Brucellosis ( Brucella Species)

History

Brucellosis is one of the earliest known diseases. Throughout history the disease has taken many different names, including Mediterranean, Maltese, or Crimean fever and Bang disease. It was named undulant fever after 1913 and then brucellosis, the name used today, from 1940 onward.

David Bruce, a military surgeon, isolated brucellae from the spleens of the patients in Malta between 1886 and 1887. The bacterium was named Micrococcus melitensis after isolation. A Danish doctor, Bang, later proved that the principal cause of abortion in bovines was a miniscule bacillus that he named Bacillus abortus. In 1914 Traum isolated the bacteria from the aborted fetuses of pigs and gave the name Bacterium abortus suis. In reference to Bruce's name, the bacterium was renamed under Brucella as Brucella melitensis , Brucella abortus, and Brucella suis in the 1920s. In 1953 van Drimmelen isolated Brucella ovis from sheep; Stoenner and Lackman recovered Brucella neotomae from rodents in 1957; and Carmichael and Bruner isolated Brucella canis from dogs in 1968. In 1994 Ewalt and Ross isolated Brucella pinnipidiae and Brucella cetaceae from sea mammals, namely dolphins and seals for B. pinnipidiae and whales for B. cetaceae . These species were later named Brucella pinnipedialis and Brucella ceti. Later, Brucella microti was identified in animals in 2008 and Brucella inopinata from a breast implant in 2010.

Microbiology

Brucella spp. are small, gram-negative, nonmotile, non–spore-forming, intracellular-reproducing, aerobic coccobacilli with a size of 0.6 to 1.5 µm in length and 0.5 to 0.7 µm in width. Three species have been divided into biovars on the basis of biologic and serologic criteria. Four species are pathogenic for humans: Brucella abortus (seven biovars), B. melitensis (three biovars), B. suis (five biovars), and B. canis. Even though all the Brucella spp. are catalase positive, their oxidase and urease activity, as well as hydrogen sulfide production, varies. Although most Brucella spp. reproduce in aerobic conditions, B. abortus and B. suis are microaerophilic and require 5% to 10% carbon dioxide _. Although biochemical reactions have long been used to identify Brucella spp., matrix-assisted laser desorption/ionization time-of-flight mass spectrometry holds promise for use in reference laboratories. Most Brucella spp. contain two annular chromosomes of approximately 2.1 and 1.2 mega–base pairs (Mbp). B. suis does not follow this structure and has a 3.1 Mbp single annular chromosome. The guanine-cytosine (GC) content of the chromosome is 57%. Genomes of different Brucella spp. have large areas of homology, leading to the suggestion that this genus should be given a single species name. Sequences of more than 30 Brucella genomes are available, including B. suis , B. melitensis, B. canis, and B. abortus.

Brucella does not possess any defined endotoxin. The lipopolysaccharide layer on the cell wall shows endotoxic activity. The O-chain polysaccharide on the lipopolysaccharide layer has a significant role in the bacterial virulence. The three epitopes on the O-chain, named A, M, and C, can vary according to their species. The cytoplasmic, periplasmic, and outer membrane structural proteins bear antigenic features and are recognized by the immune system. Changes in genes that affect the structure of the core oligosaccharide create attenuated mutants that interact more efficiently with the innate immune system (increase dendritic cell maturation and the production of inflammatory cytokines). A phospholipase A1 specific for phosphatidylethanolamine, BveA, has been identified as a factor contributing to the resistance of Brucella to polymyxins, an activity which, in other bacteria, may provide protection from polymyxin-producing bacteria in the rhizosphere. The construction of mutants in the bveA gene, which encodes a predicted esterase, modifies the lipid content of B. melitensis membranes and attenuates survival, replication, and persistence in mice.

The bacterium survives for 6 weeks at 4°C in cream, 30 days in ice cream, and 15 to 100 days in fresh cheese. Boiling and pasteurizing the milk kill the bacterium. Brucellae also die when the milk goes sour or lactic acid fermentation occurs. It is safe to consume cheese after 60 to 90 days. The bacterium is also sensitive to heating, ionized radiation, and disinfectants.

Epidemiology

The incidence of brucellosis across the world varies from less than 0.03 to 160 per 100,000 population. Brucellosis is most commonly seen in the Mediterranean countries, the Balkans, the Persian Gulf, the Middle East, and Central and South America. Since the disintegration of the former Soviet Union, the Asian continent has also emerged as a significant focus. Thus brucellosis is observed more frequently in developing countries, and it is practically eradicated in developed countries. As an example, Muslim and Druze populations in Israel suffered from outbreaks, with a report describing disease in children in the period 2005–11, noting that 50.8% were bacteremic. Hence the disease should be suspected in returning febrile travellers.

The most invasive and pathogenic type of human brucellosis is due to B. melitensis, followed by B. abortus and B. suis . The most common animal hosts for B. melitensis biovars 1 to 3 are sheep, goats, camels, and buffalo; B. abortus biovars 1 to 6 are seen mostly in bison and camels; B. suis biovars 1 to 5 are reported in pigs, reindeers, and rodents; and B. canis affects dogs.

