SUNScholar - Browsing by Author "Newton-Foot, Mae"

Browsing by Author "Newton-Foot, Mae"

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Association between fluoroquinolone resistance and MRSA genotype in Alexandria, Egypt
(Nature, 2021-02-19) Alseqely, Mustafa; Newton-Foot, Mae; Khalil, Amal; El-Nakeeb, Mostafa; Whitelaw, Andrew; Abouelfetouh, Alaa
Antimicrobial stewardship isn’t strictly observed in most Egyptian hospitals, raising antibiotic resistance. Epidemiology of Egyptian MRSA isolates, or associations with resistance to other antibiotics remain largely unknown. We identified MRSA genotypes in Alexandria Main University Hospital (AMUH) and investigated rates of moxifloxacin resistance, an alternative MRSA treatment, among different genotypes. Antibiotic susceptibility of 72 MRSA clinical isolates collected in 2015 from AMUH was determined by disc diffusion and broth microdilution. spa- and Staphylococcal Cassette Chromosome mec (SCCmec) typing were performed; with multi-locus sequence typing conducted on isolates representing major genotypes. Resistance to moxifloxacin, levofloxacin and ciprofloxacin were 69%, 78% and 96%, respectively. spa type t037 (57%) was commonest, followed by t127 (12.5%), t267 (8%) and t688 (6%). SCCmec III predominated (57%), all of these were moxifloxacin resistant and 97.6% t037 (ST241). SCCmec IV, IV E and V represented 15%, 7% and 11% of the isolates, respectively, 79% of these were moxifloxacin susceptible and of different spa types. t127 (ST-1) was associated with SCCmec V in 56% of the isolates, mostly moxifloxacin susceptible. Moxifloxacin resistance was high, most resistant isolates belonged to t037 and SCCmec III, suggesting local dissemination and antibiotic pressure. We recommend caution in treating MRSA infections with moxifloxacin.
The association between pathogen factors and clinical outcomes in patients with Staphylococcus aureus bacteraemia in a tertiary hospital, Cape Town
(Elsevier, 2019-11) Abdulgader, Shima M.; van Rijswijk, Amike; Whitelaw, Andrew; Newton-Foot, Mae
Background: Staphylococcus aureus is a serious pathogen, able to cause life-threatening infections such as bacteraemia. The association between S. aureus microbial characteristics and clinical outcomes is under-investigated in African settings. This study aimed to determine the molecular epidemiology and virulence characteristics of S. aureus isolates from bacteraemic patients at Tygerberg Hospital, South Africa, and to investigate the associations between pathogen characteristics and clinical outcomes. Methods: This study included 199 S. aureus isolates collected from blood cultures between February 2015 and March 2017. Methicillin resistance was determined using disc diffusion and all resistant isolates were further characterized by staphylococcal cassette chromosome mec (SCCmec) typing. Genotyping was done using spa and agr typing, and agr functionality was assessed using the phenotypic δ-haemolysin assay. Logistic regression models were performed to describe the associations between strain characteristics and the clinical outcomes methicillin resistance, in-hospital mortality, and length of stay (LOS). Results: Of the 199 S. aureus isolates collected, 27% were MRSA, and the overall crude in-hospital mortality rate was 29%. Seventy-three different spa types were identified, including seven new types. Agr I was the most common type, in 99 (49.7%) isolates, followed by agr II, III, and IV in 57 (28.6%), 37 (18.6%), and six (3%) isolates, respectively. Agr dysfunctionality was observed in 25 (13%) isolates, mostly belonging to spa-clonal complex (CC) 012. Methicillin resistance was significantly associated with hospital-acquired infection (odds ratio (OR) 4.77, 95% confidence interval (CI) 2.09-10.87). A significant increase in mortality was observed with increasing age (OR 7.48, 95% CI 2.82-19.8) and having a hospital-acquired infection (OR 2.26, 95% CI 1.12-4.55). S. aureus strains with a functional agr system showed an association with longer duration of stay (OR 1.66, 95% CI 0.93-2.99). Conclusions: We report the lowest MRSA prevalence at Tygerberg Hospital for the past 10 years, and agr dysfunctionality was shown to be driven by a certain genotype, spa-CC012. Despite the limited available clinical data, the study provided insights into associations between S. aureus epidemiology and agr-related virulence characteristics, and clinical outcomes.
