Analysis of virulence factors and antibiotic resistance genes in group B streptococcus from clinical samples.

BACKGROUND: Streptococcus agalacticae (Group B Streptococcus, GBS) is one of the most important causative agents of serious infections among neonates. This study was carried out to identify antibiotic resistance and virulence genes associated with GBS isolated from pregnant women. METHODS: A total of 43 GBS isolates were obtained from 420 vaginal samples collected from HIV positive and negative women who were 13-35 weeks pregnant attending Antenatal Care at Chitungwiza and Harare Central Hospitals in Zimbabwe. Identification tests of GBS isolates was done using standard bacteriological methods and molecular identification testing. Antibiotic susceptibility testing was done using the modified Kirby-Bauer method and E-test strips. The boiling method was used to extract DNA and Polymerase Chain Reaction (PCR) was used to screen for 13 genes. Data was fed into SPSS 24.0. RESULTS: Nine distinct virulence gene profiles were identified and hly-scpB-bca-rib 37.2% (16/43) was common. The virulence genes identified were namely hly 97.8% (42/43), scpB 90.1% (39/43), bca 86.0% (37/43), rib 69.8% (30/43) and bac 11.6% (5/43). High resistance to tetracycline 97.7% (42/43) was reported followed by 72.1% (31/43) cefazolin, 69.8% (30/43) penicillin G, 58.1% (25/43) ampicillin, 55.8% (24/43) clindamycin, 46.5% (20/43) ceftriaxone, 34.9% (15/43) chloramphenicol, and 30.2% (13/43) for both erythromycin and vancomycin using disk diffusion. Antibiotic resistance genes among the resistant and intermediate-resistant isolates showed high frequencies for tetM 97.6% (41/42) and low frequencies for ermB 34.5% (10/29), ermTR 10.3% (3/29), mefA 3.4% (1/29), tetO 2.4% (1/42) and linB 0% (0/35). The atr housekeeping gene yielded 100% (43/43) positive results, whilst the mobile genetic element IS1548 yielded 9.3% (4/43). CONCLUSION: The study showed high prevalence of hly, scpB, bca and rib virulence genes in S. agalactiae strains isolated from pregnant women. Tetracycline resistance was predominantly caused by the tetM gene, whilst macrolide resistance was predominantly due to the presence of erm methylase, with the ermB gene being more prevalent. Multi-drug resistance coupled with the recovery of resistant isolates to antimicrobial agents such as penicillins indicates the importance of GBS surveillance and susceptibility tests. It was also observed that in vitro phenotypic resistance is not always accurately predicted by resistance genotypes.

Human Immunodeficiency Virus Infection Is Associated With Preterm Delivery Independent of Vaginal Microbiota in Pregnant African Women.

BACKGROUND: During pregnancy, the vaginal microbiota is relatively stable. However, African women have more diverse vaginal microbiota than their European counterparts, in addition to high human immunodeficiency virus (HIV) prevalence and risk of adverse birth outcomes. Although HIV is associated with alterations in vaginal microbiota and inflammation in nonpregnant women, these relationships are underexplored in pregnant women. METHODS: In this study, we characterize the vaginal microbiota and immune factors in pregnant African women who were HIV-uninfected (n = 314) versus HIV-infected (n = 42). Mucosal samples were collected once at the enrollment visit (between 15 and 35 weeks of gestation) and women were followed until delivery. RESULTS: Vaginal microbial communities of pregnant women with HIV were significantly more diverse than women without HIV (P = .004), with community structure also differing by HIV status (P = .002, R2 = 0.02). Human immunodeficiency virus infection was also associated with increased risk of preterm birth (PTB) (31% versus 15.3%; P = .066). In a multivariate analysis, HIV infection was independently associated with diverse vaginal community state type (CST)-IVA (P = .005) and CST-IVB (P = .018) as well as PTB (P = .049). No association between HIV status and cytokine concentrations was found. CONCLUSIONS: Longitudinal studies with accurate gestational age assessment would be important to confirm these relationships.

