Brucellosis is a Disease Caused by Bacteria in the Genus Brucella

8/2/2019 Brucellosis is a Disease Caused by Bacteria in the Genus Brucella

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Brucellosisis a disease caused by bacteria in the genus Brucella. The

disease infects animals such as swine, cattle,and sheep; humans canbecome infected indirectly through contact with infected animals or by

drinking Brucella-contaminated milk. In the United States, most domesticanimals are vaccinated against the bacteria, but brucellosis remains arisk with imported animal products.Brucella are rod-shaped bacteria thatlack a capsule around their cell membranes. Unlike most bacteria,Brucella cause infection by actually entering host cells. As the bacteriacross the host cell membrane, they are engulfed by host cell vacuolescalled phagosomes. The presence of Brucella withinhost cell phagosomes initiates a characteristic immune response, inwhich infected cells begin to stick together and form aggregations called

granulomas.Bacteria of the genus Brucella cause disease primarily in domestic, feraland some wild animals and most are also pathogenic for humans. Inanimals, brucellae typically affect the reproductive organs, and abortionis often the only sign of the disorder. Human brucellosis is either anacute febrile disease or a persistent disease with a wide variety ofsymptoms. It is a true zoonosis in that virtually all human infections areacquired from animals. The disease is controlled by the routine practiceof pasteurizing milk and milk products, as well as by comprehensivecampaigns to eradicate the disease by destroying domestic animals

which exhibit positive serologic reactions to brucellae. Vaccinesproviding some protection to cattle, sheep and goats are availableIn animals, brucellosis is a self-limiting disease, and usually no treatmentis necessary for the resolution of the disease. However, for a period oftime from a few days to several weeks, infected animals may continue toexcrete brucella into their urine and milk. Under warm, moist conditions,the bacteria may survive for months in soil, milk, and evenseawater.Because the bacteria are so hardy, humans may becomeinfected with Brucella by direct contact with the acteria.

Handling or cleaning up after infected animals may put a person incontact with the bacteria. Brucella are extremely efficient in crossing thehuman skin barrier through cuts or breaks in the skin.The incubationperiod of Brucella, the time from exposure to the bacteria to the start ofsymptoms, is typically about three weeks. The primary complaints areweakness and fatigue. An infected person may also experience muscleaches,fever, and chills.The course of the disease reflects the location ofthe Brucella bacteria within the human host. Soon after the Brucella areintroduced into the bloodstream, the bacteria seek out the nearest lymph

nodes and invade the lymph node cells From the initial lymph node, theBrucella spread out to other


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organ targets, including the spleen, bone marrow, and liver.Inside these organs, the infected cells form granulomas.Diagnosingbrucellosis involves culturing the blood,liver, or bone marrow for Brucellaorganisms. Apositive culture alone does not signify brucellosis, since

persons who have been treated for the disease may continue to harborBrucella bacteria for several months. Confirmation of brucellosis,therefore,includes a culture positive for Brucella bacteria as well asevidence of the characteristic symptoms and a history of possiblecontact with infected milk or other animal products.In humans,brucellosis caused by B. abortusis a mild diseasethat resolves itself without treatment. Brucellosis caused by B.melitensisand B. suis, however, can be chronic and severe. Brucellosis is treatedwith administration of an antibiotic

that penetrates host cells to destroy the invasive bacteria.Since the invention of an animal vaccine for brucellosisin the 1970s, the disease has become somewhat rare in theUnited States. Yet the vaccine cannot prevent all incidence ofbrucellosis. The Centers for Disease Control usually reportsfewer than 100 total cases per year in the United States. Mostof these were reported in persons who worked in the meat processingindustry. Brucellosis remains a risk for those whowork in close contact with animals, including veterinarians,farmers, and dairy workers.

Brucellosis also remains a risk when animal productsfrom foreign countries are imported into the United States.Outbreaks of brucellosis have been linked to unpasteurizedfeta and goat cheeses from the Mediterranean region andEurope. In the 1960s, brucellosis was linked to bongo drumsimported from Africa; drums made with infected animal skinscan harbor Brucella bacteria, which can be transmitted tohumans through cuts and scrapes in the human skin surface.In the United States, preventive measures include a rigorous

vaccination program that involves all animals in themeat processing industry. On an individual level, people canavoid the disease by not eating animal products imported fromcountries where brucellosis is frequent, and by avoiding foodsmade with unpasteurized milk.See alsoBacteria and bacterial infection; Food safety;Infection and resistance; Pasteurization


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ETIOLOGIC AGENTHuman brucellosis is caused by strains of Brucella, a bacterial genus

considered on genetic grounds to comprise a single species, Brucellamelitensis, with a numberof biologic variants that exhibit particular host preferences. For the sakeof convenience, the traditional classification into nomen species is still ingeneral use; thisscheme, which closely follows the epidemiologic patterns of theinfection, recognizes B. melitensis, which is the commonest cause ofsymptomatic disease in humansand for which the main sources are sheep, goats, and camels; B.abortus

