Andrey Pereira Lage

Federal University of Minas Gerais, Cidade de Minas, Minas Gerais, Brazil

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Publications (108)105.72 Total impact

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    ABSTRACT: This report describes clinical, macro and microscopic changes observed in an outbreak of septicemic salmonellosis in pre-weaned calves caused by Salmonella. The necropsy of calves was performed and gross lesions were identified and characterized. The differential diagnosis included hemoparasites (Anaplasma, Babesia bigemina, Babesia bovis and Ehrlichia spp.), Leptospira and Escherichia coli. The course of infection was three days from the moment calves got sick until death. Some of the clinical signs included apathy, reduced solid food ingestion, hyperthermia and mucosal congestion. Postmortem findings were mostly congestion of oral, ocular and rectal mucosa; petechiae were observed in the vaginal, bladder and sublingual mucosa, serosa of the rumen, reticulum, omasum and small intestine; the lungs were congested. Microscopic examination of the lung was characterized by inflammatory infiltration mostly of neutrophils, fibrin and moderate multifocal haemorrhagic alveolar and bronchial lumen. Vasculitis, necrosis and thrombosis in the liver, gallbladder, spleen, and hypophysis were also observed. Salmonellosis is an important disease of dairy cattle resulting in increased morbidity and mortality of affected animals. This report offers a better understanding of the clinical and pathological characteristics of the disease and demonstrates the importance of diagnostic approaches in cases of Salmonella dublin infections.
    Full-text · Article · Oct 2015
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    ABSTRACT: This report describes clinical, macro and microscopic changes observed in an outbreak of septicemic salmonellosis in pre-weaned calves caused by Salmonella. The necropsy of calves was performed and gross lesions were identified and characterized. The differential diagnosis included hemoparasites (Anaplasma, Babesia bigemina, Babesia bovis and Ehrlichia spp.), Leptospira and Escherichia coli. The course of infection was three days from the moment calves got sick until death. Some of the clinical signs included apathy, reduced solid food ingestion, hyperthermia and mucosal congestion. Postmortem findings were mostly congestion of oral, ocular and rectal mucosa; petechiae were observed in the vaginal, bladder and sublingual mucosa, serosa of the rumen, reticulum, omasum and small intestine; the lungs were congested. Microscopic examination of the lung was characterized by inflammatory infiltration mostly of neutrophils, fibrin and moderate multifocal haemorrhagic alveolar and bronchial lumen. Vasculitis, necrosis and thrombosis in the liver, gallbladder, spleen, and hypophysis were also observed. Salmonellosis is an important disease of dairy cattle resulting in increased morbidity and mortality of affected animals. This report offers a better understanding of the clinical and pathological characteristics of the disease and demonstrates the importance of diagnostic approaches in cases of Salmonella dublin infections.
    Full-text · Article · Oct 2015
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    ABSTRACT: Background: The process for obtaining monoclonal antibodies against a specific antigen is very laborious, involves sophisticated technologies and it is not available in most research laboratories. Considering that most cytokines remain partially conserved among species during evolution, the search for antibody cross-reactivity is an important strategy for immunological studies in veterinary medicine. In this context, the amino acid sequence from human and canine cytokines have demonstrated 49-96 % homology, suggesting high probability of cross-reactivity amongst monoclonal antibodies. For this, 17 commercially available anti-human monoclonal antibodies [IL-1α, IL-1β, IL-2, IL-4, IL-5, IL-6, IL-8 (#1, #2), IL-10, IL-12, IL-13, IL-17A, IFN-γ (#1, #2), TNF-α (#1, #2) and TGF-β], were evaluated in vitro for intracellular cytokine detection in a stimulated canine blood culture by flow cytometry and confocal microscopy. Lymphocytes from peripheral blood of healthy and two unhealthy dogs were analyzed. Results: Eleven anti-human mAbs [IL-1α, IL-4, IL-5, IL-6, IL-8 (#1, #2), IL-12, IL-17A, TNF-α (#1, #2) and TGF-β] cross-reacted against canine intracellular cytokines. The specificity of the assays was not affected after Fc-blocking. Three anti-human cytokine mAbs [IL-4, IL-8 (#2) and TGF-β] when evaluated by confocal microscopy also cross-reacted with intracellular canine cytokines. The identification of human mAbs that cross-reacted with canine cytokines may support their use as immunological biomarkers in veterinary medicine studies. Conclusion: The identification of these 11 anti-human cytokine mAbs that cross-reacted with canine cytokines will be useful immunological biomarkers for pathological conditions by flow cytometry and fluorescence microscopy in dogs.