Carriage of colistin-resistant Gram-negative bacteria in children from communities in Cape Town (Tuberculosis child multidrug-resistant preventive therapy trial sub-study)
(AOSIS, 2021) Snyman, Yolandi; Whitelaw, Andrew C.; Maloba, Motlatji R. B.; Hesseling, Anneke C.; Newton-Foot, Mae
ENGLISH ABSTRACT: 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 mcr-4.3 in a colistin-resistant Acinetobacter nosocomialis clinical isolate from Cape Town, South Africa
(Elsevier, 2021) Snyman, Yolandi; Reuter, Sandra; Whitelaw, Andrew Christopher; Stein, Lisa; Maloba, Motlatji Reratilwe Bonnie; Newton-Foot, Mae
Objectives: Colistin resistance in Acinetobacter spp. is increasing, resulting in potentially untreatable noso- comial 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 function- ality 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 mech- anism 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. How- ever, 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 re- sistance 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
(Elsevier, 2020) Snyman, Yolandi; Whitelaw, Andrew Christopher; Reuter, Sandra; Dramowski, Angela; Maloba, Motlatji Reratilwe Bonnie; Newton-Foot, Mae
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.
Clonal expansion of colistin-resistant Acinetobacter baumannii isolates in Cape Town, South Africa
(Elsevier, 2020-02) Snyman, Yolandi; Whitelaw, Andrew Christopher; Reuter, Sandra; Dramowski, Angela; Maloba, Motlatji Reratilwe Bonnie; Newton-Foot, Mae
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.
Comparison of commercial assays and two-step approach to detect Clostridioides difficile in South Africa
(AOSIS, 2022-09) Singh, Sarishna; Newton-Foot, Mae; Nel, Pieter; Pienaar, Colette
Background: Clostridioides difficile is the number one cause of hospital-acquired diarrhoea. Accurate diagnosis of C. difficile is of utmost importance as it guides patient management and infection control practices. Studies evaluating the performance of commercially available nucleic acid amplification tests (NAATs) versus algorithms are lacking in resource-limited settings. Objective: This study assessed the performance of three commercially available tests and a two-step approach for the diagnosis of C. difficile infection using toxigenic culture (TC) as the gold standard. Methods: Two hundred and twenty-three non-duplicate loose stool samples were submitted to the National Health Laboratory Service Microbiology Laboratory at Tygerberg Hospital, Cape Town, South Africa, from October 2017 to October 2018. The samples were tested in parallel using the C. DIFF QUIK CHEK COMPLETE enzyme immunoassay (EIA) and two NAATs (Xpert C. difficile and BD MAX Cdiff), and the results were compared to TC. The performance of a two-step approach consisting of the C. DIFF QUIK CHEK COMPLETE followed by the Xpert C. difficile was also determined. Results: Of 223 faecal specimens tested, 37 (16.6%) were TC-positive. The sensitivity and specificity of the C. DIFF QUIK CHEK COMPLETE were 54.1% and 98.9%; Xpert C. difficile, 86.4% and 96.8%; BD MAX Cdiff, 89.2% and 96.8%; and two-step approach, 89.2% and 96.2%. Conclusion: The C. DIFF QUIK CHEK COMPLETE, in a two-step approach with the Xpert C. difficile, performed similarly to the NAATs on their own and offer advantages in terms of cost and workflow in low-resource settings.
