or other biologic fluids showing large numbers of long Gram-positive bacilli can suggest the diagnosis. In September 2001, diagnosis of the first case in Florida was speeded by an infectious disease specialist who knew such rods were extremely rare in the spinal fluid. Large Gram-positive bacilli are also unusual in sputum. Bacillus anthracis and other Bacillus species are not difficult to grow. In fact, clinical laboratories frequently isolate the nonanthrax species as environmental contaminants. The saprophytic species are usually β-hemolytic and motile, features not found in B anthracis, but most clinical laboratories are not skilled at separating Bacillus species. Blood cultures are positive in most cases of pulmonary anthrax.
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Smears with large Gram-positive rods are suggestive
Hemolysis and motility exclude B anthracis
Sputum and blood cultures are positive in pneumonia
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Antimicrobial treatment has little effect on the course of cutaneous anthrax but does protect against dissemination. Almost all strains of B anthracis are susceptible to penicillin, doxycycline, and ciprofloxacin. Although penicillin has long been the treatment of choice for all forms of anthrax, experience gained during the 2001 outbreak has caused the first-line recommendation to be changed to doxycycline or ciprofloxacin. These antibiotics are also recommended for chemoprophylaxis in the case of known or suspected exposure.
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Ciprofloxacin or doxycycline is used for treatment and prophylaxis
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The most important preventive measures are those that eradicate animal anthrax and limit imports from endemic areas. Vaccines are also useful. Pasteur's vaccine used a live strain attenuated by repeated subculture that resulted in the loss of a plasmid encoding toxin production. A similar live vaccine is still effective for animals. The human vaccine licensed in the United States is prepared by extraction from cultures of a nonencapsulated avirulent strain of B anthracis. The extract is made up of almost entirely the protective antigen component of the toxin complex. In 2002, the Institute of Medicine issued a detailed analysis of human and animal studies and declared the vaccine both safe and efficacious. Experts also feel that it is very unlikely that the architects of biologic warfare would be able to craft B anthracis strains for which this vaccine is not protective. In Russia and China, a live vaccine is used in which spores are inoculated by scarification.
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Eradication of animal anthrax is most important
Live and inactivated vaccines are available
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Other Bacillus Species
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Bacillus spores are widespread in the environment, and isolation of one of the more than 20 Bacillus species other than B anthracis from clinical material usually represents contamination of the specimen. Occasionally B cereus, B subtilis, and some other species produce genuine infections, including infections of the eye, soft tissues, and lung. Infection is usually associated with immunosuppression, trauma, an indwelling catheter, or contamination of complex equipment. The relative resistance of Bacillus spores to disinfectants aids their survival in medical devices that cannot be heat sterilized.
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Spores enhance survival in medical devices
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Bacillus cereus deserves special mention. This species is the one most likely to cause opportunistic infection, which suggests a virulence intermediate between that of B anthracis and the other species. Genes and plasmids similar to those found in B anthracis have been detected as has a destructive pyogenic toxin. Bacillus cereus can also cause food poisoning by means of enterotoxin production.
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B cereus produces pyogenic toxin and enterotoxin