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Development of OMP based indirect ELISA to gauge the antibody titers in bovines against Pasteurella multocida.

ABSTRACT: Pasteurella multocida (P. multocida) is an important pathogen of various domestic animals. The outer membrane proteins (OMPs) play a major role in pathogenesis and immunogenicity of P. multocida. The aim of the study was to develop indirect enzyme linked immuno sorbant assay (ELISA) based on OMPs to ascertain the antibody titers in animals post-infection or to gauge the potency of vaccine. The OMPs were extracted and purified from P. multocida P:52 (vaccine strain) and P. multocida B:2 isolated from natural outbreak of Haemorrhagic septicaemia (HS) and analyzed on SDS PAGE and through western blot. The OMPs profile of the vaccine strain and the isolate from the natural outbreak of HS were found to be similar. Optimization of various components viz. coating antigens, anti-species conjugate, etc. were carried out against both anti-P. multocida hyper immune and pre immune serum. Validation of OMP based indirect ELISA assay to measure immune response against P. multocida in bovine revealed 91% diagnostic sensitivity (DSN) and about 100% diagnostic specificity (DSP) at 25% cut off. OMP based indirect ELISA was found to be more specific, but less sensitive as compared to WCL based assay.

SUBMITTER: Dogra V

PROVIDER: S-EPMC4782674 | biostudies-literature | 2015

REPOSITORIES: biostudies-literature

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Development of OMP based indirect ELISA to gauge the antibody titers in bovines against Pasteurella multocida.

Dogra V V Verma S S Singh G G Wani A H AH Chahota R R Dhar P P Verma L L Sharma M M

Iranian journal of veterinary research 20150101 4

Pasteurella multocida (P. multocida) is an important pathogen of various domestic animals. The outer membrane proteins (OMPs) play a major role in pathogenesis and immunogenicity of P. multocida. The aim of the study was to develop indirect enzyme linked immuno sorbant assay (ELISA) based on OMPs to ascertain the antibody titers in animals post-infection or to gauge the potency of vaccine. The OMPs were extracted and purified from P. multocida P:52 (vaccine strain) and P. multocida B:2 isolated ...[more]

PMID: 27175202

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Project description:ObjectivePasteurella multocida is a widespread zoonotic pathogen that causes severe damage to the poultry industry. This study focused on the antibacterial effects and mechanism of action of coptisine against P. multocida.MethodsThe minimum inhibitory concentration and half maximal inhibitory concentration of coptisine against P. multocida was measured. Additionally, the effect of coptisine on growth, cell wall, activity of respiratory enzymes, soluble protein content and DNA synthesis were also analyzed. Finally, the effect of coptisine on gene transcription was determined using RNA sequencing.ResultsWe demonstrated that coptisine has a strong antibacterial effect against P. multocida, with a minimum inhibitory concentration of 0.125 mg/mL. Moreover, the measurement of the half maximal inhibitory concentration confirmed that coptisine was safe for the pathogen. The growth curve showed that coptisine inhibited bacterial growth. Measurement of alkaline phosphatase activity in the culture solution showed that coptisine affected cell wall permeability. Transmission electron microscopy revealed that coptisine chloride destroyed the cell structure. In addition, coptisine blocked the respiratory system, as measured by the levels of critical enzymes of the tricarboxylic acid cycle and glycolysis, succinate dehydrogenase and lactate dehydrogenase, respectively. Similarly, coptisine inhibited the synthesis of soluble proteins and genomic DNA. The KEGG pathway analysis of the differentially expressed genes showed that they were associated with cellular, respiratory, and amino acid metabolism, which were downregulated after coptisine treatment. Additionally, genes related to RNA degradation and the aminoacyl-tRNA pathway were upregulated.ConclusionIn this study, we demonstrated that coptisine exerts an antibacterial effect on P. multocida. These findings suggest that coptisine has a multifaceted impact on various pathways, resulting in the inhibition of P. multocida. Thus, coptisine is a potential alternative to antibiotics for the treatment of P. multocida infections in a clinical setting.

Dataset Information

Development of OMP based indirect ELISA to gauge the antibody titers in bovines against Pasteurella multocida.

Publications

Development of OMP based indirect ELISA to gauge the antibody titers in bovines against Pasteurella multocida.

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