    Full-text · Article · Sep 2015 · Acta Veterinaria Scandinavica
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    ABSTRACT: Brucella abortus S19 and RB51 strains have been successfully used to control bovine brucellosis worldwide; however, currently, most of our understanding of the protective immune response induced by vaccination comes from studies in mice. The aim of this study was to characterize and compare the immune responses induced in cattle prime-immunized with B. abortus S19 or RB51 and revaccinated with RB51. Female calves, aged 4 to 8 months, were vaccinated with either vaccine S19 (0.6-1.2 x 1011 CFU) or RB51 (1.3 x 1010 CFU) on day 0, and revaccinated with RB51 (1.3 x 1010 CFU) on day 365 of the experiment. Characterization of the immune response was performed using serum and peripheral blood mononuclear cells. Blood samples were collected on days 0, 28, 210, 365, 393 and 575 post-immunization. Results showed that S19 and RB51 vaccination induced an immune response characterized by proliferation of CD4+ and CD8+ T-cells; IFN-ɣ and IL-17A production by CD4+ T-cells; cytotoxic CD8+ T-cells; IL-6 secretion; CD4+ and CD8+ memory cells; antibodies of IgG1 class; and expression of the phenotypes of activation in T-cells. However, the immune response stimulated by S19 compared to RB51 showed higher persistency of IFN-ɣ and CD4+ memory cells, induction of CD21+ memory cells and higher secretion of IL-6. After RB51 revaccination, the immune response was chiefly characterized by increase in IFN-ɣ expression, proliferation of antigen-specific CD4+ and CD8+ T-cells, cytotoxic CD8+ T-cells and decrease of IL-6 production in both groups. Nevertheless, a different polarization of the immune response, CD4+- or CD8+-dominant, was observed after the booster with RB51 for S19 and RB51 prime-vaccinated animals, respectively. Our results indicate that after prime vaccination both vaccine strains induce a strong and complex Th1 immune response, although after RB51 revaccination the differences between immune profiles induced by prime-vaccination become accentuated.
    Full-text · Article · Sep 2015 · PLoS ONE
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    ABSTRACT: The present survey aimed at estimating the seroprevalence of brucellosis in working equines of cattle farms from Minas Gerais State, Brazil, and investigating risk factors associated with the infection. Serum samples from 6439 animals, including 5292 horses, 1037 mules and 110 donkeys, were collected from 1936 herds, between September 2003 and March 2004, in 848 municipalities from the state of Minas Gerais, Brazil. The prevalence of antibodies against smooth Brucella spp. found in equines from Minas Gerais State was 1.37% (95% CI: 0.97-1.78), resulting in a prevalence of herds with infected animals of 4.28% (95% CI: 4.21-4.36). There were differences between regions but these were not of major epidemiological relevance nor were most of them statistically significant, given the considerable overlap of confidence intervals. Nevertheless, the point estimates suggest that the three northeastern regions have slightly higher prevalence than the rest of the state, both at the herd and animal levels. No association of Brucella spp. seropositivity with sex, age or host was observed. In conclusion, the present study showed a low but widespread prevalence of antibodies against smooth Brucella in equines kept in cattle farms in Minas Gerais, a state where bovine brucellosis is also widespread albeit with low prevalence.
    Full-text · Article · Sep 2015 · Preventive Veterinary Medicine
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    ABSTRACT: This study analyzes the occurrence and distribution of phylogenetic groups of 391 strains of Escherichia coli isolated from poultry, cattle, and water buffalo. The frequency of the phylogroups was A = 19%, B1 = 57%, B2 = 2.3%, C = 4.6%, D = 2.8%, E = 11%, and F = 3.3%. Phylogroups A (í µí±ƒ < 0.001) and F (í µí±ƒ = 0.018) were associated with E. coli strains isolated from poultry, phylogroups B1 (í µí±ƒ < 0.001) and E (í µí±ƒ = 0.002) were associated with E. coli isolated from cattle, and phylogroups B2 (í µí±ƒ = 0.003) and D (í µí±ƒ = 0.017) were associated with E. coli isolated from water buffalo. This report demonstrated that some phylogroups are associated with the host analyzed and the results provide knowledge of the phylogenetic composition of E. coli from domestic animals.