Sepsis in cancer patients residing in Zimbabwe: spectrum of bacterial and fungal aetiologies and their antimicrobial susceptibility patterns.

BACKGROUND: Cancer and sepsis comorbidity is a major public health problem in most parts of the world including Zimbabwe. The microbial aetiologies of sepsis and their antibiograms vary with time and locations. Knowledge on local microbial aetiologies of sepsis and their susceptibility patterns is critical in guiding empirical antimicrobial treatment choices. METHODS: This was a descriptive cross-sectional study which determined the microbial aetiologies of sepsis from blood cultures of paediatric and adult cancer patients obtained between July 2016 and June 2017. The TDR-X120 blood culture system and TDR 300B auto identification machine were used for incubation of blood culture bottles and identification plus antimicrobial susceptibility testing, respectively. RESULTS: A total of 142 participants were enrolled; 50 (35.2%) had positive blood cultures, with 56.0% Gram positive, 42.0% Gram-negative bacteria and 2.0% yeast isolated. Common species isolated included coagulase negative Staphylococcus spp. (CoNS) (22.0%), E. coli (16.0%), K. pneumoniae (14.0%), E. faecalis (14.0%) and S. aureus (8.0%). Gram-negative isolates exhibited high resistance to gentamicin (61.9%) and ceftriaxone (71.4%) which are the empiric antimicrobial agents used in our setting. Amikacin and meropenem showed 85.7 and 95.2% activity respectively against all Gram-negative isolates, whilst vancomycin and linezolid were effective against 96.2 and 100.0% of all Gram-positive isolates respectively. We isolated 10 (66.7%) extended spectrum ß-lactamase (ESBL) amongst the E. coli and K. pneumoniae isolates. Ten (66.7%) of the Staphylococcus spp. were methicillin resistant. CONCLUSIONS: CoNS, E. coli, K. pneumoniae, E. faecalis and S. aureus were the major microbial drivers of sepsis amongst cancer patients in Zimbabwe. Most isolates were found to be resistant to commonly used empirical antibiotics, with isolates exhibiting high levels of ESBL and methicillin resistance carriage. A nationwide survey on microbial aetiologies of sepsis and their susceptibility patterns would assist in the guidance of effective sepsis empiric antimicrobial treatment among patients with cancer.

Prevalence of human immunodeficiency virus, syphilis, hepatitis B and C in blood donations in Namibia.

BACKGROUND: Transfusion Transmissible Infections (TTIs) such as Human Immunodeficiency Virus (HIV), syphilis, hepatitis B virus (HBV) and hepatitis C virus (HCV) are infections which are common in some communities in Southern Africa. It is important to screen blood donations for these infections. METHODS: This is a retrospective study which involved reviewing of previous blood donation records for the year 2012 in Namibia. The records were analyzed to determine the prevalence of HIV, syphilis, Hepatitis B and C among blood donations with regard to gender, age and geographical region of the donors. RESULTS: The findings indicated a significantly low prevalence of HIV, syphilis, HBsAg and anti-Hepatitis C among the blood donations. A low infection rate of 1.3% by any of the four tested TTIs was found among the blood donations given by the donor population in Namibia in 2012. CONCLUSION: The blood donations given by the donor population in Namibia has a low infection rate with the HIV, syphilis, HBsAg and anti-HCV. A strict screening regime must continue to be used as the infections are still present albeit in small numbers.

Coronavirus Disease 2019 (COVID-19) Pandemic across Africa: Current Status of Vaccinations and Implications for the Future.

