In Vitro Activity of Ceftolozane-Tazobactam against Escherichia coli, Klebsiella pneumoniae and Pseudomonas aeruginosa Obtained from Blood Cultures from Sentinel Public Hospitals in South Africa.

Multidrug-resistant (MDR) Gram-negative bacteria are responsible for the majority of healthcare-associated infections and pose a serious threat as they complicate and prolong clinical care. A novel cephalosporin-ß-lactamase-inhibitor combination, ceftolozane-tazobactam (C/T) was introduced in 2014, which improved the treatment of MDR pathogens. This study aimed to evaluate the activity of C/T against Escherichia coli (n = 100), Klebsiella pneumoniae (n = 100), and Pseudomonas aeruginosa (n = 100) blood culture isolates in South Africa (SA). Isolates were sequentially selected (2010 to 2020) from the Group for Enteric, Respiratory, and Meningeal Diseases Surveillance (GERMS) programme in SA. Organism identification was performed using the matrix-assisted laser desorption/ionisation-time of flight mass spectrometry (MALDI-TOF MS) instrument (Microflex, Bruker Daltonics, Bremen, Germany), and antibiotic susceptibility was performed using the Sensititre instrument (Trek Diagnostic Systems, East Grinstead, UK). C/T resistance was reported in 16 E. coli, 28 K. pneumoniae and 13 P. aeruginosa isolates. Fifty percent of the C/T resistant isolates were subjected to whole-genome sequencing (WGS). According to the whole genome multilocus sequence typing (MLST) analysis, the E. coli isolates (n = 8) belonged to sequence type (ST)10, ST131, ST405, and ST410, the K. pneumoniae isolates (n = 14) belonged to ST1, ST37, ST73, ST101, ST231, ST307, ST336 and ST6065 (novel ST), and the P. aeruginosa isolates (n = 7) belonged to ST111, ST233, ST273, and ST815. The WGS data also showed that all the E. coli isolates harboured aminoglycoside (aph (3'')-Ib, aph (6)-Id), macrolide (mdfA, mphA), and sulphonamide (sul2) antibiotic resistance genes, all the K. pneumoniae isolates harboured ß-lactam (blaCTX-M-15), and sulphonamide (sul2) antibiotic resistance genes, and all the P. aeruginosa isolates harboured aminoglycoside (aph (3')-IIb), ß-lactam (PAO), fosfomycin (fosA), phenicol (catB7), quinolone (crpP), and disinfectant (qacE) antibiotic resistance genes. It is evident that E. coli, K. pneumoniae and P. aeruginosa can adapt pre-existing resistance mechanisms to resist newer ß-lactam molecules and inhibitors, since these isolates were not exposed to ceftolozane-tazobactam previously.

Molecular characterisation of Acinetobacter baumannii isolates from bloodstream infections in a tertiary-level hospital in South Africa.

Acinetobacter baumannii is an opportunistic pathogen and causes various infections in patients. This study aimed to describe the clinical, epidemiological and molecular characteristics of A. baumannii isolated from BCs in patients at a tertiary-level hospital in South Africa. Ninety-six isolates from bloodstream infections were collected. Clinical characteristics of patients were recorded from patient files. Organism identification and AST was performed using automated systems. PCR screening for the mcr-1 to mcr-5 genes was done. To infer genetic relatedness, a dendrogram was constructed using MALDI-TOF MS. All colistin-resistant isolates (n = 9) were selected for WGS. The patients were divided into three groups, infants (<1 year; n = 54), paediatrics (1-18 years; n = 6) and adults (≥19 years; n = 36) with a median age of 13 days, 1 and 41 years respectively. Of the 96 A. baumannii bacteraemia cases, 96.9% (93/96) were healthcare-associated. The crude mortality rate at 30 days was 52.2% (48/92). The majority of the isolates were multidrug-resistant (MDR). All isolates were PCR-negative for the mcr-1 to mcr-5 genes. The majority of the isolates belonged to cluster 1 (62/96) according to the MALDI-TOF MS dendrogram. Colistin resistance was confirmed in nine A. baumannii isolates (9.4%). The colistin-resistant isolates belonged to sequence type (ST) 1 (5/6) and ST2 (1/6). The majority of ST1 isolates showed low SNP diversity (≤4 SNPs). All the colistin-resistant isolates were resistant to carbapenems, exhibited an XDR phenotype and harboured the bla OXA-23 gene. The bla NDM gene was only detected in ST1 colistin-resistant isolates (n = 5). The lpsB gene was detected in all colistin-resistant isolates as well as various efflux pump genes belonging to the RND, the MFS and the SMR families. The lipooligosaccharide OCL1 was detected in all colistin-resistant ST1 and ST2 isolates and the capsular polysaccharide KL3 and KL17 were detected in ST2 and ST1 respectively. This study demonstrated a 9.4% prevalence of colistin-resistant ST1 and ST2 A. baumannii in BC isolates. The detection of the lpsB gene indicates a potential threat and requires close prospective monitoring.

