Abortus S19 Research Articles

Detection of Brucella S19 Vaccine Strain DNA in Domestic and Wild Ungulates from Brazilian Pantanal.

The Pantanal region, the largest floodplain in the world, has a huge biodiversity and is an important livestock center. Bovine brucellosis has been reported in the region over the last three decades, posing implications for cattle industry as well as for the maintenance of biodiversity. We aimed to investigate the presence of B. abortus S19 vaccine strain DNA in unvaccinated domestic and wild ungulates from the Brazilian Pantanal. Fifty-two heifers, 63 ovine, 24 domestic pigs, 28 feral pigs, and three Pampas deer were sampled. Brucella spp. was detected through bcsp31 PCR of blood samples in 45.3% (77/170) of the sampled animals, of which 36.4% (28/77) showed positivity in ery PCR corresponding to B. abortus S19 strain. Feral pigs presented the highest occurrence of positive samples in bcsp31 PCR (75%), followed by ovine (47.6%), domestic pigs (41.7%), and unvaccinated heifers (30.8%). We did not observe positivity in Pampas deer. Our results strongly suggest that vaccination against bovine brucellosis may promote spill-over of B. abortus S19 strain in the Pantanal region. Moreover, our data indicate that wild strains of Brucella circulates in the Pantanal Biome.

Bactericidal Activity of Serum by Brucella Abortus RB51 Outer Membrane Protein's Combined by Brucella Abortus S99 Lipopolysaccharide Induction.

Brucellosis vaccines are designed to induce cellular immunity. An effective brucellosis vaccine could induce both cellular and humoral immunity. Serum Bactericidal Assay (SBA) is an important method for determining vaccine humoral immunity. This study is the first to observe humoral immunity in brucellosis by SBA. Extracted Brucella abortus (B. abortus) Lipopolysaccharide (LPS) and Outer Membrane Proteins (OMPs) were injected into rabbits. Group 1 was injected with 25 μg of LPS, Group 2 was injected with 50 μg of OMPs, and Group 3 was injected with 1 ml of combined vaccine, 3 times every 2 weeks. The groups were challenged with B. abortus 544 in the second injection. Sera were separated 2 weeks after the last injection. SBA was performed, and each well was streak-cultured into a plate of Brucella agar. A colony count was done for each plate. Results have shown, the third injection of the combined vaccine had the highest titer of , and the efficacy of the vaccine was 87.71%. As a conclusion, the results of this study showed that LPS and OMP's from B. abortus can provide acceptable immunity.

Evidence of a Viable but Nonculturable (VBNC) Phase in B. abortus S19 under Oxidative Stress (H2O2, -Fe2+, Bleach) and under Non-Oxidative Inhibitory Conditions (Isopropanol, Erythritol, Selenite).

The effect of oxidative stress on the survival of various Brucella species has not been fully investigated yet. We here conducted a study in which we investigated the effect of different types of oxidative stress (Fe2+, H2O2, bleach) versus non-oxidative inhibitory effects (selenite, erythritol, and isopropanol) on the survival of B. abortus S19, B. abortus S19 ∆mglA 3.14, and B. neotomae 5K33. The work focuses on the appearance of ATP-CFU quotient imbalances indicating the existence of a viable but nonculturable (VBNC) form of B. abortus S19, as has previously been shown.

Contrasting roles for IgM and B-cell MHCII expression in Brucella abortus S19 vaccine-mediated efficacy against B. melitensis infection.

Brucella is a neglected zoonotic pathogen with a worldwide distribution. Our study delves into B-cell effector functions in live vaccine-mediated immunity against brucellosis. Notably, we found antibody production, particularly secretory IgM, confers protection against virulent Brucella melitensis in vaccinated mice, which was associated with complement activation. By contrast, B-cell MHCII expression negatively impacted vaccine efficacy. In addition, B-cell depletion after vaccination, but before the B. melitensis challenge, enhanced protection against infection, suggesting a detrimental B-cell role during the challenge phase. Interestingly, deficiency of T follicular helper cells, which are crucial for aiding germinal center B cells, dampened vaccine efficacy at later stages of challenge independent of antibody production. This study underscores contrasting and phase-dependent roles of B-cell effector functions in vaccine-mediated immunity against Brucella.