The effect of storage conditions on microbial communities in stool
(Public Library of Science, 2020) Van Zyl, Kristien Nel; Whitelaw, Andrew C.; Newton-Foot, Mae
Microbiome research has experienced a surge of interest in recent years due to the advances and reduced cost of next-generation sequencing technology. The production of high quality and comparable data is dependent on proper sample collection and storage and should be standardized as far as possible. However, this becomes challenging when samples are collected in the field, especially in resource-limited settings. We investigated the impact of different stool storage methods common to the TB-CHAMP clinical trial on the microbial communities in stool. Ten stool samples were subjected to DNA extraction after 48-hour storage at -80˚C, room temperature and in a cooler-box, as well as immediate DNA extraction. Three stool DNA extraction kits were evaluated based on DNA yield and quality. Quantitative PCR was performed to determine the relative abundance of the two major gut phyla Bacteroidetes and Firmicutes, and other representative microbial groups. The bacterial populations in the frozen group closely resembled the immediate extraction group, supporting previous findings that storage at -80˚C is equivalent to the gold standard of immediate DNA extraction. More variation was seen in the room temperature and coolerbox groups, which may be due to the growth temperature preferences of certain bacterial populations. However, for most bacterial populations, no significant differences were found between the storage groups. As seen in other microbiome studies, the variation between participant samples was greater than that related to differences in storage. We determined that the risk of introducing bias to microbial community profiling through differences in storage will likely be minimal in our setting.
The Mycobacterium tuberculosis ESX-3 secretion system interactome
(Stellenbosch : University of Stellenbsoch, 2010-03) Newton-Foot, Mae; Van Pittius, Nicolaas Claudius Gey; Warren, Robin Mark; University of Stellenbosch. Faculty of Health Sciences. Dept. of Biomedical Sciences.
ENGLISH ABSTRACT: Mycobacterium tuberculosis is the causative agent of tuberculosis, a disease which causes approximately 2 million deaths each year. Despite extensive research on tuberculosis and M. tuberculosis, little is understood of the mechanisms of pathogenicity of the organism. The genome of M. tuberculosis contains five ESAT-6 gene cluster regions, each of which contains genes encoding proteins involved in the formation of a dedicated protein secretion system. Included in these regions are genes encoding exported T-cell antigens, serine proteases, ATP-binding proteins and other membrane-associated proteins. Although it is known that some of these secretion systems are involved in virulence and phagosomal escape of M. tuberculosis, and that deletion thereof causes attenuation of the organism, the structure, substrates and functions of the systems are largely unknown. Understanding the structure of the ESX secretion systems will advance our understanding of the mechanisms of mycobacterial pathogenicity and provide clues to ways in which to interfere with these virulence mechanisms. The ESAT-6 gene cluster region 3, encoding the ESX-3 secretion machinery, is the only ESAT-6 gene cluster region which is essential for the in vitro growth of M. tuberculosis. It is however not required for the growth of the saprophytic mycobacterium M. smegmatis. In this study we have identified proteinprotein interactions within the ESX-3 secretion system, using the Mycobacterial – Protein Fragment Complementation (M-PFC) mycobacterial two-hybrid system, and created a model of the M. tuberculosis ESX-3 secretion system. According to this model, the EsxG-EsxH and PE5-PPE4 substrate protein complexes bind to the same components of the ESX-3 secretion machinery and are secreted via the same mechanism. A knock-out of the ESX-3 secretion system in M. smegmatis was generated by homologous recombination to allow further research into the functions and properties of this secretion system. This knock-out was used, together with wild-type M. smegmatis, to investigate the secretion of the M. tuberculosis EsxH protein by the M. smegmatis ESX-3 secretion system. The ESX-3 secretion system interactome may serve as a model for the ESX secretion systems and assist in our understanding of this secretion machinery which is key to the virulence and survival of M. tuberculosis and other pathogenic mycobacteria. Improved understanding of these mechanisms and their role in pathogenicity and survival may provide means of interfering with the secretion machinery, potentially leading to developments in the prevention and treatment of tuberculosis disease.
