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Chapter 3 Bacterial zoonoses.

Objectives

After completing this chapter, the learner should be able to

* Describe how living organisms are classified

* Differentiate between prokaryotic and eukaryotic cells

* Describe properties unique to bacteria

* Identify the appearance of bacteria microscopically and by colony growth

* Briefly describe the history of specific bacteria causing zoonotic disease

* Describe the causative agents of specific bacterial zoonoses

* Identify the geographic distribution of specific bacterial zoonoses

* Describe the transmission, clinical signs, and diagnostic procedures of specific bacterial zoonoses

* Describe methods of controlling bacterial zoonoses

* Describe protective measures professionals can take to prevent transmission of bacterial zoonoses

Key Terms
bacteria
binary fission
capnophil
coliform
endotoxin
enterotoxin
eukaryote
exotoxin
facultative
microaerophil
nomenclature
obligate
prokaryote
taxonomy
thermophil
xenodiagnosis


OVERVIEW

Life on earth is incredibly diverse with new species of all types of organisms still being discovered. New technologies and scientific discoveries are allowing organisms to be classified and reclassified based upon new criteria that attempt to identify relationships among living things. Classifying organisms in an orderly fashion allows the scientific community to understand and communicate about a variety of life forms in a clear and concise manner. Describing a rabbit as "the animal with fur and short tail" can cause one person to think of a rabbit, another person thinks of a Manx cat, and yet another person thinks of a grizzly bear. By knowing classification schemes of living organisms, one can clearly understand the roles that various animals and types of microbes play in zoonotic diseases.

THE TAXONOMIC SCHEME

To effectively study living things and their relationships to each other it is necessary to understand the orderly system of classifying organisms known as taxonomy. In the 4th century B.C., Aristotle was the first person to group all organisms as either plants or animals. Although a variety of different classification schemes have been developed since Aristotle's day, the basic rules and nomenclature (naming process) that are used today have been around since the 1700s. In 1735 the Swedish scientist Carolus Linnaeus recognized that confusion would result from having several common names for the same organism. He spent many years giving scientific names to a variety of different organisms. Linnaeus developed the system of classifying organisms, called the taxonomic system, that divides every organism into seven basic categories. These seven categories are kingdom, phylum, class, order, family, genus, and species (some people use eight categories with domain being more diverse than kingdom). The seven levels in this system are arranged into descending ranks with kingdom being the most diverse and species being the most specific.

The Taxonomic Hierarchy

Kingdoms are the largest and most general taxon (category) in the Linnaean system of classification. In 1969, Robert Whittaker of Cornell University divided all life (except viruses) into five kingdoms based on cellular organization and nutritional patterns. Table 3-1 shows these five kingdoms and their characteristics.

Kingdoms contain smaller groups called phyla. Each phylum (singular form of phyla) is divided into several classes; each class is divided into several orders; each order is divided into several families; each family is divided into several genus groups; and each genus is divided into several species. Each sequential division contains more specific descriptions about living things, thus by the time an organism is given its species name little confusion exists about its characteristics. Table 3-2 shows the classification of the house cat, human, and Escherichia coli bacteria.

Nomenclature refers to the naming of organisms. Binomial nomenclature uses two parts for each organism's scientific name. The binomial nomenclature system assigns a genus and species name to each organism and allows scientists to communicate with a common language. Writing scientific names involves capitalizing the genus name and beginning the species name with a lower case letter. Both names are either italicized or underlined.

PROKARYOTIC VERSUS EUKARYOTIC MICROBES

There are two basic types of cells that differ greatly in their size and cellular organization. These two cell types are called prokaryotic and eukaryotic. Prokaryotic cells usually have a single, circular chromosome located in a part of the cytoplasm called the nucleoid (nuclear region). The prokaryotic chromosome is not surrounded by a nuclear membrane. Eukaryotic cells have a membrane-bound nucleus. Table 3-3 shows a comparison of prokaryotic cells and eukaryotic cells.

INTRODUCTION TO BACTERIA

Bacteria were among the earliest life forms on Earth billions of years ago. Many believe that more complex cells developed as free-living bacteria took up residence in other cells, eventually becoming the organelles in eukaryotic cells. Bacteria are often viewed as harmful because they cause human and animal disease; however, certain bacteria are beneficial, such as those that produce antibiotics and those that live symbiotically as normal flora of some body system of animals. Bacteria are extremely important organisms because of their adaptability and capacity for rapid growth and reproduction.

The Kingdom Monera contains bacteria. Bacteria are prokaryotic, unicellular, and reproduce asexually by a process called binary fission. Bacteria usually have one circular chromosome (some bacteria such as Borrelia burgdorferi, the causative agent of Lyme disease, has a linear chromosome) and in some cases small, circular molecules of DNA called plasmids. Table 3-4 and Figure 3-1 summarize other structures found in bacteria.

What Do Bacteria Look Like to the Naked Eye?

Bacteria can be grown outside of the host by providing the nutrients and growth factors needed by that particular organism. Bacteria are routinely grown on blood agar plates (BAP) because this agar supports the growth of most bacteria, but there are other specialized agars that allow for differentiation and selection of certain bacteria as well. When growing bacteria, their appearance on culture media varies depending upon which bacteria they are; however, most appear as colonies on the surface of the agar. Depending upon the bacterium, the colonies may appear mucoid, dry, flat, depressed, pinpoint, small, or a variety of other descriptions. Figure 3-2 shows bacteria growing on a blood agar plate.

Biochemical tests are also available for identifying specific bacteria. These tests, in addition to colony growth, allow for identification of most bacteria.

What Do Bacteria Look Like Under the Microscope?

Bacteria are extremely small (measured in micrometers) and are transparent. In order to view them under the microscope, they need to be stained. The most common stain used is a differential stain called the Gram stain. The Gram stain is based on the fact that some bacteria have a different cell wall structure than others and this difference allows some cells to retain the stain crystal violet. Bacteria described as gram-positive retain the crystal violet stain and appear dark purple, whereas gram-negative bacteria do not retain the crystal violet stain and appear pink. Gram-negative bacteria have lipids in their cell walls that prevent crystal violet stain from entering the cell. When gram-negative bacteria are rinsed with a decolorizer such as acetone, the lipids in their cell walls are removed resulting in open pores. These open pores allow the counterstain safranin to pass through the cell wall. The pink color of safranin is retained by gram-negative bacteria making them appear pink. The Gram stain and a variety of other stains allow visualization of bacteria under the microscope. Figure 3-3 describes the Gram stain process.

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There are thousands of different types of bacteria, but there are basically a few different shapes. Some bacteria are rod-shaped (called bacilli); others are shaped like little spheres (called cocci); others are comma-shaped (called vibrio); others are spiral in shape (called sphirochetes [tight] or spirilla [loose] depending upon the tightness of their spirals). Some bacterial cells exist as individuals, whereas others group together to form pairs (diplo-), chains (strepto-), clusters (staphylo-), or other arrangements. Figures 3-4, 3-5A, and 3-5B illustrate bacterial morphology.

Some bacteria, namely the gram-positive rods Bacillus and Clostridium, have the ability to produce structures called endospores. Endospores, commonly called spores, are protective structures that form when unfavorable conditions are present. Vegetative cells (bacteria that are metabolically active) of the genera Bacillus or Clostridium can form endospores when nutrients are exhausted or other conditions become unfavorable for growth. When an endospore-forming species stops growing, it starts forming endospores and when favorable conditions return, the endospores germinate to produce new vegetative cells (one endospore germinates into only one vegetative cell). Endospores are extremely resistant to drying, heat, radiation, and chemicals (such as alcohols and bleach). Endospores can be seen microscopically and can form either centrally, subterminally (near one end), or terminally (at one end). Figure 3-6 illustrates endospore formation.

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How Do Bacteria Reproduce?

Prokaryotic cell reproduction is quite simple compared to eukaryotic cell reproduction. Prokaryotic cells reproduce by binary fission, where one cell (the parent cell) splits in half to become two daughter cells identical to the parent. Prior to replication the chromosome is duplicated so that each daughter cell possesses the same genetic information as the parent.

The time it takes binary fission to occur is called the generation time. The generation time varies from one bacterial species to another and is also dependent upon growing conditions such as pH, temperature, and availability of nutrients. Under ideal conditions some bacteria can replicate every 20 minutes.

How Do Bacteria Obtain Nutrients?

Chemical nutrients such as carbon, hydrogen, oxygen, and nitrogen are needed for bacterial growth. Bacteria obtain these nutrients from a variety of environmental sources. Bacteria that use inorganic carbon source (CO2) as their sole carbon source are called autotrophs. Bacteria that break down organic molecules (such as proteins, carbohydrates, amino acids, and fatty acids) are heterotrophs and acquire these nutrients from other organisms.

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Some bacteria make their own food from sunlight. Other bacteria absorb food from the material they live on or in. Some of these bacteria produce by-products such as iron or sulfur. The microbes that live in the gastrointestinal tract absorb nutrients from digested food.

How Do Bacteria Cause Infection?

Not all bacteria cause disease. Some bacteria such as normal flora bacteria are needed by the host to maintain health of a body system. Colonization is the persistence of microbes in a body site without causing disease. Normal flora is an example of colonization. Normal flora and the host are an example of a symbiotic relationship that is mutualistic (both organisms benefit) or commensal (no harm is done to the host) rather than parasitic.

Microbes capable of causing disease are pathogens. Microbial pathogenicity is the biochemical mechanism by which microbes are able to cause disease. Infection is like a miniature battle between bacteria and the host; the bacteria are trying to remain present and multiply, whereas the host is trying to prevent bacteria from gaining access. The term infection is used to describe the persistence or multiplication of a pathogen (disease-causing organism) on or within the host. Disease occurs when an infection causes clinical signs in the host. Infection and disease are dependent on host factors as well as virulence of the microbe.

Pathogenic bacteria have certain virulence factors that help them cause disease. The first step in pathogenesis of bacterial infection is for the bacteria to reach the site of interest and to remain there. Some ways bacteria can get to and remain in an area are with the aid of fimbria and flagella. Fimbria are hair-like structures on bacteria that enable them to attach to certain body sites and help them not get washed away by body secretions. For example, E. coli produces fimbria that attach to the epithelial lining of the urinary tract allowing the bacteria to remain in the urinary bladder without getting flushed away. Flagella are structures on bacteria that help propel them from one area to another. The flagella help bacteria reach a body site where they can survive and multiply; therefore, flagella have a function in pathogenicity.

Invading microorganisms must also survive phagocytosis in order to cause disease. Some pathogens avoid direct contact with a phagocyte by producing a slippery mucoid capsule. This capsule inhibits the chemical identification of bacteria by the phagocyte. If the pathogen is not recognized as foreign by phagocytes, it has time to replicate in the body and cause disease. Pathogens that produce the thickest capsules are some of the most virulent.

The next challenge for pathogens is to compete with normal flora. One way bacteria compete with normal flora is by producing toxic compounds that cause harm to their host producing clinical signs such as vomiting, diarrhea, paralysis, pain, or fever. Some bacteria produce toxins wherever they grow and can cause illness even without being in the affected area as is the case with some forms of food poisoning. Another way bacteria compete with normal flora is by invading the host cells especially in areas such as the gastrointestinal tract and other body systems with increased numbers and varieties of normal flora. For example, Salmonella enterica serovar Typhimurium (commonly called S. typhimurium) will destroy the cells of the intestine causing them to release their cell contents that are used by bacteria as nutrients. The result in the host is diarrhea due to intestinal cell damage that alters absorption of fluid and nutrients, against the result for the pathogen is being lost from the intestine with the stool.

ANTHRAX

Overview

Anthrax, also known as black bane, fifth plague, and woolsorter's disease, has been a documented disease since biblical times as described in the Old Testament. The very severe plague cast upon the Pharaoh's cattle in the book of Exodus was believed to be anthrax, especially because the endospores that are produced by this bacterium can persist for many years in soil similar to that found in the Nile valley. In the 19th century, anthrax was known in Europe as woolsorter's and ragpicker's disease because these groups of people caught the endospores from the fibers and hides they were handling. The first cases of anthrax in the United States were reported in Louisiana in the early 1700s.

