Abortus Strain Research Articles

Quantification of Brucella abortus population structure in a natural host

Cattle are natural hosts of the intracellular pathogen Brucella abortus, which inflicts a significant burden on the health and reproduction of these important livestock. The primary routes of infection in field settings have been described, but it is not known how the bovine host shapes the structure of B. abortus populations during infection. We utilized a library of uniquely barcoded B. abortus strains to temporally and spatially quantify population structure during colonization of cattle through a natural route of infection. Introducing 108 bacteria from this barcoded library to the conjunctival mucosa resulted in expected levels of local lymph node colonization at a 1-wk time point. We leveraged variance in strain abundance in the library to demonstrate that only 1 in 10,000 brucellae introduced at the site of infection reached a parotid lymph node. Thus, cattle restrict the overwhelming majority of B. abortus introduced via the ocular conjunctiva at this dose. Individual strains were spatially restricted within the host tissue, and the total B. abortus census was dominated by a small number of distinct strains in each lymph node. These results define a bottleneck that B. abortus must traverse to colonize local lymph nodes from the conjunctival mucosa. The data further support a model in which a small number of spatially isolated granulomas founded by unique strains are present at 1 wk postinfection. These experiments demonstrate the power of barcoded transposon tools to quantify infection bottlenecks and to define pathogen population structure in host tissues.

Whole genome de novo sequencing and comparative genomic analyses suggests that Chlamydia psittaci strain 84/2334 should be reclassified as Chlamydia abortus species

BackgroundChlamydia abortus and Chlamydia psittaci are important pathogens of livestock and avian species, respectively. While C. abortus is recognized as descended from C. psittaci species, there is emerging evidence of strains that are intermediary between the two species, suggesting they are recent evolutionary ancestors of C. abortus. Such strains include C. psittaci strain 84/2334 that was isolated from a parrot. Our aim was to classify this strain by sequencing its genome and explore its evolutionary relationship to both C. abortus and C. psittaci.ResultsIn this study, methods based on multi-locus sequence typing (MLST) of seven housekeeping genes and on typing of five species discriminant proteins showed that strain 84/2334 clustered with C. abortus species. Furthermore, whole genome de novo sequencing of the strain revealed greater similarity to C. abortus in terms of GC content, while 16S rRNA and whole genome phylogenetic analysis, as well as network and recombination analysis showed that the strain clusters more closely with C. abortus strains. The analysis also suggested a closer evolutionary relationship between this strain and the major C. abortus clade, than to two other intermediary avian C. abortus strains or C. psittaci strains. Molecular analyses of genes (polymorphic membrane protein and transmembrane head protein genes) and loci (plasticity zone), found in key virulence-associated regions that exhibit greatest diversity within and between chlamydial species, reveal greater diversity than present in sequenced C. abortus genomes as well as similar features to both C. abortus and C. psittaci species. The strain also possesses an extrachromosomal plasmid, as found in most C. psittaci species but absent from all sequenced classical C. abortus strains.ConclusionOverall, the results show that C. psittaci strain 84/2334 clusters very closely with C. abortus strains, and are consistent with the strain being a recent C. abortus ancestral species. This suggests that the strain should be reclassified as C. abortus. Furthermore, the identification of a C. abortus strain bearing an extra-chromosomal plasmid has implications for plasmid-based transformation studies to investigate gene function as well as providing a potential route for the development of a next generation vaccine to protect livestock from C. abortus infection.

Bioinformatics Analysis of B. Abortus Strain 2308 Protein and Its Drug Docking

Brucellosis is among the fast-spreading disease on Earth causing casualties in livestock as well as in humans. It’s an alarming situation to kick off the study of Brucellosis causing agent. Brucellosis is an infective sickness caused by Brucella abortus. Here, we have carried out the Insilco analysis of Brucella abortus strain 2308. This strain is mainly responsible for the disease. Here, we have tried to study the B. abortus strain 2308 by the means of bioinformatics methodology. We have run several Insilco tools to predict its structure and function. Moreover, we have carried out the methodology of protein homology modelling on this strain. Furthermore, we have also carried out several protein chain analyses, protein-protein interface, structure alignment, structure prediction and active site prediction. To make our study more productive we have also performed drug docking. These results will add on a little info in the emerging bioinformatics data regarding Brucella abortus.

