Browsing by Author "Singh, Ravesh."

Now showing 1 - 12 of 12

Association of polymorphisms in the regulatory region of the cyclophilin A gene (PPIA) with gene expression and HIV/AIDS disease progression.
(Wolters Kluwer Health., 2016) Madlala, Paradise Zamokuhle.; Singh, Ravesh.; An, Ping.; Werner, Lise.; Mlisana, Koleka Patience.; Abdool Karim, Salim Safurdeen.; Winkler, Cheryl Ann.; Ndung'u, Peter Thumbi.
Abstract available in PDF file.
Association of TRIM22 with the Type 1 Interferon Response and Viral Control during Primary HIV-1 Infection.
(American Society for Microbiology., 2010) Singh, Ravesh.; Gaiha, Gaurav.; Werner, Lise.; Mlisana, Koleka Patience.; Luban, Jeremy.; Walker, Bruce D.; Abdool Karim, Salim Safurdeen.; Ndung'u, Peter Thumbi.; Brass, Abraham.; McKim, Kevin.
Type 1 interferons (IFNs) induce the expression of the tripartite interaction motif (TRIM) family of E3 ligases, but the contribution of these antiviral factors to HIV pathogenesis is not completely understood. We hypothesized that the increased expression of select type 1 IFN and TRIM isoforms is associated with a significantly lower likelihood of HIV-1 acquisition and viral control during primary HIV-1 infection. We measured IFN-a, IFN-b, myxovirus resistance protein A (MxA), human TRIM5a (huTRIM5a), and TRIM22 mRNA levels in peripheral blood mononuclear cells (PBMCs) of high-risk, HIV-1-uninfected participants and HIV-1-positive study participants. Samples were available for 32 uninfected subjects and 28 infected persons, all within 1 year of infection. HIV-1-positive participants had higher levels of IFN-b(P=0.0005), MxA (P=0.007), and TRIM22 (P=0.01) and lower levels of huTRIM5a (P< 0.001) than did HIV-1-negative participants. TRIM22 but not huTRIM5a correlated positively with type 1 IFN (IFN-a, IFN-b, and MxA) (all P<0.0001). In a multivariate model, increased MxA expression showed a significant positive association with viral load (P=0.0418). Furthermore, TRIM22 but not huTRIM5a, IFN-a, IFN-b, or MxA showed a negative correlation with plasma viral load (P=0.0307) and a positive correlation with CD4 T-cell counts (P=0.0281). In vitro studies revealed that HIV infection induced TRIM22 expression in PBMCs obtained from HIV-negative donors. Stable TRIM22 knockdown resulted in increased HIV-1 particle release and replication in Jurkat reporter cells. Collectively, these data suggest concordance between type 1 IFN and TRIM22 but not huTRIM5a expression in PBMCs and that TRIM22 likely acts as an antiviral effector in vivo.
Beyond syndromic management: opportunities for diagnosis-based treatment of sexually transmitted infections in low- and middle-income countries.
(Public Library of Science., 2018) Garrett, Nigel Joel.; Osman, Farzana.; Maharaj, Bhavna.; Naicker, Nivashnee.; Gibbs, Andrew.; Norman, Emily.; Samsunder, Natasha.; Ngobese, Hope.; Mitchev, Nireshni.; Singh, Ravesh.; Abdool Karim, Salim Safurdeen.; Kharsany, Ayesha Bibi Mahomed.; Mlisana, Koleka Patience.; Rompalo, Anne.; Mindel, Adrian.
Abstract available in pdf.
Case report: mechanisms of HIV elite control in two African women.
(BioMed Central., 2018) Moosa, Yumna.; Tanko, Ramla F.; Ramsuran, Veron.; Singh, Ravesh.; Madzivhandila, Mashudu.; Yende-Zuma, Fortunate Nonhlanhla.; Abrahams, Melissa-Rose.; Selhorst, Philippe.; Gounder, Kamini.; Moore, Penelope L.; Williamson, Carolyn.; Abdool Karim, Salim Safurdeen.; Garrett, Nigel Joel.; Burgers, Wendy A.