Anthrax, named directly from the Greek word anthrax meaning coal, is named after the cutaneous black "coal-like" lesions it can cause. Anthrax is caused by inhalation, skin exposure, and gastrointestinal absorption and is primarily a disease of herbivores with humans contracting the disease from infected animals. Anthrax is caused by Bacillus anthracis, which was the first bacterium shown to cause disease. In 1877, Robert Koch, a Polish scientist, helped launch the science of bacteriology when he became interested in this disease after several local cows died suddenly without a true explanation as to the cause of death. Koch believed that B. anthracis was the cause of the outbreak, but he did not know how the bacterium was transmitted and the mechanism by which it caused disease. Koch began his investigation by studying blood from dead cattle and found numerous rods and threads in it that did not appear in healthy cattle blood. He then used blood from cattle that had died of anthrax and applied the blood to open cuts on mice. Each mouse that came in contact with the infected blood died. To determine the impact of B. anthracis inside an animal, Koch cultured parts of swollen spleens from the infected mice and liquid from inside infected cattle eyes and watched as bacteria grew and formed the unusual rods and threads in his laboratory. He then put the samples in water and sunlight, and found that if placed in ordinary water, the rods and threads separated and disappeared. In sunlight only, the rods and threads were killed. These observations led Koch to believe that the bacterium must remain in the body to maintain its virulence.

As anthrax spread through sheep flocks in France in 1877, Louis Pasteur worked on a vaccination for this disease and by 1881 a successful anthrax vaccine for livestock was developed. Further preventative measures against anthrax were strengthened by the 1920s law that required testing of shaving brushes that were made of horse or pig bristles.
The risk of getting anthrax remains
low in developed countries with
modern animal husbandry and
industrial hygiene.


As a result of the ability of B. anthracis to produce endospores, anthrax has long been studied as a potential biological weapon. In 1925, the Geneva Protocol banned bacteriologic warfare as part of the World War I peace treaties with ratification occurring prior to World War II (except by the United States and Japan). A resolution calling on all United Nations members to ratify the ban was rejected in 1952 and all major nations maintained extensive facilities for producing and testing germ warfare (including anthrax). In 1969, a secret military test range in Utah accidentally released nerve gas killing hundreds of sheep, which lead President Nixon to renounce chemical and biological weapons in the United States. In 1975, the United States finally ratified the international ban on chemical and biological weapons. In 2001, bioterrorist-related cases of cutaneous and inhalation anthrax were seen in the United States prompting the desire for vaccination against the disease and for prophylactic antibiotics.

Causative Agent

Anthrax is an acute infectious disease caused by B. anthracis bacteria. B. anthracis are very large, nonmotile, gram-positive, encapsulated, endospore-forming bacilli. B. anthracis has two main virulence factors: the presence of a capsule and production of toxins.

* The capsule helps the bacillus avoid engulfment by phagocytes, thus allowing for establishment of infection in the animal/person. All virulent strains of B. anthracis form a capsule and are known as S or smooth variants. S variants produce mucoid or "smooth" colonies when grown on agar. R or rough variants do not produce the capsule and are relatively avirulent. The ability to produce capsules can be transferred to nonencapsulated B. anthracis via plasmid transfer.

* Anthrax toxins consist of three types including:

** Protective antigen (PA), also known as factor II, is a protein that binds to select cell receptors in the target tissue, which in turn forms a channel that permits the other factors to enter those cells.

* Edema factor (EF), also known as factor I, is a toxin that converts adenosine triphosphate (ATP) to cyclic adenosine monophosphate (cAMP). As cAMP increases cellular edema occurs in the target tissue. Build-up of fluid surrounding the lungs can inhibit immune function and can be fatal.

* Lethal factor (LF), also known as factor III, is a toxin that is believed to inhibit phagocytosis by neutrophils and release cytokines. LF can kill infected cells or prevent them from working properly.

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B. anthracis has the ability to form endospores and these endospores form in the middle of the cells and may persist for long periods in dry products such as feed, contaminated objects, or in soil. The endospores revert to the vegetative (reproductive) form when environmental conditions are optimal, including warmer seasons when temperature is above 60[degrees]F and there is heavy rainfall. Flooding allows bacteria to accumulate at the ground surface of low-lying areas and drought conditions favor the development of endospores. B. anthracis is typically found in neutral to mildly alkaline soil (pH 6 to 8.5). Figure 3-7 shows B. anthracis endospores.

Epizootiology and Public Health Significance

Anthrax occurs worldwide and is seen most commonly in agricultural regions without adequate veterinary public health programs such as South and Central America, Southern and Eastern Europe, Asia, Africa, and the Middle East. International figures are not exact because of difficulty in reporting cases in developing nations. Anthrax is endemic in Africa and Asia.

In animals, anthrax occurs sporadically throughout the United States mainly in cattle, bison, and deer herds. In humans, the incidence of anthrax in the indigenous United States averages less than one case per year. From 1955 to 1994, there were 235 human cases of anthrax in the United States (average 5 per year) with 20 of them being fatal. In the United States there are recognized areas of infection in South Dakota, Nebraska, Arkansas, Mississippi, Louisiana, Texas, and California; small areas exist in a number of other states. Anthrax is rarely seen in the United States and when seen occurs most frequently in farmers, herdsmen, butchers, veterinarians, and in wool, tannery, and slaughterhouse workers. U.S. cases of human anthrax were between 40 and 50 in 1952, but since 1962 have numbered less than 10 per year (except for the bioterrorism-related cases in 2001 and 2002). During September 2001 through November 2001, an outbreak of intentionally spread anthrax in the United States caused five deaths and a total of 22 infections (18 confirmed and 4 suspicious). Anthrax is identified by the Centers for Disease Control and Prevention (CDC) as being capable of causing death and disease in large enough numbers to devastate cities or developed areas. According to a World Health Organization (WHO) estimate, the release of 50 kilograms of anthrax over a city of 5 million people would result in 250,000 deaths, with 100,000 patients dying before receiving treatment.

Preventing and treating anthrax includes vaccination and the use of antibiotics. The U.S. Department of Defense has given more than 2 million anthrax vaccinations to more than 500,000 military personnel. The cost of the vaccine is approximately $18 for a complete immunization series (at $3 per dose). The vaccine cost is higher if it is given by individual clinicians rather than as part of mass public vaccinations. Anthrax treatment using ciprofloxacin for 60 days costs about $700 (2005 values).
Anthrax is typically a disease of
herbivores.


Transmission

B. anthracis is a facultative (capable of adapting to different conditions) organism whose cycle of vegetative growth and endospore formation occur in soil. Infection with B. anthracis occurs when animals grazing on contaminated pasture ingest endospores. The pathogenic bacillus is returned to the soil in animal excrement or when the animals die. Once in the soil the bacterium sporulates and the soil remains a long-term reservoir of infection for animals.

B. anthracis is typically transmitted between animals via ingestion of endospore-contaminated water or pasture in areas where previously infected animals lived. Ingestion of infected feedstuffs, such as bloodmeal or bonemeal, has also been implicated as a cause of infection. During an epidemic, insect transmission of disease may occur, but is not usually significant. Some animals, such as pigs, dogs, cats, mink, and captive wild animals, have acquired the disease from consumption of contaminated meat.

B. anthracis is typically transmitted to people by infected animals and their products (fur, skin, bonemeal). B. anthracis most frequently enters the body via skin abrasions, injuries, or blemishes resulting in cutaneous anthrax. Airborne infection (via shearing of infected sheep or handling of infected hides) and gastrointestinal infection (via consumption of contaminated meat or milk) can also lead to infection.

Pathogenesis

B. anthracis owes its pathogenicity to the formation of a capsule and production of toxins. The capsule protects the bacillus from bactericidal components in blood and prevents phagocytosis of the organism. The capsule plays an important role in establishment of infection because the organism is allowed to increase in number as a result of the slowing of phagocytosis by neutrophils. Toxin production plays an important role both in establishment of infection and in the later stages of disease where the toxins are responsible for the clinical signs observed in infected animals. Toxins produced by B. anthracis can have both short antiphagocytic activity and leukocidal effects on white blood cells. The PA toxin binds to cell receptors in target tissue causing exposure of a binding site. This exposed binding site combines with either EF to form edema toxin or binds with LF to form lethal toxin. Edema toxin causes cellular edema in target tissue, whereas lethal toxin may inhibit phagocytosis and may cause release of tumor necrosis factor and interleukin-1.

Once anthrax bacilli are contracted, they multiply at the site of the lesion. Phagocytic cells migrate to the site, but are unable to engulf bacilli as a result of the presence of their capsules. If some bacilli are engulfed they can resist killing and digestion by producing toxins that impair phagocytic activity and can be lethal to leukocytes. Depending upon the port of entry, bacteria and their toxins cause cutaneous, pulmonary, or gastrointestinal disease. Systemic anthrax results from the spread of these forms.

There is considerable variation in susceptibility to anthrax among animal species. The infectious dose of anthrax also varies widely among animal species with mice only requiring five bacteria to initiate disease, whereas rats require 106 bacteria to cause disease. This variation among animal species may be the result of a particular species resistance to the toxin produced or to the ability of the bacterium to establish disease in that species. Figure 3-8 depicts the pathogenesis of anthrax in humans.

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Clinical Signs in Animals

In animals there are three forms of anthrax including:

* Peracute form. The peracute form of anthrax occurs mainly in ruminants especially cattle, sheep, and goats. Signs in affected animals may include ataxia, dyspnea (difficulty breathing), trembling, and seizures. Sometimes sudden death is the only sign observed in these animals (Figure 3-9).

* Acute form. In cattle and sheep the first sign of acute anthrax may be high fever and excitement followed by stupor, respiratory depression, cardiac depression, seizures, and death. Milk production is reduced and abortion may be seen in pregnant animals. Animals may also hemorrhage from the mouth, nose, and anus. Splenomegaly may be evident and may appear as blackberry jam on necropsy. The acute form of anthrax in swine may present as sudden death or may present as progressive swelling of the throat, which may cause suffocation. In horses, the acute form of anthrax presents as fever, chills, severe colic, anorexia, muscular weakness, bloody diarrhea, and swelling in the neck, lower abdomen, and genital area.

* Chronic form. The chronic form of anthrax in ruminants produces localized, subcutaneous edema typically in the neck, thorax, and shoulders. These lesions are caused by bacteremia and are not analogous to human cutaneous lesions. In swine, lingual and pharyngeal edema and hemorrhage of the pharyngeal and cervical lymph nodes are typically seen.
In animals, anthrax typically presents
as an acute fatal septicemia with
splenomegaly and hemorrhagic
infiltration of the subcutaneous tissue.


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Clinical Signs in Humans
There are no reported cases of human-to-human
transmission of anthrax.


Humans contract B. anthracis mainly from infected animals and their products such as fur, skin, bloodmeal, or bonemeal. The symptoms of anthrax vary with the route of infection. There are several different forms of anthrax in humans including:

* Cutaneous anthrax. Cutaneous anthrax is the most common form of anthrax and occurs in about 95% of cases. Cutaneous anthrax has an incubation of 2 to 5 days. Cutaneous anthrax occurs when B. anthracis endospores enter nonintact skin through a laceration or abrasion. The endospore germinates in macrophages of the skin and toxin production results in small, red, raised skin lesions (Figure 3-10). The skin lesions enlarge and ulcerate or may become fluid filled. By 7 days these lesions develop into painless, black eschars (dark, sloughing scab) surrounded by edema and blood vessels. The eschars dry and fall off in about 1 to 2 weeks. If left untreated, the infection can become systemic producing symptoms of fever, lethargy, vomiting, and hypotension. If the portal of entry of the bacterium is on the neck or face and if septicemia or meningitis develops, the prognosis is poor. Mortality from untreated cutaneous anthrax is 10% to 20%; in treated cases mortality is about 1%.