Proposals for the Rational Control and Prevention of Bovine Brucellosis in Nigeria

In an earlier presentation and in relevant publications (Esuruoso, 1972; 1974), it was shown that Nigeria could be divided into free, moderately infected and heavily infected areas with regard to bovine brucellosis. In this paper, it is recommended that where there is little or no infection at the moment the disease could be prevented by annual serological and bacteriological surveillance and removal by culling of any positive reactions. In moderately infected areas the few reactors should be identified by branding after the appropriate tests have been performed. They should then be separated from clean animals and allowed to complete their reproductive life without contact with clean herds. Offsprings of the infected cattle which later turn out to be clean after necessary testing before they reach the breeding age, could then be mixed with the clean herd. Heifer calves in heavily infected herds should be inoculated with Brucella abortus strain 19 (S19) attenuated vaccine at the age of 3 to 6 months. Cows and inature heifers in these herds should be tested apuaily and separated into clean and infected groups. The infected group should be kept on separate pastures from the clean group. They should also be confined for calving and their manger beddings should be destroyed by incineration. At slaughter as well as during abortions and infective calvings, the Brucella abordus serotypes causing the disease should be isolated, identified completely and propagated for further study and vaccine production. Under no circumstance should indiscriminate vaccination be carried out on any herd in this country, as this will confuse the situation and confound any rational attempts at control or eradication. 

In search of a combined brucellosis and tuberculosis vaccine for cattle

Bovine brucellosis is caused by Brucella abortus. The bacterial pathogen causes economic losses because it induces abortion in cattle. Vaccination of calves with live B. abortus strain 19 induces a certain level of protection but induces persistent antibodies against cell envelope lipopolysaccharide that make it difficult to Distinguish Infected from Vaccinated Animals (DIVA). Live vaccine B. abortus strain RB51 was developed to eliminate such interfering antibodies and therefore, facilitate the differentiation of infected from vaccinated animals and help in the eradication of the disease. Vaccination with strain RB51 induces levels of protection similar to strain 19 but neither of the two vaccines give complete protection. We have been working to enhance protection induced by strain RB51 vaccine. Protective Brucella antigens can be over-expressed in strain RB51 by introducing a plasmid containing the leuB gene and the genes encoding such antigens. To avoid the expression of antibiotic resistance genes, we produced a leuB deficient strain RB51 and introduced a plasmid containing the leuB gene and the genes to be over-expressed. This new strain maintains the plasmid and has induced significantly high protection levels in mice. In addition, it allowed the construction of an RB51 vaccine strain able to express Mycobacterium bovis protective antigens so that the vaccine could protect against brucellosis and tuberculosis simultaneously.

Phylogeny of Brucella abortus strains isolated in the Russian Federation

Objective: To study Brucella (B.) abortus strains isolated in the Russian Federation, in order to identify their detailed position in the phylogenetic structure of the species global population as well as to determine genetic relationships for isolates from different geographical areas.Methods: Based on Bayesian method, the whole genome single-nucleotide polymorphism (SNP) analysis of 258 B. abortus strains from different geographical areas of the world including 20 B. abortus strains isolated in Russia was carried out.Results: The core genome SNP analysis of the B. abortus isolates allowed describing the main genetic lineages. The Russian strains entered two separate clades, including the basal branch and the C1 branch that is widely spread in Eurasia. The data on the isolation time was used for the dating of phylogenetic tree, and also the estimated time frame for the B. abortus genotype diversification was determined. There were sets of specific SNPs identified, which defined each of the genotypes and sub-genotypes.Conclusions: A significant genetic diversity of the brucellosis pathogen strains from Russia has been proven. The sets of unique specific SNPs described in our study may become one of the elements within a bio-informational analysis algorithm to be used for epidemiological study of brucellosis outbreaks, including those caused by new (atypical) genetic variants of B. abortus.