The plasmid-mediated evolution of the mycobacterial ESX (Type VII) secretion systems
(BioMed Central, 2016-03) Newton-Foot, Mae; Warren, Robin Mark; Sampson, Samantha Leigh; Van Helden, Paul David; Gey van Pittius, Nicolaas Claudius
BACKGROUND: The genome of Mycobacterium tuberculosis contains five copies of the ESX gene cluster, each encoding a dedicated protein secretion system. These ESX secretion systems have been defined as a novel Type VII secretion machinery, responsible for the secretion of proteins across the characteristic outer mycomembrane of the mycobacteria. Some of these secretion systems are involved in virulence and survival in M. tuberculosis; however they are also present in other non-pathogenic mycobacteria, and have been identified in some non-mycobacterial actinomycetes. Three components of the ESX gene cluster have also been found clustered in some gram positive monoderm organisms and are predicted to have preceded the ESX gene cluster. RESULTS: This study used in silico and phylogenetic analyses to describe the evolution of the ESX gene cluster from the WXG-FtsK cluster of monoderm bacteria to the five ESX clusters present in M. tuberculosis and other slow-growing mycobacteria. The ancestral gene cluster, ESX-4, was identified in several nonmycomembrane producing actinobacteria as well as the mycomembrane-containing Corynebacteriales in which the ESX cluster began to evolve and diversify. A novel ESX gene cluster, ESX-4EVOL, was identified in some non-mycobacterial actinomycetes and M. abscessus subsp. bolletii. ESX-4EVOL contains all of the conserved components of the ESX gene cluster and appears to be a precursor of the mycobacterial ESX duplications. Between two and seven ESX gene clusters were identified in each mycobacterial species, with ESX-2 and ESX-5 specifically associated with the slow growers. The order of ESX duplication in the mycobacteria is redefined as ESX-4, ESX-3, ESX-1 and then ESX-2 and ESX-5. Plasmid-encoded precursor ESX gene clusters were identified for each of the genomic ESX-3, -1, -2 and -5 gene clusters, suggesting a novel plasmid-mediated mechanism of ESX duplication and evolution. CONCLUSIONS: The influence of the various ESX gene clusters on vital biological and virulence-related functions has clearly influenced the diversification and success of the various mycobacterial species, and their evolution from the non-pathogenic fast-growing saprophytic to the slow-growing pathogenic organisms.
Plasmid-mediated mcr-1 colistin resistance in Escherichia coli and Klebsiella spp. clinical isolates from the Western Cape region of South Africa
(BioMed Central, 2017-08-03) Newton-Foot, Mae; Snyman, Yolandi; Maloba, Motlatji Reratilwe Bonnie; Whitelaw, Andrew Christopher
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.
The prevalence and molecular mechanisms of mupirocin resistance in Staphylococcus aureus isolates from a Hospital in Cape Town, South Africa
(BMC (part of Springer Nature), 2020-03-14) Abdulgader, Shima M.; Lentswe, Tshepiso; Whitelaw, Andrew; Newton-Foot, Mae
Abstract Background: Antimicrobial resistance is an increasingly serious problem in public health globally. Monitoring resistance levels within healthcare and community settings is critical to combat its ongoing increase. This study aimed to describe the rates and molecular mechanisms of mupirocin resistance in clinical Staphylococcus aureus isolates from Tygerberg Hospital, and to describe its association with strain types. Methods: We retrospectively selected 212 S. aureus isolates which were identified from blood samples and pus swabs during the years 2009–2011 and 2015–2017. The isolates were identified using conventional microbiological methods and genotyping was done using spa typing. Cefoxitin (30 μg) disc diffusion and the two disc strategy (5 μg and 200 μg) were used to determine susceptibility to methicillin and mupirocin, respectively. Isolates with high-level resistance were screened for the plasmid mediated genes mupA and mupB by PCR, and sequencing of the ileS gene was done for all isolates exhibiting low-level resistance to describe the mutations associated with this phenotype. Chi-square test was used to assess the associations between mupirocin resistance and S. aureus genotypes. Results: Of 212 S. aureus isolates, 12% (n = 25) were resistant to mupirocin, and 44% (n = 93) were methicillin resistant. Strain typing identified 73 spa types with spa t045 being the most predominant constituting 11% of the isolates. High-level mupirocin resistance was observed in 2% (n = 5), and low-level resistance in 9% (n = 20) of the isolates. The prevalence of high-level mupirocin resistance amongst MRSA and MSSA was 4 and 1% respectively, while the prevalence of low-level mupirocin resistance was significantly higher in MRSA (18%) compared to MSSA (3%), (p = 0.032). mupA was the only resistance determinant for high-level resistance, and the IleS mutation V588F was identified in 95% of the isolates which showed low-level resistance. A significant association was observed between spa type t032 and high-level mupirocin resistance, and