, which is usually acquired from cattle or buffalo:B. suis

, which isgenerally acquired fromswine except for one variant enzootic in reindeer and caribou andanother enzootic in rodents; and B. canis, which is most often acquiredfrom dogs. B. ovis, whichcauses reproductive disease in sheep, and B. neotomae, which isspecific for desert rodents, have not been clearly implicated in humandisease. Other brucellaehave been isolated from marine mammals and probably correspond to atleast three distinctive nomen species. At least one case of laboratory-

acquired humandisease due to these marine types has been described, and apparentcases of natural infection have been reported.All brucellae are small, gram-negative, unencapsulated, nonsporulatingrods or coccobacilli. They grow aerobically on peptone-based mediaincubated at 37C; thegrowth of some types is enhanced by supplementary CO2. In vivo,brucellae behave as facultative intracellular parasites. The organismsare sensitive to sunlight,

ionizing radiation, and moderate heat; they are killed by boiling andpasteurization but are resistant to freezing and drying. Their resistanceto drying rendersbrucellae stable in aerosol form and facilitates airborne transmission.Brucellae can survive for up to 2 months in soft cheeses made from goator sheep milk; for atleast 6 weeks in dry soil contaminated with infected urine, vaginaldischarge, or placental or fetal tissues; and for at least 6 months in dampsoil or liquid manure kept


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under cool, dark conditions. The bacteria are easily killed by a widerange of common disinfectants used under optimal conditions but arelikely to be much moreresistant at low temperatures or in the presence of heavy organic

contamination.

EPIDEMIOLOGYBrucellosis is a zoonosis whose occurrence is closely related to itsprevalence in domesticated animals. The true global prevalence ofhuman brucellosis is unknownbecause of the imprecision of diagnosis and the inadequacy of reportingand surveillance systems in many countries. Even in developedcountries, the true incidence

may be 10 to 20 times higher than the reported figures. Bovinebrucellosis has been the subject of control programs in many parts of theworld and has beeneradicated from the cattle populations of Australia, New Zealand,Bulgaria, Canada, Cyprus, Great Britain (including the Channel Islands),Japan, Luxembourg,Romania, the Scandinavian countries, Switzerland, and the Czech andSlovak Republics. Its incidence in cattle has been reduced to a low levelin the United Statesand most Western European countries, with a varied picture in other

parts of the world. There is evidence of some resurgence of brucellosisin cattle in EasternEurope following economic changes in recent years. Efforts to eradicateB. melitensisinfection from sheep and goat populations have been muchless successful.These efforts have relied heavily on vaccination programs, which havetended to fluctuate with changing economic and political conditions. Insome countries, such asIsrael, B. melitensishas caused serious outbreaks in cattle. Brucellosis

still represents a major public health problem in Mediterraneancountries; in western, central,and southern Asia; and in parts of Africa and South and CentralAmerica.Human brucellosis is usually associated with occupational or domesticexposure to infected animals or their products. Farmers, shepherds,goatherds, veterinarians,and workers in slaughterhouses and meat-processing plants in endemicareas are occupationally exposed to infection. Family members

(including children) of


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individuals involved in animal husbandry may also be at risk, although itis often difficult to differentiate food-borne infection from environmentalcontamination underthese circumstances. Laboratory workers involved in handling cultures

or infected samples are also at risk. Travelers and urban dwellersusually acquire the infectionthrough consumption of contaminated foods. In countries that haveeradicated the disease, new cases are most often acquired abroad.Dairy products, especially softcheeses, unpasteurized milk, and ice cream, are the most frequentlyimplicated sources; raw meat and bone marrow may be sources ofinfection under exceptionalcircumstances. Infection resulting from contact with cosmetic products

containing infected fetal materials has been recorded. Person-to-persontransmission isextremely rare, as is transfer of infection by blood or tissue donation.Although brucellosis is a chronic intracellular infection, there is noevidence for increasedprevalence or severity among individuals with HIV infection or otherforms of immune deficiency or immune suppression.Brucellosis may be acquired by ingestion or inhalation or throughmucosal or percutaneous exposure. B. melitensisand B. suisare knownto have been developed as

biological weapons by several countries and could be exploited asweapons of bioterrorism ( Chap. 205). This possibility should be borne inmind in the event of

sudden unexplained outbreaks


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IMMUNITY AND PATHOGENESISThe mechanisms of protective immunity against human brucellosis arepresumed to be similar to those documented in laboratory animals.Exposure to infectiongenerates both humoral and cell-mediated immune responses. Althoughantibodies promote clearance of extracellular brucellae by bactericidalaction and through thefacilitation of phagocytosis by polymorphonuclear phagocytes, theantibody response cannot eradicate the infection. Organisms taken upby macrophages and othercell types can establish persistent intracellular infections. Early in thecourse of infection, cytokines such as interleukin (IL) 12 promoteproduction of interferon ?,

which drives TH1-type responses and stimulates macrophage activation.Activated macrophages can kill intracellular brucellae (probably mainlythrough theproduction of reactive oxygen intermediates) and can clear the infection.Tumor necrosis factor a (TNF-a) is produced early in the immuneresponse and stimulatescytotoxic lymphocytes, which can achieve partial clearance; however,the ability of virulent Brucellacells to suppress the TNF-a response mayexplain its limited role


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in protection. Inflammatory cytokines, including IL-6 and IL-10,downregulate the protective response. As in other types of intracellularinfection, it is assumed thatinitial replication of brucellae takes place within cells of the lymph nodes

draining the point of entry. Subsequent hematogenous spread mayresult in chronic localizinginfection at almost any site, although the reticuloendothelial system,musculoskeletal tissues, and the genitourinary system are mostfrequently involved. Both acuteand chronic inflammatory responses develop in brucellosis, and the localtissue response may include granuloma formation, with or withoutnecrosis and caseation.Abscesses may also develop, especially in chronic localized infection.