HUMORAL IMMUNE RESPONSE IN CROSS-BRED HEIFERS IMMUNIZED WITH BRUCELLA ABORTUS STRAIN RB51 VACCINE IN MILITARY DAIRY FARM OF BANGLADESH

Background: Brucellosis is a chronic zoonotic disease with negligible mortality rate that might be the reason not to attract the concerned authority to prevent and eradicate it in low income endemic countries. Recently, it has been recognized as a re-emerging zoonotic disease not only in low income countries but also its eradicated developed world. Objective: The main objective was to determine the humoral immune response (HIR) in crossbred dairy heifers immunized with Brucella abortus strain RB51 vaccine by using indirect ELISA Materials and Methods: Each of the 20 randomly selected B. abortus sero-negative crossbred (Holstein-Friesian  Local) dairy heifers aged between 4 to 8 months old at the Military Dairy Farm received 2.0 ml imported commercial B. abortus SRB51 strain vaccine subcutaneously in the neck region at day 0 and then booster dose at 60 days after first vaccination with similar dose and route during the period from June to October 2020. Each of the collected serum samples of 20 heifers at day 0, 7, 14, 21, 28, 60, 90, 120 and 150 was tested to detect the antibody status by using commercial indirect ELISA kit. Results: The humoral immune response (HIR) in terms of antibody levels detected by OD values in the serum of immunized cross-bred dairy heifers by using B. abortus strain RB51 commercial vaccine resulted 0.097  0.0032 (mean  SE) OD value at 0 day (i.e. pre-immunization) and 0.108 ± 0.0032 at 7th day. After that, the OD value started to rise from day 14 (OD value 0.124 ± 0.0032) and reached to a peak level at 60 days (OD value 0.223  0.0032) with the initial vaccination. Booster vaccination inoculated at 60 days resulted peak antibody level in terms of OD value (0.313  0.0032) at the day 90 and then the antibody level started to decline from 120 days (OD value 0.242  0.0032) to 150 days (OD value 0.199 0.0032) in cross-bred dairy heifers. Conclusions: This study suggests that the commercial B. abortus RB51 strain vaccine has induced satisfactory HIR with initial inoculation and significantly higher HIR produced with a booster dose in crossbred heifers by using commercial I-ELISA. The presence of Brucella antibodies have importance on sero-diagnosis whereas the cell mediated immunity (CMI) plays major role in protection against brucellosis which needs further investigation in cross-bred heifers in Bangladesh. Keywords: Brucellosis, SRB51 vaccine, Humoral immune response, I-ELISA, Cross-bred heifers, Bangladesh

Isolation and molecular identification of Brucella spp. in bovine herds kept at householders in the Delta region of Egypt by MALDI-TOF and AMOS-PCR.

Brucellosis is a widespread disease in Egypt which cause huge economic losses in the dairy industry. The present study aims at isolating and identifying Brucella (B.) spp. circulating in bovine and buffalo dairy herds kept at farmers houses in four districts of the Delta region of Egypt. One hundred and five tissue specimens were collected from seropositive cattle and buffaloes. The samples included 10 vaginal swabs, 3 placentas, 3 uteri and 86 supra-mammary lymph nodes from dams, as well as 3 stomach contents from aborted fetuses. Matrix-assisted laser desorption ionization (MALDI) and the conventional biotyping techniques were used for preliminary identification of isolates into the genus level. AMOS-PCR was applied to differentiate Brucella isolates into species level. Nineteen Brucella strains have been identified, four B. abortus strains were recovered from cattle and 15 B. melitensis strains were isolated from both cattle (n = 8) and buffaloes (n = 7). The predominant occurrence of B. melitensis in bovines raises the fact that B. melitensis clone can cross species barriers and can establish a permanent reservoir in cattle and buffaloes. Presence of culture-positive animals at householders represent a high-risk factor for human infection. This knowledge is of significant importance in the control of brucellosis in bovines.

