یک بررسی در مورد روش های تشخیصی بیماری تب مالت

A Review on Diagnostic Methods of Brucellosis

Abstract

We reviewed different diagnostic methods of brucellosis in different livestock and humans. Bacteriological diagnosis, immunohistochemistry (an alternative technique for direct diagnosis) and different molecular methods for Brucella species genotyping are among the direct methods for diagnosis of brucellosis discussed in this review. The well-established indirect methods for diagnosis of brucellosis, serological and brucellin allergic skin tests were also critically conferred. Finally, for effective control and prevention of brucellosis around the world, the direct diagnostic methods are advised in order to develop vaccine against the circulating Brucella strain in the specific country.

1 Introduction

Brucellosis is an ancient disease that can possibly be traced back to the 5th plague of Egypt around 1600 BC. Recent examination of the ancient Egyptian bones, dating to around 750 BC, showed evidence of sacroilitis and other osteoarticular lesions, common complications of brucellosis [1]. David Bruce isolated Brucella melitensis (B. melitensis) (Micrococcus melitensis at that time) in 1887 from the spleen of a British soldier who died from a febrile illness (Malta fever) common among military personnel stationed on Malta. For almost 20 years aіer isolation of M. melitensis, Malta fever remained a mystery and was thought to be a vector-borne disease until Нemistocles Zammit accidentally demonstrated the zoonotic nature of the disease in 1905 by isolating B. melitensis from goat’s milk. It was believed that goats were not the source of infection since they did not become ill when inoculated with Brucella cultures. Нe discovery that healthy goats could be carriers of the disease has been termed one of the greatest advances ever made in the study of epidemiology [2,3].

Brucellosis is caused by Gram-negative coccobacilli of the genus Brucella [4,5]. In livestock, the disease results in significant economic losses due to reproductive impairment caused by abortion, stillbirth or weak calves and neonatal mortality, infertility [6]. In humans, Brucella spp. infection causes a febrile disease that may be associated with a broad spectrum of symptoms, and it may be fatal in some cases [5,7]. Currently, there are ten spp. described in the genus Brucella. Each one may infect diوٴerent host spp., but each Brucella spp. has a preference for its host spp., B. melitensis (sheep and goats), B. abortus (cattle), B. suis (pigs), B. ovis (rams), B. canis (dogs), B. microti (rodents-Microtus arvalis), B. neotomae (rodents – Neotoma lepida), B. pinnipedialis (pinnipeds), B. ceti (cetacea), and B. inopinata (originally isolated from a human patient, but its preferential host is not known) [8,9]. Нree of this Brucella spp. can be subdivided in biotypes [10,11].

Нerefore, three biotypes (1-3) have been identified in B. melitensis; eight biotypes (1-7,9) in B. abortus; and five biotypes (1-5) in B. suis [12]. All Brucella spp. are considered potentially pathogenic for humans, with the exceptions of B. neotomae, B. microti, and B. ovis [6,9].

A precise diagnosis of Brucella spp. infection is important for the control of the disease in animals and consequently in man. Clinical diagnosis is based usually on the history of reproductive failures in livestock, but it is a presumptive diagnosis [13] that must be confirmed by laboratory methods [13,14]. Нe “gold standard” in the diagnosis of brucellosis is bacterial isolation from blood or bone marrow specimens that requires long cultivation periods (4 to 7 days up to 40 days) and oіen the blood cultures are unsuccessful [15]. Serological tests, such as serum agglutination test (SAT), rose Bengal plate test (RBPT), complement fixation test (CFT), and enzyme-linked immunosorbent assay (ELISA) are still frequently used [5,16]. Since the routine identification and diوٴerentiation of brucellosis suspected specimens, based on culture isolation and phenotypic characterization, requires biosafety level-3 (BSL-3) protocols for the high risk of laboratoryacquired infections [17], molecular methods have been explored in order to overcome these diٹculties. Furthermore, the polymerase chain reaction (PCR)-based assays have shown a higher sensitivity with respect to the standard microbiological assay for the diagnosis of brucellosis [18].