The determinants of pathogenicity ofBrucella

have not been fullycharacterized, and the mechanisms underlying the manifestations ofbrucellosis are incompletelyunderstood. The survival strategy of the organism is centered onprocesses that enable it to persist within monocytic cells. The smoothBrucellalipopolysaccharide,which has an unusual O-chain composition, possesses endotoxinactivity and plays a key role in pyrogenicity and in resistance tophagocytosis and serum killing inthe nonimmune host. Specific exotoxins have not been isolated, but a

type IV secretion system responsible for secreting proteins that regulateintracellular survivaland trafficking has been identified. This system is activated by low pH,and brucellae produce acid-stable proteins that facilitate survival inphagosomes and depressactivation of the oxidative burst. Macrophage apoptosis and phagosome-lysosome fusion are also suppressed. Virulent brucellae are resistant todefensins andproduce a Cu-Zn superoxide dismutase that enhances resistance to

reactive oxygen intermediates.


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Structure

Brucellae are Gram-negative coccobacilli (short rods) measuring about0.6 to 1.5 m by 0.5-0.7 m. They are non-sporing and lack capsules orflagella and, therefore, are non-motile. The outer cell membrane closelyresembles that of other Gram-negative bacilli with a dominantlipopolysaccharide (LPS) component and three main groups of proteins.The guanine-plus-cytosine content of the DNA is 55-58 moles/cm. NoBrucella species has been found to harbor plasmids naturally althoughthey readily accept broad-host-range plasmids.

The metabolism of the brucellae is mainly oxidative and they show littleaction on carbohydrates in conventional media. They are aerobes butsome species require an atmosphere with added CO

2(5-10 percent).

Multiplication is slow at the optimum temperature of 37

C and enrichedmedium is needed to support adequate growth.

Brucella colonies become visible on suitable solid media in 2-3 days.The colonies of smooth strains are small, round and convex butdissociation, with loss of the O chains of the LPS, occurs readily to formrough or mucoid variants. These latter forms are natural in B canis and Bovis as the LPS of these lack O chains.

Classification and Antigenic Types

Distinguishing features of the six species of Brucella and their preferredhosts are shown in Table 28-1. B abortus, B melitensis and B suis areserious pathogens in humans, B canis causes mild disease and theother two species have not affected humans


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A culture can be identified as belonging to the genus Brucella on thebasis of colonial morphology, staining and slide agglutination with anti-Brucella serum, smooth or rough. Further classification is best done in aspecialized laboratory. Identification to species level may be done by the

procedures shown in Table 28-1. Further differentiation to biovars maybe useful and is illustrated in Table 28-2. As a further refinement, testsfor the oxidative metabolism of certain aminoacids and carbohydrateshave been devised. Modern DNA hybridization tests, however, show thatthe currently named species show a high degree of homology andsuggest that the genus could be appropriately reclassified as having asingle species.

The application of techniques of molecular biology have allowed thecloning and characterization of several genes coding for outermembrane proteins, the use of PCR to identify the presence of brucellarDNA at genus and species level and the demonstration of speciesspecific patterns of restriction fragment length polymorphism. It ispredictable that this work will be extended to improve diagnostic testsand even vaccine development.

Two different O chains in brucellae occur in the LPS of the brucellae with

smooth colonies. These are called A and M, nominally indicating abortusand melitensis antigens. ('Nominally', because some abortus biovars


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carry the M antigen and some common melitensis biovars the Aantigen.) Both O chains have been shown to be homopolymers of 4,6-dideoxy-4-formamido-d-mannopyranose; they differ only in that in the Achain the sugar molecules are always linked 2-1 whereas the M chain

has every fifth junction a 3-1 linkage. In routine serology, smoothspecies of brucellae cross-react almost completely with each other, butnot with rough species and vice versa. Monospecific polyclonal serareacting only to A or M antigens are prepared by cross absorption andmonoclonal antibodies specific for A and M antigens are now available,indicating that there is at least one unique epitope on each type of chain


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Pathogenesis

Brucellae are facultative intracellular parasites, multiplying mainly inmonocyte-macrophage cells. This characteristic dominates thepathology, clinical manifestations and therapy of the disease.

The organisms may gain entry into the body through a variety of portals(Fig. 28-1). Because the infection is systemic it is often not possible todetermine which portal was involved in a particular case. Oral entry, byingestion of contaminated animal products (often raw milk or itsderivatives) or by contact with contaminated fingers, probably representsthe most common route of infection even though this portal may not bethe most vulnerable one. Inhalation of aerosols containing the bacteria,or aerosol contamination of the conjunctivae, is another route. Inhalationprobably underlies some industrial outbreaks. Percutaneous infectionthrough skin abrasions or by accidental inoculation has frequently beendemonstrated.

Figure 28-1 Portals of entry for Brucella species

Brucella species differ markedly in their capacity to cause invasivehuman disease. Brucella melitensis is the most pathogenic; B abortus isassociated with less frequent infection and a greater proportion ofsubclinical cases. The virulence of B suis strains for humans varies but

is generally intermediate.


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Animal studies suggest that invading brucellae are rapidly phagocytosedby polymorphonuclear leukocytes. Brucellae are frequently able tosurvive and multiply in these cells because they inhibit the bactericidalmyeloperoxidase- peroxide-halide system by releasing 5'-guanosine and

adenine. Early in infection, macrophages are also relatively ineffectual inkilling the intracellular brucellae (Fig. 28-2). In systemic spread, it is notclear whether the bacteria are transported within neutrophils andmacrophages or in the blood stream outside cells but organisms maydisseminate widely from regional lymphoid tissue appropriate to theportal of entry and maylocalize in certain target organs such as lymph nodes, spleen, liver, bonemarrow, and (especially in animals) the reproductive organs. Thepresence of meso-erythritol in the testicles and seminal vesicles of bulls,

rams, goats, and boars and in the products of conception in pregnantruminants and pigs stimulates enormous multiplication of brucellae.Erythritol represents a potent localizing factor in the relevant species, butis absent in humans

Figure 28-2 Spread of Brucella in the body

In humans, the tissue lesions produced by Brucella species consist ofminute granulomas that are composed of epithelioid cells,polymorphonuclear leukocytes, lymphocytes and some giant cells. In

cases of infection with B melitensis these granulomas are particularlysmall although the toxemia associated with this organism is great.