Contaminated sheep and goat milk with B. melitensis appears to be the leading source of human brucellosis worldwide. Other routes of transmission to humans are direct contact with infected animals or their secretions through bruises and lacerations on the skin, inhalation of infected aerosols, and conjunctival inoculation. A child with B. canis bacteremia led to a case investigation that found that the child's dog was the probable source. Due to the high frequency of brucellosis among farmers, veterinarians, doctors, and laboratorians, it is recognized as an occupational disease. Laboratorians, in particular, are likely to acquire the microorganism through aerosols or direct contact, and laboratory-induced outbreaks are also reported. In a review of 71 reported laboratory-acquired cases, the median incubation period was 8 weeks. Four of six pregnant laboratorians with brucellosis aborted their fetus. Antimicrobial prophylaxis in highly exposed laboratorians appeared to be effective. Serologic conversion has occurred in a radiology technician whose exposure was assisting at the aspiration of a Brucella -infected prosthetic joint. Although human-to-human transmission is quite rare, congenitally and sexually transmitted cases are reported as well. Because infections related to blood transfusion and bone marrow transplantation are reported, questioning blood donors for symptoms of brucellosis and the use of diagnostic tests can be considered in endemic areas.

Pathogenesis

Brucella is an intracellular microorganism that can survive inside the macrophages, where it has specific survival mechanisms. The bacterium is protected through immune system—evading mechanisms such as blocking macrophage apoptosis, suppressing Th1-specific immune response, and inhibiting tumor necrosis factor-α (TNF-α) production. Among the significant virulence factors are the lipopolysaccharide structure on the cell membrane, VirB type-IV secretion system (T4SS), the two-component BvrR/BvrS system, and the cyclic β-1,2 glucan system. In addition, virulence factors of the bacterium include the adenine and guanine monophosphate systems, which inhibit the fusion of phagolysosomes, release of myeloperoxidase, and TNF production. A superoxide dismutase detoxifies reactive oxygen intermediates, and urease protects B. abortus and B. suis from gastric acid as they pass through the stomach.

Brucella species taken into the body arrive at local lymph nodes either inside polymorphonuclear leukocytes and macrophages or extracellular. The microorganisms reproducing intracellularly spread to the neighboring cells, local lymph nodes, or reticuloendothelial organs such as liver, spleen, and bone marrow. Brucellae form granulomas made up of epithelioid cells, polymorphonuclear leukocytes, lymphocytes, and giant cells in tissues and organs. Granulomas are known to be more frequent in B. abortus infections. Although toxemia is commonly observed in B. melitensis, abscess formation in joints and spleen is more often related to B. suis.

Immune Response

Adaptive immune responses play a crucial role in controlling the infection. Cytokines such as interferon-γ (IFN-γ) and interleukin-2 (IL-2), secreted by CD8 ⁺ T cells in particular, are the most significant agents in preventing the progression of the disease. During infection an increase is observed in the number of γδ T cells carrying Vγ9δ T receptors. These cells not only increase bactericidal activities of macrophages but also eliminate the infected cells by cytotoxic effects. Secreted cytokines, such as IL-12, IFN-γ, and TNF-α molecules, play an essential role in a natural and adaptive immune response.

Cellular immunity has a fundamental role in controlling the disease. Although the presence of specific antibodies is of utmost importance in diagnosis, they play a limited role in the immune response. The immunoglobulin M (IgM) antibodies increase in the first week and the IgG antibodies in the second. After 4 weeks of rising, both immunoglobulin levels decrease rapidly after a successful treatment. Furthermore, IgG levels decrease faster than IgM levels with treatment. Even after eradication of active infection, IgM antibodies can remain positive in low titers for months or even years. A high level of IgG and IgA antibodies for longer than 6 months is a sign of chronic infection or relapse.

Clinical Manifestations

Brucellosis can involve any organ or system in the body. Hence it mimics a myriad of human disorders and is known as a “great imitator.” The disease is a systemic infection with diverse clinical spectra, extending from asymptomatic disease to fatal illness. Clinical history of living in endemic areas, international travel, unpasteurized milk or milk products consumption, hunting, working in a laboratory, vaccinating livestock, close contact with animals or animal tissues may be suggestive of the disease. The incubation period is usually 1 to 4 weeks, although it may extend beyond several months. Although clinical differences between species are difficult to determine, B. melitensis infections are reported to present more acutely compared with disease caused by B. abortus. In addition, clinical presentation and complications of B. suis infection in humans are reported to be similar to B. melitensis or B. abortus infections.

The disease presents as either acute febrile illness or chronic infection. The onset of symptoms may be either abrupt or insidious, developing over several days to weeks. The most frequent complaints are arthralgia, fever, and fatigue seen in up to 75% to 100% of the cases, followed by sweating, malodorous perspiration, lack of appetite, myalgia, chills, and back pain. Brucellosis was formerly named undulant fever because fever waxes and wanes in due natural course of the disease. The most common clinical findings are fever and hepatomegaly in one-third to one-half of patients, followed by splenomegaly, peripheral arthritis, sacroiliitis, scrotal swelling, neck stiffness, and lymphadenopathy. Subclassifying the patients into “acute” (<8 weeks), “subacute” (8–52 weeks), and “chronic” (>52 weeks) categories according to the onset of the disease appears to be imprecise. However, fever seems to be more frequent when the onset of the disease is within 1 month.

Brucellosis is characterized by frequent organ-based complications, prolonged courses of the disease, treatment failures, and relapses. When a specific organ involvement is detected, the disease is defined as the focal form of brucellosis, for which different management strategies are required. Focal involvement in due course of brucellosis is seen in more than half of the patient population. Owing to its subtle nature, the disease is one of the leading causes of fever of unknown origin and is one of the reasons for febrile neutropenia in endemic areas.

A relapse in brucellosis is defined by the reappearance of clinical signs and symptoms with or without a positive culture. Relapse rates are frequently around 5% to 15%, depending on the regimen used. It frequently occurs within 6 months after the discontinuation of therapy and tends to be milder than the original attack. Relapse is not usually due to the emergence of antibiotic-resistant strains. Rather, it is frequently due to poor compliance with therapy. Other reasons may include inappropriate antibiotic use, microbial virulence factors, and focal infections.

Complications