    Full-text · Article · Aug 2015 · The Scientific World Journal
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    ABSTRACT: This study aimed to determine the susceptibility profile of Brazilian Brucella abortus isolates from cattle to eight antimicrobial agents that are recommended for the treatment of human brucellosis and to correlate the susceptibility patterns with origin, biotype and MLVA16-genotype of the strains. Screening of 147 B. abortus strains showed 100% sensitivity to doxycycline and ofloxacin, one (0.68%) strain resistant to ciprofloxacin, two strains (1.36%) resistant to streptomycin, two strains (1.36%) resistant to trimethoprim-sulfamethoxazole and five strains (3.40%) resistant to gentamicin. For rifampicin, three strains (2.04%) were resistant and 54 strains (36.73%) showed reduced sensitivity. Two strains were considered multidrug resistant. In conclusion, the majority of B. abortus strains isolated from cattle in Brazil were sensitive to the antimicrobials commonly used for the treatment of human bru-cellosis; however, a considerable proportion of strains showed reduced susceptibility to rifampicin and two strains were considered multidrug resistant. Moreover, there was no correlation among the drug susceptibility pattern, origin, biotype and MLVA16-genotypes of these strains.
    Full-text · Article · Jul 2015 · PLoS ONE
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    ABSTRACT: Brucella abortus vaccines play a central role in bovine brucellosis control/eradication programs and have been successfully used worldwide for decades. Strain 19 and RB51 are the approved B. abortus vaccines strains most commonly used to protect cattle against infection and abortion. However, due to some drawbacks shown by these vaccines much effort has been undertaken for the development of new vaccines, safer and more effective, that could also be used in other susceptible species of animals. In this paper, we present a review of the main aspects of the vaccines that have been used in the brucellosis control over the years and the current research advances in the development of new B. abortus vaccines.
    Full-text · Article · Jul 2015 · Veterinary Research
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    ABSTRACT: Brucella abortus live vaccines have been used successfully to control bovine brucellosis worldwide for decades. However, due to some limitations of these live vaccines, efforts are being made for the development of new safer and more effective vaccines that could also be used in other susceptible species. In this context, understanding the protective immune responses triggered by B. abortus is critical for the development of new vaccines. Such understandings will enhance our knowledge of the host/pathogen interactions and enable to develop methods to evaluate potential vaccines and innovative treatments for animals or humans. At present, almost all the knowledge regarding B. abortus specific immunological responses comes from studies in mice. Active participation of macrophages, dendritic cells, IFN-γ producing CD4(+) T-cells and cytotoxic CD8(+) T-cells are vital to overcome the infection. In this review, we discuss the characteristics of the immune responses triggered by vaccination versus infection by B. abortus, in different hosts. Copyright © 2015. Published by Elsevier Ltd.
    Full-text · Article · Jun 2015 · Vaccine
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    ABSTRACT: The aim of this study was to evaluate the growth of the B. abortus reference strains and field isolates on media containing different inhibitor agents. Reference strains were seeded on tryptose agar containing: i-erythritol (1.0 mg/mL), fuchsin (20 μg/mL and 80 μg/mL), thionin (2.5 μg/mL and 10 μg/mL), rifampicin (200 μg/mL) and safranin O (200 μg/mL). Field isolates were tested only on media containing i-erythritol, rifampicin and thionin. Furthermore, each suspension was also inoculated on tryptose agar incubated in air, to test its ability to grow without CO 2 . Sensitivity to fuchsin was similar among reference strains evaluated. Growth of S19, 544 and 2308 but not RB51 were inhibited on media containing rifampicin. Medium with safranin O showed no inhibition for RB51, 544 and 2308, but it partially inhibited the S19 growth as well as medium containing i-erythritol. Treatment/control growth ratio for 2308 on tryptose agar containing thionin (2.5 μg/mL) was approximatelly 1.0, whereas S19 and RB51 showed 0.85 and 0.89 ratios, respectively. Growth of 544, S19 and RB51 but not 2308 was completely inhibited on medium with thionin (10 μg/mL). All field strains grew on medium containing i-erythritol, but were completelly inhibited by rifampicin. With exception of A1 ( B. abortus biovar 3) all field isolates grew on medium with thionin, although some strains showed a treatment/control growth ratio of 0.75–0.80 (10 μg/mL). These results showed that tryptose agar with thionin, i-erythritol or rifampicin could be useful for differentiating vaccine, challenge and field strains of B. abortus.