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Necrosis is not common, and abscesses do not form, except in B suisinfection. The fact that humans rapidly develop hypersensitivity tobrucellar antigens suggests that many of the symptoms of humanbrucellosis result from the reaction of the host defenses

Host DefensesThe specific host defenses against brucellae resemble those againstother intracellular bacteria and are both humoral (antibody-mediated)and cell-mediated. Passively administered monoclonal antibody directedagainst LPS has been shown to reduce the numbers of brucellaesurviving in the spleens and livers of experimental mice, indicating a rolefor antibody in protection. However, the principal component in defenseagainst brucellae is cell-mediated. Macrophages have been shown toprocess brucellar antigen and present this to T lymphocytes which

produce lymphokines. These agents, of which interferon is the mostactive in this context, activate the formerly ineffective macrophages togreater bactericidal potency. Depletion of gamma interferon makesexperimental animals vulnerable to infection. T cell-derived lymphokinesare also involved in attracting cells to the foci of infection. This leads togranuloma


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formation. While this contributes to the pathology, it also delivers theactivated macrophages to the site where they are needed. Thisinflammatory response is enhanced by cytokines, such as the colony-stimulating factors, tumor necrosis factor and interleukin-1, produced by

a number of cell types.Mice which have survived brucellosis are protected against furtherchallenge, and there is clinical evidence that complete recovery from anatural infection is associated with at least a degree of residualresistance in humans.

HOST RESPONSE (animal)The alimentary tract is the major route in the transmissionof B. abortusin cattle. Licking aborted fetuses and placental membranesor ingesting contaminated milk by calves introduces brucellae to the oralmucosa, tonsils, and gastrointestinal mucosa.Passage of B. abortusthrough epithelial barriers results in acuteregional lymphadenitis and bacteremia. Epithelium covering domes ofileal Peyers patches, an important site of entry for several bovinepathogens that traverse the intestinal mucosa, is important in uptake ofbrucellae. Studies of infected ligated ileal loops in calves have shownthat transepithelial migration of B. abortusoccurs chiefly bydome lymphoepithelial cell endocytosis and transport,and that bacteriaare degraded by macrophages and neutrophils of the gut-associated

lymphoid tissue (Ackermann et al. 1988).The interaction of bacteria withserum components (i.e., antibody or complement), neutrophils,mononuclear macrophages, fibroblasts, and epithelial cells results in theproduction of a variety of biologically active substances that activatemacrophages, expand clones of antigen-recognizing T lymphocytes,stimulate lymphocytes to secrete cytokines, stimulate hematopoiesis,and induce inflammation.Several fractions have been isolated from B.abortusthat generatechemotactic factors derived from serum (Bertram et al. 1986). A

carbohydraterich,aqueous methanol fraction of B. abortusinhibitedchemotactic activity at high concentration;however, a nondialyzablecomponent of this fraction contained a potent stimulator ofchemotaxis.Preheating the serum at 56C for 30 minutes preventedgeneration of chemotactic activity by different fractions. Protein-richfractions of B. abortusstrain 2308 or B. abortusstrain 19 failed tostimulate chemotaxis.Neutrophils are considered an important line ofdefense against infection with the Brucellaspp.(Riley and Robertson1984). A component of B.abortusis capable of inhibiting release of

myeloperoxidase by dose-dependent preferential inhibition of


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primary granule release from bovine neutrophilic leukocytes (Bertram etal. 1986). Failure of polymorphonuclear leukocytes (PMNs) to kill theorganism at the primary site of infection may be responsible in part forthe dissemination of Brucellaspp. to other tissues of the body. Lysates

from granules of guinea pig, human, or bovine PMNs were highly toxic tosmooth B. abortusstrain 45/0 or to rough B. abortus45/20 (Riley andRobertson 1984).However, an oxygen-dependent killing systemappeared to be lethal to both strains. Iodine was more active thanchlorine in the presence of H2O2 and granule lysate in killing theorganism. Ingestion of either strain by PMNs failed to stimulate thehexose monophosphate shut; therefore, Brucellaspp. survive possiblybecause certain surface propertiesfail to generate a suitable stimulus to activate killing mechanisms during

interaction with the plasma membrane. Long-term protective immunityagainst intracellular pathogens such as Brucellais associated withdevelopment of strong cell-mediated responses withproduction of cytokines such as IL-2, IL-12, IL-18, and IFN-_ from CD4+T cells that are associated with Th1-type responses (Manetti et al. 1993;Ismail et al. 2002). Production of IFN-_ and TNF-_ fromactivated CD8+ cells contribute to macrophage activation and killing ofbrucellae (Ismail et al. 2002;Murphy et al. 2001). Although ntibodyresponses may be beneficial, they do not appear to be associatedwith long-term protection. Cell-mediated responses have been evaluated

in cattle infected with virulent B. abortusand in cattle vaccinated with B.abortusstrain 19. Lymphocyte-stimulationresponses were significantly greater in cattle infected with virulent B.abortusas compared to responses observed in cattle vaccinated withattenuated B.abortusstrain 19. However, no correlation wasfound between lymphocyte responses to specific antigen and humoralantibody responses in B. abortusinfected cattle.Differences insubpopulations of lymphocytes and mononuclear macrophages mayexplain in part how an organism stimulates bactericidal activity of

macrophages in one instance and suppresses this activity at other times.The chronic persistence of B.abortusinfections may be due tointracellular localization of brucellae in macrophages whose bactericidalmechanisms are resistant or refractory to activation. Overcomingbactericidal incompetence in these cells may be necessary forelimination of brucellae. Studies on the nonspecific and specificimmunity to brucellae in rodents have provided valuable information onhost cell-cell interactions and on host cell-brucellae interactions, whichcannot be adequately studied in domestic animals due to lack of

genetically identical individuals (Cheers 1984).