* Pulmonary (inhalation) anthrax. Pulmonary anthrax is rare in the United States and was historically associated with woolsorters at industrial mills. Pulmonary anthrax has an incubation of 1 to 14 days and typically develops in the textile and tanning industries among workers handling contaminated animal wool, hair, and hides. More recent cases of pulmonary anthrax in the United States are associated with biological warfare. Pulmonary anthrax occurs when aerosolized B. anthracis endospores (8,000 to 50,000 endospores needed to initiate disease) enter the respiratory tract via inhalation and deposit in the alveolar space. Macrophages will lyse and destroy some endospores and infection in the lung is rare. Surviving endospores reach the mediastinal and peribronchial lymph nodes where germination may occur up to 60 days postinfection. After germination the anthrax bacilli replicate in the lymph nodes, release toxins, and hemorrhage, edema, and necrosis occurs. Inhalation anthrax produces biphasic clinical symptoms. In the first stage of the disease symptoms resemble those of a cold (fever, dyspnea, cough, headache, chills). As endospores travel to the lungs (secondary stage), symptoms such as dyspnea, sweating, shock, cyanosis, and hypotension may appear (Figure 3-11). Rapid death may occur in 1 to 3 days following the onset of symptoms. Mortality rates without treatment are greater than 95%, and even humans treated with antibiotics rarely survive unless they are treated immediately after the onset of symptoms.

* Gastrointestinal anthrax. Gastrointestinal anthrax is associated with ingestion of contaminated meat and produces two distinct syndromes (oral-pharyngeal and abdominal). When B. anthracis endospores are deposited in the upper gastrointestinal tract, they germinate producing localized lymphadenopathy, edema, and sepsis following development of an oral or esophageal ulcer. Dysphagia (difficulty eating/swallowing) and dyspnea may also occur. When B. anthracis endospores are deposited in the lower gastrointestinal tract, they germinate producing primary intestinal lesions. Symptoms include nausea, abdominal pain, vomiting, and lethargy that may progress to bloody diarrhea or sepsis. The abdominal form of gastrointestinal anthrax is more common than the oral-pharyngeal form. Gastrointestinal anthrax is rare; however, cases have been reported in Asia and Africa.

* Systemic anthrax. Systemic infections caused by B. anthracis arise from hematogenous spread of bacteria from the cutaneous, pulmonary, and gastrointestinal forms. Spread from the cutaneous route is most common. Systemic infection usually produces meningitis; however, spread of disease from primary infection sites is rare. Symptoms of meningitis include fever, fatigue, neck pain, nausea, behavioral changes (such as agitation) and neurologic signs (Figure 3-12). Systemic anthrax is usually fatal.

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Diagnosis in Animals

The carcass of any animal suspected of having anthrax should not be necropsied because the anthrax bacilli form endospores when exposed to oxygen. Rigor mortis in these animals is typically absent or incomplete.

Diagnosis of anthrax in animals via clinical signs is difficult; therefore, laboratory confirmation is needed. Blood collected from suspect animals should be shipped to reference laboratories in leak-proof containers or as a dried specimen on either a sterile swab or dried blood smear. Tissue samples should be shipped refrigerated or frozen in leak-proof containers. All specimens shipped should be labeled as "suspected anthrax" so that proper biosafety handling measures can be followed. Common laboratory methods for identifying B. anthracis include stained blood smears, bacterial culture, animal inoculation, and serologic tests. Blood smears are typically stained with Gram stain, polychrome methylene blue stain (also called McFadyean's stain; a rapid stain that stains rods blue surrounded by pink capsular material), or Giemsa stain (demonstrates the encapsulated bacillus). Organisms stain gram-positive when stained from young cultures, but can stain gram-variable or gram-negative with age. On Gram stain endospores may be seen and appear clear because they do not retain the crystal violet or safranin stain. Endospores will stain green if the sample is stained with a malachite green spore stain using heat; the vegetative cell will appear red when counterstained with safranin.

B. anthracis bacteria can be cultured using routine media (blood agar, chocolate agar) under aerobic or anaerobic conditions and special media such as PLET (polymixin-lysozyme EDTA-thallous acetate) and bicarbonate agar. On blood agar, B. anthracis grows as nonhemolytic, medium-large, gray, flat, irregular colonies with swirling, comma-shaped projections. B. anthracis is differentiated from other gram-positive rods on culture by its lack of hemolysis and motility and by growth on phenylethyl alcohol blood agar.

Guinea pigs and mice are occasionally used to diagnose cases of anthrax by inoculation of blood or tissue from an anthrax suspect (xenodiagnosis). B. anthracis is much more pathogenic for guinea pigs and mice than other Bacillus species typically causing death within 24 hours. If the animal has died within 48 hours following inoculation, blood smears and touch preps of splenic tissue can demonstrate the characteristic large, encapsulated bacilli.

Serologic tests available for identification of B. anthracis include enzyme-linked immunoabsorbant assay (ELISA) or indirect hemagglutination antibody (IHA) tests that identify titers of antibodies directed against the capsular antigen, protective antigen, lethal factor, or edema factor (quadrupling of the titer is diagnostic). Fluorescent antibody (FA) is also available for identification of the bacterium in tissue and culture. Polymerase chain reaction (PCR) methods that amplify markers of B. anthracis have also been developed.

Diagnosis in Humans

Anthrax in people is diagnosed differently depending on the form of anthrax. Cutaneous anthrax is diagnosed when exudate from skin lesions is swabbed or aspirated and the fluid stained or cultured. Skin biopsy and immunohistochemical staining is also used to diagnose cutaneous anthrax. Pulmonary anthrax is diagnosed by radiographic lesions of the thorax that show symmetrical mediastinal widening and by computed tomography (CT) scans that show enlarged hilar lymph nodes, pleural effusion, and airway edema. Confirmation of pulmonary anthrax is by staining and culturing of sputum specimens. Gastrointestinal anthrax is diagnosed by staining and culture of fecal samples, vomitus, or hemorrhagic fluid from body cavities. Serologic testing of samples for anthrax confirmation includes PCR, ELISA, IHA, and FA. In some countries an anthraxin skin test is used to confirm cases of anthrax. The anthraxin skin test involves subdermal injection of commercially produced chemical extract of an attenuated strain of B. anthracis. A positive test indicates cell-mediated immunity. The accuracy of positive test results increases with the duration of the disease.

Treatment in Animals

Anthrax is highly fatal requiring early and vigorous treatment. All sick animals should be isolated and treated with antibiotics (the bacterium is susceptible to many antibiotics including oxytetracycline, erythromycin, and sulfonamides) and all healthy animals in the herd and on surrounding farms should be immunized. In addition to antibiotic treatment and vaccination, controlling the spread of anthrax includes the following:

* notification of the appropriate regulatory offices

* enforcement of quarantine

* prompt disposal of dead animals, manure, bedding, and other contaminated material by cremation or deep burial

* isolation of sick animals and removal of healthy animals from the contaminated area

* disinfection of stables and equipment

* improved sanitation

Treatment in Humans

Prior to October 2001, the treatment and prophylaxis for human cases of anthrax was penicillin; however, concern for genetically engineered penicillin-resistant anthrax strains prompted the CDC to recommend the use of other antibiotics. For patients with severe cases of anthrax, corticosteroid and intravenous (IV) antibiotic treatment is recommended. Pulmonary anthrax patients typically received a multidrug regimen of either ciprofloxacin or doxycycline (doxycycline is not used in patients with meningitis as a result of poor drug penetration to the central nervous system [CNS]) for 60 days and another antibiotic such as rifampin, vancomycin, or an aminoglycoside. Cases of gastrointestinal and cutaneous anthrax are treated with ciprofloxacin or doxycycline for 60 days followed by amoxicillin or amoxicillin/clavulanic acid. Despite early treatment, people infected with pulmonary, gastrointestinal, or meningeal anthrax have a very poor prognosis.

Management and Control in Animals

Prevention of anthrax is attained by annual vaccination of all grazing animals in an endemic area and by implementation of control measures during outbreaks. Vaccines for anthrax composed of killed bacilli and/or capsular antigens do not produce significant immunity; therefore, the current vaccine is a live vaccine. The Sterne's vaccine, which uses the Sterne strain of B. anthracis, produces sublethal amounts of toxin allowing for antibody production in animals and is approved for horses, cattle, sheep, and pigs. Vaccination should be done 2 to 4 weeks before the season when outbreaks may be expected in the area. Animals should not be vaccinated within 2 months of anticipated slaughter. Because it is a live vaccine, antibiotics should not be administered within 1 week of vaccination. Animals surviving naturally-acquired anthrax are immune to reinfection and second attacks are extremely rare. Permanent immunity to anthrax appears to require antibodies to both the toxin and capsule.

B. anthracis may survive for 20 to 30 years in dried cultures and remains viable in soil for many years. Freezing temperatures have little effect on the bacillus; however, endospores can be destroyed by autoclaving at standard conditions, by boiling for 30 minutes, or by exposure to dry heat at 140[degrees]F (60[degrees]C) for 3 hours. Most chemical disinfectants must be used in high concentrations over a long period of time to be effective. Cremation or deep burial (at least 6 feet or 1.8 meters) in lime (calcium oxide) is recommended for disposal of the carcasses of animals that died of anthrax.

Management and Control in Humans
Anthrax is a reportable disease.


A vaccine consisting of protective antigen of an avirulent, noncapsulated strain of B. anthracis has been used to protect U.S. military personnel and others at risk of infection such as people who work with imported animals hides, furs, bone, meat, wool, animal hair, and equipment used in grooming these animals. Multiple doses are given (three subcutaneous injections given 2 weeks apart followed by three additional subcutaneous infections given at 6, 12, and 18 months) and an annual booster is required to maintain protective immunity. The vaccine should only be used in healthy people 18 to 65 years of age and its safety in pregnant women has not been established. Passive vaccines that deliver antibody directed against protective antigen are being investigated.

Summary

B. anthracis, the causative agent of anthrax, is a rod-shaped, gram-positive, endospore-forming bacterium that typically infects herbivores such as cattle, sheep, and horses. Human disease may be contracted by handling contaminated hair, wool, hides, flesh, blood, and excretions of infected animals and from manufactured products such as bonemeal, as well as by purposeful dissemination of endospores. Infection is introduced through scratches or abrasions of the skin, wounds, inhalation of endospores, eating insufficiently cooked contaminated meat, or by flies. B. anthracis endospores are very stable and may remain viable for many years in soil and water. Anthrax endospores were weaponized by the United States in the 1950s and 1960s prior to the termination of the old U.S. offensive program. B. anthracis is easy to cultivate and endospore production is readily induced with endospores being highly resistant to sunlight, heat, and disinfectants.

Anthrax presents as three clinical syndromes in animals: peracute, acute, and chronic. The peracute form of anthrax occurs mainly in ruminants, especially cattle, sheep, and goats, with signs including ataxia, dyspnea, trembling, seizures, and sudden death. The acute form of anthrax occurs in cattle and sheep and presents with high fever and excitement followed by stupor, respiratory depression, cardiac depression, seizures, and death. Animals may also hemorrhage from the mouth, nose, and anus. Splenomegaly may be evident and may appear as blackberry jam on necropsy. The acute form of anthrax in swine may present as sudden death or may present as progressive swelling of the throat, which may cause suffocation. In horses, the acute form of anthrax presents as fever, chills, severe colic, anorexia, muscular weakness, bloody diarrhea, and swelling in the neck, lower abdomen, and genital area. The chronic form in ruminants produces localized, subcutaneous edema typically in the neck, thorax, and shoulders. The chronic form of anthrax in swine presents with lingual and pharyngeal edema and hemorrhage of the pharyngeal and cervical lymph nodes.

Anthrax presents as three distinct clinical syndromes in humans: cutaneous, pulmonary, and gastrointestinal disease. The cutaneous form occurs most frequently on the hands and forearms of persons working with infected livestock beginning as a papule followed by formation of a blister-like fluid-filled vesicle. The vesicle dries and forms a coal-black scab. Pulmonary anthrax, known as woolsorter's disease, is a rare infection contracted by inhalation of the endospores. It occurs mainly among workers handling infected hides, wool, and furs. Gastrointestinal anthrax is contracted by the ingestion of insufficiently cooked meat from infected animals. In humans, the mortality of untreated cutaneous anthrax ranges up to 25% and in pulmonary and gastrointestinal anthrax, the fatality rate is almost 100%.