Isolation of brucella strains in cattle from sedentary and nomadic communities and its public health implication

Background: Brucellosis is a highly infectious disease caused by bacteria of the genus brucella affecting animals leading to high economic loss and an impediment to livestock exportation. It also infects man with serious public health consequences. The disease is one of the world's most important neglected tropical zoonoses. Brucellosis is considered endemic in Nigeria and current information on isolation in sedentary and nomadic cattle is required. We carried out an active surveillance in sedentary cattle in Kachia Grazing Reserve (KGR), Kaduna State and in nomadic communities on the Jos Plateau to isolate brucella organisms and carry out phenotypic and molecular characterization of the isolates to species level. Methods & Materials: A total of 63 vaginal swabs, 36 milk samples, and 2 hygroma fluids were collected from cattle in KGR while 70 vaginal swabs, 50 milk samples and 2 hygroma fluids were collected from cattle on the Jos Plateau using a purposive sampling technique. Only animals that had history of abortion or infertility were sampled. They were cultured on serum dextrose agar with addition of antibiotic supplement (Oxoid) and incubated at 37 °C for 3 days in 5% CO2 atmosphere. Brucella strains isolated were biotyped along with eight archived isolates according to classical Brucella biotyping method. Isolates were further subjected to Bruce-ladder multiplex PCR according to established protocol. Results: Three and four B. abortus strains were isolated from KGR and Jos Plateau respectively. Biotyping revealed that all the 15 isolates were B. abortus biotype 3. Bruce-ladder multiplex PCR showed bands consistent with B. abortus. Conclusion: The study established the endemicity of brucellosis due to B. abortus biotype 3 in the two study areas. These findings have great veterinary and public health implications as healthy animals and the farmers are at high risk of infection due to contact with infected animals, consumption of raw milk and other products. Farmers were advised to boil their milk before consumption, isolate infected animals, maintain good level of hygiene and use protective wears in handling their animals as preventive measures. The study recommends vaccination of cattle against brucellosis in the areas.

Effect of Phage Targeting Therapy of Brucellosis on Host Antibody Response in Cattle.

Brucellosis caused by Brucella organisms is a major zoonosis globally. It causes heavy losses through abortions, delayed conception, and infertility in animals. Brucella is an intracellular bacterium. Antibiotic therapy for bovine Brucellosis is expensive and may sometimes be ineffective as Brucella can become resistant. Once infected, the animal may remain carrier and shed bacteria in milk, semen, and uterine discharges spreading infection to others for a long period. The live attenuated Brucella abortus strain S19 organisms that are commonly used as a vaccine were employed to deliver a broad acting lytic brucellaphage inside the phagocytes in vivo to reach the virulent Brucella hiding intracellularly. The phage pulsed vaccine induced sustained and significantly high titers of anti-Brucella antibodies compared with the untreated animals and animals vaccinated with S19 vaccine alone as estimated by standard tube agglutination test, microagglutination test, indirect hemagglutination assay test, and enzyme-linked immunosorbent assay. The current investigation is perhaps the first systematic attempt whereby efficacy of brucellaphage pulsed vaccine preparation in induction of specific antibody response was evaluated in cattle.

Seroprevalence, associated risk factors analysis and first molecular characterization of chlamydia abortus among Egyptian sheep

Chlamydia abortus is one of the most common abortive agents worldwide in sheep. Few studies have been reported C. abortus infection among sheep in Egypt but the available data is scarce. The objective of the present study was to determine the seroprevalence of C. abortus among sheep, the associated risk factors and its molecular characterization. The present study was conducted on 675 sheep in six Governorates at Northern Egypt. Data analysis confirmed the presence of antibodies against C. abortus in 93 out of 675 sheep. The logistic regression model was fitted to identify the associated risk factors with C. abortus infection. The results revealed that C. abortus increased significantly in ewes (OR = 4.04, 95 %CI: 1.44−11.28) during autumn season (OR = 3.6, 95 %CI: 1.64–8.28), in ewes with a history of abortion (OR = 1.4, 95 %CI: 0.87–2.50) and in farm where no lambing pen (OR = 2.2, 95 %CI: 1.30–3.94) or abscence of post abortion measures (OR = 1.96, 95%CI: 1.23-3.12). In addition, age, flock size and exchange of breeding ram had no significant effect on prevalence of chlamydiosis. Also, PCR assay was confirmed presence of C. abortus as accusative pathogen in aborted ewe and the genetic characterization of Egyptian C. abortus strain revealed 100 % identity with another strain from Iraq. A control program should be applied to reduce economic losses and risk of human infection.

