Browsing by Subject "PCR"

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    Open Access
    Analysis of virulence factors and antibiotic resistance genes in group B streptococcus from clinical samples
    (2021-01-28) Mudzana, Raymond; Mavenyengwa, Rooyen T; Gudza-Mugabe, Muchaneta
    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.
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    Development of a method to assay the microbial population in heap bioleaching operations
    (Elsevier, 2005) Coram-Uliana, Nicolette J; van Hille, Robert P; Kohr, William J; Harrison, Susan T L
    Heap bioleaching is an economically viable approach to the mining of low-grade ores. Oxidation is microbially assisted, involving a consortium of microorganisms that together span the mesophilic to extreme thermophilic range of temperatures (25–80 °C). Temperatures inside the heap are not externally regulated, making the microbial interactions difficult to predict. In order to gain insight into these interactions, a qualitative and quantitative assay of the microorganisms that colonise the ore surface or are present in the liquid phase between the ore clusters at different levels within a heap has been developed. This method was developed using crude ore and liquid samples obtained from the GeoBiotics temperature controlled mesophilic heap operation at the Agnes Gold Mine in Barberton, South Africa, and the high temperature test columns at SGS Lakefield Research, Johannesburg, South Africa. This method of sample analysis may be applied to bioheap leach operations with and without temperature control. Ease of application, reproducibility and turn around time influenced technique design in order to provide a useful assay for commercial bioleaching operations. Following microbial removal from the solid phase using successive washes with detergent and acidified water, the cells were enumerated and genetic DNA was isolated. Microbial identification was achieved via restriction endonuclease analysis of the 16S rRNA genes, as well as 16S rRNA gene sequencing where necessary. Quantification was achieved using species-and genus-specific probes through fluorescent in situ hybridisation (FISH).
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    Open Access
    A Manual of Online Molecular Biology Techniques
    (2014-09-12) Rybicki, Ed
    This resource is a comprehensive manual on practical laboratory and experimental techniques used in molecular and cell biology. This resource is useful for postgraduate students in molecular biology laboratories looking to refine or improve their experimental techniques and the proper use of laboratory equipment.