    Full-text · Article · Mar 2015 · Brazilian Journal of Microbiology
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    ABSTRACT: Eleven commercially available PE-labeled anti-human (IL-1-α, IL-6, IL-8, TNF-α, IL-17A, IL-5, IL-10, IL-12 and IL-13) and anti-mouse (IL-10, TNF-α) cytokine monoclonal antibodies (mAbs) were tested for cross-reactivity with cattle, goat, and sheep cytokines. Cross-reactivity was assessed by comparative analysis with the standard reactivity of the target species. Our data demonstrated that anti-human IL-1-α, IL-6, IL-8, IL-17A and IL-10 mAbs cross-react with all ruminant species tested. Anti-human IL-5 mAb showed a strong cross-reactivity with cattle and goat IL-5, while anti-human TNF-α mAb showed a selective cross-reactivity with goat TNF-α. No cross-reactivity with the ruminant cytokines was observed for anti-human IL-12 and IL-13 mAbs or for the two anti-mouse cytokine mAbs tested. The present study demonstrated the cross-reactivity of various anti-human cytokine mAbs with cattle, sheep, and goat cytokines, increasing the range of immunological biomarkers for studies in veterinary medicine.
    Full-text · Article · Feb 2015 · Genetics and molecular research: GMR
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    ABSTRACT: The aim of the present study was to evaluate the use of different mouse strains (BALB/c, Swiss and CD-1(®)) and different challenge strains (Brucella abortus 544 and 2308) in the study of B. abortus vaccine (S19 and RB51) immunogenicity test in the murine model. No significant difference in B. abortus spp. vaccine potency assay was found with the use of B. abortus 544 or B. abortus 2308 as challenge strain. Results of variance analysis showed an interaction between treatment and mouse strain; therefore these parameters could not be compared separately. When CD-1(®) groups were compared, those vaccinated showed significantly lower counts than non-vaccinated ones (P≤0.05), independently of the vaccine received (S19 or RB51). Similar results were observed on BALB/c groups. However, in Swiss mouse groups, S19 was more protective than RB51 (P≤0.05), which showed protection when compared to the non-vaccinated group (P≤0.05). In summary, data from the present study showed that CD-1(®), BALB/c and Swiss mice strains, as well as both challenge strains B. abortus strains 544 and 2308 can be used in immunogenicity tests of S19 and RB51 vaccines. Copyright © 2014. Published by Elsevier Ltd.
    Full-text · Article · Dec 2014 · Vaccine
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    ABSTRACT: A serological survey in free-ranging crab-eating foxes (Canidae: Cerdocyon thous) and brown-nosed coatis (Procyonidae: Nasua nasua) was performed in the Nhecolândia sub-region of the Brazilian Pantanal to evaluate the presence of anti-smooth Brucella antibodies on those wild populations. The detection of anti-smooth Brucella antibodies was performed by the Rose Bengal Test (RBT) as screening test and the Fluorescence Polarization Assay (FPA) as a confi rmatory test. The frequency of smooth Brucella Seropositive animals were 13.2% (5/38, 95% CI: 4.4% - 28.1%) for crab-eating foxes and 8.8% (3/34, 95% CI: 1.9% -23.7%) for brown-nosed coatis. No association was found between seropositivity for brucellosis and gender or age. The results of this study suggest exposure to or infection of crabeating fox and brown-nosed coati from the Brazilian Pantanal by Brucella spp.