Distribution of serotypes and antibiotic resistance of invasive Pseudomonas aeruginosa in a multi-country collection.

BACKGROUND: Pseudomonas aeruginosa is an opportunistic pathogen that causes a wide range of acute and chronic infections and is frequently associated with healthcare-associated infections. Because of its ability to rapidly acquire resistance to antibiotics, P. aeruginosa infections are difficult to treat. Alternative strategies, such as a vaccine, are needed to prevent infections. We collected a total of 413 P. aeruginosa isolates from the blood and cerebrospinal fluid of patients from 10 countries located on 4 continents during 2005-2017 and characterized these isolates to inform vaccine development efforts. We determined the diversity and distribution of O antigen and flagellin types and antibiotic susceptibility of the invasive P. aeruginosa. We used an antibody-based agglutination assay and PCR for O antigen typing and PCR for flagellin typing. We determined antibiotic susceptibility using the Kirby-Bauer disk diffusion method. RESULTS: Of the 413 isolates, 314 (95%) were typed by an antibody-based agglutination assay or PCR (n = 99). Among the 20 serotypes of P. aeruginosa, the most common serotypes were O1, O2, O3, O4, O5, O6, O8, O9, O10 and O11; a vaccine that targets these 10 serotypes would confer protection against more than 80% of invasive P. aeruginosa infections. The most common flagellin type among 386 isolates was FlaB (41%). Resistance to aztreonam (56%) was most common, followed by levofloxacin (42%). We also found that 22% of strains were non-susceptible to meropenem and piperacillin-tazobactam. Ninety-nine (27%) of our collected isolates were resistant to multiple antibiotics. Isolates with FlaA2 flagellin were more commonly multidrug resistant (p = 0.04). CONCLUSIONS: Vaccines targeting common O antigens and two flagellin antigens, FlaB and FlaA2, would offer an excellent strategy to prevent P. aeruginosa invasive infections.

Ward-specific clustering of methicillin-resistant Staphylococcus aureus spa-type t037 and t045 in two hospitals in South Africa: 2013 to 2017.

INTRODUCTION: Methicillin-resistant Staphylococcus aureus (MRSA) is a highly clonal pathogen causing infections in various settings. The aim of this study was to determine if healthcare-associated (HA) MRSA isolates with the same spa-type originating from two geographically distinct hospitals in South Africa were genetically related based on PFGE. Furthermore, a small subset of MRSA isolates were characterised with WGS and then compared to PFGE to determine if PFGE is still a reliable method to define outbreaks and/or transmission chains. METHODS: Staphylococcus aureus isolated from blood cultures (BC) were submitted to the Centre for Healthcare-Associated Infections, Antimicrobial Resistance and Mycoses (CHARM) as part of a laboratory-based surveillance programme (GERMS-SA). The identified HA-MRSA isolates underwent molecular characterisation [Staphylococcal Chromosome Cassette (SCC) mec and spa-typing]. Pulsed-field gel electrophoresis (PFGE) was performed on selected isolates with the same spa-type. Twenty-one MRSA isolates were selected for whole-genome sequencing (WGS) based on spa-type, PFGE clustering, time and place of isolation. RESULTS: Eighteen percent (n = 95/529) and 33% (n = 234/710) of isolates collected, from two public tertiary academic hospitals in the Gauteng (GAU) and the Western Cape (WC) provinces, were identified as MRSA, respectively. The most dominant clone in the GAU hospital was t037-III-MRSA (43.2%; n = 41/95). The most dominant clones in the WC hospital was t037-III-MRSA (23.9%, n = 56/234) and t045-I-MRSA (23.5%, n = 55/234). The GAU-t037-III-MRSA cases and WC-t045-I-MRSA cases occurred in the paediatric patient population, whereas the WC-t037-III-MRSA cases occurred in the adult patient population. A novel spa-type (t19935) was detected in the GAU hospital. PFGE showed that the GAU- and WC-t037-III-MRSA isolates were genetically indistinguishable, as well as most of the WC-t045-I-MRSA isolates. The Vienna/Hungarian/Brazilian clone and British EMRSA-3 clone were in circulation and a low frequency of single nucleotide polymorphisms (SNP) (≤20) differences was observed among isolates with the same spa-type. CONCLUSION: The low number of SNP differences is suggestive of uninterrupted strain transmission and the persistence of t037-III-MRSA and t045-I-MRSA from 2013 to 2017 in the two studied hospitals. Alternative infection prevention and control strategies should be considered to supplement control efforts.

