Etiology and Epidemiology: The pathogen is present worldwide, usually in spore form.  The soil is the main source of infection for herbivores.  B. anthracis sporulates with greater frequency in low-lying marshy areas with soil rich in calcium and nitrate and a pH ranging from 5.0-8.0.  Outbreaks are most often associated with neutral or alkaline, calcareous soils that serve as "incubator areas" for the organism.  The spores apparently revert to the vegetative form and multiply to infectious levels in those areas where environmental conditions of soil, moisture, temperature, and nutrition are optimal.

  Cattle, horses, mules, sheep and goats readily become infected when grazing in these areas, primarily in seasons when the minimal daily temperature is over 60°F (16°C).  Epidemics tend to occur following climatic or ecological changes such as heavy rainfall, flooding, or drought; thus, it may occur irregularly, often with many years between occurrences.

  Infection can occur from contaminated soil, water, bone meal, oil cake, tankage, offal, carrion birds, and wild animals.  States within the United States where anthrax occurs include South Dakota, Nebraska, Arkansas, Texas, Mississippi, Louisiana and California; outbreaks and sporadic cases have, however, also occurred in other locations in the United States.  Some regions of the Mississippi and Missouri River valleys harbor spores which are disseminated when flooding occurs.

   In South Africa, non-biting blowflies may contaminate vegetation by depositing vomit droplets after feeding on a carcass infected with B. anthracis.  This contamination is believed to be an important source of infection for browsing animals such as the kudu.  Pigs, dogs, cats and wild animals can acquire the disease from consumption of contaminated meat.

Morphology:  B. anthracis is a gram positive, non-motile, rectangular, aerobic, rod-shaped bacterium with square ends, measuring about 1µ x 3-5µ.  Chain formation is common.  After discharge from an infected animal, or when bacilli from an open carcass are exposed to free oxygen, spores are formed which are resistant to extremes of temperature, chemical disinfectants, and desiccation.  For this reason, the carcass of an animal that died from anthrax should not be necropsied.

Pathogenesis:  In animals, the most common mode of infection is by ingestion.  Infection may also take place via wounds, minor scratches and skin abrasions, and inhalation.  Anthrax is not transmitted horizontally (from animal to animal or human to human).

  The toxins and the capsule are the primary virulence factors of the anthrax bacillus.  Virulent strains harbor two large plasmids: pX02 codes for the capsule and pX01 codes for the exotoxin.  The anthrax toxin is complex, consisting of three protein components: I, II, and III.  Component I is the edema factor (EF), component II is the protective factor (PA) and component III is the lethal factor (LF).  Each component is a thermolabile protein.  EF and LF gain entry into target cells by competitively binding with PA that has a membrane translocation function.  These three components act synergistically to produce the toxic effects seen in anthrax.  Components I and II cause edema with low mortality, but , when component III is included, there is maximum lethality.  Only encapsulated, toxigenic strains are virulent.

  Microorganisms in infected tissue exposed to air sporulate after several hours.  After the spores enter the skin or mucosal membrane, they germinate at the site of entry.  The vegetative cells multiply and are followed by the generation of edema, a papule in 12-36 hours, a vesicle, then a pustule, and finally a necrotic ulcer.  From this lesion there is dissemination to the lymph nodes and finally the bloodstream, resulting in septicemia.  Death is attributed to respiratory failure and anoxia caused by the toxin.  Large numbers of bacilli are shed from the orifices during the terminal stage.

Clinical findings: B. anthracis is an obligate pathogen, the incubation period of which is 3-7 days (ranging from 1-14 days).

  In herbivores, the clinical course ranges from peracute to chronic.  The peracute form is characterized by sudden onset and a rapidly fatal course.  Staggering, dyspnea, trembling, collapse, and a few convulsive movements may occur in cattle, sheep or goats without any previous evidence of illness.

  In the acute form, there is an abrupt rise in body temperature and a period of excitement followed by depression, stupor, respiratory or cardiac distress, staggering, convulsions and death.  The body temperature may rise to 107°F (41.5°C), animals may abort and rumination ceases.  Bloody discharges from natural body orifices can appear.

  Chronic infections are characterized by localized, subcutaneous, edematous swelling, most frequently at the area of the ventral neck, shoulders and thorax.

  In horses, the disease is acute.  Clinical signs may include pyrexia, chills, severe colic, anorexia, depression, weakness, bloody diarrhea, and swelling at the area of the neck, sternum, lower abdomen and external genitalia.  Death usually occurs within 2-3 days of onset.

  In pigs, the disease is usually subacute and may result in pharyngitis with extensive swelling and hemorrhages of the mouth and throat; however, an acute form may occur.  An intestinal form with gastroenteritis also occurs, with non-specific clinical characteristics of anorexia, vomiting, diarrhea or constipation.  Chronic infection with localization in the tonsils and lymph nodes of the cervical region is frequent.

  In dogs and cats, the disease is rare and may resemble the clinical signs seen in pigs.

  Humans develop localized cutaneous lesions called "malignant carbuncle" or "pustule" in more than 90% of cases.  These are the result of contact of broken skin with infected blood or tissues.  The site of infection in this form is most often the face, neck, hands, or arms.  Humans can also acquire a highly fatal hemorrhagic mediastinitis ("woolsorter's disease") from spore inhalation when handling contaminated wool or hair.  Following germination of spores, there is pulmonary necrosis, bacteremia and meningitis.  Ingestion of undercooked meat contaminated with B. anthracis may lead to gastrointestinal anthrax.  Mechanical transmission by blood-feeding insects was also reported, but is of minor importance.