Abstract available in pdf.
Cytokine immune response profiles during 5 intestinal helminths and Mycobacterium 6 tuberculosis coinfection: An in vitro and human ex vivo study in KwaZulu-Natal.
(2023) Bhengu, Khethiwe Nomcebo.; Mkhize-Kwitshana, Zilungile Lynette.; Singh, Ravesh.; Naidoo, Pragalathan.
Background: There is a striking geographic overlap between helminths and tuberculosis (TB), particularly in developing countries like Africa. Underprivileged communities are more susceptible to these illnesses due to poverty, poor sanitation, and other environmental factor Helminth and tuberculosis infections exhibit distinct immune responses, which may be antagonistic in coinfected hosts and lead to poor prognosis. Helminth infections induce anti422 inflammatory Th2/Treg responses contrary to the pro-inflammatory Th1 responses triggered by Mycobacterium tuberculosis (Mtb) infection. Reduced TB protection has been associated with a strong Th2 response. Uncertainty exists on how helminth infection affects the host’s resistance to TB. This necessitates further investigation of immune responses in helminths and TB coinfection cases, particularly in KwaZulu-Natal (KZN). Aim: To determine the cytokine response profiles during intestinal helminth and TB coinfection using lymphocytic Jurkat and monocytic THP-1 cell lines for the in vitro study and TB and helminth coinfected South African adults for the human ex vivo study. Methods: Lymphocytic Jurkat and monocytic THP-1 cell lines were stimulated for 24 and 48 hours with Mtb H37Rv and Ascaris lumbricoides (A. lumbricoides) excretory-secretory protein extracts for the in vitro study. A cross-sectional study on consenting adult participants (≥18 years) (n = 414) recruited from primary health care clinics was conducted between March 2020 and August 2021 in Durban, KwaZulu Natal, for the pilot human ex vivo study. Blood and stool samples were collected from the recruited participants. The Kato-Katz and Mini-Parasep faecal parasite concentration techniques were used to detect intestinal parasite infections in stool samples. Blood samples were analysed to determine A. lumbricoides-specific immunoglobulin E (IgE) and immunoglobulin G4 (IgG4) levels to improve microscopy sensitivity. In this study, cytokine analysis was undertaken for 164 participants; 96 were HIV infected and had to be excluded, leaving 68 eligible participants. The eligible individuals were subdivided into uninfected controls (no helminth and TB infection) (n = 18), helminth only infected (n = TB only infected (n = 6), and TB and helminth co-infected (n = 6) groups. Thereafter, for both the in vitro and ex vivo study, the gene expression profiles of the T helper type 1(Th1) and transcription factors [Interferon-γ (IFN-γ), Tumour necrosis factor-α (TNF-α), Interleukin-2 (ILxvii 2), Nuclear factor of activated T cells 2 (NFATC2), Eomesodermin 446 (eomes), T helper 2 (Th2) and transcription factors (Interleukin-4 (IL-4), Interleukin5 (IL-5, Transforming growth factor-β (TGF-β), T helper type 17 (Th17) (Interleukin-17 (IL-17), immune protein and proteases (Granzyme B, Perforin), Regulatory T cells (Tregs) (Interleukin-10 (IL-10) and Fork head box P3 (FoxP3)] and the uninfected controls, TB alone, helminth alone and coinfected groups were determined using RT-qPCR. Results: (i) In vitro study: TB-stimulated Jurkat cells had significantly higher levels of IFN-γ, TNF-α, Granzyme B, and perforin compared to unstimulated controls, LPS, A. lumbricoides, and A. lumbricoides plus TB costimulated cells (p<0.0001). IL-2, IL-17, Eomes, and NFATC2 levels were also higher in TB-stimulated Jurkat cells (p<0.0001). TB alone stimulated cells had lower IL-5 and IL-4 levels compared to A. lumbricoides alone stimulated and TB plus A. lumbricoides costimulated Jurkat and THP-1 cells (p<0.0001. A. lumbricoides alone stimulated cells had higher IL-4 levels compared to TB plus A. lumbricoides costimulated Jurkat and THP- 1 cells (p<0.0001). TGF-β levels were also lower in TB alone stimulated cells compared to TB plus A. lumbricoides costimulated cells. IL-10 levels were lower in TB stimulated Jurkat and THP-1 cells compared to TB plus A. lumbricoides costimulated cells (p<0.0001. (ii) Ex vivo study: Similar results were noted for both the in vitro and the ex vivo study, although the human study had a smaller sample size. Conclusion: Data suggest that helminths induce a predominant anti-inflammatory Th2 and Treg response which may downregulate critical proinflammatory Th1 responses crucial for TB protection.
