Veterinaria Italiana (Journal)
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P3-07 Structural study of type IV secretion system protein VirJ
No full textBrucella spp. survival and replication inside host cells requires a type IV secretion system (T4SS). The T4SS translocates effectors proteins into the host cell to modulate the intracellular fate of the bacterium in order to establish a secure niche were it actively replicates. Although many VirB substrates have been identified in Brucella, we still know very little about the secretion mechanism that mediate their translocation across the two membranes and the periplasmic space. In Gram negative bacteria, T4SSs are composed of 12 proteins, VirB1 through VirB11 and VirD4 according to the nomenclature used for the prototypal Agrobacterium tumefaciens VirB/D T4SS. VirD4 serves as cytoplasmic receptor for substrate binding to facilitate entry into the translocation channel. The T4SS of Brucella is homologous to the VirB/D system, but lacks the virD4 gene raising questions about effector translocation in this system. VirJ has been identified as a periplasmic protein essential for the translocation of two Brucella effectors BPE123 and SepA. These two effectors are initially secreted in the periplasm before being translocated to the host cell in a VirJ-dependent manner by the T4SS. Indeed, VirJ also interacts with T4SS components VirB5 and VirB8 and thus could facilitate delivery of selected effectors to the T4SS machinery. In this study we have investigated the structural properties of VirJby biochemistry and modelling. We first purified the VirJ protein to investigate its biochemical properties. Multi-angle light scattering (MALS) experiments show that VirJ is monomeric in solution domain. We then predicted the structure of monomeric VirJ using alfafold 2. The structure reveals that VirJ contain 2 domains separated by a short linker. Domain 2 adopts a hydrolase fold with eight b-sheet surrounded by six α helices that appears to be conserved in A. tumefaciens VirJ. Analysis of structure-sequence conservation points towards a potential active site composed of one serine, one histidine and one aspartate. We then present the results of VirJ-effectors interactions and discuss potential mechanism for VirJ-guided translocation
P9-01 Results of serological studies on brucellosis in wild boars and hares in Ukraine in 2020
No full textBrucellosis poses a significant danger for the health of humans and animals. Wild mammals, especially wild boars and hares, are the main reservoir of infection in nature. Ukraine is free of brucellosis in farm animals, but this disease still unknown in wild fauna. The aim of our work was to conduct serological research on brucellosis among wild boars and hares in different regions of Ukraine and analyze the results of the investigation. Authors present the results of serological screening performed by complement fixation test (CFT) and enzyme-linked immunosorbent assay (ELISA, ID.vet) over 2020. In total, 411 blood serum samples from forestries in 13 regions of Ukraine and 220 samples - from 24 regions, respectively, were tested and analyzed from wild boars and hares. In addition, the differential diagnosis on yersiniosis performed by the ELISA (INDICAL Bioscience). Positive reactions on brucellosis in wild boars were registered in 11 investigated oblasts. In summary, 21 (5.1%) sera samples tested positive to Brucella spp. by CFT. By ELISA, positive reactions detected much more - 64 (15.6%) due to the significant number of hemolyzed samples (109 samples tested only by ELISA). In hares, brucellosis antibodies were detected in animals from two oblasts. Positive reactions were registered in 1.8% of samples by CFT and in 1.4% by ELISA, respectively. However, because of possible cross-reactivity between antibodies for brucellosis and yersiniosis in brucellosis serological tests, we cannot exclude that some of the animals were infected by Yersinia enterocolitica O:9 and not Brucella spp. Antibodies were not detected in hare samples. The perspectives of further research regarding brucellosis in Ukraine should improve the differential diagnosis between brucellosis and yersiniosis. Future studies may include the use of molecular methods and expansion of epizootic monitoring in wild fauna of Ukraine
P2-06 PORIFERA - Core-genome-based analytical pipeline for prediction of new markers to rapid identification of emerging Brucella species