Lesions:  Rigor mortis is frequently absent or incomplete, and thick dark blood that fails to clot may ooze from body orifices.  If the carcass is inadvertently opened, septicemic lesions are often observed.  Hemorrhages frequently occur along the gastrointestinal tract mucosa and on the serosal surfaces of the thorax, abdomen,  pericardium and endocardium.  The spleen is typically enlarged, red-black and soft.  The liver, kidneys, and lymph nodes are usually congested and enlarged.

Diagnosis:  Direct examination: Smears from tissues or blood collected aseptically from superficial vessel and stained by Gram method.  Polychrome methylene blue stain (M' Fadyean's stain) is another useful rapid presumptive diagnostic procedure (with this stain, rods appear blue surrounded by pink capsular material).  One should keep in mind, however, that clostridial organisms are found in the blood shortly after death.  They are not square-ended, lack a capsule, and do not grow aerobically.

Isolation and cultivation: on blood agar plates and incubation at 37°C.  Colonies appear within 24 hours.  When virulent strains are grown in media containing serum or bicarbonate or both, they produce capsules and the colonies appear in 24 hours.  They look flat, gray, are usually non-hemolytic, and smooth to mucoid.  Some are called "medusa-head" or "judge's wig" type colonies since the edge of the colony resembles a tangled mass of curly hair.  In the absence of serum or bicarbonate, bacteria fail to produce capsules and the colonies are rough.

Other methods of identification:  Looking for string-of-pearls-like morphology (growth in the presence of penicillin creates chains of bacteria resembling a strong of pearls) and use of bacteriophage (a gamma phage added to a diffusely inoculated plate is expected to cause lysis only of B. anthracis.

Animal tests: used for confirmation of the diagnosis (B. anthracis is much more pathogenic for guinea pigs and mice than B. cereus and other Bacillus species, causing death within 24 hours.  Large encapsulated rods are demonstrated in smears of spleen and blood of infected animals).

Immunity:  Animals that recover from the infection have permanent immunity to the bacillus.  Protective immunity is thought to be largely antitoxic and ELISA for PA, LF, and EF are used to confirm anthrax infection and monitor antibody responses.

Treatment:  Sick animals should be separated and treated; all healthy animals should be immunized.  The organism is susceptible to many antibiotics.      

Immunization:  Prevention of the disease is attained by annual vaccination of all grazing animals in the endemic area and by implementation of control measures during outbreaks.  The Sterne's vaccine is approved for horses, cattle, sheep and pigs.  It is used almost universally for livestock immunization.  Vaccination should be done 2-4 weeks before the season when outbreaks may be expected.  Animals should not be vaccinated within 2 months of anticipated slaughter.  Since it is a live vaccine, antibiotics should not be administered within one week of vaccination.

  A vaccine consisting of protective antigen from culture filtrate of an avirulent, non-capsulated strain has been used to protect U.S. military personnel and others at risk of infection.  Multiple doses are given and an annual booster is required.

  In addition to therapy and immunization, control of the disease in order to prevent its spread includes 1) notification of the appropriate regulatory offices, 2) rigid enforcement of quarantine, 3) prompt disposal of dead animals, manure, bedding and other contaminated material by cremation or deep burial, 4) isolation of sick animals and removal of healthy animals from the contaminated area, 5) disinfection of stables and equipment, and 6) improved sanitation

Bacterial and spore resistance: B. anthracis may survive for at least 2-3 decades in dried cultures.  The microorganism remains viable in soil for many years.  Freezing temperatures have little, if any, effect on the bacillus.  Spores are destroyed, however, by boiling for 30 minutes and by exposure to dry heat at 140°F (60°C) for 3 hours.  When used, most chemical disinfectants must be employed in high concentrations over a long period of time.  Cremation or deep burial (at least 6 feet or 1.8m) in lime (calcium oxide) is recommended for disposal of the carcasses of animals that  died of the disease.

Public Health Significance:  Anthrax is seen most frequently in farmers, herdsmen, butchers, veterinarians, and in wool, tannery and slaughterhouse workers.  Human infections most often result from spores entering injured skin, leading to cutaneous anthrax in more than 90% of cases.  Pulmonary anthrax resulting from inhalation of spores is almost always fatal.  Failure to diagnose human anthrax correctly and treat it adequately can result in death.

-by Inna Magner, ECFVG Student

-edited by Dr. Ingeborg Langohr, ADDL Graduate student

References:

1. Aiello SE (ed): 1998.  The Merck Veterinary Manual, 8^th ed.  Merck & Co., Philadelphia PA.  pp 432-435

2. Carter GR and Wise DJ: 2004.  Essentials of veterinary bacteriology and mycology, 6^th ed.  Iowa State Press, Ames, IA.  Pp 179-182.

3. Hirsh DC and Zee YC: 1999.  Veterinary Microbiology.  Blackwell Science, Malden, MA.  Pp 246-249.

4. Hungerford TG: 1990.  Diseases of Livestock, 9^th ed.  McGraw-Hill, Sidney.  Pp 329-332.

5. Pipkin AB: 2002.   Anthrax.  In Smith BP (ed) Large Animal Internal Medicine, 3^rd ed.  Mosby, St. Louis, MO.  pp 1074-1076