Diagnostic accuracy of the Xpert CT/NG and OSOM Trichomonas Rapid assays for point-of-care STI testing among young women in South Africa: a cross-sectional study.
(BMJ Publishing Group., 2019) Osman, Farzana.; Naidoo, Jessica.; Dorward, Jienchi.; Singh, Ravesh.; Ngobese, Hope.; Rompalo, Anne.; Mindel, Adrian.; Garrett, Nigel Joel.; Mlisana, Koleka Patience.
Abstract available in PDF.
Molecula characterization of chlamdia trachomatis isolates using sequence variation in the major outer membrane protein gene (OMP1) and evaluation of their susceptibility profile.
(2019) Khanyile, Thobile Nokwazi.; Mlisana, Koleka Patience.; Singh, Ravesh.
Chlamydia trachomatis infections are the most common bacterial sexually transmitted infections (STIs) in humans, worldwide. Due to asymptomatic nature of C. trachomatis, the need for sensitive, reliable and affordable laboratory methods for diagnosis is critical. The aim of this study was to ascertain if the genetic profiles of different C. trachomatis isolates associate with antibiotic resistance.Two hundred and sixty-five Eswab™ clinical samples were screened for Ct using Anyplex™ II STI-7 Detection. We have applied High Resolution Melting Analysis (HRMA) for the genotyping of the Ct and applied it specifically to the 14 sexually transmitted infection-related genotypes: AC, D-K and L1-L3. Based on the genotype of the OMP1 (Outer Membrane Protein) gene C. trachomatis is grouped into different serovars, which present in different clinical manifestations; with type A, B, Ba, and C causing trachoma, D-K cause urogenital infections and LI, LII & LIII associated with lymphogranuloma venereum (LGV). We confirmed the presence of the OMP1 gene with the conventional PCR. HRMA was performed to identify the C. trachomatis serovars on a Quantstudio 5 real – time PCR instrument and CDC control strains were included in the analysis. HRM analysis was done on the High-Resolution Melt Softwarev3.1. We identified the following serovars A, B, C, D, E, F, G, I, J, L3 and our prevalence for the above serovars were as follows 3.2%, 6.4%, 3.2%, 9.7%, 16.1%, 29%, 9.7%, 12.9%, 3.2% and 6.4%, respectively. None of the serovars: H, K, L1, L2 were observed. A TaqMan real time PCR assay was also performed to measure the bacterial concentration of each C. trachomatis positive sample to elucidate if there is any association with the serovar type. D-K serovars had higher bacterial load compared to A-C and L3 serovars, (p =0.0045). We also performed sanger sequencing on ribosomal proteins (L4 and L22) to determine the presence of mutations that have been previously associated with drug resistance. The ribosomal protein L4 had mutations located in 7 different positions, significant mutations associated with macrolides resistance were observed at amino acid number 109 and 151. Ribosomal protein L22 had 21 samples with mutation at amino acid number 24, that has not been associated with resistance before. Based on our study and previous studies, it is clear that macrolide resistance in C.trachomatis is multifactorial besides changes in the amino acids.
Prevalence and molecular susceptibility of Mycoplasma genitalium in KwaZulu-Natal population.
(2021) Mvuna, Londeka Desire.; Mlisana, Koleka Patience.; Singh, Ravesh.