Non-infectious outer membrane vesicles derived from Brucella abortus S19Δper as an alternative acellular vaccine protects mice against virulent challenge

The prime human and animal safety issues accentuate the search of promising newer alternative vaccine candidates to resolve complications associated with the live attenuated Brucella abortus strain19 (S19) vaccine. Outer membrane vesicles (OMVs S19 Δper) extracted from Brucella abortus S19Δper (S19Δper) as an alternative subunit vaccine candidate has been explored in the present study as OMVs are endowed with immunogenic molecules, including LPS and outer membrane proteins (OMPs) and do not cause infection by virtue of being an acellular entity. The LPS defective S19Δper released a higher amount of OMVs than its parent strain S19. Under transmission electron microscopy (TEM), OMVs were seen as nano-sized outward bulge from the surface of Brucella. Dynamic light scattering analysis of OMVs revealed that OMVs S19Δper showed the less polydispersity index (PDI) than OMVs S19 pointing towards relatively more homogenous OMVs populations. Both OMVs S19Δper and OMVs S19 with or without booster dose and S19 vaccine were used for immunization of mice and subsequently challenged with 2 × 105 CFU virulent Brucella abortus strain 544 (S544) to assess protective efficacy of vaccines. The less splenic weight index and less S544 count in OMVs immunized mice in comparison to unimmunized mice after S544 challenge clearly indicated good protective efficacy of OMVs. OMVs S19 Δper induced relatively high titer of IgG than OMVs S19 but conferred nearly equal protection against brucellosis. An ELISA based determination of IgG and its isotype response, Cytometric Bead Array (CBA) based quantitation of serum cytokines and FACS based enumeration of CD4+ and CD8+ T cells revealed high titer of IgG, production of both Th1 (IgG2a) and Th2 (IgG1) related antibodies, stimulation of IL-2, TNF (Th1) and IL-4, IL-6, IL-10 (Th2) cytokines, and induced T cell response suggested that OMVs S19Δper elicited Th1 and Th2 type immune response and ensured protection against S544 challenge in murine model.

Comparative study of in vitro prepared Rose Bengal Plate Test (RBPT) antigen with commercially available antigens

Background and aim: Brucellosis is one of the world most common zoonotic diseases. The current study was aimed to prepare the Rose Bengal Plate Test (RBPT) antigen for the diagnosis of brucellosis and to determine its specificity and sensitivity. Material and methods: The Rose Bengal plate test antigen prepared from Brucella abortus (B. abortus) strain 99 was compared with two commercial Rose Bengal Plate Test antigens and its specificity and sensitivity are determined. Results: The were Rose Bengal plate test and I-ELISA result show that the in vitro antigen was superior to RBPT antigen University Diagnosis Laboratory (UDL) Lahore Pakistan, and RBPT antigen Veterinary Laboratory Agency (VLA) UK. Out of 196 samples analyzed by in vitro RBPT antigen, RBPT antigen (UDL), RBPT antigen (VLA), and an indirect enzyme-linked immunosorbent assay (I-ELISA) 56.63 %, 53.57%, 41.84%, 35.71% were found B. abortus positively. The sensitivity calculated for the in vitro RBPT antigen was 96.62, while RBPT antigen (UDL) and RBPT antigen (VLA) were 89.77, 63.91 correspondingly. However, the specificity of the in vitro RBPT antigen was lower (77.57%), than the commercial RBPT antigen (VLA) (79.79%). Conclusions: A very sensitive and low-cost in vitro RBPT antigen compared to commercial RBPT was magnificently developed in the current study. It was determined that the in vitro RBPT antigen could substitute the available commercial RBPT antigen, which is comparatively expensive and less sensitive in the detection of brucellosis. Therefore, it is concluded that the in vitro RBPT antigen could be used for epidemiological surveillance of brucellosis.

Serological detection of brucellosis among febrile, malaria-negative children and domesticated dogs in an urban African setting.

