Large Outbreaks of Fungal and Bacterial Bloodstream Infections in a Neonatal Unit, South Africa, 2012-2016.

Candidemia is a major cause of healthcare-associated infections. We describe a large outbreak of Candida krusei bloodstream infections among infants in Gauteng Province, South Africa, during a 4-month period; a series of candidemia and bacteremia outbreaks in the neonatal unit followed. We detected cases by using enhanced laboratory surveillance and audited hospital wards by environmental sampling and epidemiologic studies. During July-October 2014, among 589 patients, 48 unique cases of C. krusei candidemia occurred (8.2% incidence). Risk factors for candidemia on multivariable analyses were necrotizing enterocolitis, birthweight <1,500 g, receipt of parenteral nutrition, and receipt of blood transfusion. Despite initial interventions, outbreaks of bloodstream infection caused by C. krusei, rarer fungal species, and bacterial pathogens continued in the neonatal unit through July 29, 2016. Multiple factors contributed to these outbreaks; the most functional response is to fortify infection prevention and control.

Characterisation and antimicrobial susceptibility pattern of non-tuberculous mycobacteria.

Background: Non-tuberculous mycobacteria (NTM) management comprises prolonged therapy that includes macrolides. Non-tuberculous mycobacteria can cause disease in patients with predisposing conditions such as HIV and structural lung disease. Local data on NTM disease and macrolide resistance are scarce, and routine antimicrobial susceptibility testing is currently not performed for NTM in South Africa. Objectives: This study aims to characterise NTM isolated at Tshepong National Health Laboratory Service (NHLS) according to species and antimicrobial susceptibility pattern. Method: A retrospective data analysis of NTM isolates from Tshepong NHLS was performed from January to June 2020. GenoType® NTM-DR was performed on selected isolates where the assay can confirm the species and determine resistance to macrolides and aminoglycosides. Results: Of the 194 collected NTM isolates, 183 were included in the study. Patients' ages ranged from 1 day to 81 years (median 36 years). The most common specimen was sputum (84.7%), followed by gastric aspirate (6.6%). The most common NTM isolated were Mycobacterium (M.) intracellulare (67.6%), M. fortuitum (12.6%), M. species (4.3%), M. kansasii (3.9%), and M. scrofulaceum (3.9%). Macrolide resistance occurred in 2.8% of tested isolates; no aminoglycoside resistance was detected. Although most isolates were from males (62.3%), resistance was observed only in females. Conclusion: M. intracellulare predominated, with only two M. intracellulare and two M. abscessus isolates showing macrolide resistance; aminoglycoside resistance was absent. Contribution: This study highlights the need for increased awareness of NTM, regular nationwide NTM surveillance, and monitoring of resistance trends to guide future patient management and ensure good treatment outcomes.

Corrigendum: Carriage of colistin-resistant Gram-negative bacteria in children from communities in Cape Town (Tuberculosis child multidrug-resistant preventive therapy trial sub-study).

[This corrects the article DOI: 10.4102/sajid.v36i1.241.].

Exudative pharyngitis and Corynebacterium pseudodiphtheriticum: A case report and review of the literature.

Corynebacterium pseudodiphtheriticum is an established member of the normal flora of the respiratory tract. This organism is an emerging cause of respiratory tract infection, as well as infection of the skin and skin structures, urinary tract and other sterile sites. The syndrome of C. pseudodiphtheriticum exudative pharyngitis is a diagnostic challenge of particular relevance in recent times as this organism can be confused with Corynebacterium diphtheriae in the clinical setting and in the laboratory. We report a case of exudative pharyngitis, possibly due to C. pseudodiphtheriticum, in a 14-month old, incompletely vaccinated, human immunodeficiency virus (HIV)-positive infant and review the role of this organism in terms of its microbiological profile and identification, disease spectrum and antimicrobial susceptibility pattern.

Carriage of colistin-resistant Gram-negative bacteria in children from communities in Cape Town (Tuberculosis child multidrug-resistant preventive therapy trial sub-study).