Background: Mycoplasma genitalium is a recently classified sexually transmitted pathogen associated with causing urethritis and cervicitis, potentially causing reproductive complications. Mycoplasma genitalium has been shown to develop resistance to the currently recommended drug regimens, namely, azithromycin used as first-line therapy and moxifloxacin (second-line). The resistance prevalence of M. genitalium to the current treatment is diverse across the world. In South Africa, few studies have evaluated the prevalence of resistance to azithromycin and other fluoroquinolones. This study was conducted to evaluate the presence of macrolide and fluoroquinolone-resistant gene mutations in a KwaZulu-Natal population infected with M. genitalium. Methods: Samples from the CAPRISA HIPPS cohort study were used for this analysis. Deoxyribonucleic acid was extracted from 100 stored M. genitalium positive self-collected vaginal swab samples. Real-time PCR was performed to confirm M. genitalium positivity. Genes associated with resistance to macrolides (23S rRNA, L4, L22) and fluoroquinolones (gryA) were sequenced and analysed. Results: All 100 samples were confirmed genotypically to be M. genitalium positive and were sequenced. From the 100 samples tested for M. genitalium, 73 were successful for 23S rRNA, 99 for L4, 91 for L22, and 80 for gryA. Of the seventy-three 23S rRNA sequences, five samples carried mutations associated with macrolide resistance. A total of 12 mutations coding for macrolide resistance (5 for 23S, 4 for L4, and 3 for L22) were identified following sequencing. The prevalence of resistance mutations to macrolides was thus 7% (5 of 73) for 23 S rRNA, 4% (4of 99) for L4, and 3% (3 of 91) for L22 protein. In the five samples that harboured 23S rRNA gene mutation one had: A2071T, A2072C, A2072T, and two with A2072G mutation. L4 and L22 harboured silent mutations at positions: T327C, G429A, C438T, C438A, G81A, and C351T. In the 80 samples successful for gryA, six carried mutations with a prevalence of 8% (6/80 x100%). Conclusion: Ongoing antimicrobial surveillance needs to be performed in our local populations as in our study we have seen the presence of mutations to macrolides and fluoroquinolones. This study was performed on samples collected during the early stages of introducing azithromycin as the first-line regimen for M. genitalium infection. More current studies need to be performed at a later time point to evaluate the antimicrobial resistance burden in M. genitalium.
The prevalence and risk factors for genital mycmoplasmas in Human immunodeficiency virus infected pregnant women from King Edward Vlll hospital.
(2021) Nundlall, Nikita.; Abbai, Nathlee Samantha.; Singh, Ravesh.
Background: Genital mycoplasmas can be found amongst the normal human flora mostly in the respiratory, reproductive and urinary tracts as commensal or pathogenic organisms. These bacteria are sexually transmitted and can be linked to sexually transmitted diseases and other conditions. There are a limited number of studies conducted in South African pregnant women especially from KwaZulu-Natal which have assessed the prevalence, co-infection rates and risk factors for genital mycoplasmas. In this study, the prevalence, co-infection rates and risk factors for Mycoplasma genitalium, M. hominis, Ureaplasma urealyticum and U. parvum were investigated in a cohort of Human immunodeficiency virus (HIV) infected pregnant women. The data generated in this study, therefore adds to the growing body of knowledge on these pathogens. Methods: This study included 264 HIV infected pregnant women attending the King Edward VIII antenatal clinic in eThekwini, South Africa. The women were recruited between October 2020 and April 2021. Each enrolled women provided self-collected vaginal swabs (dry swabs) for detection of the vaginal infections. The consenting women had also completed a questionnaire on socio-demographic, behavioural and clinical factors. DNA was extracted from the vaginal samples using the PureLink Microbiome kit. The individual pathogens were detected