Parenteral Vaccination with a Live Brucella melitensis Mutant Protects against Wild-Type B. melitensis 16M Challenge.

Susceptibility to brucellosis remains prevalent, even in herds vaccinated with conventional vaccines. Efforts are underway to develop an improved brucellosis vaccine, and possibly a universal vaccine, given that Brucella species are highly homologous. To this end, two B. melitensis mutants were developed, znBM-lacZ (znBMZ) and znBM-mCherry (znBM-mC), and were tested for their ability to confer systemic immunity against virulent B. melitensis challenge. To assess the extent of their attenuation, bone-marrow-derived macrophages and human TF-1 myeloid cells were infected with both mutants, and the inability to replicate within these cells was noted. Mice infected with varying doses of znBM-mC cleared the brucellae within 6-10 weeks. To test for efficacy against systemic disease, groups of mice were vaccinated once by the intraperitoneal route with either znBMZ or B. abortus S19 vaccine. Relative to the PBS-dosed mice, znBMZ vaccination greatly reduced splenic brucellae colonization by ~25,000-fold compared to 700-fold for S19-vaccinated mice. Not surprisingly, both znBMZ and S19 strains induced IFN-γ+ CD4+ T cells, yet only znBMZ induced IFN-γ+ CD8+ T cells. While both strains induced CD4+ effector memory T cells (Tems), only znBMZ induced CD8+ Tems. Thus, these results show that the described znBM mutants are safe, able to elicit CD4+ and CD8+ T cell immunity without a boost, and highly effective, rendering them promising vaccine candidates for livestock.

Brucella abortus Rough-Type Mutant Induces Ferroptosis and More Oxidative Stress in Infected Macrophages.

Brucella is an intracellular parasitic bacterium that uses multiple strategies to evade the host's defense mechanisms. However, how Brucella manipulates the host-induced oxidative stress and relevant biological processes are still poorly understood. In this study, a comparative transcriptome assay of macrophages infected with Brucella abortus S2308 and its rough mutant RB14 was performed to investigate the differentially expressed genes which might be associated with the pathogenic mechanism of Brucella. Our results showed that numerous host pro-oxidative and antioxidative stress genes were differentially expressed in macrophages infected with B. abortus S2308 and mutant RB14 at 4, 8, 24, and 48 h post-infection. Interestingly, we found that several ferroptosis-associated genes were differentially expressed during B. abortus RB14 infection. Moreover, we found that the rough mutant RB14-induced macrophage death was associated with reduced levels of host glutathione and glutathione peroxidase 4, together with increased free iron, lipid peroxidation, and ROS, all of which are important hallmarks of ferroptosis. The ferroptosis occurring during infection with RB14 was reduced by treatment with the inhibitor ferrostatin-1. However, B. abortus S2308 infection did not induce these hallmarks of ferroptosis. Taken together, our results demonstrate that ferroptosis is involved in rough B. abortus infection. Investigating how Brucella manipulates oxidative stress and ferroptosis in its host will be helpful to clarify the pathogenicity of B. abortus.

Evaluation of an In-house Indirect ELISA for Differential Detection of IgM and IgG anti-Brucella Antibodies in Human Brucellosis

Brucellosis caused by various species of the genus Brucella is one of the most important zoonotic diseases of global importance with veterinary, public health, and economic concerns. The study aimed to standardize IgM and IgG-based iELISA to detect anti-Brucella antibodies for serodiagnosis of acute and chronic human brucellosis. The test was standardized using 1:320 dilution of smooth lipopolysaccharide (sLPS) antigen from B. abortus S99 strain, 1:80 serum dilution, 1:4000 anti-human IgM and IgG conjugates, respectively for both IgM and IgG iELISA. The cut-off using 50 each brucellosis positive and negative human sera panel samples was set at ≥ 42 for both IgM and IgG iELISA. A total of 700 human sera samples were evaluated (137 veterinary doctors, 157 artificial inseminators, and 406 veterinary assistants). Overall, the study detected 8.3%, 8.1%, 8%, and 6.1% positivity by in-house IgG iELISA, RBPT, IgM iELISA, and SAT tests, respectively. Considering commercial iELISA kit as a gold standard, the sensitivities of IgM and IgG iELISA were 90% and 97.9%, respectively, whereas, specificities were >99%. The study established >98% specificity and >90% sensitivity for differential detection of immunoglobulin classes in the standardized iELISA. The developed assay outperformed the other evaluated tests with a shorter assay time and can be implemented in both endemic and non-endemic regions for surveillance and diagnosis of human brucellosis.