Colistin is a last-resort antibiotic against multidrug-resistant, Gram-negative bacteria. Colistin resistance has been described in the clinical settings in South Africa. However, information on carriage of these bacteria in communities is limited. This study investigated gastrointestinal carriage of colistin-resistant Escherichia coli and Klebsiella spp. and mcr genes in children from communities in Cape Town. Colistin-resistant E. coli was isolated from two participants (4%, 2/50), and mcr-1-mcr-9 genes were not detected. Gastrointestinal carriage of colistin-resistant Enterobacterales was rare; however, continuous extensive surveillance is necessary to determine the extent of carriage and its contribution to resistance observed in clinical settings.

Characterisation of mobile colistin resistance genes (mcr-3 and mcr-5) in river and storm water in regions of the Western Cape of South Africa.

BACKGROUND: Colistin is regarded as a last-resort antimicrobial against multi-drug resistant Gram-negative bacteria (GNB), therefore the dissemination of colistin resistance in the environment is of great concern. Horizontal transfer of mobile colistin resistance (mcr) genes to potential pathogens poses a serious problem. This study aimed to describe the presence of colistin resistant GNB and mcr genes in river and storm water in regions of the Western Cape. METHODS: Water samples were collected from three rivers during May 2019 and January 2020 and two storm water samples were collected in November 2019. Colistin resistant GNB were cultured on MacConkey agar containing colistin and identified by MALDI-TOF. Colistin resistance was confirmed using broth microdilution (BMD). mcr-1-5 genes were detected by PCR performed directly on the water samples and on the colistin resistant isolates. mcr functionality was assessed by BMD after cloning the mcr genes into pET-48b(+) and expression in SHuffle T7 E. coli. RESULTS: mcr-5.1 and various mcr-3 gene variants were detected in the Plankenburg-, Eerste- and Berg rivers and in storm water from Muizenberg, and only mcr-5.1 was detected in storm water from Fish Hoek. Colistin resistant GNB were isolated from all of the water sources. Aeromonas spp. were the most common colistin resistant organisms detected in the water sources; 25% (6/24) of colistin resistant Aeromonas spp. isolated from the Berg river contained novel mcr-3 variants; mcr-3.33 (n = 1), mcr-3.34 (n = 1) mcr-3.35 (n = 1) mcr-3.36 (n = 2) and mcr-3.37 (n = 1), which were confirmed to confer colistin resistance. CONCLUSIONS: The mcr-5.1 and mcr-3 colistin resistance gene variants were present in widely dispersed water sources in regions of the Western Cape. The mcr genes were only detected in water sampled downstream of and alongside communities, suggesting that their presence is driven by human influence/contamination. This is the first documentation of mcr-3 and mcr-5 gene variants in any setting in South Africa. Spill-over of these genes to communities could result in horizontal gene transfer to pathogenic bacteria, exacerbating the challenge of controlling multidrug resistant GNB infections.

Colistin Resistance Mechanisms in Clinical Escherichia coli and Klebsiella spp. Isolates from the Western Cape of South Africa.

Objectives: Colistin is a last-resort antibiotic for the treatment of carbapenem-resistant Gram-negative infections. Colistin resistance thus poses a threat to human health. Colistin resistance is most commonly encoded by mutations in chromosomal pmrA, pmrB, phoP, phoQ, ccrB, and mgrB genes, and the presence of plasmid-mediated mcr genes. This study describes colistin resistance mechanisms in clinical Enterobacterales isolates from the Western Cape, South Africa. Results: Escherichia coli (n = 22) and Klebsiella spp. (n = 7) isolates, from nine health care facilities, were confirmed to be colistin resistant during 2016 and 2017. mcr-1 was present in 55% (12/22) of E. coli and 71% (5/7) of Klebsiella spp. isolates. Colistin resistance mutations in pmrB were identified in 8/10 mcr-negative E. coli isolates using whole-genome sequencing, with pmrB Pro-94→Gln being the most frequent with presence in 4 isolates. One mcr-negative Klebsiella spp. isolate had a complete deletion of the mgrB and one contained an insertion sequence (IS1) in mgrB. Conclusion: A reduction in the proportion of colistin-resistant isolates harboring mcr-1 from 2016 to 2017 was observed. Colistin-resistant E. coli attributed by chromosomal mutations in pmrB in 2017 were mostly clonal related, which contrasts with the 2016 unrelated mcr-1-positive isolates. The diverse strains, hospitals, and resistance mechanisms may suggest that selective pressure is the main driver of colistin resistance.