Diagnosis of anthrax in animals via clinical signs is difficult; therefore, laboratory confirmation is needed. Common laboratory methods for identifying B. anthracis include stained blood smears, bacterial culture, animal inoculation, and serologic tests. Diagnosis of anthrax in humans includes blood smears, bacterial culture, serologic tests, and clinical tests such as thoracic radiographs (for identification of pulmonary anthrax). Treatment of anthrax in both animals and humans includes antibiotics that need to be administered early in the disease course to be successful. Prevention of anthrax includes vaccination of animals using the Sterne's vaccine and selective vaccination of high-risk humans with vaccine developed from attenuated strains of B. anthracis.

BITE WOUNDS

Overview

Approximately half of the U.S. population will incur an animal bite sometime in their life. Every year, about 330,000 people are seen in emergency rooms in the United States for dog bites; about 4% of these cases are hospitalized and approximately 20 deaths are estimated to occur in the United States from animal bites. Dog bites compose approximately 80% of animal bite wounds and are typically seen in children (peak incidence is between the ages of 5 and 9 years). Dog bites typically occur on the extremities and an estimated 4% to 25% of dog bite wounds become infected. Cat bites occur more commonly in women with half of all victims of cat bites older than 20 years of age. Cat bites also typically occur on the extremities. Because cats have thin, sharp teeth they tend to cause puncture wounds (about 85% are puncture wounds). Approximately 30% to 50% of cat bite wounds become infected. The first symptoms associated with cat bite wounds typically occur within 12 hours following the bite. The first symptoms associated with dog bite wounds typically occur approximately 24 hours following the bite. The focus in this section will be dog and cat bite wounds.
A large dog can exert more than
450 psi of pressure with its jaws,
causing significant injury and tissue
devitalization to a bite area.


Causative Agent

The microbes causing infection following dog and cat bites may consist of normal flora from the animals' mouths or from the skin of the person/animal being bitten. At least 30 different infectious agents have been reported to be transmitted from dog or cat bites and most infections as a result of dog and cat bites consist of multiple bacteria (a median of five isolates per infected wound). About 50% of dog bites and 60% of cat bites involve both aerobic and anaerobic bacteria. Bacteria found in bite wounds include:

* Pasteurella spp. are the most common pathogens found in dog and cat bite wounds (typically P. multocida and P. canis). Wounds containing Pasteurella bacteria tend to show signs of infection more rapidly than wounds containing other bacteria. Pasteurella bacteria are commonly found in abscesses and non-purulent wounds. P. multocida is normal oral flora in 50% to 70% of healthy cats and is found in about 45% of all cat bite wounds. Other animals that transmit Pasteurella bacteria through their bites are dogs, horses, sheep, and pigs.

* Staphylococcus aureus bacteria are commonly isolated from nonpurulent wounds. Animals that transmit St. aureus bacteria through bite wounds are dogs, cats, horses, camels, pigs, lizards, and rodents.

* Streptococcus mitis bacteria are commonly isolated from nonpurulent wounds. Animals that transmit Str. mitis are dogs and cats; other species of Streptococcus are spread through the bites of horses, camels, pigs, simians, squirrels, and birds.

* Moraxella spp., Corynebacterium spp., and Neisseria spp. are commonly isolated aerobic bacteria from the bite wounds of dogs, cats, horses, simians, rodents, and squirrels.

* Bergeyella zoohelcum (formerly known as Weeksella zoohelcum) is an uncommon zoonotic pathogen that causes acute cellulitis from dog and cat bites.

* Capnocytophaga canimorsus and C. cynodegmi are normal flora of the canine and feline mouth. These bacteria may cause local wound infections after a dog or cat bite and may lead to sepsis, meningitis, and disseminated coagulopathy. Capnocytophaga bacteria are also associated with rabbit bites.

* Fusobacterium spp., Bacteroides spp., Porphyromonas spp., and Prevotella spp. are anaerobic bacteria cultured from dog, cat, horse, camel, simian, rodent, bird, and reptile wounds, but they are rarely cultured alone.

Epizootiology and Public Health Significance

Animal bite wounds are seen throughout the world and are common in the United States (about 1% of all emergency room visits are related to animal bites). In the United States an estimated 1 to 3 million animal bites occur annually with approximately 80% to 90% from dogs, 5% to 15% from cats, and 2% to 5% from rodents (the remainder coming from other small animals, such as rabbits and ferrets, farm animals, monkeys, and reptiles). Internationally, it is difficult to get accurate numbers as a result of the variety of animals inflicting animal bites including large cats (tigers, lions, and leopards), wild dogs, hyenas, wolves, crocodiles, and other reptiles. In England and Wales, 200,000 people seek medical care for dog bites; in France, 500,000 people seek medical care for dog bites; and in Germany, 35,000 people are bitten by dogs. Most bites seen worldwide are from domestic dogs. Young children tend to get bitten by dogs, whereas adult women are more frequently bitten by cats. Other groups of people at risk include veterinary professionals, animal keepers, breeders, and trainers. In developing countries, dog bites carry a high risk of rabies infection.

Animal bites result in approximately 0.4% to 1.5% of all emergency room visits with an annual cost of approximately $100 to $165 million dollars in health care expenses and lost income. Most bites are the result of a family's own pet or a neighbor's animal. Because 58% of all households in the United States having at least one pet, the potential public health significance of animal bites is enormous.
Capnocytophaga spp. and Bergeyella
zoohelcum bacteria can be transmitted
to people through close contact with
animals as well as through a bite
wound.


Transmission

Most of these organisms are transmitted by bite wounds, but can also be transmitted through close animal contact such as the animal licking an open wound of a person/animal (especially immunocompromised individuals).

Pathogenesis

The pathogenesis of bite wounds is variable depending on the type of animal that produces the bite. Dog bites typically cause crushing-type wounds, because their rounded teeth and strong jaws cause injury to deeper tissues such as bones, vessels, tendons, muscle, and nerves. The pointed teeth of cats usually cause puncture wounds and lacerations that may inoculate bacteria into deep tissues. Other animals such as monkeys and herbivores can cause injury from bite wounds as a result of the trauma they cause followed by infection.
In general, the better the vascular
supply and the easier the wound is to
clean (i.e., laceration versus puncture),
the lower the risk of infection. Bites on
the hand have a high risk for developing
infection because of the relatively poor
blood supply in the hand and difficulty
of adequately cleansing the wound.


Wound infections from dog and cat bites can cause abscess formation, septic arthritis, osteomyelitis, endocarditis, and CNS infections. The incubation period from animal bites varies with the animal producing the bite, the organism or organisms causing the infection, and health factors of the person/animal being bitten. Patients who present within 8 to 12 hours of a bite typically show local lesions without significant signs of local inflammation. Those patients that present after 12 hours may present with localized cellulitis, pain at the bite site, discharge, and enlarged lymph nodes. If septicemia develops, signs include fever, chills, vomiting, diarrhea, abdominal pain, lethargy, dyspnea, and headache. Immunocompromised people (splenectomized people, alcoholics, people on corticosteroids or chemotherapy) may develop more severe signs associated with animal bites and signs may include endocarditis, pneumonia, meningitis, peripheral gangrene, and shock.

Clinical Signs in Animals

Bite wounds inflicted from one animal to another are usually caused by fighting.

* Cats. Intact male cats fight more than neutered male cats, which fight more than female cats. Feline fight wounds typically occur on the face, legs, back, tail, and rump. The most common complication of fight wounds is infection because cat bites create small puncture wounds in the skin that quickly close and are difficult to find in affected animals. Microbes from the biting cat's mouth enter the other animal's skin and multiply rapidly. An abscess will form if loose skin surrounds the bite site. Cellulitis may develop on less fleshy areas, such as the foot or tail (Figure 3-13). Both abscesses and cellulitis trap pus, causing swelling and pain. If a cat is bitten by a cat with feline leukemia virus (FeLV) or feline immunodeficiency virus (FIV), it could contract these viruses. Transmission of rabies is also a concern when any animal is bitten by a cat.

* Dogs. Intact male dogs fight more than neutered male dogs. Female dogs tend to fight with other female dogs. Dog bite wounds tend to cause more trauma from ripping or damaging of tissue followed by infection once the microbes from the oral cavity of the dog enter the wound and multiply. Abscesses and cellulitis are common findings following dog bite wounds. If a dog is bitten by an animal with rabies virus it could contract this illness.

[FIGURE 3-13 OMITTED]

Clinical Signs in Humans

Clinical signs of animal bite wounds in people depend on the area in which the bite occurs. Hands are common locations of bite wounds in people. The skin is thin over most of the hand, offering little protection (especially over the joints). Hand wounds are prone to soft tissue, joint, and tendon sheath infection (Figure 3-14). Complications of animal bite wounds include cellulitis, septic arthritis, osteomyelitis, and sepsis.

[FIGURE 3-14 OMITTED]

Diagnosis in Animals

A tissue sample obtained surgically or at necropsy could show the causative organism with final confirmation based on bacterial culture. Samples are typically obtained via swabbing of the wound area. Diagnosis of bacterial pathogens causing infection following animal bites varies with the disease-causing microbe. Identification of specific bacteria such as Pasteurella, Staphylococcus, and Streptococcus is covered under separate sections; those covered here are only found in bite wounds.

* Capnocytophaga spp. are gram-negative, fusiform-shaped bacteria with one rounded end and one tapered end. The species found in dogs may be curved. Capnocytophaga spp. are slow growers and are identified by culture on enriched media (blood agar), where after 48 to 72 hours the colonies are small- to medium-size, opaque, shiny, nonhemolytic pale beige or yellow in color. The colonies demonstrate gliding motility and may produce swarming similar to Proteus spp. This bacterium grows best at 35[degrees]C to 37[degrees]C in an aerobic environment containing 5% to 10% CO2 or anaerobically (they are known as capnophiles or bacteria that require additional carbon dioxide). The species of Capnocytophaga found in the oral cavity of dogs are C. canimorsus and C. cynodegmi and can be differentiated from each other via biochemical tests with both being oxidase-positive, catalase-positive, and indole-positive; C. cynodegmi is positive for esculin hydrolysis, whereas C. canimorsus is variable on this agar. Other species of Capnocytophaga are oxidase- and catalase-negative. Various selective media and PCR tests have been developed for identification of this bacterial genus.

* Bergeyella zoohelcum are gram-negative, short, straight, nonendospore forming, aerobic rods. This bacterium does not grow on MacConkey agar and on blood agar produces yellow colonies that are circular, shiny, sticky, and smooth. B. zoohelcum is oxidase, catalase, and indole-positive. PCR tests are available for identifying this bacterium.

Diagnosis in Humans

Diagnosis in humans is the same as in animals.

Treatment in Animals

Bite wounds in animals are typically cleaned, irrigated with saline under high pressure, and debrided (any necrotic tissue is removed). Antibiotics given within 24 hours of obtaining the wound will often prevent infection. Fight wounds and infections usually heal within a few days with proper treatment. More aggressive treatment may be needed if the therapy is started after infection has set in. Evaluation of rabies status needs to be performed when animals are presented with bite wounds; evaluation of other infectious diseases is dependent on the species. Treatment of carrier animals is not warranted.

Treatment in Humans

People are treated with a variety of antibiotics for bite wounds. A complete history should include questioning whether the person is immunocompromised. The person's tetanus immunization status needs to be checked as well as rabies immunization status of the animal. Any simian (monkey, apes, and humans) bite should have viral cultures done for Herpesvirus simiae. Wounds are typically cleaned, irrigated with saline under high pressure, and the necrotic tissue debrided. Infected wounds seen within 24 hours of being bitten could be sutured following irrigation and debridement. If the wound is over 24 hours old it should be left open. Wounds in areas that typically develop infection are usually left open no matter when they present for medical care. Antibiotics are typically prescribed for 7 to 14 days unless the wounds are complicated by joint or bone involvement, which warrants 3 to

6 weeks of treatment.

Management and Control in Animals

Almost half of the U.S. population will be bitten by an animal in their lifetime. In some areas animal leash laws and other animal ordinances try to limit the general public's exposure to animals. Required vaccination for rabies can help limit the spread of this virus secondary to a bite wound. Species specific vaccines may limit the spread of other diseases between animals of the same species.