Evaluación de los niveles de factor de necrosis tumoral alfa, interferón gamma, procalcitonina y neopterina en bovinos seropositivos para Brucella

Brucellosis is a zoonotic disease that affects a large number of people and animals, causing physical disability, workforce loss and significant economic losses in the livestock industry. In the current study, it was aimed to determine and compare the levels of tumor necrosis factor alpha (TNF–α), interferon gamma (IFN–γ), Procalcitonin (PCT) and Neopterin in the blood serums of cattle with brucellosis and vaccinated against brucellosis. The materials of this study consisted of a total 48 blood serums belonging to three basic groups, each consisting of 16 animals. Disease group (1st group) were divided into two subgrups each consisting of 8 animals that 21st day after abortion and seropositive 7 months pregnant, the vaccinated (2nd group) and the control (3rd group) groups were divided into two subgroups, each consisting of 8 animals that gave birth 21 days ago and 7 months pregnant. IFN–γ and PCT levels were determined by sandwich enzyme immunoassay, TNF–α and Neopterin levels were determined using competitive inhibition enzyme immunoassay method by using ELISA device. In this study, TNF–α, PCT and Neopterin levels measured in the blood serums of the Brucella seropositive (1st), conjunctival Brucella abortus S19 vaccine administered (2nd) and unvaccinated Brucella seronegative control groups were compared and no significant difference could be determined between the subgroups of the groups (P>0.05). There were a significant differences between 1st, 2nd, and 3rd groups (P<0.05). IFN–γ levels determined in the blood serums of 1st, 2nd and 3rd groups were compared and nosignificant differences were found between the subgroups of 2nd and 3rd groups (P>0.05), but there were a significant differences between the subgroups of the 1st group (P<0.05). Similarly, a significant differences were determined between 1st, 2nd and 3rd groups in terms of IFN–γ levels (P<0.05). As a result, it was thought that detecting very high serum TNF–α, IFN–γ, neopterin levels in cattle with brucellosis would be helpful in the diagnosis and follow–up of brucellosis. However, it was concluded that there is a need for controlled studies comparing more herds with brucellosis to determine whether the relevant cytokines can be used in the diagnosis of brucellosis.

Characterization of Brucella abortus Mutant A19mut2, a Potential DIVA Vaccine Candidate with a Modification on Lipopolysaccharide.

Brucella abortus is the main causative agent for bovine brucellosis. B. abortus A19 is a widely used vaccine strain to protect cows from Brucella infection in China. However, A19 has a similar lipopolysaccharide (LPS) antigen to that of the field virulent Brucella strain, whose immunization interferes with the serodiagnosis of vaccinated and infected animals. [Aim] To develop a novel Brucella DIVA vaccine candidate. The B. abortus mutant A19mut2 with the formyltransferase gene wbkC is replaced by an acetyltransferase gene wbdR from E. coli O157 using the bacterial homologous recombination technique, generating a modified O-polysaccharide that cannot induce antibodies in mice against wild-type Brucella LPS. The biological phenotypes of the A19mut2 were assessed using a growth curve analysis, agglutination tests, Western blotting, and stress resistance assays. Histopathological changes and bacterial colonization in the spleens of vaccinated mice were investigated to assess the residual virulence and protection of the A19mut2. Humoral and cellular immunity was evaluated by measuring the levels of IgG, IgG subtypes, and the release of cytokines IFN-γ and IL10 in the splenocytes of the vaccinated mice. ELISA coated with wild-type LPS can distinguish mouse antibodies induced by A19 and A19mut2 immunization. The A19mut2 showed a decreased residual virulence in mice, compared to the A19 strain, but induced significant humoral and cellular immune responses, as the A19 immunization did. The protection efficacy of A19mut2 immunization against B. abortus S2308 NalR infection was similar to that of A19 immunization. The A19mut2 has potential as a novel DIVA vaccine candidate in the future.