Diversity of SCCmec elements and spa types in South African Staphylococcus aureus mecA-positive blood culture isolates.

BACKGROUND: The prevalence of Staphylococcus aureus varies depending on the healthcare facility, region and country. To understand its genetic diversity, transmission, dissemination, epidemiology and evolution in a particular geographical location, it is important to understand the similarities and variations in the population being studied. This can be achieved by using various molecular characterisation techniques. This study aimed to provide detailed molecular characterisation of South African mecA-positive S. aureus blood culture isolates by describing the SCCmec types, spa types and to lesser extent, the sequence types obtained from two consecutive national surveillance studies. METHODS: S. aureus blood culture isolates from a national laboratory-based and enhanced surveillance programme were identified and antimicrobial susceptibility testing was performed using automated systems. A real-time PCR assay confirmed the presence of the methicillin-resistance determinant, mecA. Conventional PCR assays were used to identify the SCCmec type and spa type, which was subsequently analysed using the Ridom StaphType™ software. Multilocus sequence typing was performed on selected isolates using conventional methods. MRSA clones were defined by their sequence type (ST), SCCmec type and spa type. RESULTS: A detailed description of findings is reported in this manuscript. SCCmec type III predominated overall followed by type IV. A total of 71 different spa types and 24 novel spa types were observed. Spa type t037 was the most common and predominated throughout followed by t1257. Isolates were multidrug resistant; isolates belonging to all SCCmec types were resistant to most of the antibiotics with the exception of type I; isolates with spa type t045 showed resistance to all antibiotics except vancomycin. The most diverse SCCmec-spa type complex was composed of the SCCmec type IV element and 53 different spa types. CONCLUSION: Although ST data was limited, thereby limiting the number of clones that could be identified, the circulating clones were relatively diverse.

An outbreak of cutaneous abscesses caused by Panton-Valentine leukocidin-producing methicillin-susceptible Staphylococcus aureus among gold mine workers, South Africa, November 2017 to March 2018.

BACKGROUND: We aimed to describe an outbreak of cutaneous abscesses caused by Panton-Valentine leukocidin (PVL)-producing methicillin-susceptible Staphylococcus aureus (MSSA) among gold mine workers. METHODS: In February 2018, we retrospectively reviewed a random sample of 50 medical records from 243 cases and conducted face-to-face interviews using a structured questionnaire. Pus aspirates were sent to the National Institute for Communicable Diseases from prospectively-identified cases (November 2017-March 2018). Nasopharyngeal swabs were collected during a colonisation survey in February 2018. Staphylococcus aureus isolates were screened with a conventional PCR for lukS/F-PV. Pulsed-field gel electrophoresis (PFGE) was performed to determine the genetic relatedness among the isolates. A sample of isolates were selected for whole genome sequencing (WGS). We conducted an assessment on biological risks associated with mining activities. RESULTS: From January 2017 to February 2018, 10% (350/3582) of mine workers sought care for cutaneous abscesses. Forty-seven medical files were available for review, 96% were male (n = 45) with a mean age of 43 years (SD = 7). About 52% (24/46) were involved in stoping and 28% (13/47) worked on a particular level. We cultured S. aureus from 79% (30/38) of cases with a submitted specimen and 14% (12/83) from colonisation swabs. All isolates were susceptible to cloxacillin. Seventy-one percent of S. aureus isolates (30/42) were PVL-PCR-positive. Six PFGE clusters were identified, 57% (21/37) were closely related. WGS analysis found nine different sequence types. PFGE and WGS analysis showed more than one cluster of S. aureus infections involving closely related isolates. Test reports for feed and product water of the mine showed that total plate counts were above the limits of 1000 cfu/ml, coliform counts > 10 cfu/100 ml and presence of faecal coliforms. Best practices were poorly implemented as some mine workers washed protective clothing with untreated water and hung them for drying at the underground surface. CONCLUSIONS: PVL-producing MSSA caused an outbreak of cutaneous abscesses among underground workers at a gold mining company. To our knowledge, no other outbreaks of PVL-producing S. aureus involving skin and soft tissue infections have been reported in mining facilities in South Africa. We recommend that worker awareness of infection prevention and control practices be strengthened.