Management and Control in Humans

There are an estimated 52 to 68 million dogs and 57 million cats kept as pets in the United States. Educating people about the risk of infection following an animal bite is important in patients receiving prompt treatment of animal bite injuries. Teaching children to be cautious around unfamiliar animals and to properly handle animals is essential in preventing animal bites in people. Immunocompromised people need to be especially careful around animals.

Summary

Almost half of the U.S. population will be bitten by an animal in their lifetime. The microbes that can cause infection following dog and cat bites may consist of normal flora from the animals' mouths or from the skin of the person/animal being bitten. At least 30 different infectious agents have been reported to be transmitted from a dog or cat bite and most infections as a result of dog and cat bites consist of multiple bacteria. Bacteria found in bite wounds include Pasteurella spp., Staphylococcus aureus, Streptococcus mitis, Moraxella spp., Corynebacterium spp., Neisseria spp., Bergeyella zoohelcum (formerly known as Weeksella zoohelcum), Capnocytophaga canimorsus and Capnocytophaga cynodegm, Fusobacterium spp., Bacteroides spp., Porphyromonas spp., and Prevotella spp. Wound infections from dog and cat bites can cause abscess formation, septic arthritis, osteomyelitis, endocarditis, and CNS infections. The incubation period from animal bites varies with the animal producing the bite, the organism or organisms causing the infection, and health factors of the person/animal being bitten. Animal lesions from bites tend to be isolated to abscesses, whereas human lesions can range from abscesses to septicemia to CNS infection. Educating people about medical care regarding animal bites and ways to avoid bites is crucial in lowering the incidence of animal bites. Rabies immunization status and tetanus immunization status should also be determined in treating human cases of animal bites. Methods of identifying particular bacteria vary with each bacterium.

BRUCELLOSIS

Overview

Brucellosis, also known as Bang's disease and contagious abortion in animals, and Malta fever, Mediterranean fever, and undulant fever in humans, is a disease named after Sir David Bruce, an English army surgeon who identified the cause of this disease in 1887. Bruce found the causative agent of brucellosis, Bacillus melitensis, in the spleens of British soldiers who died of undulant fever on the Mediterranean island of Malta. Several years later the infection in these British soldiers was traced to the soldier's drinking contaminated goat's milk. In 1897, Brucella abortus was isolated and identified from an aborted bovine fetus by Danish veterinarian, Dr. Fredrick Bang. The infection in cattle became known as Bang's disease and was eventually proven to be ubiquitous in many animals. Brucellosis is one of the most serious diseases of livestock because of the damage it causes, including decreased milk production, weight loss, loss of young, infertility, and lameness.

Brucella spp. was the first microbe that the United States chose to develop as a biological weapon. It was considered as a biological weapon by the United States in World War II until the time of the destruction of its stockpile in the 1970s. The reasons it was chosen as a biological weapon include its low lethality, ease of manufacture, susceptibility to sunlight, and its ability to be spread by aerosol dispersion or by contamination of food or milk. It has the advantage of being debilitating to people without being fatal. Under optimal storage conditions it has a half-life of a few weeks. Field tests with live bacteria were performed in the early 1950s and it was effectively disseminated in 4-pound bombs. In 1954, it became the first biological agent developed by the old U.S. offensive biological weapons program with field testing on animals beginning soon afterwards. By 1955, the United States was producing Br. suis--filled cluster bombs for the U.S. Air Force at the Pine Bluff Arsenal in Arkansas. Development of Brucella as a biological weapon was halted in 1967, and President Nixon later banned development of all biological weapons on November 25, 1969.

Causative Agent

Brucellosis is a contagious bacterial disease that typically affects cattle and bison (Br. abortus), swine (Br. suis), dogs (Br. canis), and sheep and goats (Br. melitensis). Br. neotomae (from desert wood rats), Br. ovis (mainly from sheep), and Br. maris (from dolphins) have not been isolated from people. Brucellosis only occasionally affects horses, and cats are relatively resistant to Brucella infections. Depending upon how an animal contracts brucellosis, a different species of Brucella may be causing the infection. For example, pigs, sheep, and goats that are in contact with infected cattle can be infected with Br. abortus. Dogs that ingest placentas from farm animals may be infected with Br. abortus, Br. suis, and Br. melitensis.

Brucella spp. are gram-negative coccobacilli that infect the placenta, uterus, and fetus, causing abortion in females, and infect the testes and accessory sex glands, causing orchitis and accessory sex gland infection in males. Brucellosis can cause infertility in both sexes. Br. abortus and Br. canis cause mild disease in humans, whereas Br. suis and Br. melitensis can be fatal.

Epizootiology and Public Health Significance

Brucellosis has a worldwide distribution and can affect a variety of animals, including reindeer in Alaska and Siberia, camels in the Middle East, and livestock throughout the world.

Concentrations of brucellosis can be seen in Europe, Africa, India, Mexico, and Central and South America. In the United States and Europe brucellosis is uncommon as a result of its elimination from cattle herds. In unvaccinated herds, infection spreads quickly causing many abortions; however, after the first exposure cattle typically develop antibodies and subsequent gestations and lactations appear normal.

Br. melitensis in sheep and goats represents the most important source of brucellosis in humans. Br. melitensis is not enzootic in the United States, Canada, northern Europe, Australasia, or Southeast Asia, but is prevalent in Latin America, the Mediterranean area, Central Asia and, especially, in the countries around the Arabian Gulf. Humans are infected by the handling of animals during the birthing process and the consumption of raw milk and milk products, especially fresh soft cheeses.

Br. suis affects both sexes of swine, causing infertility, abortion, orchitis, and bone and joint lesions. The prevalence is generally low except in parts of South America and Southeast Asia. Br. suis occurs in areas in which pigs are kept, including the southeastern United States and Australia where populations of feral swine are heavily infected. Human infections with Br. suis occur in people handling pigs on farms and during slaughtering and processing feral and domestic swine.

Bovine brucellosis, caused by Br. abortus, has been eradicated from Canada, Japan, northern Europe, and Australia. Cases in humans tend to be sporadic and are acquired by drinking unpasteurized milk, by working with infected cattle at a slaughter facility, by attending infected parturient cattle, and by accidental inoculation with live vaccine.

Br. canis infection in humans tends to occur in dog handlers because close, frequent contact seems to be necessary for transmission.

In the United States, the frequency of brucellosis is related to the number of infected animals. Infected animals are rare in the United States and pasteurization of milk has eliminated that mode of transmission. Occupational exposure (cattle-workers, veterinarians, slaughterhouse workers) is the main transmission route in the United States. The incidence is approximately 200 cases per year or 0.04 per 100,000 persons. People with brucellosis in the United States are primarily found in Texas, California, Virginia, and Florida. Worldwide the frequency of brucellosis varies across nations but is higher in places where handling of animal products and dairy products is less stringently monitored.

Transmission
Brucellosis is commonly transmitted
to susceptible animals by direct
contact with infected animals or
with an environment that has been
contaminated with discharges from
infected animals.


Brucella spp. are facultative, intracellular bacteria that are able to establish infection because the virulent strains are able to survive inside phagocytes. Animal brucellosis is transmitted by contact, by mechanical vectors such as contaminated food, water, and excrement, or by ingestion of bacteria present in large numbers in aborted fetuses and uterine discharges. Cattle can contract brucellosis from contaminated feed or water, licking of contaminated genitals or aborted fetuses, venereal transmission from infected bulls during natural copulation or artificial insemination, or through mucous membranes, lacerations, and rarely intact skin. Replacement cattle or bison that are infected or have been exposed to infection prior to purchase or when wild animals or animals from an affected herd mingle with a brucellosis-free herd are common ways that cattle become infected with Brucella. Goats and sheep can contract brucellosis from ingestion of bacteria and through conjunctiva, vaginal, and subcutaneous wounds. In sheep, transmission occuring between rams is especially common during mating season when healthy rams acquire the infection by servicing ewes previously serviced by infected rams. Swine contract brucellosis by animal-to-animal contact, usually through ingestion of infected material and sexually transmitted fluids like semen. In dogs, transmission is congenital or venereal or by ingestion of infective materials. Humans get brucellosis by direct contact with infected animals or their secretions. People who handle diseased animals can be infected through a break in the skin, across mucous membranes such as conjunctiva, or by inhalation. Drinking infected milk is an important mode of transmission because Brucella bacteria concentrate in mammary glands of infected animals. Pasteurization kills Brucella spp. and has helped decrease cases of brucellosis where milk is pasteurized. Veterinarians may become infected as a result of accidental vaccination of themselves while vaccinating animals. Figure 3-15 summarizes the transmission of Brucella bacteria.

Pathogenesis
Person-to-person spread of brucellosis
is rare.


Once in the body, Brucella spp. are engulfed by neutrophils and are carried in the lymphatic fluid to the lymph nodes draining the infected area. The infected neutrophils release bacteria into the blood and bacteria localize in certain organs such as the liver, spleen, bone marrow, and kidney. The affected organs vary with the animal/human and the species of Brucella. The gross lesions seen in an animal are subtle and rarely diagnostic. In cows, placental lesions include edema, necrosis, and a brownish odorless discharge. In aborted bovine fetuses, edema and bronchopneumonia may be seen. In cows, mammary glands and supramammary lymph nodes may show diffuse inflammation, whereas in bulls the scrotum becomes enlarged and thick connective tissue may compress the testes. In swine, Br. suis causes the formation of white nodules on the uterus of females and testes of males, and similar lesions in the spleen, liver, kidney, lymph nodes, and bone of both sexes. In sheep, brucellosis causes edema and inflammation of the epididymis in rams, necrosis of the placenta in ewes, and inflammatory changes in the lung, liver, lymph nodes, spleen, and kidneys of lambs. Br. canis causes uterine and placental lesions in bitches, orchitis in males, and bronchopneumonia in pups. In humans, the lung, spleen, liver, CNS, bone marrow, and synovium are more frequently affected with disease manifestation reflecting this distribution.

[FIGURE 3-15 OMITTED]

Clinical Signs in Animals

There is no distinct appearance to animals with brucellosis. The signs of brucellosis are spontaneous abortion and inability to conceive in females and inflammation of sex organs in male animals. In susceptible animals, primarily cattle, swine, and goats, brucellosis causes infertility and death. The incubation period of brucellosis is variable ranging from 2 weeks to 1 year or longer (the typical length is 30 to 60 days). Some animals will abort prior to developing a positive reaction to the diagnostic test making determination of an incubation period difficult. Some infected animals never abort. Species specific clinical manifestations are described as follows.
Brucellosis in animals typically causes
late-term abortion in females and
inflammatory lesions in the male
reproductive tract.


* Cattle. The severity of the clinical signs of brucellosis is dependent upon the immune status of the herd or animals. In nonvaccinated pregnant cattle, abortion rates are high (up to 90%) and abortions occurring after the fifth to sixth month of pregnancy causing autolysis of the fetus is a common finding. Brucella bacteria also colonize the mammary glands causing mastitis. A carrier state is a common sequela in cows leading to a reduction in milk yield, retained placenta, and metritis. Figure 3-16 shows a placenta with Br. abortus lesions. In bulls, orchitis and epididymitis are common in Brucella infected animals. Bilateral or unilateral scrotal swelling may persist for a considerable amount of time resulting in necrosis of the testes and infertility. Brucella organisms may also be isolated from arthritic joints and lameness.

* Sheep and goats. In rams the first sign of brucellosis is the deterioration of semen quality. Scrotal edema and inflammation (Figure 3-17), epididymitis, fever, and an increased respiratory rate may also be seen in rams. Abortion or the birth of weak or stillborn lambs is only rarely seen in ewes. In goats, abortion during late pregnancy, birth of weak kids, and mastitis are the most common findings with brucellosis. Fever, weight loss, and diarrhea may also be seen in affected goats.

* Swine. In sows irregular estrous cycles, infertility, and abortion typically in the third month may be seen with brucellosis. Orchitis, testicular necrosis, sterility, lameness, and incoordination may be observed in boars. Heavy mortality may be seen with piglets.