Research progress in the therapy of brucellosis

AbstractBrucellosis is a zoonotic bacterial disease posing serious threats to the health of humans and animals. Currently, effective therapeutics available for brucellosis are in urgent demand. Brucellosis is transmitted to humans through diet or contact with infected animals. Several live‐attenuated vaccine strains are commercially available for animals, such as Brucella melitensis Rev.1, Brucella abortus S19, and RB51. However, no secure and effective vaccine for human beings has been developed yet against brucellosis, for which antibiotic therapy is the most frequently used and valid. Nevertheless, excessive antibiotics could prohibit the sensitivity of Brucella and develop resistance in humans. In addition, other clinical treatments, such as surgical treatment, have little effect on brucellosis. Therefore, novel treatment protocols should be explored. Based on numerous experimental studies, it is speculated that brucellosis can be fundamentally treated by inhibiting the activation of the type IV secretion system and changing the acidic environment of eBCV. In addition, previous literature showed that traditional Chinese medicine therapy also had certain efficacy, providing a new perspective angle for the exploration of treatment options.

Immunoassay-based evaluation of rOmp28 protein as a candidate for the identification of Brucella species.

Introduction. Brucellosis is an important bacterial zoonosis, re-emerging as a serious public health concern in developing countries. Two major species, Brucella melitensis and Brucella abortus, cause recurrent facile infection in human. Therefore, rapid and accurate diagnosis for early disease control and prevention is needed in areas with low disease burden.Hypothesis. This study evaluated the sandwich enzyme-linked immunosorbent assay (ELISA) (S-ELISA) immunoassay for potential use of whole-cell (WC) and recombinant outer-membrane protein (rOmp28)-derived IgG polyclonals in sensitive detection of Brucella.Aim. Immunoassay-based WC detection of Brucella species in important sub-clinical matrices at lower limits of detection.Methodology. We purified recombinant rOmp28 with Ni-NTA gel affinity chromatography and produced IgG polyclonal antibodies (pAbs) using BALB/c mice and New Zealand white female rabbits against different antigens (Ags) of Brucella. Checkerboard sandwich ELISA and P/N ratio (optical density of 'P' positive test sample to 'N' negative control) were used for evaluation and optimization of the study. The pAbs were characterized using Western blot analysis and different matrices were spiked with WC Ag of Brucella.Results. Double-antibody S-ELISA was developed using WC Ag-derived rabbit IgG (capture antibody at 10 µg ml-1) and rOmp28-derived mice IgG (detection antibody at 100 µg ml-1) with a detection range of 102 to 108 cells ml-1 and a limit of detection at 102 cells ml-1. A P/N ratio of 1.1 was obtained with WC pAbs as compared to 0.6 and 0.9 ratios with rOmp28-derived pAbs for detecting B. melitensis 16M and B. abortus S99, respectively. An increased P/N ratio of 4.4 was obtained with WC Ag-derived rabbit IgG as compared to 4.2>4.1>2.4 ratios obtained with rabbit IgGs derived against cell envelope (CE), rOmp28 and sonicated antigen (SA) of Brucella with high affinity for rOmp28 Ag analysed on immunoblots. The rOmp28-derived mice IgG revealed two Brucella species at P/N ratios of 11.8 and 6.3, respectively. Upon validation, S-ELISA detected Brucella WCs in human whole blood and sera samples with no cross-reactivity to other related bacteria.Conclusion. The developed S-ELISA is specific and sensitive in early detection of Brucella from different matrices of clinical and non-clinical disease presentation.

Effect of the Lacticaseibacillus paracasei JLM Strain Against Brucella abortus Strains in Ripened Cheese

This study evaluated the antagonistic effect of the Lacticaseibacillus paracasei JLM strain isolated from aguamiel, against Brucella abortus RB51, S19, and 2308 strains, during the manufacture of soft-ripened cheese. First, the tolerance of Lc. paracasei JLM was tested with pH values and bile salt concentrations for 3 h to simulate digestive tract conditions. The antagonistic effect against B. abortus strains was evaluated through double-layer diffusion and agar well diffusion assays. In addition, the stability of the cell-free supernatant (CFS) was tested with the agar well diffusion method under different conditions of temperature, pH, and treatment with digestive enzymes. Finally, the antagonistic effect against B. abortus strains was observed during the manufacture of ripened cheese for 31 days at 4°C and 25°C using the Lc. paracasei JLM strain as starter culture. The results showed that the Lc. paracasei JLM strain remains viable after exposure to different pH values (from 3.00 to 7.00) and concentrations of bile salts (from 0.5% to 7%). Moreover, the results demonstrate that the growth of the three B. abortus strains was inhibited in both antagonism tests and that CFS maintained 86% activity after heat treatment at 100°C, 121°C, or enzymatic digestion (proteinase K, trypsin, chymotrypsin), but it was inactivated at pH levels above 6. Finally, Lc. paracasei JLM completely inhibited the growth of B. abortus in ripened cheese at 25°C from day 17 and showed greater inhibition on the B. abortus RB51 strain in the ripened cheese at 4°C, showing statistical differences for the B. abortus S19 and B. abortus 2308 strains. The current research concluded that the Lc. paracasei JLM strain has an antagonistic effect on B. abortus, enhancing the potential of its use in the future as a probiotic.