The introduction of effective vaccines in December 2020 marked a significant step forward in the global response to COVID-19. Given concerns with access, acceptability, and hesitancy across Africa, there is a need to describe the current status of vaccine uptake in the continent. An exploratory study was undertaken to investigate these aspects, current challenges, and lessons learnt across Africa to provide future direction. Senior personnel across 14 African countries completed a self-administered questionnaire, with a descriptive analysis of the data. Vaccine roll-out commenced in March 2021 in most countries. COVID-19 vaccination coverage varied from low in Cameroon and Tanzania and up to 39.85% full coverage in Botswana at the end of 2021; that is, all doses advocated by initial protocols versus the total population, with rates increasing to 58.4% in Botswana by the end of June 2022. The greatest increase in people being fully vaccinated was observed in Uganda (20.4% increase), Botswana (18.5% increase), and Zambia (17.9% increase). Most vaccines were obtained through WHO-COVAX agreements. Initially, vaccination was prioritised for healthcare workers (HCWs), the elderly, adults with co-morbidities, and other at-risk groups, with countries now commencing vaccination among children and administering booster doses. Challenges included irregular supply and considerable hesitancy arising from misinformation fuelled by social media activities. Overall, there was fair to reasonable access to vaccination across countries, enhanced by government initiatives. Vaccine hesitancy must be addressed with context-specific interventions, including proactive programmes among HCWs, medical journalists, and the public.

Group B Streptococcus colonization during pregnancy and maternal-fetal transmission in Zimbabwe.

OBJECTIVE: To explore risk factors for group B Streptococcus (GBS) colonization during pregnancy and at delivery, estimate the predictive value of early GBS colonization for colonization at delivery and in the newborn, and explore the relationship to adverse perinatal factors. DESIGN AND SETTING: Cohort study of pregnant women from three communities in Zimbabwe. METHODS: Information collected by questionnaire at inclusion and from delivery records. Vaginal and rectal swabs collected for GBS culture at 20 and 26 weeks gestation, at delivery and from the newborn infant. MAIN OUTCOME MEASURES: GBS colonization in pregnancy, colonization of mother and newborn, and perinatal factors. RESULTS: GBS culture results were obtained at one or more occasion for 780 (75.2%) of 1,037 women recruited. Altogether, 470/780 women (60.3%) tested positive for GBS, with colonization rates at 20, 26 weeks and delivery of 47%, 24.2% and 21%, respectively. Positive GBS culture at 20 and 26 weeks gestation had a low positive predictive value on colonization at delivery and in the newborn. Women living in rural areas were significantly more often colonized than those who lived in urban areas (p < 0.001). Other socio-economic, demographic and obstetric factors were not statistically associated with GBS colonization. GBS transmission was not statistically significantly associated with adverse perinatal outcomes. CONCLUSIONS: GBS colonization was common among pregnant women in Zimbabwe. Dwelling in a rural area was significantly associated with GBS colonization while other risk factors were not. Early GBS colonization had a low predictive value for colonization at delivery and colonization was not associated with adverse perinatal outcome.

Evaluation of the Xpert® MTB/RIF assay and microscopy for the diagnosis of Mycobacterium tuberculosis in Namibia.

BACKGROUND: Tuberculosis (TB) kills approximately two million people and infects around nine million worldwide annually. Its proper management, especially in resource-limited settings, has been hindered by the lack of rapid and easy-to-use diagnostic tests. Sputum smear microscopy remains the cheapest, readily available diagnostic method but it only identifies less than half of the patients with a HIV/TB co-infection because the bacilli would have disseminated from the lungs to other areas of the body. The fully automated Xpert® MTB/RIF assay is a promising innovation for diagnosing TB and detecting resistance to rifampicin. This study aimed to evaluate the use of Xpert® MTB/RIF assay and microscopy in the diagnosis of Mycobacterium tuberculosis in Namibia, by determining the disease's epidemiology and calculating the proportion of cases infected just with TB and those with a resistance to rifampicin among the total suspected cases of TB in the country. METHODS: This retrospective study analysed TB cases that were diagnosed using both the Xpert® MTB/RIF assay and microscopy. Data were collected from patient records from the Meditech laboratory information system of the Namibia Institute of Pathology for the time period of July 2012-April 2013. Data from 13 regions were collected. RESULTS: The total number of specimens collected from patients with symptoms of pulmonary TB was 1 842. Of these, 594 (32.20%) were found to be positive for MTB by Xpert® MTB/RIF assay, out of which 443 (24.05%) were also found to be positive by microscopy. The remainder was negative. The male patients were more resistant to rifampicin when compared to the female patients. CONCLUSIONS: Tuberculosis is widely distributed throughout Namibia, with slightly more males infected than females. Most TB patients are also co-infected with HIV. Both microscopy and Xpert® MTB/RIF assay are crucial for the diagnosis of TB in the country. Screening diagnostic efforts should focus on the sexually active HIV positive male population who could be the source of more RIF-resistant TB than females to prevent its spread.

