Papers by Charlotte Mienie
The South African Journal of Plant and Soil, 2000
DNA from six dry bean cultivars was analysed with a total of thirty two primers in RAPD analysis ... more DNA from six dry bean cultivars was analysed with a total of thirty two primers in RAPD analysis to develop a method to distinguish between the different cultivars. The varieties are cultivated in three African countries and belong to two different gene pools. The cultivars analysed were Lyamunga 85 and Lyamunga 90 from Tanzania, CAL 96 and K 20 from Uganda, and Awash I and Mexican 142 from Ethiopia. The two types of beans, the Mesoamerican white seeded and large seeded Andean calima types, were separated in the two genetic pools. RAPD analysis revealed a close relationship between Lyamunga 85 and CAL 96. All the cultivars tested could be distinguished with a minimum of six primers.
International Journal of Food Microbiology, Mar 1, 2016
Soy-daddawa, a fermented soybean (Glycine max (L.) Merr.) condiment, plays a significant role in ... more Soy-daddawa, a fermented soybean (Glycine max (L.) Merr.) condiment, plays a significant role in the culinary practice of West Africa. It is essential to understand the microbial community of soy-daddawa for a successful starter culture application. This study investigated the microbial community structure of soy-daddawa samples collected from Nigerian markets, by PCR-denaturing gradient gel electrophoresis (DGGE) targeting the V3-V5 region of the 16S rRNA gene of bacteria and internal transcribed spacer 2 (ITS2) region of fungi. Six bacterial and 16 fungal (nine yeasts and seven molds) operational taxonomic units (OTUs)/species were obtained at 97% sequence similarity. Taxonomic assignments revealed that bacterial OTUs belonged to the phyla Firmicutes and Actinobacteria, and included species from the genera Atopostipes, Bacillus, Brevibacterium and Nosocomiicoccus. Densitometric analysis of DGGE image/bands revealed that Bacillus spp. were the dominant OTU/species in terms of population numbers. Fungal OTUs belonged to the phyla Ascomycota and Zygomycota, and included species from the genera, Alternaria, Aspergillus, Candida, Cladosporium, Dokmaia, Issatchenkia, Kodamaea, Lecythophora, Phoma, Pichia, Rhizopus, Saccharomyces and Starmerella. The majority of fungal species have not been previously reported in soy-daddawa. Potential opportunistic human pathogens such as Atopostipes suicloacalis, Candida rugosa, Candida tropicalis, and Kodamaea ohmeri were detected. Variation in soy-daddawa microbial communities amongst samples and presence of potential opportunistic pathogens emphasises the need for starter culture employment and good handling practices in soy-daddawa processing.
Genome Announcements, Jun 28, 2018
Salmonella enterica subsp. enterica serovar Typhimurium WG49 is widely used for enumeration of F-... more Salmonella enterica subsp. enterica serovar Typhimurium WG49 is widely used for enumeration of F-specific RNA (F-RNA) coliphages in water. Escherichia coli WG5 is broadly used for the detection and enumeration of somatic coliphages in water samples. We report here the genome sequences of these bacterial strains used in South Africa under ISO methods 10705-1 and 10705-2.
PLOS ONE, 2021
Mammals, born with a near-sterile intestinal tract, are inoculated with their mothers’ microbiome... more Mammals, born with a near-sterile intestinal tract, are inoculated with their mothers’ microbiome during birth. Thereafter, extrinsic and intrinsic factors shape their intestinal microbe assemblage. Wastewater treatment works (WWTW), sites synonymous with pollutants and pathogens, receive influent from domestic, agricultural and industrial sources. The high nutrient content of wastewater supports abundant populations of chironomid midges (Diptera), which transfer these toxicants and potential pathogens to their predators, such as the banana bat Neoromicia nana (Vespertilionidae), thereby influencing their intestinal microbial assemblages. We used next generation sequencing and 16S rRNA gene profiling to identify and compare intestinal bacteria of N. nana at two reference sites and two WWTW sites. We describe the shared intestinal microbiome of the insectivorous bat, N. nana, consisting of seven phyla and eleven classes. Further, multivariate analyses revealed that location was the m...
South African Journal of Science, 2020
The adaptation of Zulu (Nguni) sheep (Ovis aries) to environmental stress and survival under exte... more The adaptation of Zulu (Nguni) sheep (Ovis aries) to environmental stress and survival under extensive conditions makes them uniquely important to rural Nguni farmers of South Africa. Here, the faecal bacterial community of five Zulu sheep populations managed under extensive conditions across summer and winter seasons was investigated in order to understand the influence of prevailing seasonal factors. Bacterial operational taxonomic units (OTUs)/species (at 97% 16S rRNA gene similarity) in Zulu sheep faeces were more diverse in winter than in summer at most (80%) sites and varied between seasons at specific sites. Firmicutes was the most abundant phyla in both summer and winter seasons, while the relative abundance of Actinobacteria reduced in 80% of sites from summer to winter. The genera (or family) such as Akkermansia, Eubacterium coprostanoligenes group, Intestinibacter, R-7 group (family Christensenellaceae), Ruminococcus, Ruminoclostridium, Treponema and UCG-005 (family Rumin...
