A phenotypic and genotypic characterisation of strain types, virulence factors and agr groups of colonising Staphylococcus aureus associated with bloodstream infection

Karayem, Karayem (2015-03)

Thesis (PhD)--Stellenbosch University, 2015

Thesis

ENGLISH ABSTRACT : Several studies investigating the molecular characteristics of Staphylococcus aureus have been conducted worldwide, however, in South Africa, most of these have focused on Methicillinresistant S. aureus (MRSA). This study investigated the phenotypic and genotypic characteristics of isolates of S. aureus collected from the blood and nasal cavity of patients admitted to Tygerberg Hospital, South Africa. Investigations included determining the association between blood and nasal isolates, describing the molecular epidemiology of the population, determining the prevalence of various virulence factor genes among the different clones and descibing the accessory gene regulator (agr) functionality of S. aureus clones. Pulsed-field gel electrophoresis (PFGE), performed on 208 blood and nasal isolates from 162 patients with S. aureus bacteraemia, showed that 93 (57.4%) of the patients were colonised with the same strain type (p =0.061). MRSA was significantly associated with endogenous bacteraemia (same strain obtained from the blood and the nose) (p = 0.042). Molecular typing of the 208 blood and nasal isolates (43.3% MRSA) revealed that the majority of strains were ST239-t37-agr I (25.5%) which harboured different SCCmec types including SCCmec type III and a potentially novel type presumed to consist of ccrC/Class A mec. ST612-MRSA-IV was the second most predominant clone (10.2%). Other MRSA clones included ST5-t045 with a potentially novel variant of SCCmec type I consisting of ccrA1B1 and a ccrC/Class B mec; and ST461-MRSA-IV, reported for the first time in South Africa. All 18 (8.7%) pvl-positive isolates were MSSA except one isolate (ST612-MRSA-IV). The identification of novel MRSA clones (ST641-MRSA-IV), MSSA STs (ST2122, ST2126), and the potentially novel SCCmec type and type I variant suggest the local emergence of new clones. Twenty-one isolates (representing nine clonal complexes (CCs)) previously characterised by Multi-locus Sequence Typing (MLST) were analysed for the prevalence of 38 virulence factor genes. There was an association between different enterotoxin gene cluster (egc) gene combinations and CC5, CC22, CC30, and CC45. Both CC15 and CC97 were negative for Superantigen (SAg) genes. The intracellular adhesion locus A (icaA) gene was common (90.4%) and detected in all CCs (except CC30) and the enterotoxin I (sei) gene was significantly more widespread in MRSA isolates (77.8% in MRSA; 25.0% MSSA; p = 0.03). Accessory gene regulator dysfunction was significantly higher amongst MRSA than MSSA isolates and was more commonly associated with ST36-MRSA-II, ST239-MRSA-III and ST239-MRSA- ccrC/Class A mec. Shifting of agr in the same host was not common. Key findings in this study relate to the likely emergence of populations at Tygerberg Hospital, as evidenced by novel STs and potentially novel SCCmec types. The identification of a circulating clone within the burns unit both illustrates the potential for organisms to spread within the hospital, as well as reinforcing the value of molecular typing for infection control purposes. The association of different agr types, agr functionality, and virulence factors with typing data has shown results consistent with other studies, as well as some unusual results. However, the clinical relevance of these associations is not yet well understood, and should form the basis of further research.

AFRIKAANSE OPSOMMING : Verskeie studies wat die molekulêre eienskappe van Staphylococcus aureus ondersoek, is al wêreldwyd uitgevoer, maar in Suid-Afrika het die meeste gefokus op Methicillinweerstandige S. aureus (MRSA). Hierdie studie ondersoek die fenotipiese en genotipiese eienskappe van bloed en nasale S. aureus isolate van pasiënte wat by Tygerberg-hospitaal opgeneem is. Ondersoeke sluit in die bepaling van die verwantskap tussen bloed en nasale isolate, die beskrywing van die molekulêre epidemiologie van die bevolking, die bepaling van die teenwoordigheid van verskeie virulensiefaktorgene tussen die verskillende klone; en die beskrywing van die “accessory gene regulator” (agr) funksie van S. aureus klone. “Pulsed-Field Gel Electrophoresis” (PFGE), wat op 208 bloed en nasale isolate (afkomstig van 162 pasiënte met S. aureus bacteriemie) uitgevoer is, toon dat 93 (57.4%) van die pasiënte gekoloniseer is met dieselfde stam tipe (p = 0.061). MRSA vertoon ‘n betekenisvolle verwantskap met endogeniese bakteremie (dieselfde stam teenwoordig in bloed en nasale isolate) (p = 0.042). Molekulêre tipering van die 208 bloed en nasale isolate (43.3% MRSA) het getoon dat die meerderheid van die stamme deel van ST239-T37-agr I (25.5%) is en uit SCCmec tipe III en 'n potensiële nuwe tipe (ccrC/KlasA mec) bestaan. ST612-MRSA-IV was die tweede mees oorheersende kloon (10.2%). Ander MRSA klone het ST5-t045 ingesluit, wat 'n potensiële nuwe variant van SCCmec tipe I (bestaande uit ccrA1B1 en 'n ccrC / Klas B mec); en ST461- MRSA-IV wat vir die eerste keer in Suid-Afrika gevind is. Al 18 (8.7%) PVL-positiewe isolate was MSSA behalwe een isolaat (ST612-MRSA-IV). Die identifisering van nuwe MRSA klone (ST641-MRSA-IV), MSSA STs (ST2122, ST2126), en die potensiële nuwe SCCmec tipes reflekteer die plaaslike ontstaan van hierdie nuwe klone. Een-en-twintig isolate wat voorheen beskryf is deur “Multi-locus Sequence Typing” (MLST) is bestudeer vir die teenwoordigheid van 38 virulensiefaktor gene. 'n Assosiasie tussen verskillende “enterotoxin gene cluster” (EGC) geen kombinasies en CC5, CC22, CC30 en CC45 is gevind. Beide CC15 en CC97 was negatief vir die “Superantigen” (SAg) gene. Die “intracellular adhesion locus A” (icaA) geen was algemeen (90.4%) en in al die CCe waargeneem (behalwe CC30). Die “enterotoxin I” (sei) geen was aansienlik meer teenwoordig in MRSA isolate (77.8% in MRSA; 25,0 % MSSA; p = 0.03). “Accessory gene regulator” disfunksie was aansienlik hoër in die MRSA groep (in vergelyking met die MSSA groep) en was geassosieer met ST36-MRSA-II, ST239-MRSA-III en ST239-MRSA-ccrC/Klas A mec. Die verskuiwing van agr funksionaliteit tussen isolate afkomstig van die bloed of neus was skaars.

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