using the TaqMan Real-time PCR assays using commercially available primers and probes on a QuantStudio 5 Real-time polymerase chain reaction (PCR) platform. The statistical data analysis was conducted in a freely available Statistical Computing Environment, R software, version 3.6.3 using the RStudio platform. Results: The most prevalent infection in the study population was U. urealyticum, 236/264 (89.4%), followed by M. hominis, 215/264 (81.4%), U. parvum, 203/264 (76.9%) and lastly M. genitalium, 7/264 (2.70%). A total of five women (1.90%) were coinfected with all four microorganisms. Within the group of women who tested positive for Mycoplasma genitalium (M. genitalium), partner having other partners was the only significant behavioral factor in relation with being positive, p=0.031. However, a smaller proportion of positive women reported that their partner had other partners (28.6%) when compared to 57.1% who reported that their partner did not have other partners and 14.3% who did not know if their partner had other partners. Within the group of women who tested positive for Mycoplasma hominis (M. hominis), partner having STI symptoms was a significant clinical factor in relation with being positive, p=0.027. Women that experienced current symptoms of STIs was significantly associated with being positive, p<0.001. In addition, of the M. hominis positive women, a higher proportion, 80.5% tested positive for U. parvum infection compared to 19.5% who tested negative and this was significant, p=0.004. Partner being circumcised was a significant clinical factor in relation with being positive for Ureaplasma urealyticum (U. urealyticum), p=0.028. In addition, partner having symptoms of STIs was a significant clinical factor in relation with being positive, p=0.027. The majority of the positive women were in the third trimester of pregnancy and trimester of pregnancy was significantly associated with being positive for infection, p=0.040. Of the women who tested positive for U. urealyticum, a higher proportion of women also tested positive for M. hominis and this association was significant, p=0.051. Within the group of women who tested positive for Ureaplasma parvum (U. parvum), partners HIV status was significant in relation with being positive, p=0.049. Lifetime number of sex partners was significantly associated with being positive, p=0.012. Partner having other partners was also a significant factor in relation with being positive, p=0.023. Of the U. parvum positive women, a higher proportion of women (85.2%) tested positive for M. hominis. This association was significant, p=0.004. In the adjusted analysis, being employed increased the risk of getting infected with M. hominis p=0.012. In the adjusted analysis, current STI symptoms increased the risk for M. hominis by 95.27 fold, p<0.001. Being U. parvum positive increased the risk for M. hominis by 8.19 fold, p=0.001. Being U. urealyticum positive also increased the risk for M. hominis, p=0.039. In the adjusted analysis, having >4 lifetime sex partners increased the risk of infection with U. parvum by 88.02 fold. This factor was significant, p<0.001. Partner having other partners increased the risk of infection with U. parvum, p=0.008. In the adjusted analysis, being M. hominis positive increased the risk for U. parvum by 4.33 fold, p=0.008. Conclusion: The present study provides information on the risk factors associated with genital mycoplasma infections. The identification of risk factors provides the foundation for the development of prevention interventions. In this study, clinical and behavioral factors were shown to be significantly associated with the risk for infection. Based on this finding, it is evident that a single prevention strategy will not be sufficient, what will be needed is a combination prevention strategy for this vulnerable population.
The prevalence of Methicillin-resistant Staphylococcus aureus in blood product hampers at the South African National Blood Service.
(2021) Seoraj, Varsha.; Chuturgoon, Anil Amichund.; Singh, Ravesh.; Van den Berg, Karin.