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The protective effects of submucosal immune responses andinflammatory reactions against invading organisms may substantiallyalter the ability of Brucellaspp. to colonize the local lymph nodes.Bacterial and host factors that allow brucellae to penetrate intact mucosa

should be examined.Submucosal inflammatory responses composed ofmacrophages, lymphocytes, plasma cells, and largenumbers of eosinophils and neutrophils have been observed as early as2 and 4 days after conjunctival inoculation of cattle with B. abortus.These reactions are present in the submucosa of theconjunctiva,lachrymal duct, and in tonsils. Alteration of submucosalinflammatory reactions by presensitization and their effectiveness indestroying brucellae are unknown.Some pregnancy-associated orpregnancyspecific factors that suppress immune responses may

alter the effectiveness of vaccines. The responses of apregnant host toBrucellaspp. may represent a useful model to elucidate variousmechanisms of immune suppression during pregnancy.

CLINICAL FEATURES:

Brucellosis almost invariably causes fever, which may be associatedwith profuse sweats, especially at night. In endemic areas, brucellosismay be difficult todistinguish from the many other causes of fever. However, two featureswere recognized in the nineteenth century to distinguish brucellosis fromother tropical fevers,such as typhoid and malaria: (1) Left untreated, the fever of brucellosisshows an undulating pattern that persists for weeks before thecommencement of an afebrileperiod that may be followed by relapse, and (2) the fever of brucellosis isassociated with musculoskeletal symptoms and signs in about one-halfof all patients.The clinical syndromes caused by the different nomen species aresimilar, although B. melitensistends to be associated with a more acuteand aggressivepresentation and B. suiswith focal abscess induction. B. abortusinfections may be more insidious in onset and more likely to becomechronic.


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The incubation period varies from 1 week to several months, and theonset of fever and other symptoms may be abrupt or insidious. Inaddition to fever and sweats,patients become increasingly apathetic and fatigued; lose appetite and

weight; and have nonspecific myalgia, headache, and chills. Overall, thepresentations ofbrucellosis often fit into one of three patterns: febrile illness thatresembles typhoid but is less severe; fever and acute monarthritis,typically of hip or knee, in a youngchild; or long-lasting fever, misery, and low-back pain or hip pain in anolder man. In an endemic area (e.g., much of the Middle East), a patientwith fever anddifficulty walking into the clinic would be regarded as having brucellosis

until it was proved otherwise.Diagnostic clues in the patient's history include travel to an endemicarea, employment in a diagnostic microbiology laboratory, consumptionof unpasteurized milkproducts (including soft cheeses), contact with animals, and in anendemic setting a history of similar illness in the family (documentedin almost 50% of cases).Focal features are present in the majority of patients. The most commonis musculoskeletal pain and physical findings in the peripheral and axialskeleton (~40% of

cases). Osteomyelitis more commonly involves the lumbar and lowerthoracic vertebrae than the cervical and high thoracic spine. Individual

joints that are mostcommonly affected by septic arthritis are the knee, hip, sacroiliac,shoulder, and sternoclavicular joints, and the pattern may be one ofeither monarthritis orpolyarthritis. Osteomyelitis may also accompany septic arthritis.In addition to the usual causes of vertebral osteomyelitis or septicarthritis, the most important differential diagnosis is tuberculosis. This

point has an impact on thetherapeutic approach as well as on the prognosis, given that severalantimicrobial agents used to treat brucellosis are also used to treattuberculosis. Septic arthritisin brucellosis progresses slowly, starting with small pericapsularerosions. In the vertebrae, anterior erosions of the superior end plate aretypically the first features tobecome evident, with eventual involvement and sclerosis of the wholevertebra. Anterior osteophytes eventually develop, but vertebral

destruction or impingement onthe spinal cord is rare and usually suggests tuberculosis ( Table 141-1).


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Other systems may be involved in a manner that resembles typhoid.About one-quarter of patients have a dry cough, usually with fewchanges visible on the chestx-ray, although pneumonia, empyema, intrathoracic adenopathy, or lung

abscess can occur. One-quarter of patients have hepatosplenomegaly,and 10 to 20% havesignificant lymphadenopathy; the differential diagnosis includesglandular fever-like illness such as that caused by Epstein-Barr virus,Toxoplasma, andcytomegalovirus; HIV1 infection; or tuberculosis. Up to 10% of men haveacute epididymoorchitis, which must be distinguished from that due tomumps or surgicalproblems such as torsion. Prostatitis, inflammation of the seminal

vesicles, salpingitis, and pyelonephritis all occur. There is an increasedincidence of fetal lossamong infected pregnant women, although teratogenicity has not beendescribed and the tendency to cause abortions is much less pronouncedin humans than infarm animals.Neurologic involvement is common, with depression and lethargy whoseseverity may not be fully appreciated by either the patient or thephysician until aftertreatment. A small proportion of patients develop lymphocytic

meningoencephalitis that mimics neurotuberculosis or noninfectiousconditions and that may becomplicated by intracerebral abscess, a variety of cranial nerve deficits,and ruptured mycotic aneurysms.Endocarditis occurs in ~1% of cases, most often affecting the aorticvalve (natural or prosthetic). Any site in the body may be involved inmetastatic abscess formationor inflammation; the female breast and the thyroid gland are affectedparticularly often. Nonspecific maculopapular rashes and other skin

manifestations areuncommon and are rarely noticed by the patient even if they are present

BRUCELLA ABORTUS

Brucella abortusinfection in pregnant cattle involves the placenta, fetus,and mammary gland.One investigation indicated initial lesions occurredwithin the placenta as early as 4 weeks after challenge;at 5 weeks, a more extensive endometritis with erosion of endometrialepithelium was evident (Payne 1959; Anderson and Cheville 1986).