    Full-text · Article · Dec 2014 · Ciência Rural
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    ABSTRACT: This report describes the clinical manifestation of different Salmonella serovars in dairy calves. S. typhimurium was identified in faecal samples of a calf with rectal prolapse and in faecal samples and mesenteric lymph nodes of a calf with necrosis at the tip of the tail. S. agona was identified in faecal samples of a calf with diarrhoea but no other clinical manifestation, and S. dublin was observed in faecal and organ samples from a calf with neurological symptoms. Assays to differentiate between the main enteric pathogens (Enteropathogenic E. coli, rotavirus and coronavirus) were performed and were negative. Due to the negative impact of salmonellosis and occurrence of different serovars and clinical manifestations in calves, the correct diagnosis is important to identify control and prophylactic measures in a dairy herd.
    Full-text · Article · Nov 2014 · SEMINA: CIENCIAS AGRARIAS
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    ABSTRACT: Mycobacterium tuberculosis is the major cause of tuberculosis in humans. This bacillus gained prominence with the occurrence of HIV, presenting itself as an important opportunistic infection associated with acquired immunodeficiency syndrome (AIDS). The current study aimed to develop a real-time PCR using Eva Green technology for molecular identification of M. tuberculosis isolates. The primers were designed to Rv1510 gene. Ninety nine samples of M. tuberculosis and sixty samples of M. bovis were tested and no sample of the bovine bacillus was detected by the qPCR. Statistical tests showed no difference between the qPCR and biochemical tests used to identify the Mycobacterium tuberculosis. The correlation between tests was perfect with Kappa index of 1.0 (p < 0.001, CI = 0.84 - 1.0). The diagnostic sensitivity and specificity were 100% (CI = 95.94% - 100%) and 100% (CI = 93.98% - 100%). This qPCR was developed with the goal of diagnosing the bacillus M. tuberculosis in samples of bacterial suspension. TB reference laboratories (health and agriculture sectors), public health programs and epidemiological studies probably may benefit from such method.
    Full-text · Article · Oct 2014 · Brazilian Journal of Microbiology
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    ABSTRACT: This prospective longitudinal study investigated the epidemiology of enteric disease associated with infections in calves aging up to 70 days. A total of 850 fecal samples were collected from 67 calves. Seventeen isolates of Salmonella spp. were recovered from feces of 11 calves (16.4 %), and statistical analysis revealed no association between the presence of Salmonella spp. and clinical signs of diarrhea or age. Virulence factors of Escherichia coli were identified in 103 strains: eae (7), K99/STa (7), Stx1 (7), Stx1/eae (36), Stx1/Stx2/eae (2), Stx2 (43), and Stx2/eae (1). There was statistical association between diarrheic animals carrying E. coli Stx1/eae + in their feces at 2 and 4 weeks of age (P = 0.003) and E. coli Stx2 + at 5 weeks of age (P = 0.03). Rotavirus was detected in 49 (5.76 %) fecal samples collected from 33 calves (49.2 %). The presence of rotavirus was correlated with diarrheic feces (P < 0.0001) rather than feces with normal consistency. There was a significant relationship between age group and diarrhea (P = 0.001). Bovine coronavirus (BCoV) was detected in 93 fecal samples collected from 46 calves (68.6 %). There was an association (P < 0.0001) between diarrheic animals positive for BCoV and age groups. The results demonstrate the importance of the pathogens studied in the etiology of diarrhea in calves.
    Full-text · Article · Sep 2014 · Tropical Animal Health and Production
  • Fernanda M. Coura · Andrey P. Lage · Marcos B. Heinemann
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    ABSTRACT: Diarrhea is one of the most frequent diseases in calves up to 30 days of age and is a major cause of economic losses. Its etiology is complex and involves the interaction of various infectious, nutritional, immunological, environmental and managerial factors. The main clinical signs are diarrhea, progressive dehydration, metabolic acidosis, electrolyte imbalance and negative energy balance with or without hypoglycemia, which if left untreated, results in death of the animal. Escherichia coli stands as an important enteropathogen involved in diarrheal syndrome. Pathogenic E. coli strains are classified into groups or pathotypes based on the production of virulence factors and on the mechanisms by which they cause diarrhea. There are five E. coli pathotypes associated with diarrhea in calves: enterotoxigenic E. coli (ETEC), enteropathogenic E. coli (EPEC), enterohemorrhagic E. coli (EHEC), Shiga toxin - producing E. coli (STEC) and necrotoxigenic E. coli (NTEC). In this article, we present the main characteristics and an update on E. coli pathotypes causing calf diarrhea.
