Genotypic and epidemiological characterization of Methicillin resistant Staphylococcus aureus (MRSA) and Coagulase Negative Staphylococcal (CoNS) strains isolated at Tygerberg Hospital

Van Zyl, Kristien Nel (2016-12)

Thesis (MScMedSc)--Stellenbosch University, 2016.

Thesis

Background The global burden of methicillin resistant Staphylococcus aureus (MRSA) can be largely attributed to S. aureus’ ability to acquire the resistance element, SCCmec (Staphylococcal chromosome complex mec). Classification of SCCmec types is an essential component of staphylococcal epidemiology and is based on the arrangement and classes of the mec and ccr gene complexes and Open Reading Frames (ORFs) in the joining (J) regions. At least twelve SCCmec types and numerous subtypes have been described to date. We identified potentially novel and novel variant SCCmec types in MRSA isolates from a tertiary hospital in Cape Town, South Africa. This study aimed to describe the molecular structure and possible origin of these novel elements in our setting, and to determine the prevalence of these SCCmec types. Methods We screened 87 clinical MRSA and 100 methicillin resistant coagulase negative Staphylococci (MR-CoNS) isolates using a multiplex PCR for SCCmec typing. Additional typing employed a combination of six multiplex PCRs on 3 MRSA isolates each from the novel and novel variant types. Whole genome sequencing (WGS) was performed on representative isolates using the Illumina next-generation sequencing platform. Results Among the MRSA isolates, 36% contained the novel SCCmec type (ccrC/Class A mec), followed by SCCmec IV (24%), the novel variant SCCmec type (ccrA1B1, ccrC/Class B mec) (17%) and SCCmec III (11%). Only one MR-CoNS isolate contained the novel type. Preliminary genomic analysis support the PCR findings, and show the presence of a possible truncated SCCHg element in the novel, novel variant and SCCmec III isolates. Discussion We have successfully optimized and implemented two SCCmec typing assays on MRSA and MR-CoNS isolates in our setting and in doing so, identified possible novel and novel variant SCCmec elements. The novel SCCmec type is common among local MRSA isolates, and may reflect clonal spread within the hospital. Preliminary WGS analysis showed that these isolates are composites of SCCHg and SCCmec elements, however further phylogenetic analysis is required to provide insights into how these elements emerged.

Agtergrond Die wêreldwye las van metisillienweerstandige Staphylococcus aureus (MRSA) kan toegeskryf word aan S. aureus se vermoë om die antibiotiese weerstandbiedende element, “Staphylococcal chromosome complex mec” (SCCmec) op te neem. Klassifikasie van SCCmec is ‘n essensiële komponent in die epidemiologiese ondersoek van stafilokokkus-infeksies en is gebaseer op die kombinasies van verskillende “mec” en “ccr” geenkomplekse en oopleesrame (OLR) in die “joining (J) regions” van die element. Ten minste twaalf SCCmec-tipes en verskeie subtipes is al beskryf en gepubliseer en kom in beide MRSA en metisillienweerstandige koagulase negatiewe stafilokokke (MR-KoNS) voor. Ons het ‘n potensiële nuwe SCCmec-tipe en variant tipe in MRSA-isolate van Tygerberg Akademiese Hospitaal ontdek. In hierdie studie het ons gepoog om die molekulêre struktuur, moontlike oorsprong en voorkoms van die nuwe tipe SCCmec-isolate in hierdie instelling te ondersoek. Metodes ‘n veelvoudige polimerasekettingreaksie (vPKR) is op 87 MRSA en 100 MR-KoNS-isolate uitgevoer. Verdere epidemiologiese ondersoek is vir drie isolate elk van die nuwe tipe en nuwe variant tipe gedoen met ‘n protokol wat die gebruik van ses mPKR-reaksies insluit. Heel-genoomvolgordebepaling is op verteenwoordigende isolate met Illumina tegnologie gedoen. Resultate Ses-en-dertig persent van die MRSA isolate het die nuwe SCCmec tipe (ccrC/Klas A mec) bevat, gevolg deur SCCmec IV (24%), die nuwe variant (ccrA1B1, ccrC/Class B mec) (17%) en SCCmec III (11%). Slegs een MR-KoNS-isolaat het die nuwe tipe bevat. Voorlopige analise van die genome van hierdie nuwe tipes ondersteun die PKR resultate, en dui op die voorkoms van ‘n verkorte SCCHg element in die nuwe, variant en SCCmec III-isolate. Bespreking Ons het twee SCCmec-tiperingsmetodes in ons instelling suksesvol geöptimiseer en geïmplementeer op MRSA- en MR-KoNS-isolate en daardeur ‘n nuwe SCCmec-tipe en nuwe variant SCCmec-tipe ontdek. Die nuwe SCCmec tipe is algemeen in die plaaslike MRSA-isolate en dit mag ‘n aanduiding wees dat hierdie tipe op ‘n klonale wyse versprei in die hospitaal. Heel-genoomvolgordebepalings-analise wys daarop dat hierdie nuwe elemente samestellings van SCCHg en SCCmec elemente is, maar verdere filogenetiese ondersoeke moet uitgevoer word, om insig te gee oor die oorsprong en verspreiding van hierdie nuwe element.

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