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Comparative genome analysis of Salmonella enterica serovar Gallinarum biovars Pullorum and Gallinarum decodes strain specific genes.

ABSTRACT: Salmonella enterica serovar Gallinarum biovar Pullorum (bvP) and biovar Gallinarum (bvG) are the etiological agents of pullorum disease (PD) and fowl typhoid (FT) respectively, which cause huge economic losses to poultry industry especially in developing countries including India. Vaccination and biosecurity measures are currently being employed to control and reduce the S. Gallinarum infections. High endemicity, poor implementation of hygiene and lack of effective vaccines pose challenges in prevention and control of disease in intensively maintained poultry flocks. Comparative genome analysis unravels similarities and dissimilarities thus facilitating identification of genomic features that aids in pathogenesis, niche adaptation and in tracing of evolutionary history. The present investigation was carried out to assess the genotypic differences amongst S.enterica serovar Gallinarum strains including Indian strain S. Gallinarum Sal40 VTCCBAA614. The comparative genome analysis revealed an open pan-genome consisting of 5091 coding sequence (CDS) with 3270 CDS belonging to core-genome, 1254 CDS to dispensable genome and strain specific genes i.e. singletons ranging from 3 to 102 amongst the analyzed strains. Moreover, the investigated strains exhibited diversity in genomic features such as virulence factors, genomic islands, prophage regions, toxin-antitoxin cassettes, and acquired antimicrobial resistance genes. Core genome identified in the study can give important leads in the direction of design of rapid and reliable diagnostics, and vaccine design for effective infection control as well as eradication. Additionally, the identified genetic differences among the S. enterica serovar Gallinarum strains could be used for bacterial typing, structure based inhibitor development by future experimental investigations on the data generated.

SUBMITTER: Vaid RK

PROVIDER: S-EPMC8375982 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Salmonella enterica serovar Gallinarum biovars Gallinarum and Pullorum cause severe chicken salmonellosis, a disease associated with high mortality and morbidity among chickens worldwide. The conventional serotyping and biochemical reactions have been used to identify Salmonella serovars. However, the conventional methods are complicated, time-consuming, laborious, and expensive. Furthermore, it is challenging to distinguish S. Gallinarum and S. Pullorum via biochemical assays and serotyping because of their antigenic similarity. Although various PCR methods were established, a PCR protocol to detect and discriminate S. Gallinarum and S. Pullorum simultaneously is lacking. Herein, a one-step multiplex PCR method was established for the accurate identification and discrimination of S. Pullorum and S. Gallinarum. Three specific genes were used for the multiplex PCR method, with the I137_14445 and ybgL genes being the key targets to identify and differentiate S. Gallinarum and S. Pullorum, and stn being included as a reference gene for the Salmonella genus. In silico analysis showed that the I137_14445 gene is present in all Salmonella serovars, except for S. Gallinarum, and could therefore be used for the identification of S. Gallinarum. A 68-bp sequence deficiency in ybgL was found only in S. Pullorum compared to other Salmonella serovars, and this could therefore be used for the specific identification of S. Pullorum. The developed PCR assay was able to distinguish S. Gallinarum and S. Pullorum among 75 various Salmonella strains and 43 various non-Salmonella pathogens with excellent specificity. The detection limit for the genomic DNA of S. Gallinarum and S. Pullorum was 21.4 pg./μL, and the detectable limit for bacterial cells was 100 CFU. The developed PCR method was used for the analysis of Salmonella isolates in a chicken farm. This PCR system successfully discriminated S. Gallinarum and S. Pullorum from other different Salmonella serovars. The PCR results were confirmed by the conventional serotyping method. The newly established multiplex PCR is a simple, accurate, and cost-effective method for the timely identification and differentiation of S. Pullorum and S. Gallinarum.

Project description:Salmonella enterica serovar Gallinarum biovars Pullorum (S. Pullorum) and Gallinarum (S. Gallinarum) can result in pullorum disease and fowl typhoid in avian species, respectively, and cause considerable economic losses in poultry in many developing countries. Conventional Salmonella serotyping is a time-consuming, labor-intensive and expensive process, and the two biovars cannot be distinguished using the traditional serological method. In this study, we developed a rapid and reliable one-step multiplex polymerase chain reaction (PCR) assay to simultaneously identify and discriminate the biovars Pullorum and Gallinarum. The multiplex PCR method focused on three specific genes, stn, I137_08605 and ratA. Based on bioinformatics analysis, we found that gene I137_08605 was present only in S. Pullorum and S. Gallinarum, and a region of difference in ratA was deleted only in S. Pullorum after comparison with that of S. Gallinarum and other Salmonella serovars. Three pairs of primers specific for the three genes were designed for the multiplex PCR system and their selectivity and sensitivity were determined. The multiplex PCR results showed that S. Pullorum and S. Gallinarum could be identified and discriminated accurately from all tested strains including 124 strains of various Salmonella serovars and 42 strains of different non-Salmonella pathogens. In addition, this multiplex PCR assay could detect a minimum genomic DNA concentration of 67.4 pg/μL, and 100 colony forming units. The efficiency of the multiplex PCR was evaluated by detecting natural-occurring Salmonella isolates from a chicken farm. The results demonstrated that the established multiplex PCR was able to identify S. Gallinarum and S. Pullorum individually, with results being consistent with traditional serotyping and biochemical testing. These results demonstrated that a highly accurate and simple biovar-specific multiplex PCR assay could be performed for the rapid identification and discrimination of Salmonella biovars Gallinarum and Pullorum, which will be useful, particularly under massive screening situations.