    No preview · Article · Sep 2014 · Pesquisa Veterinária Brasileira
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    ABSTRACT: The aims of this study were to address the protective immune response induced by S19 vaccination (n = 10) and RB51 revaccination, in pregnant (n = 9) and non-pregnant (n = 10) S19 calfhood-vaccinated cattle as follows: evaluate the in vitro CD4+ and CD8+ T-lymphocytes specific proliferation, and in vitro expression of IFN-γ by CD4+ and CD8+ T-cells and IL-4 by CD4+, CD8+ and CD21+ lymphocytes subset. Upon in vitro stimulation with γ-irradiated Brucella abortus 2308, blood mononuclear cells from S19 vaccinated and RB51 revaccinated cows exhibited significantly higher proliferation of CD4+ and CD8+ T-lymphocytes and CD4+IFN-γ+ T-cells compared to non-vaccinated animals. RB51 revaccination, regardless of the pregnancy status, did not enhance the proliferation of CD4+ or CD8+ T-cells nor IFN-γ or IL-4 production. Data from the present study suggest that cattle's cellular immune response induced after brucellosis vaccination and revaccination is due to CD4+ and CD8+ T-lymphocytes, being CD4+ T-cells the main source of IFN-γ.
    Full-text · Article · Sep 2014 · Vaccine
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    ABSTRACT: This study aimed to develop and validate real-time PCR for the diagnosis of Mycobacterium bovis isolates. Two hundred and seventy-four M. bovis isolates and 156 M. tuberculosis isolates were tested. Both qPCRs amplified all of the 274 M. bovis samples, but none of the 156 M. tuberculosis samples. The qPCR for PE-PGRS 20 had 91% efficiency and a detection limit of 0.32 ng (sensitivity and specificity for qPCR "Mbovis.100" were 99.64 and 100%, respectively). The qPCR for RD4 had 100% efficiency, and a detection limit of 4 pg (diagnostic sensitivity and specificity were 100 and 100%. The qPCR tests were performed using 4 extraction sets, 3 qPCR kits, and with a range of equipment; yet, all combinations produced similar results in a diagnostic test, demonstrating the robustness of this method. The techniques proved to be efficient, robust, sensitive, and specific for the diagnosis of M. bovis.
    Full-text · Article · Jul 2014 · Genetics and molecular research: GMR
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    ABSTRACT: Background Ovine epididymitis is predominantly associated with Brucella ovis infection. Molecular characterization of Brucella spp. achieved by multi-locus variable number of tandem repeats (VNTR) analyses (MLVA) have proved to be a powerful tool for epidemiological trace-back studies. Thus, the aim of this study was to evaluate the genetic diversity of Brucella ovis isolates from Rio Grande do Sul State, Brazil, by MLVA16. Findings MLVA16 genotyping identified thirteen distinct genotypes and a Hunter-Gaston diversity index of 0.989 among the fourteen B. ovis genotyped strains. All B. ovis MLVA16 genotypes observed in the present study represented non-previously described profiles. Analyses of the eight conserved loci included in panel 1 (MLVA8) showed three different genotypes, two new and one already described for B. ovis isolates. Among ten B. ovis isolates from same herd only two strains had identical pattern, whereas the four isolates with no epidemiologic information exhibited a single MLVA16 pattern each. Analysis of minimal spanning tree, constructed using the fourteen B. ovis strains typed in this study together with all nineteen B. ovis MLVA16 genotypes available in the MLVAbank 2014, revealed the existence of two clearly distinct major clonal complexes. Conclusions In conclusion, the results of the present study showed a high genetic diversity among B. ovis field isolates from Rio Grande do Sul State, Brazil, by MLVA16.
    Full-text · Article · Jul 2014 · BMC Research Notes