Characterisation of mcr-4.3 in a colistin-resistant Acinetobacter nosocomialis clinical isolate from Cape Town, South Africa.

OBJECTIVES: Colistin resistance in Acinetobacter spp. is increasing, resulting in potentially untreatable nosocomial infections. Plasmid-mediated colistin resistance is of particular concern due to its low fitness cost and potential transferability to other bacterial strains and species. This study investigated the colistin resistance mechanism in a clinical Acinetobacter nosocomialis isolate from Cape Town, South Africa. METHODS: A colistin-resistant A. nosocomialis isolate was identified from a blood culture in 2017. PCR and Illumina whole-genome sequencing (WGS) were performed to identify genes and mutations conferring resistance to colistin. Plasmid sequencing was performed on an Oxford Nanopore platform. mcr functionality was assessed by broth microdilution after cloning the mcr gene into pET-48b(+) and expressing it in SHuffle® T7 Escherichia coli and after curing the plasmid using 62.5 mg/L acridine orange. RESULTS: The colistin minimum inhibitory concentration (MIC) of the A. nosocomialis isolate was 16 mg/L. The mcr-4.3 gene was detected by PCR and WGS. No other previously described colistin resistance mechanism was found by WGS. The mcr-4.3 gene was identified on a 24 024-bp RepB plasmid (pCAC13a). Functionality studies showed that recombinant mcr-4.3 did not confer colistin resistance in E. coli. However, plasmid curing of pCAC13a restored colistin susceptibility in A. nosocomialis. CONCLUSION: We describe the first detection of a plasmid-mediated mcr-4.3 gene encoding colistin resistance in A. nosocomialis and the first detection of mcr-4.3 in a clinical isolate in Africa. Recombinant expression of mcr-4.3 did not confer colistin resistance in E. coli, suggesting that its functionality may be RepB plasmid-dependent or species-specific.

Clonal expansion of colistin-resistant Acinetobacter baumannii isolates in Cape Town, South Africa.

OBJECTIVES: To describe colistin-resistant Acinetobacter baumannii isolates in Cape Town, South Africa. METHODS: A. baumannii isolates identified on Vitek 2 Advanced Expert System were collected from Tygerberg Hospital referral laboratory between 2016 and 2017. Colistin resistance was confirmed using broth microdilution and SensiTest. mcr-1-5 were detected using PCR and strain typing was performed by rep-PCR. Whole genome sequencing (WGS) was performed on a subset of isolates to identify chromosomal colistin resistance mechanisms and strain diversity using multilocus sequence typing (MLST) and pairwise single nucleotide polymorphism analyses. RESULTS: Twenty-six colistin-resistant and six colistin-susceptible A. baumannii were collected separately based on Vitek susceptibility; 20/26 (77%) were confirmed colistin-resistant by broth microdilution. Four colistin-resistant isolates were isolated in 2016 and 16 in 2017, from five healthcare facilities. Thirteen colistin-resistant isolates and eight colistin-susceptible isolates were identical by rep-PCR and MLST (ST1), all from patients admitted to a tertiary hospital during 2017. The remaining colistin-resistant isolates were unrelated. CONCLUSIONS: An increase in colistin-resistant A. baumannii isolates from a tertiary hospital in 2017 appears to be clonal expansion of an emerging colistin-resistant strain. This strain was not detected in 2016 or from other hospitals. Identical colistin-susceptible isolates were also isolated, suggesting relatively recent acquisition of colistin resistance.

Decreasing fluconazole susceptibility of clinical South African Cryptococcus neoformans isolates over a decade.