* Dogs. In bitches the main clinical sign is abortion in the last trimester of pregnancy, typically between 40 and 60 days of gestation. Prolonged vaginal discharge typically follows the abortion and repeated abortions in subsequent pregnancies are common. Stillbirths and conception failures are also seen in females. In affected males, epididymitis, orchitis, scrotal dermatitis, and prostatitis are frequently observed. Brucellosis is an important differential diagnosis for diskospondylitis in dogs of either sex.

* Horses. In horses, Br. abortus or Br. suis can be found in the bursa of the neck and withers, muscles, tendons, and joints. Brucellosis is not common in horses.

[FIGURE 3-16 OMITTED]

[FIGURE 3-17 OMITTED]
Brucellosis is incapacitating rather than
fatal (fatality is about 2%).


Clinical Signs in Humans

Brucella spp. are able to establish an infection by surviving phagocytosis and are passed from the lymph to blood and then to organs throughout the body, mainly the liver, spleen, and bone marrow. In humans, the incubation period is typically 5 to 60 days (or longer) and the most prominent symptoms are weakness, loss of appetite, chills, headache, back pain, and intermittent (undulating) fever. Brucellosis persists for weeks to months if left untreated but is seldom fatal in humans. In the chronic form, symptoms may persist for years, either continuously or intermittently, and may assume an undulant nature with periods of normal temperature between acute attacks. Chronic infection can damage joints and the spinal cord.

Diagnosis in Animals

Tissues infected with Brucella spp. do not provide pathognomonic (distinctive of a disease) findings. In some fetuses, pneumonia may be found. In females the placenta is edematous, wheras in males inflammation of the testes may be found. Histologic samples include the uterus which shows nodular inflammatory thickening and abscessation. The placenta of affected ruminants may show firm, yellow-white plaques in the cotyledons of the placenta. Arthritis and vertebral body necrosis may be found in lame swine and dogs. Testicular necrosis and scrotal inflammation may be seen in males of all species. In fetuses, findings consistent with pneumonia may be found.
Abortions seen with brucellosis may be
described as a "storm" of abortions. The
storm of abortions is the high number
of abortions seen when the disease is
introduced into a herd or flock, followed
by a period of resistance during which
abortions do not occur.


Diagnosis in animals consists of bacteriologic or serologic identification. Bacteriologic identification involves the Gram stain and cellular morphology. Brucella is faintly staining, tiny, gram-negative coccobacilli. They are nonendospore forming and lack capsules or flagella; therefore, they are nonmotile. To ensure safety, all tiny gram-negative coccobacilli should be processed in a biosafety cabinet until Brucella can be ruled out. Brucella bacteria are aerobes but some species require an atmosphere with added C[O.sub.2] (5% to 10%). Multiplication is slow at the optimum temperature of 37[degrees]C and enriched medium is needed to support adequate growth. Brucella colonies become visible on suitable solid media in 2 to 3 days, but should be incubated at least 21 days before discarding the samples. Many laboratories now rely on commercial identification systems to identify Brucella or send suspect samples to a reference lab.

Serum agglutination and ELISA tests are standard for diagnosing bovine cases of brucellosis. Antibodies to the bacteria are present in the blood serum, vaginal mucus, seminal fluid, and milk and serve as important diagnostic factors. ELISA tests have been developed for detecting antigens in vaginal discharge and antibodies in milk and serum. Standard plate or tube agglutination tests are used to identify positive animals within a herd. In cattle serum dilutions of 1:100 or above for unvaccinated animals and 1:200 for vaccinated animals between 3 and 9 months of age are considered positive (the animals are classified as reactors). In goats a titer of 1:100 in any goat in a herd is positive and all goats at 1:50 and 1:25 are considered reactors and culled. In sheep a variety of tests may be used. A complement fixation test is run on ram serum and bacterial culture is done on semen or aborted tissue. FA staining is another highly specific diagnostic test in sheep. In swine, identification of Brucella organisms is done with the brucellosis card test. In dogs, qualitative agglutination tests such as latex agglutination are used as a screen to diagnose brucellosis and quantitative agglutination tests such as slide agglutination or agar gel immunodiffusion (AGID) are used to definitively diagnose the disease.

Diagnosis in Humans

Brucellosis diagnosis is primarily dependent on clinical suspicion, adequate history of possible exposure including travel, and isolation of the organism. Clinical presentation can be highly variable and focal lesions may present decades after exposure to the bacterium making brucellosis difficult to diagnose. An unequivocal diagnosis requires isolation of the organism using blood culture as the method of choice. Specimens need to be obtained early in the disease and cultures may need to be incubated for up to 28 days to ensure accurate identification of the organism. Isolation rates of only 20% to 50% are reported even from experienced laboratories and failure to grow the organism is common, especially in cases of Br. abortus infection. Commercial identification systems are hampered by the small amount of CO2 produced during the incubation of organisms. Culture from bone marrow, blood, and affected organs may be successful. Presumptive identification of cultures based on colony morphology and slide agglutination with specific antiserum should be followed by further work in a reference facility. Molecular techniques for identification of Brucella organisms are being developed.

Serology is the preferred method of laboratory diagnosis in humans, but the interpretation of results is difficult. The standard serum agglutination test (SAT) and a modified Coombs' (antiglobulin) test have been used to identify Brucella organisms, whereas ELISA tests have been used to differentiate between specific IgM and IgG antibodies. The use of antibody tests has been limited because the species of Brucella have many common antigens and cross-reacting antigens are seen with many gram-negative bacteria. In addition, cases of brucellosis are often investigated late in their course and rising antibody titers may be missed. The variability of individual responses and the frequency of subclinical infections make the interpretation of single high titers difficult. PCR allows for a rapid diagnosis, but the technique has not been standardized.

Treatment in Animals

Treatment of infected animals is not attempted because animals may recover from the disease signs but do not clear the infection. Efforts to control the disease are aimed at eradication. Dogs may be isolated and management procedures such as individual cages may be attempted; however, these animals remain a source of infection for others.

Treatment in Humans

Humans are treated with antibiotic combinations for 4 to 6 weeks with doxycycline and rifampin (adults) or trimethoprim-sulfamethoxazole and rifampin (children).

Management and Control in Animals

Measures for prevention and control of brucellosis include vaccination of calves, periodic testing of bulk milk from farms, blood testing of adults, and slaughtering of infected animals. The level of enzootic disease can be reduced by intensive use of live, attenuated vaccines (Br. abortus RB51 in cattle, Br. melitensis strain Rev. 1 for sheep and goats). Detection of infected herds (by skin tests in sheep; serologic tests on milk or blood samples taken at sale or slaughter in cattle) and individual animals (by serologic tests) also reduces cases of brucellosis. Finally, the elimination of infected animals by slaughter effectively reduces the source of Brucella infections.

In the United States a federal program for brucellosis eradication called the Cooperative State Federal Brucellosis Eradication Program has existed since 1934. This program has minimum standards (called the Uniform Methods and Rules) for states to achieve eradication. States are deemed brucellosis free when none of their cattle or bison is found to be infected for 12 consecutive months under an active surveillance program. There are different class statuses for rates of infection (Class A _0.25% of herds are infected; Class B 0.26% to 1.5% of herds are infected; Class C _1.5% of herds are infected). In June 2000, 44 U.S. states, plus Puerto Rico and the U.S. Virgin Islands, were brucellosis free.

There are two surveillance procedures used to locate infection without having to test every animal: milk from dairy herds is checked two to four times annually by testing a sample from creameries or the bulk milk tank and blood tests on animals upon change of ownership.

* Milk is tested by the brucellosis ring test (BRT) in which milk from each cow in a herd is pooled and a sample taken for testing. A suspension of stained, killed Brucella organisms is added to the milk and if any cow is positive a bluish ring forms at the cream line.

* Animals (cattle and bison) are tested using market cattle identification (MCI). MCI involves using U.S. Department of Agriculture (USDA)-approved numbered tags (backtags) placed on the shoulders of adult breeding animals being marketed and blood samples are collected from these animals at livestock markets and slaughter facilities. If a sample reacts, it is traced to the back-tag number of the herd and the herd owner is contacted by a state or federal animal health official to arrange for herd testing. All eligible animals in the herd are tested at no cost to the owner. At slaughter, eligible animals are all cattle and bison 2 years of age or older except steers and spayed heifers. At market, eligible animals are all beef cattle and bison older than 24 months of age and all dairy cattle older than 20 months of age except steers and spayed heifers. Pregnant and postparturient heifers are tested regardless of age. Eligible animals for herd tests include all cattle and bison older than 6 months of age except steers and spayed heifers. MCI provides a means of determining the brucellosis status of animals marketed from a large area and eliminates the need to round up all animals from a herd for testing. Blood collected can either be tested by blood agglutination tests or brucellosis card tests.

* Blood agglutination detects nonspecific antibodies to Brucella using serum taken from each animal and mixing it with a test fluid containing killed Brucella organisms (antigen). If the organisms agglutinate, the test is positive.

* The brucellosis card test, also known as the Rose Bengal test, is a compact test kit in which serum on a white card has Brucella antigen added to it. The test is read 4 minutes after the serum and antigen are mixed. Agglutination is a positive result. False-positive results can be seen with the card test especially as a result of residual antibody in calves from vaccination, colostral antibody in calves, and cross-reaction with other bacteria.
Brucellosis in free-ranging bison in
Yellowstone National Park and Grand
Teton National Park threatens the
brucellosis status of livestock herds in
that area. State and federal agencies are
working toward containing the spread
of brucellosis from bison to domestic
livestock.


Control of brucellosis in cattle and bison is through vaccination. Vaccination is about 65% effective in preventing cattle from becoming infected by an average exposure to Brucella. RB51 is the newer, live, attenuated vaccine strain used today (it was licensed in February 1996). Strain 19 Brucella vaccine was the original vaccine developed in 1941 that caused postvaccination reactions in cattle such as abortions and localized inflammation at the vaccine injection site. Unlike the strain 19 vaccine, RB51 does not stimulate the same type of antibodies that could be produced by actual infection causing confusion on interpretation of standard diagnostic tests. The organism in the RB51 vaccine is cleared from blood within 3 days and is not present in nasal secretions, saliva, or urine and the organism is not spread from vaccinated to nonvaccinated cattle. The vaccine is safe in all cattle older than 3 months of age. Live vaccines can only be administered by an accredited veterinarian or state or federal animal health official. In case of human exposure, strain RB51 is sensitive to a range of antibiotics used in the treatment of human brucellosis, but is resistant to rifampin and penicillin. Guidelines for vaccinating cattle include:

* Female dairy calves are vaccinated between 4 and 8 months of age. Female beef calves are vaccinated between 4 and 10 months of age. At the time of vaccination, a tattoo is applied in the right ear that identifies the animal as an official vaccinate and identifies the year in which the vaccination took place. RV/5 would indicate that the calf was vaccinated with RB51 (RV) in 2005 (5). A brucellosis tag is also placed in the right ear of the vaccinate.

* Vaccination is not done in pregnant animals because of the risk of vaccine-induced abortion.

* Males are not vaccinated because the live vaccine may cause bacteria to colonize the male reproductive tract resulting in venereal spread during coitus. Any vaccinated male is neutered.

Swine brucellosis is controlled through serologic testing, inspection at slaughter, and tracing infections back to the farm of origin. Swine are not vaccinated for brucellosis.

The cost of maintaining the brucellosis eradication program is offset by the financial savings to the livestock and dairy industries. Losses from lowered milk production, aborted calves, and reduced breeding efficiency have decreased from $400 million in 1952 to less than $1 million today. The number of infected herds has dropped from 124,000 (1956) to 700 (1992) to 6 (2000) to 7 (2004).

Management and Control in Humans

Human brucellosis is an occupational disease among farmers, veterinarians, slaughterhouse workers, meat packers, laboratory workers, and others who come in direct contact with infected animals or their products (raw meat or unpasteurized dairy products). Individuals who are occupationally exposed can be protected by wearing impermeable clothing, rubber boots, gloves and face masks, and by practicing good personal hygiene. Adequate containment of the organisms to reduce aerosol spread in a laboratory setting is essential. Failure of laboratory kits to identify Brucella spp. quickly and accurately has also caused infection in unsuspecting laboratory workers.