The Diversity of Lipopolysaccharide (O) and Capsular Polysaccharide (K) Antigens of Invasive Klebsiella pneumoniae in a Multi-Country Collection.

Klebsiella pneumoniae is a common cause of sepsis and is particularly associated with healthcare-associated infections. New strategies are needed to prevent or treat infections due to the emergence of multi-drug resistant K. pneumoniae. The goal of this study was to determine the diversity and distribution of O (lipopolysaccharide) and K (capsular polysaccharide) antigens on a large (>500) global collection of K. pneumoniae strains isolated from blood to inform vaccine development efforts. A total of 645 K. pneumoniae isolates were collected from the blood of patients in 13 countries during 2005-2017. Antibiotic susceptibility was determined using the Kirby-Bauer disk diffusion method. O antigen types including the presence of modified O galactan types were determined by PCR. K types were determined by multiplex PCR and wzi capsular typing. Sequence types of isolates were determined by multilocus sequence typing (MLST) targeting seven housekeeping genes. Among 591 isolates tested for antimicrobial resistance, we observed that 19.3% of isolates were non-susceptible to carbapenems and 62.1% of isolates were multidrug resistant (from as low as 16% in Sweden to 94% in Pakistan). Among 645 isolates, four serotypes, O1, O2, O3, and O5, accounted for 90.1% of K. pneumoniae strains. Serotype O1 was associated with multidrug resistance. Fifty percent of 199 tested O1 and O2 strains were gmlABC-positive, indicating the presence of the modified polysaccharide subunit D-galactan III. The most common K type was K2 by both multiplex PCR and wzi capsular typing. Of 39 strains tested by MLST, 36 strains were assigned to 26 known sequence types of which ST14, ST25, and ST258 were the most common. Given the limited number of O antigen types, diverse K antigen types and the high multidrug resistance, we believe that an O antigen-based vaccine would offer an excellent prophylactic strategy to prevent K. pneumoniae invasive infection.

Molecular diagnostics in South Africa and challenges in the establishment of a molecular laboratory in developing countries.

The laboratory plays a significant role in public health surveillance, outbreak investigation and infection prevention and control strategies. Microbiology laboratories are moving towards incorporating molecular biology techniques for the surveillance and identification of pathogens causing infectious diseases as well as the genotypic characterisation of these organisms. These methods are accurate, rapid, reliable, and provide a wealth of information that are not available using conventional phenotypic methods. However, establishing such a laboratory can be challenging in developing countries due to poor infrastructure, the lack of funding and the required expertise. This manuscript discusses the essential issues that need to be addressed when establishing a molecular microbiology laboratory and the usefulness of molecular techniques in public health surveillance and outbreaks in developing countries. Molecular data on South African findings obtained from surveillance and outbreak studies are also presented in this manuscript.

Complete Genome Sequence of a Staphylococcus aureus Isolate from a Nasopharyngeal Swab from a Mine Worker in South Africa.

Here, we present the complete genome sequence of Staphylococcus aureus NP66, isolated from a South African mine worker.