Tropical Plant Pathology, 2019
Identification of Meloidogyne spp. is challenging when using only the classical techniques since ... more Identification of Meloidogyne spp. is challenging when using only the classical techniques since some species share similar characteristics. In this study, 37 Meloidogyne populations were obtained from four provinces in South Africa during 2015 and 2016 and molecularly characterised. We used SCAR-PCR markers and sequencing of four DNA fragments, viz. the D2-D3 segment (28S rDNA), COI, viable region between COII and 16S rDNA and NADH5 (mtDNA) genes. Using SCAR-PCR, M. enterolobii, M. javanica, M. incognita and M. hapla (in descending order of prevalence) were identified. According to D2-D3, COI, COII/16S and NADH5 sequence analyses, the 23 M. enterolobii populations identified clustered together in a well-supported clade, while M. javanica (9 populations) and M. incognita (5 populations) grouped in another. Results showed that D2-D3 and COI genes were appropriate for accurate identification of M. enterolobii, but not to differentiate among M. arenaria, M. incognita and M. javanica. Conversely, SCAR-PCR and NADH5 were more accurate in distinguishing among different Meloidogyne spp. In conclusion, SCAR-PCR was best for the situation of mixed populations.
South African Journal of Plant and Soil, 2000
Genetic engineering is an important tool which can be used together with conventional breeding me... more Genetic engineering is an important tool which can be used together with conventional breeding methods in transferring certain economically important traits into cultivated varieties of almost any crop species. One of the methods for gene transfer is the particle bombardment technique. Several primary explants can be used with this technique for the transformation of grQundnut. Tl:1i,s study explored the use of mature embryo axes as explants for somatic embryogenesis, and determined the effect of medium composition on four South African genotypes. A modified Murashige and Skoog-medium was used with various concentrations of picloram. Mature embryo axes were found to be a suitable explant for induction of somatic embryogenesis. A significant difference was found in the response of the different genotypes to medium composition.
International Journal of Food Microbiology, 2022
Atypical enteropathogenic E. coli (aEPEC) strains are emerging pathogens responsible for fatal di... more Atypical enteropathogenic E. coli (aEPEC) strains are emerging pathogens responsible for fatal diarrhoea in humans worldwide. The purpose of this study was to investigate genetic diversity, virulence and antimicrobial resistance profiles of aEPEC O177 strains isolated from faeces of cattle reared in intensive and extensive production systems in South Africa. A total of 96 multidrug resistant (MDR) aEPEC O177 isolates were typed using enterobacterial repetitive intergenic consensus (ERIC) and random amplified polymorphism DNA (RAPD) typing. The resistome, virulome and mobilome of two aEPEC O177 isolates were investigated using WGS analysis. The ERIC typing was efficient and reproducible with a discriminatory index of 0.95. RAPD typing had poor reproducibility with satisfactory discriminatory power of 0.859. The dendrograms constructed based on ERIC and RAPD banding patterns produced 9 and 8 clusters, respectively, which indicate genetic variation among E. coli O177 isolates. WGS analysis revealed that CF-154-A and CF-335-B) isolates belonged to the O177 serotype with H7 and H21, respectively. Both isolates harboured several virulome genes such as intimin (eaeA), haemolysin (hlyA and hlyE), translocated iron receptor (tir), Type III secretion system (eprH, gspL and prgH), bssR and bssS. However, genes encoding shiga toxins were not found in either isolate. Antibiotic resistance genes such as ampC, tet, ermB, sul2, strB AcrD, aph(6)-Ic, aph(6)-Ib, aph(3″)-I, ant (3″)-1a AcrA and acrE were found in the E. coli O177 strains. Furthermore, genome annotation results indicated that both isolates carried plasmids, insertion sequences, prophages and cluster of regularly interspaced short palindromic repeats (CRISPR) type I. Based on in silico multi locus typing (MLST) analysis, the two isolates were assigned to different sequence types (CF-154-A, ST-1308 and CF-335-B, ST-58). Whole genome multi locus typing tree showed that our isolates clustered with E. coli O177:H21 (reference), suggesting the close genomic relatedness among the strains. Overall, these findings showed that cattle carry genetically diverse E. coli O177 strains, which harbour a repertoire of virulome, resistome and mobilome genes. This highlights a need for multidrug resistant E. coli O177 strain surveillance in cattle.