Нerefore, the objectives of this paper are to review a diagnostic methods that are used for isolation, screening, monitoring or epidemiological surveillance and complementary or confirmatory for brucellosis in livestock and humans.

اهمیت تشخیص آزمایشگاهی در درمان بیماری تب مالت

Implications of laboratory diagnosis on brucellosis therapy

Brucellosis is a worldwide zoonosis with a huge economic impact on animal husbandry and public health. The diagnosis of human brucellosis can be protracted because the disease primarily presents as fever of unknown origin with unspecific clinical signs and symptoms. The isolation rate of the fastidious etiologic agent from blood cultures is low, and therefore laboratory diagnosis is mainly based on serologic and molecular testing. However, seronegative brucellosis patients have been described, and antibody titers of diagnostic significance are difficult to define. Whether the molecular detection of Brucella DNA in clinical samples should be followed by long-term antibiotic treatment or not is also a matter of debate. The aim of this article is to review and discuss the implications of laboratory test results in the diagnosis of human brucellosis on disease therapy.

Brucellosis is one of the world’s most widespread bacterial zoonoses, leading to tremendous economic losses in endemic regions and serious complaints in affected patients. The infection may be transmitted by direct animal contact, but is usually acquired through the consumption of contaminated food products of animal origin, mainly via unpasteurized goat’s milk and cheese. Furthermore, brucellosis is the most common bacterial laboratory-acquired infection worldwide [1]. Although many national and international programs have been established to eradicate the pathogen and control its spreading in animal husbandry, brucellosis is still a re-emerging disease. The surveillance of animal brucellosis is difficult due to bacterial persistence in wildlife and environmental reservoirs, with consecutive spill-over to domestic animals [2].

Brucellosis is caused by members of the genus Brucella (B.), which are gram-negative, facultative intracellular coccobacilli that were historically differentiated by their preferred animal host, varying pathogenicity and a few selected phenotypic traits. The genus comprises six classical species: B. melitensis bv 1–3 (primarily isolated from sheep and goats); B. abortus bv 1–6 and 9 (from cattle and other Bovidae); B. suis bv 1–3 (from pigs), bv 4 (from reindeer), bv 5 (from small rodents); B. canis (from dogs); B. ovis (from sheep); and B. neotomae (from desert wood rats). Recently, two novel species of marine origin, B. pinnipedialis (isolated from seals) and B. ceti (from dolphins and whales) [3], B. microti isolated from the common vole (Microtus arvalis) [4], red foxes (Vulpes vulpes) [5] and from soil [6], and B. inopinata isolated from a breast implant wound of a 71-year-old female patient [7] have been described. In the past, a lot of atypical Brucella strains arose. These could represent novel species or lineages of already described species, for example various Brucella strains originating from wild native rodent species in North Queensland, Australia [8], a novel Brucella isolate in association with two cases of stillbirth in nonhuman primates [9], and a B. inopinata-like strain (BO2), which was isolated from a lung biopsy of a 52-year-old Australian patient suffering from chronic destructive pneumonia [10].

Physicians’ awareness of the infection is very poor in many countries, and most cases correctly identified are clinically advanced. Because of its protean clinical manifestations, human brucellosis can be easily confused with other infectious and noninfectious diseases, leading to diagnostic delays and late onset of therapy. The isolation of the fastidious organisms is often unsuccessful or takes a long time, which is why the presumptive clinical diagnosis is usually confirmed by serologic tests. However, seronegative cases, cross-reactivity of anti-Brucella antibodies with many other clinically relevant bacteria, poorly defined cutoffs in serologic test systems, and so on can make the interpretation of the titers measured very difficult. Alternatively, molecular techniques can be used for the laboratory diagnosis of human brucellosis, but the detection of Brucella DNA does not prove an active infection with viable bacteria, and therefore does not effectively support therapeutic decision making.

The scope of this article is to review up-to-date laboratory techniques in the diagnosis of human brucellosis and to discuss if positive serologic or molecular tests should result in long-term antibiotic therapy.