BACKGROUND: Fluconazole is used in combination with amphotericin B for induction treatment of cryptococcal meningitis and as monotherapy for consolidation and maintenance treatment. More than 90% of isolates from first episodes of cryptococcal disease had a fluconazole minimum inhibitory concentration (MIC) ≤4 µg/ml in a Gauteng population-based surveillance study of Cryptococcus neoformans in 2007-2008. We assessed whether fluconazole resistance had emerged in clinical cryptococcal isolates over a decade. METHODOLOGY AND PRINCIPAL FINDINGS: We prospectively collected C. neoformans isolates from 1 January through 31 March 2017 from persons with a first episode of culture-confirmed cryptococcal disease at 37 South African hospitals. Isolates were phenotypically confirmed to C. neoformans species-complex level. We determined fluconazole MICs (range: 0.125 µg/ml to 64 µg/ml) of 229 C. neoformans isolates using custom-made broth microdilution panels prepared, inoculated and read according to Clinical and Laboratory Standards Institute M27-A3 and M60 recommendations. These MIC values were compared to MICs of 249 isolates from earlier surveillance (2007-2008). Clinical data were collected from patients during both surveillance periods. There were more males (61% vs 39%) and more participants on combination induction antifungal treatment (92% vs 32%) in 2017 compared to 2007-2008. The fluconazole MIC50, MIC90 and geometric mean MIC was 4 µg/ml, 8 µg/ml and 4.11 µg/ml in 2017 (n = 229) compared to 1 µg/ml, 2 µg/ml and 2.08 µg/ml in 2007-2008 (n = 249) respectively. Voriconazole, itraconazole and posaconazole Etests were performed on 16 of 229 (7%) C. neoformans isolates with a fluconazole MIC value of ≥16 µg/ml; only one had MIC values of >32 µg/ml for these three antifungal agents. CONCLUSIONS AND SIGNIFICANCE: Fluconazole MIC50 and MIC90 values were two-fold higher in 2017 compared to 2007-2008. Although there are no breakpoints, higher fluconazole doses may be required to maintain efficacy of standard treatment regimens for cryptococcal meningitis.

Plasmid-mediated mcr-1 colistin resistance in Escherichia coli and Klebsiella spp. clinical isolates from the Western Cape region of South Africa.

BACKGROUND: Colistin is a last resort antibiotic for the treatment of carbapenem-resistant Gram negative infections. Until recently, mechanisms of colistin resistance were limited to chromosomal mutations which confer a high fitness cost and cannot be transferred between organisms. However, a novel plasmid-mediated colistin resistance mechanism, encoded by the mcr-1 gene, has been identified, and has since been detected worldwide. The mcr-1 colistin resistance mechanism is a major threat due to its lack of fitness cost and ability to be transferred between strains and species. Surveillance of colistin resistance mechanisms is critical to monitor the development and spread of resistance.This study aimed to determine the prevalence of the plasmid-mediated colistin resistance gene, mcr-1, in colistin-resistant E. coli and Klebsiella spp. isolates in the Western Cape of South Africa; and whether colistin resistance is spread through clonal expansion or by acquisition of resistance by diverse strains. METHODS: Colistin resistant E. coli and Klebsiella spp. isolates were collected from the NHLS microbiology laboratory at Tygerberg Hospital. Species identification and antibiotic susceptibility testing was done using the API® 20 E system and the Vitek® 2 Advanced Expert System™. PCR was used to detect the plasmid-mediated mcr-1 colistin resistance gene and REP-PCR was used for strain typing of the isolates. RESULTS: Nineteen colistin resistant isolates, including 12 E. coli, six K. pneumoniae and one K. oxytoca isolate, were detected over 7 months from eight different hospitals in the Western Cape region. The mcr-1 gene was detected in 83% of isolates which were shown to be predominantly unrelated strains. CONCLUSIONS: The plasmid-mediated mcr-1 colistin resistance gene is responsible for the majority of colistin resistance in clinical isolates of E. coli and Klebsiella spp. from the Western Cape of South Africa. Colistin resistance is not clonally disseminated; the mcr-1 gene has been acquired by several unrelated strains of E. coli and K. pneumoniae. Acquisition of mcr-1 by cephalosporin- and carbapenem-resistant Gram negative bacteria may result in untreatable infections and increased mortality. Measures need to be implemented to control the use of colistin in health care facilities and in agriculture to retain its antimicrobial efficacy.

