Distribution of virulence genes and the molecular epidemiology of Streptococcus pyogenes clinical isolates by emm and multilocus sequence typing methods.

BACKGROUND: Streptococcus pyogenes has a variety of virulence factors and the predominant invasive strains differ according to specific emm types and geographical orientation. Although emm typing is commonly used as the gold standard method for the molecular characterisation, multilocus sequence typing (MLST) has become an important tool for comparing the genetic profiles globally. This study aimed to screen selected virulence genes from invasive and non-invasive clinical samples and to characterise the molecular epidemiology by emm typing and MLST methods. MATERIALS AND METHODS: A total of 42 S. pyogenes isolates from invasive and non-invasive samples collected from two different tertiary hospitals were investigated for the distribution of virulence factors and their molecular epidemiology by emm and multilocus sequence typing methods. Detection of five virulence genes (speA, speB, speJ, ssa and sdaB) was performed using multiplex polymerase chain reaction (PCR) using the standard primers and established protocol. Phylogenetic tree branches were constructed from sequence analysis utilised by neighbour joining method generated from seven housekeeping genes using MEGA X software. RESULTS: Multiplex PCR analysis revealed that sdaB/speF (78.6%) and speB (61.9%) were the predominant virulence genes. Regardless of the type of invasiveness, diverse distribution of emm types/subtypes was noted which comprised of 27 different emm types/subtypes. The predominant emm types/subtypes were emm63 and emm18 with each gene accounted for 11.8% whereas 12% for each gene was noted for emm28, emm97.4 and emm91. The MLST revealed that the main sequence type (ST) in invasive samples was ST402 (17.7%) while ST473 and ST318 (12% for each ST) were the major types in non-invasive samples. Out of 18 virulotypes, Virulotype A (five genes, 55.6%) and Virulotype B (two genes, 27.8%) were the major virulotypes found in this study. Phylogenetic analysis indicated the presence of seven different clusters of S. pyogenes. Interestingly, Cluster VI showed that selected emm/ST types such as emm71/ST318 (n=2), emm70.1/ST318 (n=1), emm44/ST31 (n=1) and emm18/ST442 (n=1) have clustered within a common group (Virulotype A) for both hospitals studied. CONCLUSION: The present study showed that group A streptococcci (GAS) are genetically diverse and possess virulence genes regardless of their invasiveness. Majority of the GAS exhibited no restricted pattern of virulotypes except for a few distinct clusters. Therefore, it can be concluded that virulotyping is partially useful for characterising a heterogeneous population of GAS in hospitals.

Presence of methicillin resistance and heteroresistance among Coagulase Negative Staphylococci (CoNS) isolates obtained from Health Sciences students at a Public University.

This study was carried out to investigate the Coagulase Negative Staphylococci (CoNS) nasal carriage and the presence of methicillin resistant Coagulase Negative Staphylococci (MR-CoNS) among health sciences students at Faculty of Medicine and Health Sciences, Universiti Putra Malaysia. A total of 120 isolates of CoNS (62.5%) was isolated from 192 student volunteers. The mecA gene was detected in 15 isolates of CoNS (12.5%). Eight out of the 15 isolates of mecA positive CoNS were resistant to cefoxitin in disc diffusion test whereas the remaining seven isolates of mecA positive CoNS were susceptible to cefoxitin. Analysis of questionnaires showed no significant association between CoNS nasal carriage and the socio-demographic and risk factors except for the genders and history of cold (P < 0.050). Generally, this finding showed a relatively low level of methicillin resistance among CoNS nasal carriage from student volunteers.

Presence of methicillin resistance and heteroresistance among Coagulase Negative Staphylococci (CoNS) isolates obtained from Health Sciences students at a Public University

This study was carried out to investigate the Coagulase Negative Staphylococci (CoNS) nasal carriage and the presence of methicillin resistant Coagulase Negative Staphylococci (MR-CoNS) among health sciences students at Faculty of Medicine and Health Sciences, Universiti Putra Malaysia. A total of 120 isolates of CoNS (62.5%) was isolated from 192 student volunteers. The mecA gene was detected in 15 isolates of CoNS (12.5%). Eight out of the 15 isolates of mecA positive CoNS were resistant to cefoxitin in disc diffusion test whereas the remaining seven isolates of mecA positive CoNS were susceptible to cefoxitin. Analysis of questionnaires showed no significant association between CoNS nasal carriage and the socio-demographic and risk factors except for the genders and history of cold (P < 0.050). Generally, this finding showed a relatively low level of methicillin resistance among CoNS nasal carriage from student volunteers.

Multiple ambler class A ESBL genes among Klebsiella pneumoniae isolates in a Malaysian district hospital.

Detailed reports regarding the distribution and activity of extended-spectrum beta-lactamase (ESBL)-producing Klebsiella pneumoniae isolates are currently not widely available in the Malaysian setting. This study was conducted to determine the ESBL genes distribution rate, phenotypic detection, and antimicrobial susceptibility patterns among betalactam resistant Klebsiella pneumoniae isolated from a Malaysian district hospital. K. pneumoniae isolates were collected from a microbiology laboratory at Hospital Pakar Sultanah Fatimah, Malaysia. Following exclusion and inclusion criteria, 141 isolates were selected for this study. K. pneumoniae was identified by phenotypic method, whilst antibiotics' susceptibility patterns were determined by the Kirby-Bauer method, as described in Clinical Laboratory Standard Institute (CLSI) guidelines (Oxoid, UK; Becton-Dickenson, USA). Detection of Ambler Group A ESBL gene (blaSHV, blaTEM, blaCTX-M-1, blaCTX-M-2, blaCTX-M-8, blaCTX-M-9, and blaCTX-M-25) was done using polymerase chain reaction (PCR). ESBL genes were found in 85.8% of K. pneumoniae (121 of 141) isolates. Only blaSHV, blaTEM, blaCTX-M-1, and blaCTX-M-9 were detected among K. pneumoniae isolates with distribution rates of 75.2% (106 of 141), 41.1% (58 of 141), 44% (62 of 141), and 0.7% (1 of 141), respectively. There was no blaCTX-M-2, blaCTX-M-8, or blaCTX-M-25 detected from any isolates in this study. Sequencing of representative amplicons revealed blaSHV as SHV-12, blaTEM as TEM-1, blaCTX-M-1 as CTX-M-15, and blaCTX-M-9 as CTX-M-18. The phenotypic detection rate of ESBL was 71.6% (101 of 141), whilst 9.2% (13 of 141) were positive for carbapenemase. AmpC betalactamase was detected in 22% (31 of 141) of all isolates. Antibiotic resistance was between 44.6% (netilmicin) and 97.2% (cefotaxime). Based on ESBL genes distribution, blaSHV was a predominant gene found in one of Malaysian district hospitals, notwithstanding having blaTEM, blaCTX-M-1, and blaCTX-M-9. Despite carrying multiple ESBL genes, some strains were positive for carbapenemase or AmpC betalactamase, which resulted in high antimicrobial resistance rates.