Cases of human brucellosis are usually caused by Br. melitensis in travelers to areas such as Mexico and the Mediterranean region where this organism is prevalent, and by the importation of infected (unpasteurized) dairy products. Pasteurization of milk and other dairy products is effective in protecting people from brucellosis. Unpasteurized milk and cheese is still available in foreign countries such as France and can serve as a source of infection to foreign travelers.

Brucellosis is a less significant problem in the United States where approximately 200 human deaths from this disease occur annually compared with more than 500,000 human deaths per year worldwide. Eradication of brucellosis from domestic animals has greatly reduced the threat of disease to humans in the United States and several other countries. No widely accepted vaccines for humans have been developed.

Summary

Brucellosis is a costly, contagious, zoonotic disease that can cause considerable damage in livestock. The organisms of greatest concern are Br. abortus, Br. melitensis, and Br. suis. Brucellosis in animals typically affects the reproductive organs and udder, causing abortion, inflammation of the reproductive organs, and mastitis. Bacteria are shed in milk and are found in aborted fetuses, placentas, and reproductive discharges. Signs of brucellosis in animals are unremarkable and are typified by late-term abortions in females and inflammatory lesions in the male reproductive tract. Human brucellosis also presents with vague signs such as headache, backache, and an undulating fever. Brucellosis is identified with bacterial culture and serologic tests. Eradication programs in livestock, aimed at testing milk and animals, have greatly reduced the incidence of brucellosis in the United States. Vaccination of cattle has also greatly reduced the incidence of brucellosis.
Brucellosis is a reportable disease in
the United States and state and federal
health authorities must be notified
within 7 days of diagnosis.


CAMPYLOBACTERIOSIS

Overview

Campylobacteriosis, also known as vibriosis and vibrionic abortion in animals, is a zoonotic disease caused by the genus of bacteria Campylobacter, a gram-negative curved rod-shaped bacterium (kampter is Greek for bend or angle and bakterion is Greek for little rod). In 1886, Theodor von Escherich (of E. coli fame) observed organisms resembling Campylobacter in the stool of children with diarrhea. In 1913, two English veterinarians named McFaydean and Stockman identified campylobacters in fetal tissues of aborted sheep and named them Vibrio fetus. Since that time, campylobacters have been recovered in blood samples of children with diarrhea and stool samples of patients with diarrhea. For many years, bacteria of the genus Campylobacter were thought to be Vibrio organisms and it was not until the 1970s that the name of Vibrio bacteria was changed to Campylobacter. In 1972 the development of selective growth media allowed laboratories to test stool specimens for Campylobacter.

Campylobacteriosis is one of the most common bacterial causes of human diarrheal disease in the United States and the enteric disease in people is caused by Ca. jejuni. Campylobacteriosis produces symptoms in people that range from loose stools to dysentery that commonly present as diarrhea, fever, and abdominal cramping. Ca. jejuni can also produce bacteremia, septic arthritis, and can be a trigger for Guillain-Barre syndrome (a neuromuscular paralyzing disease). Ca. coli, Ca. lari, and Ca. upsaliensis are also associated with human disease. A variety of Campylobacter species inhabit the gastrointestinal tract of animals such as poultry, dogs, cats, sheep, and cattle, as well as the reproductive organs of several animal species. Ca. jejuni is found in many foods of animal origin and ingestion of raw agricultural products is implicated in many cases of human infection.

Causative Agent

Campylobacter are pleomorphic, helical (curved), gram-negative rods that are commonly referred to as having a "gull-winged" appearance. They have a long flagellum at one or both ends of the cell that may be several times the length of the cell and is responsible for its rapid motility (Figure 3-18). This motility contributes to Campylobacter's ability to colonize and infect the intestinal mucosa. Campylobacter spp. are microaerophilic (optimal growth occurs with reduced oxygen levels at about 5% to 7%) and capnophilic (optimal growth occurs with increased carbon dioxide at about 10%). Ca. jejuni and Ca. coli are thermophilic species of Campylobacter and grow better at about 42[degrees]C. Cultures are typically incubated for 42 to 78 hours and grow as fine, pinpoint colonies on blood agar. Most pathogenic species of Campylobacter are oxidase--and catalase-positive.

Campylobacter spp. have been isolated from dogs, cats, hamsters, ferrets, nonhuman primates, rabbits, swine, sheep, cattle, birds, and wildlife. There are more than 13 species of Campylobacter, but only some are pathogenic to animals or are zoonotic in nature. Strains of Campylobacter that cause disease include:

* Ca. jejuni was first identified as a human diarrheal pathogen in 1973 and is the most frequently diagnosed bacterial cause of human gastroenteritis in the United States. Ca. jejuni is a commensal in the intestinal tract of many species of domestic animals, including poultry, dogs, cats, cattle, goats, pigs, sheep, mink, and ferrets. Ca. jejuni subspecies jejuni is found in animals, whereas Ca. jejuni subspecies doylei is found in children with diarrhea and in gastric biopsies from adults. Some strains of Ca. jejuni produce a heat-labile (unstable in the presence of heat) enterotoxin thought to be responsible for causing diarrhea.

* Ca. coli is a commensal in the intestinal tract of poultry, swine, and humans. It can cause enteritis in humans and piglets. Dogs and pigs can be carriers of Ca. coli. Ca. coli produces a heat-labile toxin and is difficult to differentiate from Ca. jejuni (hippurate hydrolysis is one way to differentiate these two species).

* Ca. upsaliensis is found in the feces of both healthy and diarrheic dogs and cats. Dogs are the major reservoir of Ca. upsaliensis worldwide. This species of Campylobacter can also be found in the feces of healthy children. Ca. upsaliensis causes gastroenteritis (acute, watery diarrhea), septicemia, and abscesses in humans.

* Ca. hyointestinalis is a species of Campylobacter mainly found in swine, but can also be found in cattle, hamsters, and deer. In humans Ca. hyointestinalis causes gastrointestinal disease (watery or bloody diarrhea and vomiting).

* Ca. fetus is a major veterinary pathogen causing reproductive problems in ruminants. There are two subspecies of Ca. fetus found worldwide: Ca. fetus subspecies fetus and Ca. fetus subspecies venerealis. Ca. fetus subspecies venerealis is found in the prepuce of bulls and the genital tract of cows and heifers producing both infections and carriers in these animals. Ca. fetus subspecies venerealis mainly causes infertility (irregular estrous cycles, resorption of embryos, and infected semen) and may cause low levels of abortions in cattle. This bacterium is spread through coitus (natural or through artificial insemination). Ca. fetus subspecies venerealis can cause septicemia in humans. Ca. fetus subspecies fetus causes abortion mainly in sheep, but also cattle and pigs producing both infected and carrier animals. Contaminated tissue such as feces, uterine discharge, and aborted fetuses and membranes are sources of infection with secondary sources being scavenger birds. In humans Ca. fetus subspecies fetus causes gastroenteritis, septicemia, abortion, and meningitis. Both subspecies are believed to cause zoonotic disease; however, direct and indirect transmission of these bacteria from animals to humans has not been proven.

* Ca. lari is found in birds (mainly seagulls), poultry, dogs, and river and seawater animals. In humans, Ca. lari rarely causes gastroenteritis and septicemia and its identification is sometimes confused with Ca. jejuni.

* Ca. mucosalis is found in pigs with swine proliferative enteritis and can be cultured from the oral cavity and intestinal content of healthy pigs. Ca. mucosalis is not believed to cause human disease.

[FIGURE 3-17 OMITTED]
Ca. jejuni grows best at about 42[degrees]C,
which is approximately the same
temperature as the body temperature
of chickens.


Epizootiology and Public Health Significance

Campylobacter is the most common cause of acute infectious diarrhea in developed countries and is increasing worldwide particularly during the warm summer months in temperate climates (with a secondary peak occurring in the late fall in the United States). In the United States about 2 million cases of gastroenteritis caused by Campylobacter occur annually. The peak incidence of this disease is found in children younger than 1 year of age and in young adults. Other people at high risk for developing campylobacteriosis are those with frequent contact with animals. In developing countries Campylobacter infections can be endemic with symptomatic disease occurring in young children and persistent carriers found in adults. Ca. jejuni accounts for greater than 80% of all gastroenteritis cases caused by Campylobacter. Ca. coli accounts for only 2% to 5% of total cases of campylobacteriosis in the United States; however, it accounts for a higher percentage of cases in developing countries. Ca. fetus infections in animals occur worldwide and can be an unrecognized cause of abortion on farms for long periods of time.

In response to the increase in cases of campylobacteriosis, the CDC began a national surveillance program in 1982 to understand the number of cases and the spread of Campylobacter. In 1996 a more detailed program was initiated to learn the frequency of this disease and the risk factors for acquiring it. In addition to the CDC, the USDA conducts research on how to prevent the Campylobacter infection in chickens (in 1999 up to 88% of poultry in supermarkets tested positive for Campylobacter). No vaccines are currently available for Campylobacter spp.

Transmission

Ca. jejuni is spread to humans by oral ingestion of contaminated food or water or by contact with the excretions of infected animals (poultry and cattle are the main sources of human infection). Campylobacter bacteria can survive at 40[degrees]C in feces and milk for up to 3 weeks, in water for 4 weeks, and in urine for 5 weeks. In animals Campylobacter can be shed in feces for at least 6 weeks post-infection. Person-to-person transmission via the fecal-oral route has also been documented, as well as person to animal (kittens and puppies).
In contrast to other bacteria that
can cause foodborne gastroenteritis,
Campylobacter does not multiply in food.


Ca. jejuni can be transmitted between animals in a variety of ways. Many chicken flocks have Campylobacter carriers and the bacterium can be easily spread from bird to bird through a common water source or through contact with infected feces. During slaughter an infected bird can transfer Campylobacter from the gastrointestinal tract to the meat. Campylobacter is also present in the giblets (especially the liver) and the skin.

Transmission in cattle may occur from unpasteurized milk from a cow that has Campylobacter in her udder, from Campylobacter manure-contaminated milk, and from contaminated surface water and mountain streams.

Ca. fetus is spread between animals via coitus, artificial insemination, peneal contact, and contaminated bedding.

Pathogenesis
A very small number of Campylobacter
bacteria (fewer than 500) can cause
clinical disease in humans.


Campylobacter bacteria have adhesins (that help with attachment to the mucosa), cytotoxins, and endotoxins that appear to help them colonize and invade the jejunum, ileum, and colon, producing hemorrhagic lesions that trigger inflammation. Motility by means of a polar flagellum also contributes to Campylobacter's ability to colonize the intestinal mucosa. Campylobacter cells adhere to the intestinal lining by receptors on its cell wall. Bacteria then penetrate the intestinal mucosa by burrowing, causing ulcerative lesions in the gastrointestinal tract. Pathology appears to involve a heat-labile enterotoxin (Ca. jejuni enterotoxin (CJT) that stimulates a secretory diarrhea) but the understanding of Campylobacter's pathogenicity is not completely understood.

Clinical Signs in Animals

Many animals maintain Campylobacter spp. without producing clinical signs. Campylobacter can cause diarrhea typically 1 to 7 days after exposure in a variety of animals including dogs and cats (particularly puppies and kittens), calves, sheep, ferrets, mink, hamsters, and nonhuman primates. These animals can also be carriers of the disease. Younger animals are more likely to acquire this infection and shed the organism. Most species infected with Campylobacter develop mild to moderate watery diarrhea that may contain mucus, bile, or blood. Some animals such as ferrets, hamsters, and Guinea pigs can develop proliferative lesions of the gastrointestinal tract. Campylobacter can also cause hepatitis in poultry and abortion in ruminants. Reproductive lesions found in ruminants include diffuse, mucopurulent endometritis, infiltration of the uterine mucosa with white blood cells, and necrotic and autolytic changes of the placenta. Most aborted fetuses are resorbed, but if passed vaginally they will appear macerated and may contain necrotic liver lesions.