Project description:Salmonella enterica subspecies enterica serovar Gallinarum biovar Pullorum (S Pullorum) is the etiological agent of pullorum disease, causing white diarrhea with high mortality in chickens. There are many unsolved issues surrounding the epidemiology of S Pullorum, including its origin and transmission history as well as the discordance between its phenotypic heterogeneity and genetic monomorphism. In this paper, we report the results of whole-genome sequencing of a panel of 97 S Pullorum strains isolated between 1962 and 2014 from four countries across three continents. We utilized 6,795 core genome single nucleotide polymorphisms (SNPs) to reconstruct a phylogenetic tree within a spatiotemporal Bayesian framework, estimating that the most recent common ancestor of S Pullorum emerged in ?914 CE (95% confidence interval [95%CI], 565 to 1273 CE). The extant S Pullorum strains can be divided into four distinct lineages, each of which is significantly associated with geographical distribution. The intercontinental transmissions of lineages III and IV can be traced to the mid-19th century and are probably related to the "Hen Fever" prevalent at that time. Further genomic analysis indicated that the loss or pseudogenization of functional genes involved in metabolism and virulence in S Pullorum has been ongoing since before and after divergence from the ancestor. In contrast, multiple prophages and plasmids have been acquired by S Pullorum, and these have endowed it with new characteristics, especially the multidrug resistance conferred by two large plasmids in lineage I. The results of this study provide insight into the evolution of S Pullorum and prove the efficiency of whole-genome sequencing in epidemiological surveillance of pullorum disease.IMPORTANCE Pullorum disease, an acute poultry septicemia caused by Salmonella Gallinarum biovar Pullorum, is fatal for young chickens and is a heavy burden on poultry industry. The pathogen is rare in most developed countries but still extremely difficult to eliminate in China. Efficient epidemiological surveillance necessitates clarifying the origin of the isolates from different regions and their phylogenic relationships. Genomic epidemiological analysis of 97 S Pullorum strains was carried out to reconstruct the phylogeny and transmission history of S Pullorum. Further analysis demonstrated that functional gene loss and acquisition occurred simultaneously throughout the evolution of S Pullorum, both of which reflected adaptation to the changing environment. The result of our study will be helpful in surveillance and prevention of pullorum disease.

Project description:Pullorum disease is one of the most common diarrhea-related diseases caused by Salmonella enterica subspecies enterica serovar Gallinarum biovar Pullorum (S Pullorum); it negatively affects the poultry industry. However, limited studies have explored the association between the gut microbiota and S Pullorum infection in chickens. In the present study, we performed a microbiome comparison and a microbiome genome-wide association study (mGWAS) to investigate the association among the host genetics, the gut microbiota, and pullorum disease in chickens. We found that S Pullorum infection in chickens could alter the abundance of 39 bacterial genera (P < 0.05). The altered structure and composition of the gut microbiota were also detected in the offspring. mGWAS results revealed host genetic variants to be prominently associated with gut microbial diversity and individual microbes. The pathogens Pelomonas and Brevundimonas, which had a high abundance in positive parent chickens and their offspring, were significantly associated with several genetic mutations in immunity-related genes, such as TGIF1, TTLL12, and CCR7 This finding explained why Pelomonas and Brevundimonas were heritable in S Pullorum-infected chickens. The heritable gut microbes and identified genetic variants could provide references for the selection of resistant chickens and the elimination of pullorum disease.IMPORTANCE The present study investigated the association among the host genome, the gut microbiome, and S Pullorum infection in chickens. The results suggested that the gut microbial structure is altered in S Pullorum-infected chickens. The diversity and abundance of the gut microbiota remarkably differed between the offspring coming from S Pullorum-positive and S Pullorum-negative chickens. Heritable gut microbiota were detected in the offspring. Moreover, host genetic variants were associated with microbial diversity and individual gut microbes. The pathogens Pelomonas and Brevundimonas, which exhibited a high heritability in S Pullorum-positive parents and their offspring, were associated with several genetic mutations in immunity-related genes.

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Comparative genome analysis of Salmonella enterica serovar Gallinarum biovars Pullorum and Gallinarum decodes strain specific genes.

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