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Identification, Cloning, and Characterization of Staphylococcus pseudintermedius Coagulase.

ABSTRACT: Coagulase activation of prothrombin by staphylococcus induces the formation of fibrin deposition that facilitates the establishment of infection by Staphylococcus species. Coagulase activity is a key characteristic of Staphylococcus pseudintermedius; however, no coagulase gene or associated protein has been studied to characterize this activity. We report a recombinant protein sharing 40% similarity to Staphylococcus aureus coagulase produced from a putative S. pseudintermedius coagulase gene. Prothrombin activation by the protein was measured with a chromogenic assay using thrombin tripeptide substrate. Stronger interaction with bovine prothrombin than with human prothrombin was observed. The S. pseudintermedius coagulase protein also bound complement C3 and immunoglobulin. Recombinant coagulase facilitated the escape of S. pseudintermedius from phagocytosis, presumably by forming a bridge between opsonizing antibody, complement, and fibrinogen. Evidence from this work suggests that S. pseudintermedius coagulase has multifunctional properties that contribute to immune evasion that likely plays an important role in virulence.

SUBMITTER: Sewid AH

PROVIDER: S-EPMC6056873 | biostudies-literature | 2018 Aug

REPOSITORIES: biostudies-literature

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Identification, Cloning, and Characterization of Staphylococcus pseudintermedius Coagulase.

Sewid Alaa H AH Hassan M Nabil MN Ammar A M AM Bemis David A DA Kania Stephen A SA

Infection and immunity 20180723 8

Coagulase activation of prothrombin by staphylococcus induces the formation of fibrin deposition that facilitates the establishment of infection by Staphylococcus species. Coagulase activity is a key characteristic of Staphylococcus pseudintermedius; however, no coagulase gene or associated protein has been studied to characterize this activity. We report a recombinant protein sharing 40% similarity to Staphylococcus aureus coagulase produced from a putative S. pseudinter ...[more]

PMID: 29891539

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Project description:BackgroundThe emergence of methicillin-resistant Staphylococcus aureus (MRSA) and methicillin-resistant Staphylococcus pseudintermedius (MRSP) have a significant health impact on people with direct or supportive occupations in veterinary medicine including veterinarians, animal handlers, laboratory personnel and pet owners.ObjectivesThis cross-sectional survey was conducted to determine the prevalence of and risk factors for S. aureus, S. pseudintermedius, MRSA and MRSP in dogs in Bangladesh.MethodsA total of 358 swab samples were collected from different body sites of 150 dogs attending a university teaching veterinary hospital between January and June 2018. Standard bacteriological procedures were followed to isolate Staphylococcus, and identification was confirmed to the species level by PCR to detect the nuc gene. MRSA and MRSP were confirmed by the presence of the mecA gene.ResultsThe prevalence of coagulase-positive S. aureus and S. pseudintermedius in dogs were 16% and 45.3%, respectively. S. aureus and S. pseudintermedius isolates displayed the highest resistance against nalidixic acid (95.2% and 91%, respectively) and erythromycin (89.3% and 84.7%, respectively). Notably, all the staphylococcal isolates showed resistance to ≥3 antimicrobial classes. The prevalence of MRSA and MRSP in dogs was 8.7% and 6%, respectively. Multivariable logistic regression analysis identified the following variables as risk factors for MRSA colonisation in dogs: dogs with dermatitis (odds ratio [OR], 12.24, 95% CI: 3.12-57.33; p < 0.001) and history of antibiotic use (OR 8.73, 95% CI: 2.23-43.10; p < 0.001). Presence of otitis (OR 14.22; 95% CI: 1.64-103.58; p = 0.008) and oral lesions (OR 9.48, 95% CI: 1.14-64.82; p = 0.002) were identified as the significant risk factors for the carriage of MRSP.ConclusionsThe circulation of multidrug-resistant S. aureus and S. pseudintermedius is a serious concern to dogs and humans. To our knowledge, this is the first report of S. pseudintermedius and MRSP affecting dogs in Bangladesh.

Project description:BackgroundStaphylococcus pseudintermedius is an opportunistic pathogen that is the major cause of pyoderma affecting dogs. Conventional antimicrobial treatment for infections caused by this organism have failed in recent years due to widespread resistance and alternative treatment strategies are a high priority. Protein A encoded in Staphylococcus aureus by spa protects the bacterium by binding IgG and acts as a superantigen. Staphylococcus pseudintermedius possess two genes orthologous to S. aureus spa, spsP, and spsQ.MethodsSpsQ and SpsQ-M, a non-toxigenic SpsQ, were cloned and expressed as recombinant proteins and their cytotoxic effect on canine B cells was measured. The neutralizing ability of antibody raised against them in clinically healthy dogs was evaluated.ResultsS. pseudintermedius SpsQ induced apoptosis of canine B cells. Specific amino acid substitutions diminished SpsQ-M binding to immunoglobulin and its super-antigenic activity, while its antigenicity was maintained. This recombinant, non-toxigenic S. pseudintermedius SpsQ stimulated the production of antibodies in dogs that specifically reacted with SpsQ and greatly diminished its cytotoxic effect on canine B cells.ConclusionsThe production of neutralizing antibody suggests that attenuated, non-toxic SpsQ produced in this study is a good candidate for inclusion in a vaccine for use in the treatment and prevention of S. pseudintermedius infections.AbbreviationsSpA: Staphylococcus aureus protein A; SpsP: Staphylococcus pseudintermedius protein A; SpsQ: Staphylococcus pseudintermedius protein A; SpsQ-M: attenuated Staphylococcus pseudintermedius protein A; MRSP: methicillin resistant Staphylococcus pseudintermedius; IgA: immunoglobulin A; IgG: immunoglobulin G; IgM: immunoglobulin M; VH: variable region of immunoglobulin heavy chain; IgBD: immunoglobulin binding domains; MFI: mean fluorescent intensity; SEM: standard error of the mean; PBMC: Peripheral blood mononuclear cells; CD21: complement receptor type 2; ST: Sequence type; OD: Optical density; ORF: open reading frame; PBS: Phosphate buffered saline; Tween 20: Polyethylene glycol sorbitan monolaurate 20; HRP: horseradish peroxidase; TMB- 3,3',5,5'-Tetramethylbenzidine.

Dataset Information

Identification, Cloning, and Characterization of Staphylococcus pseudintermedius Coagulase.

Publications

Identification, Cloning, and Characterization of Staphylococcus pseudintermedius Coagulase.

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