types t037 and t012 and low-level resistance (p < 0.0001). Conclusion: The study reported higher rates of low-level mupirocin resistance compared to high-level resistance, and in our setting, mupirocin resistance was driven by certain genotypes. Our study advocates for the continuous screening for mupirocin resistance in S. aureus in clinical settings to better guide treatment and prescribing practices. Background Antimicrobial resistance is an increasingly serious problem in public health globally. Monitoring resistance levels within healthcare and community settings is critical to combat its ongoing increase. This study aimed to describe the rates and molecular mechanisms of mupirocin resistance in clinical Staphylococcus aureus isolates from Tygerberg Hospital, and to describe its association with strain types. Methods We retrospectively selected 212 S. aureus isolates which were identified from blood samples and pus swabs during the years 2009–2011 and 2015–2017. The isolates were identified using conventional microbiological methods and genotyping was done using spa typing. Cefoxitin (30 μg) disc diffusion and the two disc strategy (5 μg and 200 μg) were used to determine susceptibility to methicillin and mupirocin, respectively. Isolates with high-level resistance were screened for the plasmid mediated genes mupA and mupB by PCR, and sequencing of the ileS gene was done for all isolates exhibiting low-level resistance to describe the mutations associated with this phenotype. Chi-square test was used to assess the associations between mupirocin resistance and S. aureus genotypes. Results Of 212 S. aureus isolates, 12% (n = 25) were resistant to mupirocin, and 44% (n = 93) were methicillin resistant. Strain typing identified 73 spa types with spa t045 being the most predominant constituting 11% of the isolates. High-level mupirocin resistance was observed in 2% (n = 5), and low-level resistance in 9% (n = 20) of the isolates. The prevalence of high-level mupirocin resistance amongst MRSA and MSSA was 4 and 1% respectively, while the prevalence of low-level mupirocin resistance was significantly higher in MRSA (18%) compared to MSSA (3%), (p = 0.032). mupA was the only resistance determinant for high-level resistance, and the IleS mutation V588F was identified in 95% of the isolates which showed low-level resistance. A significant association was observed between spa type t032 and high-level mupirocin resistance, and types t037 and t012 and low-level resistance (p < 0.0001). Conclusion The study reported higher rates of low-level mupirocin resistance compared to high-level resistance, and in our setting, mupirocin resistance was driven by certain genotypes. Our study advocates for the continuous screening for mupirocin resistance in S. aureus in clinical settings to better guide treatment and prescribing practices.
Two promoters in the esx-3 gene cluster of Mycobacterium smegmatis respond inversely to different iron concentrations in vitro
(Biomed Central, 2017-08) Fang, Zhuo; Newton-Foot, Mae; Sampson, Samantha Leigh; Gey Van Pittius, Nicolaas Claudius
Background The ESX secretion system, also known as the Type VII secretion system, is mostly found in mycobacteria and plays important roles in nutrient acquisition and host pathogenicity. One of the five ESXs, ESX-3, is associated with mycobactin-mediated iron acquisition. Although the functions of some of the membrane-associated components of the ESX systems have been described, the role of by mycosin-3 remains elusive. The esx-3 gene cluster encoding ESX-3 in both Mycobacterium tuberculosis and Mycobacterium smegmatis has two promoters, suggesting the presence of two transcriptional units. Previous studies indicated that the two promoters only showed a difference in response under acid stress (pH 4.2). This study aimed to study the effect of a mycosin-3 deletion on the physiology of M. smegmatis and to assess the promoter activities in wildtype, mycosin-3 mutant and complementation strains. Results The gene mycP 3 was deleted from wildtype M. smegmatis via homologous recombination. The mycP 3 gene was complemented in the deletion mutant using each of the two intrinsic promoters from the M. smegmatis esx-3 gene cluster. The four strains were compared in term of bacterial growth and intracellular iron content. The two promoter activities were assessed under iron-rich, iron-deprived and iron-rescued conditions by assessing the mycP 3 expression level. Although the mycP 3 gene deletion did not significantly impact bacterial growth or intracellular iron levels in comparison to the wild-type and complemented strains, the two esx-3 promoters were shown to respond inversely to iron deprivation and iron rescue. Conclusion This finding correlates with the previously published data that the first promoter upstream of msmeg0615, is upregulated under low iron levels but downregulated under high iron levels. In addition, the second promoter, upstream of msmeg0620, behaves in an inverse fashion to the first promoter implying that the genes downstream may have additional roles when the iron levels are high.