Background: Methicillin-resistant Staphylococcus aureus (MRSA), are strains of the Gram-positive cocci known to cause various health conditions. Patients suffer from abscesses, skin infections and more severe conditions such as osteomyelitis and septicaemia. These bacteria are highly resistant to antibiotics and bacteria such as these are a great risk to the public, especially since Staphylococcus aureus is an opportunistic bacterium. In 2016, a donor unit received for the production of eye serum at South African National Blood Service (SANBS), when quality controlled, tested positive for MRSA. The bacterial contamination was traced to a staff member at the clinic where the blood was donated. Little research has been conducted to determine if MRSA is a problem and if it could negatively affect the blood supply. Aim: The aim of this study was to determine whether blood services contribute to the spread of MRSA and other bacterial pathogens through the blood product hamper system. Materials and Methods: A cross-sectional study to determine the prevalence of MRSA on 850 blood product hampers moving between SANBS inventory laboratories and blood banks, was conducted at SANBS between August 2020 and May 2021. Hampers were swabbed with a Sigma-Transwab containing liquid Amies transport medium for the detection of MRSA. The swabs were cultured onto CHROMagar MRSA where a rose or mauve coloured colony confirmed the presence of MRSA. Bacterial contaminants which were detected during the testing procedure were isolated, and loaded onto the Vitek 2 Compact for bacterial identification. Results: A total of 696 hampers were processed as per the study protocol (81.9%). Out of the 850 hampers planned to be swabbed, 143 (16.8%) hampers were not swabbed as a result of staff not performing the procedure and swabs from 11 hampers were omitted (1.3%) as they did not comply too protocol requirements. Of the 696 hampers swabbed, MRSA was not detected (0%) however, bacterial growth other than MRSA was observed. The most common isolates detected were Aerococcus viridans, Rothia dentocariosa, followed by Bacillis spp as well as Stenotrophomonas maltophilia. Conclusion: The study findings have shown that an effective hamper cleaning system is needed to safeguard the integrity of our blood supply. The findings of this study should be taken into consideration throughout all provinces at SANBS, for the consistent and regular cleaning of hampers, which carry blood and blood products.
Regulation of TRIM E3 Ligases and Cyclophilin A and the impact on HIV-1 replication and pathogenesis.
(2011) Singh, Ravesh.; Ndung'u, Peter Thumbi.
No abstract available.
TRIM5α and TRIM22 are differentially regulated according to HIV-1 infection phase and compartment.
(American Society for Microbiology., 2014) Singh, Ravesh.; Patel, Vinod B.; Mureithi, Marianne W.; Naranbhai, Vivek.; Ramsuran, Duran.; Tulsi, Sahil.; Hiramen, Keshni.; Werner, Lise.; Mlisana, Koleka Patience.; Altfeld, Marcus.; Luban, Jeremy.; Kasprowicz, Victoria.; Dheda, Keertan.; Abdool Karim, Salim Safurdeen.; Ndung'u, Peter Thumbi.
The antiviral role of TRIM E3 ligases in vivo is not fully understood. To test the hypothesis that TRIM5α and TRIM22 have differential transcriptional regulation and distinct anti-HIV roles according to infection phase and compartment, we measured TRIM5α, TRIM22, and type I interferon (IFN-I)-inducible myxovirus resistance protein A (MxA) levels in peripheral blood mononuclear cells (PBMCs) during primary and chronic HIV-1 infection, with chronic infection samples being matched PBMCs and central nervous system (CNS)-derived cells. Associations with biomarkers of disease progression were explored. The impact of IFN-I, select proinflammatory cytokines, and HIV on TRIM E3 ligase-specific expression was investigated. PBMCs from individuals with primary and chronic HIV-1 infection had significantly higher levels of MxA and TRIM22 than did PBMCs from HIV-1-negative individuals (P < 0.05 for all comparisons). PBMCs from chronic infection had lower levels of TRIM5α than did PBMCs from primary infection or HIV-1-uninfected PBMCs (P = 0.0001 for both). In matched CNS-derived samples and PBMCs, higher levels of MxA (P = 0.001) and TRIM5α (P = 0.0001) in the CNS were noted. There was a negative correlation between TRIM22 levels in PBMCs and plasma viral load (r = -0.40; P = 0.04). In vitro, IFN-I and, rarely, proinflammatory cytokines induced TRIM5α and TRIM22 in a cell type-dependent manner, and the knockdown of either protein in CD4(+) lymphocytes resulted in increased HIV-1 infection. These data suggest that there are infection-phase-specific and anatomically compartmentalized differences in TRIM5α and TRIM22 regulation involving primarily IFN-I and specific cell types and indicate subtle differences in the antiviral roles and transcriptional regulation of TRIM E3 ligases in vivo.