Characterization of humoral and cellular immune responses elicited by reduced doses of Brucella abortus S19 (calfhood) vaccine in cattle calves of India

Brucella abortus S19 vaccine is a stable attenuated smooth strain, globally used as calfhood vaccine for the prevention of bovine brucellosis. Various agencies demonstrated different doses for vaccinating cattle and buffalo calves leading to ambiguity in selecting a suitable immune vaccine dose. The current study aimed at evaluating four graded doses of S19 vaccine to arrive at the dose which could produce comparable effectiveness as that of full dose prescribed by Indian Pharmacopeia among the Indian calves. Four vaccine doses of which the first dose consisted of full dose (40 × 109 CFU/dose) and the other three were 1/10th, 1/20th, 1/100th reduced doses along with control were tested. Each vaccine dose was administered to 13 cattle calves of 4–5 months of age maintained in separate groups. The blood samples were collected on 0 to 240 days post-vaccination (DPV) at the intervals of 0, 14, 28, 45, 60, 90, 150, 180 and 240 for assessment of vaccine-induced innate, humoral and cell-mediated immune responses. The sero-conversion of all vaccinated animals on DPV 45 and persistence of antibody till DPV 240 were noticed. No significant differences were observed in antibody response between animal groups that received full and 1/10th reduced doses. Innate and cell-mediated response by IL-6, TNF-α¸ IFN-γ, CD4+ and CD8+ cell counts showed dose-dependent responses with no significant difference between full dose and 1/10th reduced doses. The results suggest a possible one log reduction of full dose without compromising immune responses to aid larger vaccination coverage for creating herd immunity.

Brucella abortus in Kazakhstan, population structure and comparison with worldwide genetic diversity.

Brucella abortus is the main causative agent of brucellosis in cattle, leading to severe economic consequences in agriculture and affecting public health. The zoonotic nature of the infection increases the need to control the spread and dynamics of outbreaks in animals with the incorporation of high resolution genotyping techniques. Based on such methods, B. abortus is currently divided into three clades, A, B, and C. The latter includes subclades C1 and C2. This study presents the results of whole-genome sequencing of 49 B. abortus strains isolated in Kazakhstan between 1947 and 2015 and of 36 B. abortus strains of various geographic origins isolated from 1940 to 2004. In silico Multiple Locus Sequence Typing (MLST) allowed to assign strains from Kazakhstan to subclades C1 and to a much lower extend C2. Whole-genome Single-Nucleotide Polymorphism (wgSNP) analysis of the 46 strains of subclade C1 with strains of worldwide origins showed clustering with strains from neighboring countries, mostly North Caucasia, Western Russia, but also Siberia, China, and Mongolia. One of the three Kazakhstan strains assigned to subclade C2 matched the B. abortus S19 vaccine strain used in cattle, the other two were genetically close to the 104 M vaccine strain. Bayesian phylodynamic analysis dated the introduction of B. abortus subclade C1 into Kazakhstan to the 19th and early 20th centuries. We discuss this observation in view of the history of population migrations from Russia to the Kazakhstan steppes.

Safe Subunit Green Vaccines Confer Robust Immunity and Protection against Mucosal Brucella Infection in Mice.