Prevalence of bacterial contamination in blood and blood products at the National Blood Service Zimbabwe.

INTRODUCTION: Advances in screening for infections improve the safety of donated blood. Transfusion-related bacterial sepsis, although not established in Zimbabwe, stills makes bacterial contamination of blood clinically relevant. METHODOLOGY: This cross-sectional study was conducted in Harare. Bacteriological and antibiotic susceptibility testing were done using standard methods. RESULTS: Of the 196 samples analyzed, 6 (3.1%) were contaminated with bacteria. Platelets had a significantly high contamination rate compared to other blood products. Bacteria showed varying patterns of susceptibility to the antibiotics tested. CONCLUSIONS: The prevalence of bacterial contamination in blood products suggests that patients who receive blood products are at risk of developing infection.

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.

Novel aspects of the Z and R3 antigens of Streptococcus agalactiae revealed by immunological testing.

Group B streptococci (GBS) are important human and bovine pathogens which can be classified by a variety of phenotype- and gene-based techniques. The capsular polysaccharide and strain-variable, surface-anchored proteins are particularly important phenotypic markers. In an earlier study, a previously unrecognized protein antigen called Z was described. It was expressed by 27.2% of GBS strains from Zimbabwe, usually in combination with R3 protein expression. In this study, a putative Z-specific antiserum actually contained antibodies against two different antigens named Z1 and Z2; Z1 was >250 kDa in molecular mass. Z1, Z2, and R3 generated multiple stained bands on Western blots and showed similar chromatographic characteristics with respect to molecular mass, aggregate formation, and charge. Of 28 reference and prototype GBS strains examined, 8/28 (28.5%) isolates expressed one, two, or all three of the Z1, Z2, and R3 antigens; 4/28 expressed all three antigens; 2/28 expressed Z2 and R3; 1/28 expressed Z1 only; and 1/28 expressed R3 only. Twenty (71.5%) of the 28 isolates expressed none of the three antigens. Expression of one or more of these antigens was shown by isolates of the capsular polysaccharide types Ia, Ib, V, and IX and NT strains and occurred in combination with expression of various other strain-variable and surface-localized protein antigens. When used as serosubtype markers, Z1, Z2, and R3 affected existing GBS serotype designations for some of the isolates. For instance, the R3 reference strain Prague 10/84 (ATCC 49447) changed serotype markers from V/R3 to V/R3, Z1, and Z2. Other isolates may change correspondingly, implying consequences for GBS serotyping and research.

Streptococcus agalactiae alpha-like protein 1 possesses both cross-reacting and Alp1-specific epitopes.