Clinical Signs in Humans

Campylobacter produces acute gastrointestinal disease in humans presenting with diarrhea (with or without blood), abdominal pain, lethargy, and fever. The incubation period is 1 to 7 days. The degree of diarrhea can range from loose stools to profuse watery diarrhea producing bowel movements ten or more times daily. Although campylobacteriosis is self-limiting, symptoms in people may last a week or longer. A potential sequela to campylobacteriosis is Guillain-Barre syndrome (GBS), an autoimmune neuropathy that can occur in approximately 1 out of 1,000 cases of Campylobacter infection. GBS is most frequently found with Ca. jejuni infections that produce an acute inflammatory demyelinating neuropathy from cross-reaction of antibodies to Ca. jejuni antigens with Schwann cells or myelin. Twenty to 40% of all cases of GBS are preceded by Ca. jejuni infection. Ca. fetus subspecies fetus is most commonly associated with systemic infections particularly in immunocompromised people and range from bacteremia, arthritis, septic abortion, and meningitis.

Diagnosis in Animals

Tissue samples obtained from biopsy or aborted fetuses containing Campylobacter bacteria may show ulcerated intestinal mucosa with crypt abscesses and infiltration of the tissue with neutrophils, monocytes, and eosinophils. In swine, hamsters, and ferrets proliferative lesions are seen in the intestinal mucosa.

Diagnosis of Campylobacter is made via fecal culture from liquid stool or rectal swabs. Samples should be examined within a few hours and transported in semisolid transport media such as Cary-Blair transport medium or Wang's medium. It is important for fecal samples not to be exposed to oxygen. Campylobacter grows well on CAMPY-BAP (a selective media with antibiotics in a Brucella agar base with sheep blood) or Skirrow's agar (a selective media with peptone and soy protein base agar with lysed horse blood and antibiotics). Campylobacter is slow-growing and produces pinpoint colonies when incubated in a microaerophilic atmosphere (5% to 7% oxygen, 5% to 10% carbon dioxide). Ca. jejuni and Ca. coli are thermophilic (grow better at higher temperatures) and require 42[degrees]C to 43.5[degrees]C for 24 to 72 hours for growth. A filtration method can also be used with a nonselective medium, but is not as sensitive as direct culture. Direct examination under the microscope reveals gram-negative "gull-wing" rods. Darting motility using a wet mount preparation is seen using phase-contrast or darkfield microscopy. Direct examination is best done on samples collected during the acute stage of clinical diarrhea when large numbers of the organism are being shed in the feces. Heat-stable or heat-labile antigen tests are available to check rising antibody titers.

Diagnosis in Humans

Stool cultures are used in humans as they are in animals. Blood cultures are routinely set up for humans in addition to fecal cultures. In humans latex agglutination kits and complement fixation tests are also available, but have provided variable results as a result of the large number of Campylobacter serovars. PCR tests for identification of Campylobacter have been developed.

Treatment in Animals

Treatment of Campylobacter infection in animals depends on the severity of the disease and the zoonotic potential. Antibiotics such as erythromycin, gentamicin, and doxycycline are effective against Campylobacter bacilli in both animals and humans (penicillins are ineffective against Campylobacter) and should be prescribed for adequate amounts of time (anywhere from 7 to 28 days of treatment). Animals will continue to shed the bacteria in the stool despite antibiotic treatment; therefore, follow-up fecal cultures are important after antibiotic treatment. Fluid and electrolyte replacement is important in animals that are dehydrated, especially young animals.

Treatment in Humans

Campylobacter infections in humans are typically self-limiting within 2 to 5 days (up to 10 days in some cases). Fluid and electrolyte replacement is important in humans with more severe cases of Campylobacter gastroenteritis. Antibiotics should only be used when diarrhea is persistent or recurs. Antibiotics may not shorten the duration of disease but can shorten the length of bacterial shedding. Antibiotics effective against Campylobacter include erythromycin, gentamicin, and doxycycline. Resistance to ciprofloxacin has been demonstrated in some strains of Campylobacter.

Management and Control in Animals

Control of diarrhea-causing bacteria is discussed in the chapter on Escherichia coli and those precautions apply to preventing Campylobacter outbreaks. The use of antibiotics, particularly fluoroquinolones, in feed has been associated with drug-resistant strains of Campylobacter in poultry. Within 2 years of the 1995 approval of fluoroquinolone use in poultry the rate of domestically-acquired human cases of fluoroquinolone-resistant campylobacteriosis increased. The use of the fluoroquinolone antibiotic enrofloxacin (Bayril[R] 3.23%) has been banned for use in poultry by the FDA in July 2005.

Management and Control in Humans

Prophylaxis of Campylobacter infections is aimed at domestic animal reservoirs and interrupting transmission of the bacterium to people. Most human cases of Campylobacter are caused by fecal contamination of food making proper food hygiene a key factor in preventing campylobacteriosis. Proper food hygiene involves proper cooking of pork and poultry (170[degrees]F for breast meat and 180[degrees]F for thigh meat), preventing cross-contamination between utensils and cutting surfaces and foods, and drinking pasteurized milk and treated water. Proper hand washing after handling animals (livestock, wildlife, and pets) is also important in controlling spread of this bacterium from animals to people. Physicians and laboratories who diagnose campylobacteriosis in people should report their findings to the local health department.

Summary

Campylobacter are gram-negative bacteria that normally reside in the intestines of animals; however, some species appear to be pathogenic in humans and animals. Some species inhabit the reproductive system of animals. Ca. jejuni and Ca. coli are the two species some commonly associated with infections in humans and are usually transmitted by contaminated food, milk, or water. Poultry and cattle are the main sources of human campylobacteriosis. Ca. jejuni causes about 2 million cases of gastroenteritis annually in the United States and is the number one cause of gastroenteritis in the United States. Ca. jejuni is typically spread to humans by oral ingestion of contaminated food or water or by contact with the excretions of infected animals. Campylobacter can cause diarrhea typically 1 to 7 days after exposure in a variety of animals including dogs and cats (particularly puppies and kittens), calves, sheep, ferrets, mink, hamsters, and nonhuman primates; however, many animals do not show clinical signs while harboring this bacterium. These animals can also be carriers of the disease. Campylobacter produces acute gastrointestinal disease in humans, producing diarrhea (with or without blood), abdominal pain, lethargy, and fever. The incubation period in humans is also 1 to 7 days. Diagnosis of Campylobacter is made via fecal culture from liquid stool or rectal swabs. In humans latex agglutination kits and complement fixation tests are also available. Treatment of Campylobacter infection in animals depends on the severity of the disease and the zoonotic potential, and may include antibiotics such as erythromycin, gentamicin, and doxycycline. These antibiotics are also effective against Campylobacter bacilli in humans. The use of antibiotics, particularly fluoroquinolones, in feed has been associated with drug-resistant strains of Campylobacter in poultry and these drugs have been banned for use in poultry by the FDA. Prophylaxis of Campylobacter infections is aimed at domestic animal reservoirs and interrupting transmission of the bacterium to people. Using proper food hygiene techniques is a key factor in preventing campylobacteriosis.
Table 3-1 The Five Kingdom System

Kingdom    Cellular           Nutrition Type    Examples
           Organization

Monera     * Prokaryotic      * Varies          Bacteria
           * Unicellular

Fungi      * Eukaryotic       * Heterotroph     Molds, yeasts,
           * Unicellular or     (cannot make    mushrooms,
             multicellular      its own food)   smuts, rusts
                              * Absorptive

Protista   * Eukaryotic       * Heterotroph     Protozoa
           * Unicellular        (a few
                                autotrophic
                                species
                                exist)

Animalia   * Eukaryotic       * Heterotroph     Invertebrates,
           * Multicellular    * Ingestive       vertebrates

Plantae    * Eukaryotic       * Autotroph       Plants, mosses,
           * Multicellular      (can make its   ferns
                                own food)

Table 3-2 Classification of the House Cat, Human,
and E. coli Bacteria

Category   House cat   Human       E. coli bacteria

Kingdom    Animalia    Animalia    Monera

Phylum     Chordata    Chordata    Proteobacteria

Class      Mammalia    Mammalia    Gamma proteobacteria

Order      Carnivore   Primate     Enterobacteriales

Family     Felidae     Hominidae   Enterobacteriaceae

Genus      Felis       Homo        Escherichia

Species    domestica   sapien      coli

Table 3-3 Comparison of Prokaryotic and Eukaryotic Cells

Characteristic     Prokaryotic           Eukaryotic

Average size       0.20-2.0 [micro]m     10-100 [micro]m
of cells           in diameter           in diameter

Nucleus            No nuclear envelope   Membrane-bound,
                   or nucleoli           nucleoli present

Location/type of   Single, circular      Multiple, linear
genetic material   chromosome in         chromosomes in
                   cytoplasm; some       nucleus; other DNA
                   have plasmids         in organelles

Membrane-bound     Not present           Present (examples
organelles                               include mitochondria
                                         and endoplasmic
                                         reticulum)

Flagella           Hollow, made of       Complex, 9+2
                   protein, attached     arrangement of
                   by basal body         microtubules

Glycocalyx         Exists as capsule     Exists in animal
                   or slime layer        cells

Cell wall          Usually present;      Present in plant
                   many contain          cells, no
                   peptidoglycan         peptidoglycan

Plasma membrane    No carbohydrates,     Sterols and
                   most lack sterols     carbohydrates
                                         present

Cytoskeleton       Not present           Present

Ribosomes          70S                   80S
                                         (70S in organelles)

Cell division      Binary fission        Mitosis

Sexual             Transfer of DNA       Involves meiosis
reproduction       fragments by
                   conjugation,
                   transformation,
                   or transduction

Table 3-4 Structures Found in a Typical Prokaryotic Cell

Structure             Characteristics         Function

Glycocalyx (capsule   Gelatinous              Surrounds the cell
or slime layer)       polysaccharide          wall
                      polypeptide layer       May protect against
                                              phagocytosis and
                                              dessication (drying)
                                              Aids in adherence

Fimbriae and pili     Short, thin, hollow     Fimbriae--attachment
                      appendages attached     to surfaces
                      to the cell wall        Pili--conjugation

Flagella              Long, thin, hollow      Flagella rotate to
                      structures consisting   push the cell
                      of a filament, hook,    Attach to the cell
                      and basal body          wall

Axial filaments       Similar to flagella     Provides motility
                      but wrapped around      to spirochetes
                      the cell, associated
                      with  spirochetes

Cell wall             Two types,              Surrounds the cell
                      gram-positive and       membrane and
                      gram-negative           protects cell
                                              from environmental
                                              stress Contains
                                              peptidoglycan

Cell membrane         Selectively             Surrounds cytoplasm
                      permeable,              and contains enzymes
                      phospholipid bilayer    involved in metabolic
                      and protein             reactions

Cytoplasm             Gelatinous matrix       Made of water and
                      located inside the      organic and inorganic
                      cell membrane           molecules

Ribosomes             70S, contain rRNA       Site of protein
                      and protein             synthesis

Nucleoid              Contains the            Area in the cytoplasm
                      bacterial chromosome    where the main
                      (DNA molecule)          chromosome is located

Plasmids              Small, circular,        Found in some cells
                      extrachromosomal        in addition to the
                      DNA molecules           main chromosome

Inclusions            Reserve deposits of     Examples include
                      various materials       sulfur granules and
                      found in the            metachromatic
                      cytoplasm               granules

Endospores            The dormant,            Assist survival in
                      resistant stage         adverse conditions
                      of some bacteria
                      (6 genera of
                      gram-positive
                      bacteria; 2
                      genera of medical
                      significance)

Capsule               Protective structure    Protects against or
                      outside the cell wall   delays phagocytosis
                      in some bacteria
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Title Annotation:Part 1: OVERVIEW-CAMPYLOBACTERIOSIS
Author:Romich, Janet Amundson
Publication:Understanding Zoonotic Diseases
Article Type:Disease/Disorder overview
Geographic Code:1USA
Date:Jan 1, 2008
Words:17159
Previous Article:Chapter 2 Principles of immunity and diagnostic techniques.
Next Article:Chapter 3 Bacterial zoonoses.
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