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Detection and sequence analysis of accessory gene regulator genes of Staphylococcus pseudintermedius isolates.

ABSTRACT: BACKGROUND:Staphylococcus pseudintermedius (SP) is the major pathogenic species of dogs involved in a wide variety of skin and soft tissue infections. The accessory gene regulator (agr) locus of Staphylococcus aureus has been extensively studied, and it influences the expression of many virulence genes. It encodes a two-component signal transduction system that leads to down-regulation of surface proteins and up-regulation of secreted proteins during in vitro growth of S. aureus. The objective of this study was to detect and sequence analyzing the AgrA, B, and D of SP isolated from canine skin infections. MATERIALS AND METHODS:In this study, we have isolated and identified SP from canine pyoderma and otitis cases by polymerase chain reaction (PCR) and confirmed by PCR-restriction fragment length polymorphism. Primers for SP agrA and agrBD genes were designed using online primer designing software and BLAST searched for its specificity. Amplification of the agr genes was carried out for 53 isolates of SP by PCR and sequencing of agrA, B, and D were carried out for five isolates and analyzed using DNAstar and Mega5.2 software. RESULTS:A total of 53 (59%) SP isolates were obtained from 90 samples. 15 isolates (28%) were confirmed to be methicillin-resistant SP (MRSP) with the detection of the mecA gene. Accessory gene regulator A, B, and D genes were detected in all the SP isolates. Complete nucleotide sequences of the above three genes for five isolates were submitted to GenBank, and their accession numbers are from KJ133557 to KJ133571. AgrA amino acid sequence analysis showed that it is mainly made of alpha-helices and is hydrophilic in nature. AgrB is a transmembrane protein, and AgrD encodes the precursor of the autoinducing peptide (AIP). Sequencing of the agrD gene revealed that the 5 canine SP strains tested could be divided into three Agr specificity groups (RIPTSTGFF, KIPTSTGFF, and RIPISTGFF) based on the putative AIP produced by each strain. The AIP of SP contains serine and produce lactone ring structured AIP. CONCLUSION:Presence of AgrA, B, and D in all SP isolates implies the importance of this regulatory system in the virulence genes expression of the SP bacteria. SP isolates can be typed based on the AgrD auto-inducible protein sequences as it is being carried out for typing of S. aureus isolates. However, further studies are required to elucidate the mechanism of controlling of virulence genes by agr gene locus in the pathogenesis of soft tissue infection by SP.

SUBMITTER: Chitra MA

PROVIDER: S-EPMC4774685 | biostudies-literature | 2015 Jul

REPOSITORIES: biostudies-literature

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Detection and sequence analysis of accessory gene regulator genes of Staphylococcus pseudintermedius isolates.

Chitra M Ananda MA Jayanthy C C Nagarajan B B

Veterinary world 20150723 7

<h4>Background</h4>Staphylococcus pseudintermedius (SP) is the major pathogenic species of dogs involved in a wide variety of skin and soft tissue infections. The accessory gene regulator (agr) locus of Staphylococcus aureus has been extensively studied, and it influences the expression of many virulence genes. It encodes a two-component signal transduction system that leads to down-regulation of surface proteins and up-regulation of secreted proteins during in vitro growth of S. aureus. The obj ...[more]

PMID: 27047173

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Project description:Staphylococcus pseudintermedius is a major bacterial colonizer and opportunistic pathogen in dogs. Methicillin-resistant S. pseudintermedius (MRSP) continues to emerge as a significant challenge to maintaining canine health. We sought to determine the phylogenetic relationships of S. pseudintermedius across five states in the New England region of the United States and place them in a global context. The New England dataset consisted of 125 previously published S. pseudintermedius genomes supplemented with 45 newly sequenced isolates. The core genome phylogenetic tree revealed many deep branching lineages consisting of 142 multi-locus sequence types (STs). In silico detection of the mecA gene revealed 40 MRSP and 130 methicillin-susceptible S. pseudintermedius (MSSP) isolates. MRSP were derived from five structural types of SCCmec, the mobile genetic element that carries the mecA gene conferring methicillin resistance. Although many genomes were MSSP, they nevertheless harbored genes conferring resistance to many other antibiotic classes, including aminoglycosides, macrolides, tetracyclines and penams. We compared the New England genomes to 297 previously published genomes sampled from five other states in the United States and 13 other countries. Despite the prevalence of the clonally expanding ST71 found worldwide and in other parts of the United States, we did not detect it in New England. We next sought to interrogate the combined New England and global datasets for the presence of coincident gene pairs linked to antibiotic resistance. Analysis revealed a large co-circulating accessory gene cluster, which included mecA as well as eight other resistance genes [aac (6')-Ie-aph (2″)-Ia, aad (6), aph (3')-IIIa, sat4, ermB, cat, blaZ, and tetM]. Furthermore, MRSP isolates carried significantly more accessory genes than their MSSP counterparts. Our results provide important insights to the evolution and geographic spread of high-risk clones that can threaten the health of our canine companions.

Project description:BackgroundStaphylococcus aureus (S. aureus) with accessory gene regulator (agr) dysfunction occurs in health care settings. This study evaluated the prevalence and the molecular and drug resistance characteristics of S. aureus with dysfunctional agr in a pediatric population in Beijing, China.ResultsA total of 269 nonduplicate S. aureus clinical isolates were isolated from Beijing Children's Hospital, including 211 methicillin-resistant S. aureus (MRSA) from September 2010-2017 and 58 methicillin-sensitive S. aureus (MSSA) from February 2016-2017. Only 8 MRSA and 2 MSSA isolates were identified as agr dysfunction, and the overall prevalence rate was 3.7%. For MRSA isolates, ST59-SCCmec IV and ST239-SCCmec III were the most common clones, and the prevalence rate of agr dysfunction in ST239-SCCmec III isolates (17.39%) was significantly higher than in ST59-SCCmec IV (1.69%) and other genotype strains (P = 0.006). Among the agr dysfunctional isolates, only one MRSA ST59 isolate and one MSSA ST22 isolate harbored pvl. No significant difference was detected between agr dysfunction and agr functional isolates regarding the biofilm formation ability (P = 0.4972); however, 9/10 agr dysfunctional isolates could effectuate strong biofilm formation and multidrug resistance. Among MRSA, the non-susceptibility rates to ciprofloxacin, gentamicin, and trimethoprim-sulfamethoxazole were significantly higher in agr dysfunctional isolates than in isolates with functional agr (P < 0.05). Two isolates belonging to ST239 had no mutations in agr locus, but a synonymous mutation was found in agrA in another ST239 isolate. The inactivating mutations were detected in other seven agr dysfunctional isolates. The variants were characterized by non-synonymous changes (n = 5) and frameshift mutations (insertions, n = 2), which mainly occurred in agrC and agrA.ConclusionsThe results showed that agr dysfunctional S. aureus was not common in Chinese children, and ST59-SCCmec IV was associated with lower prevalence of agr dysfunction as compared to ST239-SCCmec III isolates. The agr dysfunctional isolates were healthcare-associated, multidrug resistant and form strong biofilm, which suggested that agr dysfunction might offer potential advantages for S. aureus to survive in a medical environment.

Dataset Information

Detection and sequence analysis of accessory gene regulator genes of Staphylococcus pseudintermedius isolates.

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Detection and sequence analysis of accessory gene regulator genes of Staphylococcus pseudintermedius isolates.

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