Unconventional SCCmec types and low prevalence of the Panton-Valentine Leukocidin exotoxin in South African blood culture Staphylococcus aureus surveillance isolates, 2013-2016.

Staphylococcus aureus is a healthcare-associated pathogen that can harbour multiple antimicrobial resistance determinants and express multiple virulence factors e.g. Panton-Valentine Leukocidin (PVL). Unknown staphylococcal cassette chromosome mec (SCCmec) typing patterns were previously observed among 11% (n = 52) of methicillin-resistant S. aureus (MRSA) isolates; we further investigated these as well as the proportion of PVL, encoded by lukS/F-PV, in 761 S. aureus isolates from patients with a diagnosis of pneumonia/lower respiratory tract, skin/soft tissue, bone and joint infection. S. aureus isolates from blood culture were identified and antimicrobial susceptibility testing was performed using automated systems. Conventional PCR assays were used to identify the ccr and mec gene complexes in mecA-positive isolates with an unknown SCCmec type and screen for lukS/F-PV. Epidemiological data was used to classify isolates as healthcare- or community-associated infections. Antimicrobial susceptibility profiles according to SCCmec type and PVL were reported. Of the unknown SCCmec types, isolates were interpreted as type I-like (86%, 38/44), type II-like (9%, 4/44) and type III-like (5%, 2/44). Eight isolates did not produce definitive results. Of all MRSA isolates, majority were multidrug-resistant as indicated by their non-susceptibility to most antimicrobial agents; 92% were healthcare-associated. PVL was seen in 14% of the isolates (MRSA: 25%, MSSA: 75%); 56% were classified as healthcare-associated infection. The SCCmec typing method did not definitively classify all unknown isolates into clearly defined types. It showed that majority of these isolates were not the conventional types; untypeable elements appeared to be composite SCCmec elements, consisting of multiple ccr gene complexes. Majority of the MRSA isolates were non-susceptible to most antibiotics indicating that multiple resistance genes are present in our population. Furthermore, the proportion of PVL was low and more prevalent in MSSA.

An overview of antimicrobial resistance surveillance among healthcare-associated pathogens in South Africa.

Healthcare-associated infections are a serious public health concern resulting in morbidity and mortality particularly in developing countries. The lack of information from Africa, the increasing rates of antimicrobial resistance and the emergence of new resistance mechanisms intensifies this concern warranting the need for vigorous standardised surveillance platforms that produce reliable and accurate data which can be used for addressing these concerns. The implementation of national treatment guidelines, policies, antimicrobial stewardship programmes and infection prevention and control practices within healthcare institutions require a platform from which it can draw information and direct its approach. In this review, the importance of standardised surveillance systems, the challenges faced in the application of a surveillance system and the condition (existence and nonexistence) of such systems in African countries is discussed. This review also reports on some South African data.

Laboratory based antimicrobial resistance surveillance for Pseudomonas aeruginosa blood isolates from South Africa.

INTRODUCTION: Antimicrobial resistant bacterial infections are widespread globally and increases in antimicrobial resistance presents a major threat to public health. Pseudomonas aeruginosa is an opportunistic healthcare-associated pathogen with high rates of morbidity and mortality and an extensive range of resistance mechanisms. This study describes the antibiotic susceptibility profiles of P. aeruginosa isolates from patients with bacteraemia submitted by sentinel laboratories in South Africa from 2014 to 2015. METHODOLOGY: Organism identification and antimicrobial susceptibility testing were done using automated systems. Molecular methods were used to detect common resistance genes and mechanisms. RESULTS: Overall the susceptibility was high for all antibiotics tested with a decrease over the two-year period. There was no change in the MIC50 and MIC90 breakpoints for all antibiotics from 2014 to 2015. The MIC50 was within the susceptible breakpoint range for most antibiotics and the MIC90 was within the susceptible breakpoint range for colistin only. Phenotypically carbapenem non-susceptible isolates harboured the following plasmid-mediated genes: blaVIM (n = 81, 12%) and blaGES (n = 6, 0.9%); blaNDM (n = 4, 0.6%) and blaOXA-48 and variants (n = 3, 0.45%). Porin deletions were observed in one meropenem non-susceptible isolate only, and multi-drug resistance efflux pumps were expressed in the majority of the non-susceptible isolates investigated. BlaVEB-1, blaIMP and blaKPC were not detected. CONCLUSION: The prevalence of resistance to commonly used antibacterial agents was low for P. aeruginosa isolates and similarly, tested resistance mechanisms were detected in a relatively small proportion of isolates. Findings in this study represent baseline information for understanding antimicrobial susceptibility patterns in P. aeruginosa isolates from blood. Our surveillance report may assist in contributing to hospital treatment guidelines.