Most isolates of group B streptococci (GBS) express an alpha-like protein (Alp), Cα (encoded by bca), Alp1 (also called epsilon; alp1), Alp2 (alp2), Alp3 (alp3), Alp4 (alp4), or R4/Rib (rib). These proteins are chimeras with a mosaic structure and with antigenic determinants with variable immunological cross-reactivities between the Alps, including Alp1 and Cα cross-reactivity. This study focused on antigenic domains of Alp1, studied by using rabbit antisera in immunofluorescence, Western blotting, and enzyme-linked immunosorbent assay (ELISA)-based tests and whole cells of GBS or trypsin-extracted and partially purified antigens from the strains A909 (serotype Ia/Cα, Cß) and 335 (Ia/Alp1). Alp1 and Cα shared an antigenic determinant, Alp1/Cα common, not harbored by other Alps, probably located in the Alp1 and Cα repeat units, as these units are nearly identical in genomic sequence. An antigenic Alp1 determinant was Alp1 specific and was most likely located in the N-terminal unit of Alp1 in which an Alp1-specific primer site for PCR is also located. In addition, Alp1 possessed a domain with low immunogenicity which cross-reacted immunologically with Alp2 and Alp3, with unknown location in Alp1. Alp1 was partially degraded by trypsin during antigen extraction but with the antigenic domains preserved. The results indicate that Cα and Alp1 are immunologically related in the same manner that R4 (Rib) and Alp3 are related. The domain called Alp1 specific should be important in GBS serotyping as a surface-anchored serosubtype marker. The Alp1/Cα common determinant may be of prime interest as an immunogenic domain in a GBS vaccine.

Putative novel surface-exposed Streptococcus agalactiae protein frequently expressed by the group B streptococcus from Zimbabwe.

Group B streptococci (GBS) express a variety of surface-exposed and strain-variable proteins which function as phenotypic markers and as antigens which are able to induce protective immunity in experimental settings. Among these proteins, the chimeric and immunologically cross-reacting alpha-like proteins are particularly important. Another protein, R3, which has been less well studied, occurred at a frequency of 21.5% in GBS from Zimbabwe and, notably, occurred in serotype V strains at a frequency of 75.9%. Working with rabbit antiserum raised against the R3 reference strain ATCC 49447 (strain 10/84; serotype V/R3) to detect the expression of the R3 protein, we recorded findings which suggested that strain 10/84 expressed a strain-variable protein antigen, in addition to R3. The antigen was detected by various enzyme-linked immunosorbent assay-based tests by using acid extract antigens or GBS whole-cell coats and by whole-cell-based Western blotting. We named the putative novel antigen the Z antigen. The Z antigen was a high-molecular-mass antigen that was susceptible to degradation by pepsin and trypsin but that was resistant to m-periodate oxidation and failed to show immunological cross-reactivity with any of a variety of other GBS protein antigens. The Z antigen was expressed by 33/121 (27.2%) of strains of a Zimbabwean GBS strain collection and by 64.2% and 72.4% of the type Ib and type V strains, respectively, and was occasionally expressed by GBS of other capsular serotypes. Thus, the putative novel GBS protein named Z showed distinct capsular antigen associations and presented as an important phenotypic marker in GBS from Zimbabwe. It may be an important antigen in GBS from larger areas of southern Africa. Its prevalence in GBS from Western countries is not known.

Distinctive features of surface-anchored proteins of Streptococcus agalactiae strains from Zimbabwe revealed by PCR and dot blotting.

The distribution of capsular polysaccharide (CPS) types and subtypes (serovariants) among 121 group B streptococcus (GBS) strains from Zimbabwe was examined. PCR was used for the detection of both CPS types and the surface-anchored and strain-variable proteins Calpha, Cbeta, Alp1, Alp2, Alp3, R4/Rib, and Alp4. The R3 protein was detected by an antibody-based method using monoclonal anti-R3 antibody in dot blotting. The CPS types detected, Ia (15.7% of strains), Ib (11.6%), II (8.3%), III (38.8%), V (24.0%), and nontypeable (1.7%), were essentially as expected on the basis of data from Western countries. The type V strains showed distinctive features with respect to protein markers in that Alp3 was detected in only 6.9% of the isolates while R3 occurred in 75.9% and R4/Rib occurred in 37.9% of the isolates. R3 occurred nearly always in combination with one of the alpha-like (Alp) proteins, and it was the third most common of the proteins studied. These results show that type V GBS strains from Zimbabwe differed from type V strains from other geographical areas and also emphasize the importance of the R3 protein in GBS serotyping and its potential importance in the immunobiology of GBS, including a potential role in a future GBS vaccine.