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Following experimental infection of pregnant cattle via the conjunctivalroute, bacteria were first detected in the uterus 45 weekspostinoculation.Chorionic epithelium becomes infected andinfection extends into the placenta. Considerable variation occurs in

distribution and severity of placental lesions in natural and experimentalB. abortusinfections. Seldom are all placentomes involved,andfrequently only portions of individual placentomesdevelop lesions. Moreover, fetuses that become infected during lategestation may be aborted,with an absence of grossly recognizableplacental lesions. In such cases, microscopic examinationof placental tissues often reveals mild focal inflammation(Enright et al. 1984). Other investigators report that extensive fetalinflammatory disease involving multiple organs leads to fetal stress,

which results in premature deliveries. Bovine fetuses of a knowngestation age were inoculated per os or intramuscularly with low dosesof B. abortusstrain 2308 and examined after abortion or delivery bycesarean section at intervals of 616 days postinoculation (Enright et al.1984). Infected fetuses developed lymphoreticular hyperplasia of lymphnodes as early as 6 days postinoculation. Progression of lymph nodechanges to primary-follicle formation and germinal center formation wasnoted at 9 and 12 days postinoculation, respectively.Morphologicalterations of lymph nodes correlatedwith elevations of IgM and IgG immunoglobulin levels by day 9

postinoculation. Granulomatous responses, composed of acrophages,epithelioid cells, and giant cells, were present in lymphatic tissues,liver, and occasionally within the kidney interstitium as early as 6 and 9days postinoculation.Inflammatory lesions were extensive in fetal lungscharacterized by thickened interlobular septae and focal to diffuse,exudative interstitial pneumonia.Pulmonary lesions were composedprimarily of macrophages and a few neutrophils.Measurement of fetal cortisol levels may serve as an indicator of fetalstress. Also, as fetal corticosteroids can induce the physiological

process of fetal expulsion/parturition (Gaverick and Smith 1993),they may also serve as an early indicator of a physiologic process thatcould lead to abortion. Fetal cortisol levels were elevated substantiallyabove normal values and were present as early as 6 dayspostinoculation (Enright et al. 1984). Experimentally infectedbovine fetuses were aborted between 7 and 19 days postinoculation.This relatively short and consistent interval between infection andabortion suggests that the bovine fetus is a sensitive indicator of in uteroinfection with B. abortus. The character and early development of

granulomatous inflammatory reaction and a rapid development of thefetal immune response demonstrate that the fetus is capable of


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reaction to B. abortusinfection similar to that of mature animals.Mammary gland infection is often clinically inapparent, and gross lesionsmay not be present.Histologically, there is a lobular andperiductal,lymphoplasmacytic, and histiocytic interstitial mastitis

with leukocytes in alveoli and ducts. Brucellae localize and replicateprimarily in macrophages in mammary secretions or in phagocytes in theinter- stitium. Lesions are not seen in all infected ammals(Meador et al.1989a). Supramammary lymph nodes are typically enlarged and arecharacterized by edema, lymphofollicular hyperplasia, medullaryplasmacytosis, and sinus histiocytosis. Localization of B. abortusin themammary gland is markedly influenced by nursing, that is, milk retentionin the mammary gland correlates with the degree of infection.Studies ingoats have shown that failure to nurse or release milk enhances

localization and replication of bacteria in mammary glands afterparturition (Meador et al. 1989b). In turn, mammaryinfection may result in increased systemic spread and persistence ofbrucellae in the host.BRUCELLA MELITENSIS

Due to its high pathogenicity in humans, B. melitensisis one of the mostserious zoonoses in the world with distribution in the Mediterranean,Arabian peninsula, and Asia, and parts of Africa, Latin America, andSouth America (Alton 1990).Brucella melitensisis the principal cause ofbrucellosis in sheep and goats, is endemic in camels in some areas

(Abbas and Agab 2002), and is becomingproblematic as a cause of cattle brucellosis in some countries (Corbel1997). It is the least species specific of the brucellae, and transmits tomany other species (Alton 1990). With the exception of small ruminants,camels, and cattle, most species are considered to be end hosts for B.melitensis(Alton 1990). The pathogenesis of B. melitensisinfection ingoats and early localization within the mammarygland and pregnant uterus of sheep is similar to that of B. abortusincattle. In sheep and goats, excretion in milk is a significant route of B.

melitensistransmission to offspring (Grill et al. 1997). Followingintravenous injection in sheep, B. melitensisinduced necrosis andedema of placentomes and glandular endometritis (Mollelo et al. 1963).In general, B.melitensisis considered to cause more necrosis inplacental tissue and less exudation than B. abortus.