Draft Genome Sequence of a Multidrug-Resistant Serratia marcescens Strain, Isolated from a Patient with Peritoneal Cancer in South Africa.

We report here the draft genome sequence of Serratia marcescens ML2637, isolated from a South African pediatric patient in the intensive care unit with peritoneal cancer. The genome comprised 5,718,350 bp, with a 59.1% G+C content. There were 5,594 predicted genes, including 5,301 protein-coding genes, 199 pseudogenes, and 94 RNA genes.

Antimicrobial susceptibility testing in predicting the presence of carbapenemase genes in Enterobacteriaceae in South Africa.

BACKGROUND: Carbapenem-resistant Enterobacteriaceae (CRE) is a concern in South Africa and worldwide. It is therefore important that these organisms be accurately identified for infection prevention control purposes. METHOD: In this study 1193 suspected CREs from 46 laboratories from seven provinces in South Africa were assessed to confirm the prevalence of carbapenemase genes from our referral diagnostic isolates for the period 2012 to 2015. We compared the antimicrobial susceptibility testing method used in the reference laboratory to the polymerase chain reaction (PCR) which is used as the gold standard. Organism identification and antimicrobial susceptibility testing were performed using automated systems and DNA was extracted using a crude boiling method. The presence of carbapenemase-producing genes (bla NDM, bla KPC, bla OXA-48&variants, bla GES, bla IMP and bla VIM) was screened for using a multiplex real-time PCR. RESULTS: Sixty-eight percent (n = 812) of the isolates harboured a carbapenemase-producing gene; the three most common genes included: bla NDM, bla OXA-48&variants and bla VIM. Majority of the carbapenemase producing Enterobacteriaceae (CPE) isolates were Klebsiella species (71 %). The Microscan® Walkaway system used for the screening of carbapenemase production was 98 % sensitive with a minimal inhibitory concentration (MIC) breakpoint of less than 0.5 as susceptible for ertapenem and a low specificity (13 %). CONCLUSION: From this study we can conclude that carbapenemase-producing Enterobacteriaceae is increasing in South Africa and the use of phenotypic methods for detection of CPEs showed good sensitivity but lacked specificity.

Staphylococcus aureus bacteraemia in Gauteng academic hospitals, South Africa.

INTRODUCTION: Methicillin-resistant Staphylococcus aureus (MRSA) infections are responsible for longer hospital stays, increased hospital costs, and poorer outcomes compared to methicillin-sensitive S. aureus (MSSA) infections. We aimed to describe the epidemiology of S. aureus bacteraemia (SAB) and to determine factors associated with MRSA infection in South Africa. METHODS: Cases of SAB were reported from September 2012 to September 2013 from three sentinel sites. A case was defined as the isolation of S. aureus from a blood culture during a 21-day period. Detailed clinical information was collected. Multivariable logistic regression was done to determine factors associated with MRSA infection and mortality. RESULTS: There were 442 cases of SAB reported; antimicrobial susceptibility testing was performed on 240 isolates (54%). Thirty-six percent (86/240) of cases had an MRSA infection. A longer hospital stay before positive specimen collection (odds ratio (OR) 1.08, 95% confidence interval (CI) 1.02-1.13, p=0.004), hospitalization in the last year (OR 15.7, 95% CI 2.5-99.5, p=0.003), HIV infection (OR 4.9, 95% CI 1.05-22.90, p=0.044), and antibiotic use in the previous 2 months (OR 0.1, 95% CI 0.01-0.68, p=0.022) were independent predictors of MRSA. Older age, and in particular age 25-44 years (OR 22.2, 95% CI 2.7-185.5, p=0.004, compared to those aged<5 years), was the only independent predictor of mortality amongst cases with SAB. MRSA isolates were non-susceptible to more antimicrobial agents compared to MSSA isolates. CONCLUSIONS: HIV infection was an independent risk factor for MRSA infection. The selection of appropriate empirical antimicrobial treatment is essential in patients with MRSA infections because of non-susceptibility to many other antimicrobial classes.