No full textSix new Brucella species isolated from humans, wild animals and environmental sources, were added to the six classical Brucella species potentiating an additional threat for humans. Other atypical Brucella strains isolated from wild rodents, frogs and fishes, will likely be proposed as new species in the future. This work aims to implement a core-genome-based analytical pipeline, PORIFERA, for prediction of new markers to rapid identification of emerging Brucella species. The comparison of core-genome at greater resolution of closely related Brucella spp. may provide information on specific genetic markers, such as single-nucleotide polymorphism (SNP). Twenty-three Brucella genomes (10 B. melitensis, seven B. suis and six B. abortus) were sequenced using Illumina technology. Genome assemblies were performed using de novo assembler SPAdes. The analysis involved the 23 newly sequenced genomes and 25 Brucella spp. complete genomes publicly available from eight of the 12 recognized species B. suis, B. abortus, B.canis, B. ovis, B. microti, B. pinnipedialis and B. ceti. Two Brucella sp. strains isolated from amphibians (09RB8910; 09RB8471) were used as outgroup. The alignment of core-genome was performed using Parsnp and the evolutionary history was inferred using Neighbor-Joining and Maximum Likelihood methods available in the Harvest suite (vs 1.1.2). Functional annotation had been accomplished to each SNP, as well as exclusion of intragenic SNPs in order to identify novel discriminatory and informative biomarkers. The evolutionary history was inferred from a total of 256 667 putative SNPs shared among the 46 more closely related genomes and the genomes of the amphibians isolates 09RB8910 and 09RB8471. From these, a total of 31034 SNPs were significantly associated with B.canis (n=2127), B. ovis (n=2018), B. suis (n=1782), B. abortus (n=1466), B. suis (n=1365), B. ceti (n=1463), B. pinnipedialis (n=891) and B. microti (n=786). These data will be further used for development of novel molecular methods to identify, genotype or direct assignment of Brucella species
O2-6 In silico pipeline for protein comparisons in Brucella genus
No full textAlthough species of Brucella genus share more than 90% of their genomic content, their phenotypic characteristics may vary greatly among species and even strains. To explain some of those differences, e.g. zoonotic potential, pathogenicity or immune response specificity, we devised the supervised machine-learning algorithm that compares the 3D protein structure, molecular weight, hydrophobicity, isoelectric point and structure stability. Based on the available whole chromosome sequences of B. abortus, B. melitensis, B. suis, B. ovis, B.canis and B. microti, the database was created, and for the efficacy of the model, only annotated proteins were used to predict the 3D structure based on the Swiss model system. Further, using the Bio.SeqUtils the protein structure characteristics were calculated and used for the comparisons. Our in silico results showed two membrane (cyst, cysW), urease and one heat-chock chaperon proteins characteristic for B. melitensis strains, while in other species, they have 100% homology. In B. ovis strains two cytochrome oxidase subunits II and two sugar metabolic proteins were constantly truncated and their structure stability, iso-electric point and hydrophobicity varied greatly compared to other Brucella species. Also, B. ovis strains in 95% of the cases were missing and the remaining 5% had very shortened transcriptional regulator; iron-sulfur, NAD binding and ATP binding enzymes; dsbA family protein. Carbohydrate hydrolase is characteristically truncated in B. microti, while imidazolonepropionase has B.canis specific properties. All strains of B. ovis and B. microti have similar ATP-dependent helicase HrpB that differs in its’ hydrophilic characteristics and iso-electric point compared to others, while these species completely miss lipids catabolismand cardiolipin sintase enzymes. Two proteins alcohol dehydrogenase AdhP and ornithine/lysine decarboxylase, were found to be present only in B. abortus, B. suis and B.canis, while only Cu(I)-responsive transcriptional regulator is 100% similar between three species. Differences in OMP 2a and OMP 31 among species were observed regarding molecular weights and protein stability. These in silico analyses are in concordance with published results, but further laboratory confirmations are necessary. However, our analysis used intra species differ-ences and, for now, we have not included differences among biovars, nor comparisons of active sites
P8-14 Brucellosis in Cameroon: Isolation, characterization, and prevalence of a zoonotic biothreat
No full textBrucellosis negatively affects agricultural economies as well as animal and human health throughout Africa. Historically, the presence of Brucella in Cameroon has not been confirmed, and the disease situation not fully understood, as previous studies were based solely on serological tests. In this study, we attempt to better understand the disease extent in the major animal species of agricultural and public health concern including cattle, sheep, goats, and pigs within the most important regions of Cameroon in terms of trade, movement, and animal numbers (Far North, North, and West). Currently, blood, milk, urine, vaginal swab, hygroma fluid, placenta, and lymph node samples are being collected from each of the livestock species at slaughterhouses across each region. Serum and milk are subject to the Rose