BRUCELLA OVIS

B. ovisis an important cause of epididymitis in rams, but appears to be

the least pathogenic of the Brucellathat affects animals. Venerealexposure is probably the most frequent route of transmission (Buddle


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1955). Lesions in rams are most often located in the tail of theepididymis. Initial localization in the epididymis is accompanied byperivascular edema and infiltration of peritubular tissue by lymphocytesand monocytes; subsequently, neutrophils

infiltrate the exudate. Previously inflamedtubular epithelium developspapillary hyperplasia and local hydropic degeneration, with subsequentformation of inflammatory reaction leading to anextravasation of spermatozoa. Host responses to extravasatedspermatozoa lead to the formation of large spermatic granulomas, whichmay result in complete blockage of the epididymis; testiculardegeneration and fibrosis are secondary to this blockage. B. oviscan becultured from spleens of infected ewes in which no lesions areobserved.Placental pathology experimentally induced by B.ovistends to

localize in the intercotyledonary placenta and is often less severe thanthe placentitis caused by B. melitensisor B. abortus.Pregnant ewesinoculated with B. ovisin the conjunctiva on days 30 to 90 of gestationusually develop uterine infections and abort or deliver infected lambs. Incontrast to the limited period of susceptibilityof fetuses in the intraconjunctivally infected ewes, fetuses exposed to B.ovis in uteroare susceptible to infection throughout pregnancy. Intervalsbetween in uteroinfection with B. ovisand abortionrange from 23 to 80 days postinoculation; by comparison,bovine fetusesinfected in uteroby B. abortususually abort by 12 days postinoculation.

This difference may be attributed to the low pathogenicity of B. ovis. Theimmunologic and granulomatousinflammatory responses of B. ovis-infected bovine fetuses are similar tothose observed in B. abortusinfected bovine fetuses.

BRUCELLA SUIS

Brucellosis in swine is usually caused by B. suis,which may betransmitted by ingestion, although venereal transmission occurs(McMillan 1992).Boars are as susceptible to infection as sows, and

many infected boars develop lesions in the testicles and accessoryreproductive organs and shed B. suisorganisms in semen for extendedperiods. There is atendency for focal granulomatous inflammation todevelop in the endometrium and extend to the entire nongravid uterus.B. suisalso tends to secondarily localize in a greater variety of tissuesthan does B.abortus. The organism demonstrates a predilectionfor localization in bone and joints, spleen, liver, kidney,and brain.Prolonged bacteremia often occurs with B. suisinfection and maycontribute to localization in a wide variety of tissues.The early stages in

pathogenesis of B. suisinfection in swine are comparable to the early


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stages of Brucella313 B. abortusinfection in cattle. However, thecharacter of the response of swine to B. suisdiffers slightlyfrom the response of cattle to B. abortus. B. suismultiplies inmononuclear phagocytes and produces granulomatous lesions

composed of macrophages and epitheliod cells. The granulomas tend toundergo caseous necrosis and become encapsulated by fibrousconnective tissue.

BRUCELLA CANIS

Brucellosis in dogs is characterized by abortions in females, andepididymitis, testicular atrophy, and infertility in males (Carmichael1990). Brucella canishas a limited host range with canids mostsusceptible and with cats, laboratory animals, and humans being less

susceptible to infection. The disease is rapidly transmitted via vaginaldischarges,semen, urine, milk, and other body fluids. The principleclinical sign is abortion, however, many dogs do not demonstrateprominent clinical signs.Diagnosis is based on serologic tests. There isno acceptable vaccine for canine brucellosis.

Diagnosis

The diagnosis of brucellosis is primarily dependent on clinical suspicionallied with the taking of an adequate history of possible exposure -including during travel. Presentation can, however, be highly atypicaland focal lesions may present decades after exposure.

Unequivocal diagnosis requires isolation of the organism. Blood culture

is the method of choice but specimens need to be obtained early in the

disease and cultures may need to be incubated for up to four weeks.

Even so, failure to grow the organism is common, especially in cases of

B abortus infection, and isolation rates of only 20-50% are reported even

from experienced laboratories. Modern commercial systems are

hampered by the small amount of CO2 produced during growth. Culture

from bone marrow and from presenting foci may be successful.

Presumptive identification of cultures from morphology and slide

agglutination with specific antiserum should be followed by further work

in a reference facility. Molecular techniques for typing are being

developed


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Figure 28-4 Diagnosis of brucellosis.

Serology remains the mainstay of laboratory diagnosis, but theinterpretation of results is fraught with difficulties. The large number oftechniques in use is evidence of the problems. The standard serumagglutination test (SAT) has been augmented by the modified Coombs'(antiglobulin) technique and the use of 2-mercaptoethanol to separatethe actions of specific IgG and IgM. These classical methods may, intime, be supplanted by EIA (enzyme immunoassay) tests, designed todifferentiate between specific IgM and IgG antibodies. While the SATtiters commonly decline after recovery from infection and antiglobulintest levels are maintained much longer, the IgM antibody that iscommonly measured by the SAT does not fall away as regularly as insome infections. Nevertheless, persisting levels of antibody may indicatea remaining focus of infection and specific IgG levels rise again with atrue relapse.

Further, because cases often are investigated late in their course, risingtiters are frequently missed; the variability of individual responses andthe frequency of subclinical infections make the interpretation of singlehigh titers subject to error. All serologic tests have to be interpreted withcaution in the light of clinical data and in the context of the local


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prevalence of brucellosis. Moreover, serum from persons with tularemiamay show cross-reactions with Brucella antigen.