Prevalence and Trends of Staphylococcus aureus Bacteraemia in Hospitalized Patients in South Africa, 2010 to 2012: Laboratory-Based Surveillance Mapping of Antimicrobial Resistance and Molecular Epidemiology.

INTRODUCTION: We aimed to obtain an in-depth understanding on recent antimicrobial resistance trends and molecular epidemiology trends of S. aureus bacteraemia (SAB). METHODS: Thirteen academic centres in South Africa were included from June 2010 until July 2012. S. aureus susceptibility testing was performed on the MicroScan Walkaway. Real-time PCR using the LightCycler 480 II was done for mecA and nuc. SCCmec and spa-typing were finalized with conventional PCR. We selected one isolate per common spa type per province for multilocus sequence typing (MLST). RESULTS: S. aureus from 2709 patients were included, and 1231 (46%) were resistant to methicillin, with a significant decline over the three-year period (p-value = 0.003). Geographical distribution of MRSA was significantly higher in Gauteng compared to the other provinces (P<0.001). Children <5 years were significantly associated with MRSA with higher rates compared to all other age groups (P = 0.01). The most prevalent SCCmec type was SCCmec type III (531 [41%]) followed by type IV (402 [31%]). Spa-typing discovered 47 different spa-types. The five (87%) most common spa-types were t037, t1257, t045, t064 and t012. Based on MLST, the commonest was ST612 clonal complex (CC8) (n = 7) followed by ST5 (CC5) (n = 4), ST36 (CC30) (n = 4) and ST239 (CC8) (n = 3). CONCLUSIONS: MRSA rate is high in South Africa. Majority of the isolates were classified as SCCmec type III (41%) and type IV (31%), which are typically associated with hospital and community- acquired infections, respectively. Overall, this study reveals the presence of a variety of hospital-acquired MRSA clones in South Africa dominance of few clones, spa 037 and 1257. Monitoring trends in resistance and molecular typing is recommended to detect changing epidemiological trends in AMR patterns of SAB.

National sentinel site surveillance for antimicrobial resistance in Klebsiella pneumoniae isolates in South Africa, 2010 - 2012.

BACKGROUND: The increasing rates of antimicrobial resistance observed in the nosocomial pathogen Klebsiella pneumoniae are of major public health concern worldwide. OBJECTIVES: To describe the antibiotic susceptibility profiles of K. pneumoniae isolates from bacteraemic patients submitted by sentinel laboratories in five regions of South Africa from mid-2010 to mid-2012. Molecular methods were used to detect the most commonly found extended-spectrum beta-lactamase (ESBL) and carbapenemase resistance genes. METHODS: Thirteen academic centres serving the public healthcare sector in Gauteng, KwaZulu-Natal, Free State, Limpopo and Western Cape provinces submitted K. pneumoniae isolates from patients with bloodstream infections. Vitek 2 and MicroScan instruments were used for organism identification and susceptibility testing. Multiplex polymerase chain reactions (PCRs) were used to detect blaCTX-M, blaSHV and blaTEM genes in a proportion of the ESBL isolates. All isolates exhibiting reduced susceptibility to carbapenems were PCR tested for blaKPC and blaNDM-1 resistance genes. RESULTS: Overall, 68.3% of the 2,774 isolates were ESBL-positive, showing resistance to cefotaxime, ceftazidime and cefepime. Furthermore, 46.5% of all isolates were resistant to ciprofloxacin and 33.1% to piperacillin-tazobactam. The major ESBL genes were abundantly present in the sample analysed. Most isolates (95.5%) were susceptible to the carbapenems tested, and no isolates were positive for blaKPC or blaNDM-1. There was a trend towards a decrease in susceptibility to most antibiotics. CONCLUSION: The high proportion of ESBL-producing K. pneumoniae isolates observed, and the prevalence of ESBL genes, are of great concern. Our findings represent a baseline for further surveillance in SA, and can be used for policy and treatment decisions.