Brucellosis is a zoonotic disease that causes significant negative impacts on the animal industry and affects over half a million people worldwide every year. The limited safety and efficacy of current animal brucellosis vaccines, combined with the lack of a licensed human brucellosis vaccine, have led researchers to search for new vaccine strategies to combat the disease. To this end, the present research aimed to evaluate the safety and efficacy of a green vaccine candidate that combines Brucella abortus S19 smooth lipopolysaccharide (sLPS) with Quillaja saponin (QS) or QS-Xyloglucan mix (QS-X) against mucosal brucellosis in BALB/C mice. The results of the study indicate that administering two doses of either sLPS-QS or sLPS-QS-X was safe for the animals, triggered a robust immune response, and enhanced protection following intranasal challenge with S19. Specifically, the vaccine combinations led to the secretion of IgA and IgG1 in the BALF of the immunized mice. We also found a mixed IgG1/IgG2a systemic response indicating evidence of both Th1 and Th2 activation, with a predominance of the IgG1 over the IgG2a. These candidates resulted in significant reductions in the bioburden of lung, liver, and spleen tissue compared to the PBS control group. The sLPS-QS vaccination had conferred the greatest protection, with a 130-fold reduction in Brucella burdens in lung and a 55.74-fold reduction in the spleen compared to PBS controls. Vaccination with sLPS-QS-X resulted in the highest reduction in splenic Brucella loads, with a 364.6-fold decrease in bacterial titer compared to non-vaccinated animals. The study suggests that the tested vaccine candidates are safe and effective in increasing the animals' ability to respond to brucellosis via mucosal challenge. It also supports the use of the S19 challenge strain as a safe and cost-effective method for testing Brucella vaccine candidates under BSL-2 containment conditions.

Molecular characterization of Brucella melitensis detected in blood and Serum samples of sheep and Goats

The incidence of brucellosis was investigated in sheep and goats with a history of abortions. Out of 550 blood samples collected from small ruminants, a total of 113 samples including 44 blood and 69 serum samples tested positive for brucellosis by using i-enzyme linked immunosorbent assay. Further, these samples were subjected to molecular detection of Brucella spp by BCSP31 and IS711/BM polymerase chain reaction (PCR) assays. The reference strains B. abortus S19, B. abortus 544 and B. melitensis Rev 1 were included in PCR assay as controls. The reference strains B. abortus S19 and B. abortus 544 produced amplification product of 223 bp. Moreover, 223 bp amplicon was observed in 12 (27.27%) blood and 14 (20.29%) serum samples confirming the presence of Brucella spp. In IS711/BM PCR B. melitensis Rev 1, 10 (22.73%) blood and 10 (14.49%) serum samples produced amplicons of 731 bp confirming the presence of B. melitensis. Phylogenetic analysis of four IS711/BM gene sequences of B. melitensis detected the ancestral origin of organism. It was inferred that BCSP31 and IS711/BM PCR assays were found useful in direct detection of Brucella organisms from whole blood and serum samples of sheep and goats.

Diagnosis of canine brucellosis using Rose Bengal plate test

Canine brucellosis or beagle fever is a zoonotic bacterial reproductive disease of dogs, caused by Brucella canis and occasionally by Brucella abortus, B. melitensis and B. suis. The actual seroprevalence of canine brucellosis in India is unknown and not yet studied in Kerala. A total of 131 animals presented to the outpatient unit of medicine, gynaecology and obstetrics of the two University Veterinary Hospitals at Mannuthy and Kokkala with clinical signs of epididymitis, orchitis, abortion, still birth, foetal resorption, foetal mummification, foetal maceration, neonatal death and infertility were randomly selected for the study. Paired sera samples were collected on the day of presentation and after three weeks of presentation for Rose Bengal Plate Test (RBPT). In this study, sera samples from forty-seven (35.88 per cent) infected dogs showed agglutination on RBPT using B. abortus S99 antigen. Out of forty-seven RBPT positive samples, 80.85 per cent (38/47) were female and 19.15 per cent (9/47) were male dogs. The high seroprevalence of canine brucellosis in this study is attributed to the endemicity of bovine brucellosis in the study area.

Differentially expressed long noncoding RNAs in RAW264.7 macrophages during Brucella infection and functional analysis on the bacterial intracellular replication.