BackgroundChildhood brucellosis and malaria are co-endemic febrile illnesses in some sub-Saharan African countries. Malaria and brucellosis co-infection or brucellosis sole infections are often missed due to an over emphasis on malaria and the lack of appropriate diagnostic infrastructure. Brucellosis in dogs is usually overlooked and yet there is extensive contact between humans and their pets.ObjectiveThis study investigated brucellosis in children and dogs using a confirmatory serological testing series that screens for three Brucella sp.MethodsResidual blood samples from malaria smear-negative febrile children were collected and tested for Brucella sp and malaria parasite. During the same period, residual blood samples presented to a veterinary microbiology laboratory in the same area were tested for brucellosis using the same approach.ResultsA total of 105 human and 80 canine blood samples were tested for brucellosis antibodies. The seroprevalence of brucellosis was 22.86% (25/105) in children and 1.3% (1/80) in dogs using the Card, buffered acidified plate antigen, and standard plate agglutination tests but was 0% using the rivanol precipitation plate agglutination test.ConclusionGiven that brucellosis can be caused by both smooth and rough colony strains, there is a need to modify the current serological surveillance strategy (targeted at only Brucella abortus and other smooth colony Brucella strains) to figure out the relative contribution of rough colony Brucella strains (B. ovis and B. canis). Since Uganda is endemic for brucellosis there is a need to modify the brucellosis surveillance strategy.

Immunization efficacy and safety of Brucella 104M against aerosol challenge in BALB/c mice

Objective: To evaluate the protective efficacy and safety of Brucella 104M against aerosol challenge in BALB/c mice and characterize its immunological effects. Methods: Female mice of 6-8 weeks old were immunized with Brucella abortus strain 104M by intratracheal aerosol delivery or intranasal instillation or subcutaneous injection route. Six mice of each group were sacrificed at 4, 8, 16, 24 weeks after immunization. At each time point, the clinical manifestations of mice were investigated, the serum, spleen and lung samples of mice were collected, body weight, spleen weight, bacteria loads in spleens, the anti-Brucella antibodies titers in serum and the cytokines concentrations of IFN-γ, IL-18 in serum or lung homogenate of the mice were detected. Twenty two weeks after immunization, all the mice were challenged with Brucella A19 through intratracheal aerosol delivery. Results: Compared with the control group, neither abnormal clinical symptoms nor significant changes in body weight were found in 104M immunization groups, at each time point when immunized through either nose dropping route, subcutaneous injection or aerosol routes; and the spleen weight of immunization groups were lower than control group after challenge (P<0.05): *M1 (0.26±0.16)g<M2 (0.40±0.19)g; *M3(0.21±0.11)g<M4(0.28±0.19)g; *M5(0.14±0.02)g<M6 (0.30±0.18)g. The spleen bacterium load of the mice in 104M immunization groups decreased until 20 weeks later, the cells were completely clear out, 2 weeks after challenge (24 weeks), the bacterium load increased again, the bacterium load in immunization groups were significantly lower than that in control group (P<0.05): *M1(4.49±0.13)log(10) (Colony-Forming Units) CFU/g<M2(6.90±0.46)log(10)CFU/g; *M3(3.59±1.06)log(10)CFU/g<M4(7.08±0.14) log(10)CFU/g; *M5(3.00±2.03)log(10)CFU/g<M6(6.81±0.34)log(10)CFU/g. The high titer of Brucella 104M specific antibodies were detected at week 4, and the peak was reached at week 8 of immunization groups. At each time point, the serum antibody titers IgG 2a was higher than that of IgG 1; before challenge, the cytokines concentrations of IFN-γ and IL-18 in all immunization groups were significantly higher than that in control group (P<0.05), after challenge, cytokines concentrations of IFN-γ and IL-18 in the serum were lower in all immunization groups than in control group (P<0.05). At each time point, IFN-γ, IL-1βand IL-18 concentrations in lung homogenate were higher in immunization groups than in control group (P<0.05). There were no significant differences among the three immunization routes in all sample detection indexes (P<0.05). Conclusions: Intratracheal aerosol delivery is a safe and effective immunization route in BALB/c mice. Both humoral immunity and cellular immunity were stimulated by Brucella 104M, showing obvious protective efficacy against aerosol challenge in BALB/c mice. No significant weight loss was detected. However, the colonizing of Brucella 104M in the spleen of mice was too long (20 weeks), and slighter spleen swelling of the mice were detected, which illustrated that the attenuated strain 104M has residual virulence as a vaccine strain in mice.