Antigenic distribution of Streptococcus agalactiae isolates from pregnant women at Garankuwa hospital - South Africa.

INTRODUCTION: Streptococcus agalactiae (group B streptococcus; GBS) is globally recognised as one of the leading causes of neonatal sepsis and meningitis. It also causes adverse pregnancy outcomes such as stillbirth and miscarriages. Incidence of invasive disease is increasing in non-pregnant adults with underlying medical conditions (e.g., diabetes mellitus). Epidemiological studies of GBS infections are based on capsular serotyping. Genotyping of the surface anchored protein genes is also becoming an important tool for GBS studies. Currently ten different GBS serotypes have been identified. This study was performed to determine the prevalence of GBS capsular types (CTs) and surface anchored protein genes in isolates from colonized pregnant women attending antenatal clinic, at Dr George Mukhari Academic Hospital, Garankuwa, Pretoria, South Africa. METHODS: The samples were collected over 11 months and cultured on selective media. GBS was identified using different morphological and biochemical tests. Capsular typing was done using latex agglutination test and conventional PCR. Multiplex PCR with specific primers was used to detect the surface anchored protein genes. RESULTS: Of the 413 pregnant women recruited, 128 (30.9%) were colonized with GBS. The capsular polysaccharide (CPS) typing test showed that CPS type III (29.7%) was the most prevalent capsular type followed by CPS type Ia (25.8%), II (15.6%), IV (8.6%), V (10.9%) and Ib (8.6%); 0.7% of the isolates were nontypeable. Multiplex PCR revealed that the surface proteins genes were possessed by all the capsular types: rib (44.5%), bca (24.7%), alp2/3 (17.9%), epsilon (8.6%) and alp4 (4.7%). CONCLUSION: The common capsular types found in this study are Ia, III, and II. The most common protein genes identified were rib and bca, and the distribution of the surface protein genes among the isolates of different capsular types showed similar trends to the distribution reported from previous studies.

Antibiotic resistance of Streptococcus agalactiae isolated from pregnant women in Garankuwa, South Africa.

BACKGROUND: This study was undertaken to determine the susceptibility profile and the mechanism of antibiotic resistance in Group B streptococcus (GBS) isolates detected in vaginal and rectal swabs from pregnant women attending Dr George Mukhari Academic Hospital, a University Teaching Hospital in Pretoria, South Africa. METHODS: The samples were collected over an 11-month period, cultured on selective media (colistin and nalidixic acid agar and Todd-Hewitt broth), and GBS positively identified by using different morphological and biochemical tests. The susceptibility testing was done using the Kirby-Bauer and E test methods according to CLSI guidelines 2012. The D test method was used for the detection of inducible clindamycin resistance. Multiplex PCR with specific primers was used to detect different genes coding for resistance. RESULTS: Out of 413 samples collected, 128 (30.9%) were positive with GBS. The susceptibility testing revealed that 100% of isolates were sensitive to penicillin, ampicillin, vancomycin and high level gentamicin. Erythromycin and clindamycin resistance was 21.1 and 17.2%, respectively, in which 69% had harboured constitutive macrolide, lincosamide and streptogramin B (MLS(B)), 17.4% had inducible MLS(B). The M and L phenotypes were present in 6.8% each. The methylation of target encoded by ermB genes was the commonest mechanism of resistance observed in 55% of isolates, 38% of isolates had both ermB and linB genes and efflux pump mediated by mefA genes was also distributed among the isolates. CONCLUSIONS: The study reaffirmed the appropriateness of penicillin as the antibiotic of choice for treating GBS infection. However it identified the challenges of resistance to macrolides and lincosamides used as alternative drugs for individuals allergic to penicillin. More GBS treatment options for penicillin allergic patients need to be researched on.