Long noncoding RNAs (lncRNAs) are a group of functional RNA molecules without protein-coding potential and play vital roles in majority of biological processes. To date, the expression profiles of lncRNAs and their influence on Brucella replication in RAW264.7 cells are poorly understood. In this study, we performed high-throughput transcriptome analysis to investigate the differentially expressed lncRNAs associated with Brucella abortus S2308 infection. Of these, 8, 6, 130 and 94 cellular lncRNAs were differentially expressed at 4, 8, 24 and 48h post-infection, respectively. Moreover, 1918 protein-coding genes are predicted as potential cis target genes of differentially expressed lncRNAs by searching protein-coding genes located at upstream and downstream of lncRNA loci on the chromosome DNA of Mus musculus. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes analyses indicated that majority of lncRNA target genes were associated with B. abortus infection. Fourteen lncRNAs from transcriptome data were selected for qRT-PCR verification, confirming 13 were differentially expressed. Animal experiments revealed three were differentially expressed in vivo by qRT-PCR analysis. Furthermore, knockdown of LNC_000428 by CRISPR/dCas9 inhibition or Locked Nucleic Acids transfection downregulated Tnfrsf8 expression at mRNA level and increased Brucella intracellular replication. Thus, we provide a novel evidence that lncRNAs induced by Brucella-infection function on Brucella intracellular replication.

Brucellosis in the Brazilian Pantanal wetland: threat to animal production and wildlife conservation.

This review was performed to gather knowledge about brucellosis in livestock and wildlife in the Brazilian Pantanal, a biome with a huge biodiversity and extensive livestock production. Following the preferred reporting items for narrative review guidelines and using the terms "Brucella" and "Pantanal," we explored the PubMed, SciELO, Jstor, Science Direct, and Scholar Google databases. Information on host species, diagnostic test, number of positive animals, and positivity rates were acquired. Articles dating from 1998 to 2022 registered 14 studies including cattle, dogs, and the following wildlife species: Ozotoceros bezoarticus, Sus scrofa, Tayassu peccari, Nasua nasua, Cerdocyon thous, Panthera onca, Dasypus novemcintus, Cabassous unicinctus, Euphractus sexcinctus, Priodontes maximus, Myrmecophaga tridactyla and Hydrochoerus hydrochaeris. Brucella occurrence in cattle was demonstrated through the serological confirmatory test 2-mercaptoetanol. Molecular diagnosis detected Brucella abortus in dogs, smooth Brucella in O. beoarticus, and Brucella spp. in T. peccari. Cattle may have a pivotal importance in maintenance and spreading of Brucella spp. due to their high population density, environmental contamination from abortion of infected cows, and eventual excretion of B. abortus S19 strain from vaccinated heifers. The occurrence of Brucella spp. in O. bezoarticus and T. peccari represent a risk for wildlife conservation. These data indicate that Brucella spp. are enzootic in the Pantanal wetland, sustained by a reservoir system including domestic and wild mammals. Due to marked seasonality and large populations of wildlife species sharing same environments with livestock, brucellosis acquires great complexity in Pantanal and, therefore, must be analyzed considering both animal production and conservation.

Smooth and Rough Brucella Infections in Dogs

Background: B. canis infections have been reported in many countries of the world. B.canis can cause infections both in dogs and human beings. It can be transmitted to human via laboratory accidents during handling and contact with sick dogs. is an infectious agent for humans and it is transmitted to humans by various ways including laboratory accidents and contact with sick dogs. Methods: In this study serum samples were collected by using stratified random sampling method from the Diyarbakýr Metropolitan Municipality Animal Care and Rehabilitation Center, where approximately 600 dogs were housed. Result: Out of 83 samples, 9.6% and 26.5% of serum samples were found as positive by B.canis RSAT and B.canis ELISA, respectively. While 6% and 9.6% of the samples were positive by B. abortus S99 RSAT and B. abortus ELISA, respectively. Sensitivity and specificity of B. abortus S99 ELISA was 98% and 95.5%, respectively. On the other hand, sensitivity and specificity of B.canis M(-) ELISA was 96.6% and 94%. As a result, it was concluded that B. canis infection is common in dogs and this should be considered both for dogs and public health. Unfortunately, there is no standardized commercial serological test for diagnosis of B.canis and other rough Brucella species currently. This is the main reason that rough brucella infections might not represent the real B.canis infection situation. Therefore, most of the cases have the possibility of being undetected. For this reason it is utmost important that B.canis RSAT and ELISA can be used as routine tests in the hospitals and veterinary clinics in the serological diagnosis of brucellosis.