Molecular epidemiology of Brucella abortus strains from cattle in Inner Mongolia, China

Although the prevalence of brucellosis in Inner Mongolia Autonomous Region currently remains high, data available on the epidemiological of circulating Brucella abortus strains were limited. A total of 75 isolates obtained from cattle, sheep, and humans were analysed using both the classical method and multiple locus variable-number tandem repeat analysis (MLVA). There are at least three B. abortus biovars (1, 3 and 6) in this region, and B. abortus biovar 3 is the predominant one. Ten known MLVA-11 genotypes were identified, of which five genotypes (72, 75, 78, 82 and 210) were shared among strains from this study and others previously collected in two to seven different nations, suggesting that this population has multiple geographic origins. An MLVA-16 assay sorted the 75 B. abortus strains into two groups (I and II), 5 clusters (A–E) and 44 genotypes (GT1-44), with 26 unique genotypes represented by single isolates, indicating that these B. abortus brucellosis cases were not directly epidemiologically related. The remaining 18 shared genotypes (among a total of 47 isolates) were represented by two to eight isolates, suggesting that there were epidemiologically related pathogens from each shared genotype among the cases. Importantly, the cluster B1 branch including 22 cluster isolates with identical or similar genotypes confirmed the occurrence of a concentrated outbreak epidemic in the eastern region during 1988–1995. This work will contribute to better understanding of B. abortus brucellosis epidemiology in Inner Mongolia.

Prospects for Improving Quality Evaluation of the Live Brucellosis Vaccine in Terms of Specific Activity

Prophylactic immunisation against brucellosis is part of the National Immunisation Schedule for Epidemic Settings. The immunisation is performed with a live vaccine—a lyophilized suspension of the Brucella abortus strain 19 BА in a stabilizing medium. The paper presents the results of quality evaluation of 9 batches of live brucellosis vaccine that were submitted to the Testing Centre for Evaluation of Medicinal Immunobiological Products’ Quality of the Federal State Budgetary Institution “Scientific Centre for Expert Evaluation of Medicinal Products” of the Ministry of Health of the Russian Federation for assessment of the product’s compliance with the established specifications. The paper also presents the results of evaluation of the passport information provided by the manufacturer for these batches. There is no doubt about the need for objective quality evaluation of brucellosis vaccines as well as about the significance of its improvement.The aim of study was to assess the prospects for improving quality evaluation of live brucellosis vaccines in terms of Specific activity (concentration of microbial cells, number of living microbial cells, number of cutaneous doses).Materials and methods: specific activity (concentration of microbial cells and number of living microbial cells) was determined by visual and microbiological methods using the industrial reference standard of brucellosis vaccine OSO 42-28-396-2018, batch 6 and the bacterial suspension of the Brucella abortus strain 19 BА acquired from the joint stock company Scientific and Production Association “Microgen” in 2016. The number of cutaneous doses in the brusellosis vaccine was determined by the calculation method. Statistical processing of the results was performed using Microsoft Excel.Results: there was a mismatch between the brucella concentration coefficient of 1.7×109 microbial cells/mL determined by comparison with the industrial reference standard of bacterial suspension turbidity, 10 IU and the actual concentration of microbial cells obtained in the study. According to preliminary results, the brucella concentration coefficient corresponding to the industrial reference standard of bacterial suspension turbidity, 10 IU can reach 3.0×109 microbial cells/mL.Conclusions: the obtained results can serve as a basis for amending the data on the brucella concentration coefficient in the Passport and the Instructions for use of the industrial reference standard of bacterial suspension turbidity, 10 IU, as well as the Specific activity section (concentration of microbial cells, number of living microbial cells, number of cutaneous doses) of the established specifications for the brucellosis vaccine. Before amending the information on the brucella concentration corresponding to 10 IU in the Passport and the Instructions for use of the reference standard of bacterial suspension turbidity (OSO 42-28-85P), additional studies should be performed with other types of brucella.

MLVA-16 Genotyping of Brucella abortus and Brucella melitensis Isolates from Different Animal Species in Egypt: Geographical Relatedness and the Mediterranean Lineage.