Bengal Test (RBT) for serology. All samples are subject to culture, as well as real-time PCR (RT-PCR). Countrywide, approximately 88% of sample collection, 82% RBT, 75% culture, and 9% RT-PCR has been completed. So far, RBT suggests a countrywide seropositivity of 6.79% for cattle, sheep (1.10%), goats (2.24%), and pigs (2.81%). Culture has resulted in 0.65% positivity for cattle and 0% in the other species. Although RT-PCR has only been conducted on 393 total animals, 10 cattle (5 North, 3 West, and 2 Far North) and 1 goat (West) have tested positive for Brucella abortus. To this point in our study, it appears that B. abortus is endemic across the country. Interestingly, despite the initial assumption that B. melitensis and B. suis are present, we have not yet identified the presence of these species. Furthermore, generalization that B. melitensis is a significant problem throughout Africa should not be assumed. Therefore, this study reveals that control strategies in Cameroon should likely be directed at cattle farming and the associated supply chain countrywide. Forthcoming, we will complete sampling and diagnostic assays, sequence DNA using whole genome sequencing, and conduct a region-specific epidemiologic risk investigation. Our overall findings within this project will serve to enhance the capabilities of the Cameroonian government, as well as human and animal health services to prevent, detect, and respond to the apparent brucellosis biothreat
P8-15 Sustainable brucellosis control programme
No full textBrucellosis is endemic in the Republic of Azerbaijan. There was a testing and slaughter program in the country; but with about 740,000 small farms with livestock scattered in about 4,300 villages, it was very difficult to enforce. Since 2015, within the framework of the Agricultural Competitiveness Improvement Project, the strategy has been changed to universal vaccination of small ruminants and cattle. We carried out a series of cross-sectional epidemiologic studies of the disease in livestock. The series started in 2009 with a pilot study in four regions, and its efficacy was evaluated in 2015 when a seroprevalence study was conducted in 51 regions which revealed a reduction in small ruminant seroprevalences from 3.3% to 0.14% in pilot regions. Another seroprevalence study was conducted in early 2020 in 25 randomly selected regions. The pilot study program of vaccination with conjunctival Rev.1 of all non-pregnant female small ruminants of breeding age and all females between 3 and 8 months was expanded to the whole country in 2015 and was again associated with a reduction in small ruminant seroprevalences when assessed in 2020. The overall RR for all species in a comparison of the 2015 and 2020 surveys in 25 regions was 0.8% (0.7, 0.9). Differences between overall prevalence in individual regions in both surveys were small in most cases. This may havebeen due to sampling variability in selection of livestock owners at different locations and variability in prevalence and proportions of species of livestock. Small ruminant seroprevalences were lower in 2020 than in 2015. The pilot study program of annual vaccination with conjunctival administered Rev.1 to all female small ruminants between 3 and 8 months old and all non-pregnant females of breeding age in the first two years conducted between 2009 and 2015 resulted in lower brucellosis seroprevalence leading to the adoption of a national control program using the same vaccination strategy. Vaccination of cattle began in 2017 with vaccination of female calves 3 to 8 months of age and was expanded in 2020 to include vaccination of adult non-pregnant females
P10.2 Beyond mapping hotspots: using geospatial models to guide surveillance against antimicrobial resistance in animals.
No full textThe rise of antimicrobial resistance (AMR) in food animals is a growing threat for animal health, and potentially for human health. China, as the world’s largest user of veterinary antimicrobials could play a pivotal role in leading the international response to AMR, and maps of AMR could help coordinate this response (Zhao et al., 2021). Here, we used geospatial models to identify regions where future surveillance effort could be targeted in priority.
We extracted resistance rates from 446 point-prevalence surveys on Escherichia coli, Salmonella, Campylobacter, and Staphylococcus aureus. We mapped the proportion of drugs tested with resistance rates higher than 50%. The uncertainty map associated with these predictions was used to identify locations where surveillance efforts could be intensified in the future. We calculated an index of ‘necessity for additional surveillance’ - the product between uncertainty in AMR level and animal densities- to identify locations where 50 surveys could be conducted such as to minimize uncertainty on future AMR levels in China. We computed both exact and approximate solution for the spatially optimal allocation of surveys to improve AMR surveillance in China.