Understanding the evolution and function of the mycobacterial Type VII ESX secretion systems (T7SSs) and their substrates
(Stellenbosch : Stellenbosch University, 2013-03) Newton-Foot, Mae; Gey van Pittius, Nicolaas Claudius; Warren, Robin Mark; Stellenbosch University. Faculty of Medicine and Health Sciences. Dept. of Biomedical Sciences. Division of Molecular Biology and Human Genetics.
ENGLISH ABSTRACT: Mycobacterium tuberculosis, the causative agent of tuberculosis disease, contains five copies of the ESAT-6 gene cluster, each encoding a dedicated ESX protein secretion system which has been defined as a novel Type-VII secretion system. The ESX have been implicated in virulence and survival of M. tuberculosis, and as such present a promising target for novel treatment interventions. This study has investigated the evolution, regulation, functions and substrates of the ESX secretion systems. The evolutionary history of the ESX secretion systems was established using in silico and phylogenetic analyses of the sequenced mycobacteria, closely related actinomycetes and WXG-FtsK clusters from other bacteria. The ESX-4 gene cluster appears to have evolved with the start of the evolution of the mycomembrane, followed by the duplication of ESX-3, which marks the evolution of the genus Mycobacterium. The ESX-1 duplication occurred next, followed by ESX-2 and ESX-5 which occur only in the slow growing mycobacteria. Five additional ESX gene clusters were newly identified and named ESX-P1 to - P5. These additional ESX clusters occur, or are predicted to occur, on plasmid DNA, and appear to be progenitors of the genomic ESX-1 to -5 gene clusters, possibly indicating a plasmid-mediated mechanism of ESX duplication and evolution. The promoters expressing the M. tuberculosis ESX-1 to ESX-5 secretion systems were investigated using a promoter probe assay, and characterised using in silico analyses. Promoters were identified for ESX-1, -2, -3 and -5. The functions of the mycobacterial ESX secretion systems were investigated using whole proteomic, secretomic and metabolomic analyses of the fast growing, non-pathogenic M. smegmatis, which contains three of the ESX secretion systems, ESX-1, 3, and 4. ESX knockout strains of M. smegmatis were generated and used in comparative analyses with wild-type M. smegmatis. ESX-1 was highly expressed in wild-type M. smegmatis, however no specific pathways showed considerable variation when ESX-1 was deleted. Deletion of ESX-3 resulted in substantial variation to multiple cellular pathways, including amino acid, carbohydrate and fatty acid metabolism and oxidative stress. These and other differences indicate possible perturbed polyamine metabolism in the absence of ESX-3. Although no ESX-4 protein components were detected in wild type M. smegmatis, the ESX-4 knockout displayed substantial proteomic variation. Reduced levels of ESX-3 component proteins in the ESX-4 knockout suggest that ESX-4 influences ESX-3 expression. Other variation linked ESX-4 to cell division and molybdenum metabolism. Secretomic analyses of wild-type and ESX knockout M. smegmatis strains were used to search for novel substrates of the M. smegmatis ESX secretion systems. No prototype ESX substrates were identified in the culture filtrates, however 10 possible substrates of the ESX-1, -3 and -4 secretion systems, containing the general ESX secretion signal, YxxxD/E, were identified. The functions of some of these proteins correlate with the ESX functions identified in the proteomic and metabolomic analyses. This study sets the groundwork for future work in understanding the functional roles and expression patterns of these ESX secretion systems and in using global proteomic and metabolomic analyses to understand cellular changes in response to specific signals or genomic changes.

Browsing by Author "Newton-Foot, Mae"

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