Brucellosis is a common zoonotic disease in Egypt. However, there are limited data available on the genetic diversity of brucellae circulating in Egypt and other Mediterranean areas. One hundred and nine Brucella (B.) strains were isolated from different animal species in thirteen Egyptian governorates. Multi-locus variable number tandem repeats (VNTRs) analysis (MLVA-16) was employed to determine the geographical relatedness and the genetic diversity of a panel of selected Egyptian strains (n = 69), with strains originating from Italy (n = 49), Portugal (n = 52), Greece (n = 63), and Tunisia (n = 4). Egyptian B. melitensis strains clustered into two main clusters containing 21 genotypes. Egyptian B. abortus strains clustered into three main clusters containing nine genotypes. The genotypes were irregularly distributed over time and space in the study area. Egyptian strains of B. melitensis showed MLVA-16 patterns closer to that of Italian strains. Egyptian B. abortus strains isolated from cattle share the same genotype with strains from Portugal and similar to strains from Italy with low genetic diversity. Strains with similar MLVA patterns isolated from different governorates highlight the movement of the pathogen among governorates. Hence, it may also reflect the long endemicity of brucellosis in Egypt with earlier dispersal of types and great local genetic diversity. Open markets may contribute to cross-species transmission and dissemination of the new types nationwide. The presence of West Mediterranean lineages of B. melitensis and relatedness of B. abortus strains from the studied countries is a result of the socio-historical connections among the Mediterranean countries. Transnational eradication of brucellosis in the Mediterranean basin is highly demanded.

Immune responses and clinical effects of experimental challenge of elk with Brucella abortus strain 2308

To assess the effects of challenge dose and stage of gestation on infection and abortion, 35 elk were conjunctivally challenged with virulent Brucella abortus strain 2308 (S2308) during pregnancy. Seventeen elk were experimentally challenged early in the second trimester of gestation (December) with high (approximately 108 CFU) or low challenge (approximately 107 CFU) treatments having 8 and 9 pregnant elk, respectively. Other pregnant elk were experimentally challenged at a later challenge time (approximately early third trimester, February), with high and low challenge treatments having 8 and 10 elk, respectively. Conjunctival swabs from all animals were culture positive for the S2308 strain at 7 days after experimental challenge. All animals seroconverted on a B. abortus ELISA but optical density readings were not influenced (P > 0.05) by time of challenge or by challenge dosage. In the early challenge group, abortions occurred in 2 of 9 (22%) in the low challenge treatment and 3 of 8 (37%) in the high challenge treatment, whereas in the later challenge group, 1 of 8 (12.5%) in the low challenge treatment and 2 of 10 (20%) in the high challenge treatment aborted. The ability to recover B. abortus from samples obtained at necropsy did not differ (P > 0.05) between early and late challenges or between high and low challenge treatments. Despite the lack of abortions observed after experimental challenge, recovery from maternal tissues ranged from 50% (low dose, late challenge) to 77% (low dose, early challenge). Our data suggests that naïve elk do not abort as frequently after experimental infection with B. abortus strain 2308 as compared to similar data in cattle and bison.

First identification of a Brucella abortus biovar 4 strain from yak in Tibet, China

Brucellosis is one of the major zoonotic diseases in the world. In China, understanding on its causative agent Brucella is still limited. Recently, we isolated a Brucella strain XZ19-1 from yak in Lhasa, Tibet. Phenotypical characterization proved that it belongs to B. abortus biovar 4, a biotype that has never been reported in China. MLVA-16 genotyping revealed a novel profile (4-5-3-12-2-2-3-3-8-32-8-5-4-3-3-3) in this strain, while MLST sequence typing demonstrated that it belongs to ST 71. Furthermore, the whole genome of XZ19-1 strain was sequenced. Subsequent phylogenetic analysis demonstrated that XZ19-1was genetically more closely related to B. abortus strains originated from European countries rather than to those collected from China previously. Isolation and identification of XZ19-1 strain may thus indicate a unique Brucella lineage existing in Qing-Tibet plateau. These findings will help to improve the diagnosis and epidemiological studies of brucellosis in animals and human in this part of China.