The diagnosis of the chronic brucellosis syndrome, without specificlocalization, is often very unsatisfactory. When cultures are negative and

the results of serologic tests are equivocal a confident diagnosis is oftenimpossibleTREATMENTThe broad aims of antimicrobial therapy for brucellosis are to treatcurrent infection and relieve its symptoms and to prevent relapse. Focaldisease presentations mayrequire surgical intervention (e.g., cardiac valve replacement, abscessdrainage, joint replacement) in addition to more prolonged and tailoredantibiotic therapy. In

addition, tuberculosis must always be excluded, or to prevent theemergence of resistance the regimen must be tailored to specificallyexclude monotherapy withagents active against tuberculosis (e.g., rifampin) or to include a fullantituberculous regimen.Early experience with streptomycin monotherapy for brucellosis showedthat relapse was common; thus dual therapy with streptomycin andtetracyclines became thenorm. This is still the most effective combination, but alternatives may be

used, with the options depending on local or national policy about theuse of rifampin for thetreatment of nonmycobacterial infection. Antimicrobial efficacy canusually be predicted by in vitro testing. The efficacy of fluoroquinolonemonotherapy has beendisappointing, with a high relapse rate, despite the good in vitro activityand white-cell penetration of most agents of this class.For adults with acute nonfocal brucellosis (duration,


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The "gold standard" for the treatment of brucellosis in adults isintramuscular streptomycin (750 mg to 1 g daily for 14 to 21 days)together with doxycycline (100 mgtwice daily for 6 weeks). In both clinical trials and observational studies,

relapse follows such treatment in 5 to 10% of patients. The usualalternative regimen (and thecurrent World Health Organization recommendation) is rifampin (600 to900 mg/d) plus doxycycline (100 mg twice daily) for 6 weeks. In trialconditions, therelapse/failure rate is ~10%, but this rate rises to >20% in many nontrialsituations, possibly because doxycycline levels are reduced andclearance rates increased byconcomitant rifampin administration. Patients who cannot tolerate or

receive tetracyclines (children, pregnant women) can be given high-doseTMP-SMX3 instead (2or 3 standard-strength tablets twice daily for adults, depending onweight).Evidence is beginning to accumulate that other aminoglycosides can besubstituted for streptomycin e.g., netilmicin or gentamicin given at adosage of 5 to 6 mg/kgper day for at least 2 weeks. (Shorter courses have been associatedwith high failure rates in adults.) A 5- to 7-day course of therapy withgentamicin (and a 3-week

course of TMP-SMX3) is probably adequate for children withuncomplicated disease. Early experience with fluoroquinolonemonotherapy was disappointing, buthigh-dose ofloxacin (400 mg twice daily) or ciprofloxacin (500 mg twicedaily), given for 6 weeks with rifampin, may become accepted as analternative to the other6-week regimens for adults.Significant neurologic disease due to Brucellarequires prolongedtreatment (i.e., for 6 to 12 months), usually with ceftriaxone

supplementation of a standard regimen.Brucellaendocarditis is treated with at least three drugs (anaminoglycoside, a tetracycline, and rifampin), and many experts addceftriaxone and/or a fluoroquinoloneto reduce the need for valve replacement. Treatment is usually given forat least 6 months, and clinical end points for its discontinuation are oftendifficult to define.Surgery is still required for the majority of cases of infection of prostheticheart valves and prosthetic joints.


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There is no evidence base to guide prophylaxis after exposure tobrucellae in the laboratory, inadvertent immunization with live vaccineintended for use in animals, orexposure to deliberately released brucellae. Most authorities

recommend the administration of rifampin plus doxycycline for 3 weeksafter a low-risk exposure (e.g., anonspecific laboratory accident) and for 6 weeks after a major exposureto aerosol or injected material.PROGNOSIS AND FOLLOW-UPRelapse occurs in up to 30% of poorly compliant patients. Thus patientsideally should be followed clinically for up to 2 years to detect relapse,which responds to aprolonged course of the same therapy that was originally used. The

general well-being and body weight of the patient are more useful guidesthan serology to lack ofrelapse. IgG antibody levels detected by SAT4 and variants of this testcan remain in the diagnostic range for >2 years after successfultreatment. Complementfixation titers usually fall to normal within 1 year of cure. Immunity is notsolid; patients can be reinfected after repeated exposures. Fewer than1% of patients die ofbrucellosis. When the outcome of this infection is fatal, death is usually aconsequence of cardiac involvement; more rarely, it results from severe

neurologic disease.Despite the low mortality rate, recovery from brucellosis is slow, and theillness in humans can cause prolonged inactivity, with consequentdomestic difficulties andeconomic losses.The existence of a prolonged chronic brucellosis state after successfultreatment remains controversial. Evaluation of patients in whom thisstate is considered (oftenthose with work-related exposure to brucellae) includes careful exclusion

of malingering, nonspecific chronic fatigue syndromes and other causesof excessivesweating, such as alcohol abuse and obesity. In the future, theavailability of more sensitive assays to detect Brucellaantigen or DNAmay help to identify patientswith ongoing infection.PREVENTIONVaccines based on live attenuated Brucellastrains, such as B. abortusstrain 19BA or 104M, have been used in some countries to protect high-

risk populations but


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have displayed only short-term efficacy and a high incidence of local andsystemic side effects (local inflammation, pain, lymphadenopathy, fever,malaise, nausea).Subunit vaccines have been developed but are of uncertain value and

cannot be recommended at present. Interest in biodefense hasstimulated research in this area(Chap. 205). The mainstay of veterinary prevention is a nationalcommitment to testing and slaughter of infected herds and flocks (withcompensation for owners),control of animal movement, and active immunization of animals. Thesemeasures are usually sufficient to control human disease as well. In theirabsence,pasteurization of all milk products before consumption is sufficient to

prevent animal-to-human transmission. All cases ofBrucella

infection inanimals and humansshould be reported to the appropriate public health authorities.

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Brucellosis is a Disease Caused by Bacteria in the Genus Brucella

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