In China, AMR levels were the highest in the east and lowest in the southwest. Regions that would benefit the most from increased surveillance were the southwest (21/50 surveys) and northeast (11/50 surveys). Using geographically targeted surveillance could reduce uncertainty of AMR level by 104% compared with an equal surveillance effort across administrative divisions. Our findings help outline priorities for AMR surveillance in China, and identify where future surveys could best improve the accuracy of AMR maps
S01 Communication Breakdown – Of disease clusters, a trillium and One Health
No full textOne health is a concept and an approach to promote the health of human and animal populations in a shared and healthy environment. This interdisciplinary endeavour requires collaboration across professions and a common language. Geographic epidemiology is concerned with the identification of spatial patterns of health related states and events in a population defined via the boundary of the study area. Such spatial patterns (trend, cluster and clustering) require clear definition to be meaningful in science communication. However, the term “disease cluster” has been defined in the literature in various and rather different ways. When geographic epidemiology is unable to make sense of its own concepts it is questionable how respective research results can benefit one health. The goal of this study is to clarify the concept of disease cluster within geographic epidemiology.Examples of disease cluster definitions from the literature will be compared. The term disease cluster will be semantically distinguished from similar concepts: clustering, hot spot and outbreak. The epidemiological triangle of causation (agent, host and environment) will be used to conceptualize geographic epidemiological data analysis. The semantics and statistical meaning of expectation and prediction will be reviewed and play a key role. The true meaning of a disease cluster will be identified as embedded within the well-known Besag-York-Mollié (BYM) regression model. All the spatial patterns identifiable from the BYM model will be conceptualized in a new paradigm: the geographic epidemiological trillium of causation. The concept of a disease cluster is clarified and a definition proposed. The paradigm of the geographic epidemiological trillium will help guiding causal epidemiological research. And researchers will be equipped to ask more precise questions and communicate their results more clearly to those who need to know
S03 Impact of climate change on soil-borne and vector-borne diseases
No full textVector-borne, water-borne and soil-borne diseases are climate sensitive. Temperature affects the development of arthropods and helminths. Rainfall provides suitable breeding sites for mosquitoes and suitable soil moisture conditions for the development of helminths on pasture. Climate change, through its impact on global temperature and weather extremes is already affecting the health of humans and animals directly, and will have indirect impacts on the transmission of vector-borne, water-borne and soil-borne diseases. Over the past decade, various mathematical risk models driven by climate change scenarios have been used to simulate and anticipate future changes in disease burden. This presentation will review recent advances in our understanding of the impact of climate change on vector-borne diseases affecting animals.
Bluetongue disease is a viral disease transmitted by Culicoides midges to ruminants. A Bluetongue epidemic significantly affected livestock in Northern Europe in 2006-07. Modeling research highlighted the impact of a heatwave on increased risk of Bluetongue disease transmission in Northern Europe. Future climate change could enhance this risk further over this region (Guis et al., 2012); however, the use of standard control zones and subsequent restriction of animal movements could mitigate such risk (Jones et al., 2019). Fasciola hepatica, also known as the common liver fluke, is a parasitic trematode that affects ruminants in Europe. Fasciolosis has large economical impacts on the livestock sector. The parasite requires the presence of a freshwater snail to complete its life cycle. Infectious metacercariae and the intermediate snail host are favoured by wet weather conditions and soil moisture on grassland. Modelling studies highlight that climatic suitability for fasciolosis increased during the 2000s over northern Europe. Recent trends are likely to continue in future. The liver fluke transmission season could lengthen by 3-4 months in some locations (Caminade et al., 2015). Other examples of climate sensitive animal diseases, such as lungworms and Rift Valley Fever, as well as other non-climatic drivers will also be discussed. Finally, future research directions for this field of research will be suggested
P10.3 Multicriteria geospatial analysis of the risk of avian influenza in Cuba
No full textInfluenza A viruses, especially because of their capacity for genetic reassortment and mutagenesis as RNA viruses, and the possibility of cross-species transmission, are considered among the most challenging viruses that threaten human and animal health. The persistence of Highly Pathogenic Avian Influenza A (H5N1) virus and the emergence of multiple novel Avian Influenza viruses in Asia, the Americas, and Europe in 2022 raised concerns in the global community about their pandemic potential (Mosaad et al., 2023). This work aimed to identify opportunities for improvements in the active surveillance system for avian influenza established in Cuba. Risk factors associated with the occurrence of the disease (Leon et al., 2009; Steven et al., 2013; Ferrer et al., 2013) were used to perform a geospatial multi-criteria analysis with a resolution of one km2. Additionally, the existence of contiguous areas between poultry farms (< 3 km2) where the spread of the causal agent could be favored in case of introduction was taken into account. As a result, areas with a very high risk of occurrence were identified, either by exposure or by spread, which was sometimes favored by the contiguity between commercial poultry farms. Accuracy and management of the risk of spread are of great importance because they are often the main determinant of the magnitude of the epidemic. It is concluded that there are strategic and highly important areas where significant resources should be prioritized to strengthen biosecurity surveillance and early warning