KEYWORDS: host plant resistance; legume; molecular markers; plant-parasitic nematodes; root-knot ... more KEYWORDS: host plant resistance; legume; molecular markers; plant-parasitic nematodes; root-knot nematodes
African Journal of Biotechnology, 2013
One of the major shortcomings in cultivated groundnut in South Africa is the short shelf-life of ... more One of the major shortcomings in cultivated groundnut in South Africa is the short shelf-life of the product due to rancidity of the oil rich seeds. Polyunsaturated fatty acids are more susceptible to oxidation than mono-unsaturated residues. Thus, it would be preferable to adjust the oleic acid: linoleic acid ratio to a more favourable one. The high-oleic acid trait in groundnut was reported to be dependent on two homeologous oleoyl-PC desaturase genes, ahFAD2A and ahFAD2B. Breeding of new cultivars with this characteristic can be time-consuming and expensive when doing fatty acid analysis in every generation for selection of the progeny with the highest oleic acid: linoleic acid ratio. Marker-assisted selection was applied to the local groundnut breeding program with the utilization of real-time polymerase chain reaction (PCR). The inheritance of the high oleic trait was followed in the 4th backcross progeny and revealed that all combinations of the two genes were found, except th...
The area planted to soybean in South Africa has increased by 54% since the 2009 growing season, m... more The area planted to soybean in South Africa has increased by 54% since the 2009 growing season, mainly as a result of the increasing demand for protein-rich food and fodder sources. Moreover, the introduction of advanced technology, namely the availability of genetically modified herbicide tolerant soybean cultivars also contributed towards increased soybean production. The omnipresence of plant-parasitic nematodes in local agricultural soils, however, poses a threat to the sustainable expansion and production of soybean and other rotation crops. Meloidogyne incognita and M. javanica are the predominant nematode pests in local soybean production areas and those where other grain-, legume- and/or vegetable crops are grown. The lack of registered nematicides for soybean locally, crop production systems that are conducive to nematode pest build-ups as well as the limited availability of genetic host plant resistance to root-knot nematode pests, complicate their management. Research aim...
igh-density lipoprotein (HDL)-cholesterol is one of the well-established independent risk factors... more igh-density lipoprotein (HDL)-cholesterol is one of the well-established independent risk factors for cardiovascular disease, and an inverse association between circulating HDL-cholesterol levels and the risk of coronary heart disease has been consistently demonstrated in epidemiological studies. 1-5 Because HDL-cholesterol is a routinely measured quantitative trait with substantial heritability, genetic determinants of HDL-cholesterol have been previously studied by a candidate gene approach. 6 Common polymorphisms in CETP, LPL, LIPC, LIPG and APOA1 have been reported to be significantly associated with HDLcholesterol levels in Japanese. 7-11 Editorial p 1016 There have been an increasing number of genome-wide association studies (GWAS) being conducted, followed by replication studies in independent populations. This GWAS approach has led to the identification of previously unrecognized associations between genetic variants and common diseases 12-16 or phenotypic traits. 17,18 In the previously performed genome-wide association analyses of HDL-cholesterol, the most strongly and consistently associated single nucleotide polymorphisms (SNPs) with genome-wide significance, have been reported to be near or in the CETP gene at 16q13. 18-21 The genome-wide screening of 341,518 SNPs in 6,382 white women has found rs3764261 located upstream of CETP to be the most strongly associated SNP (P =1.05×10-41), accounting for 3% of the residual variance in HDL-cholesterol. 19 A recent meta-analysis based on 3 GWAS results of 8,816 white individuals, and the subsequent replication involving 11,569 individuals, have not only confirmed the previously reported associations between HDL-cholesterol and genetic variations in CETP, LPL, LIPC, LIPG and ABCA1 genes but have also identified novel genetic loci for HDL-cholesterol; near MVK-MMAB and GALNT2. 18 While GWAS have been increasingly utilized to identify
Journal of Water and Health, 2020
Until recently, research has focused on Clostridium perfringens in clinical settings without cons... more Until recently, research has focused on Clostridium perfringens in clinical settings without considering environmental isolates. In this study, environmental genomes were used to investigate possible antibiotic resistance and the presence of virulence traits in C. perfringens strains from raw surface water. In silico assembly of three C. perfringens strains, DNA generated almost complete genomes setting their length ranging from 3.4 to 3.6 Mbp with GC content of 28.18%. An average of 3,175 open reading frames was identified, with the majority associated with carbohydrate and protein metabolisms. The genomes harboured several antibiotic resistance genes for glycopeptides, macrolide–lincosamide–streptogramin B, β-lactam, trimethoprim, tetracycline and aminoglycosides and also the presence of several genes encoding for polypeptides and multidrug resistance efflux pumps and 35 virulence genes. Some of these encode for haemolysins, sialidase, hyaluronidase, collagenase, perfringolysin O ...
Microorganisms, 2021
Soybean is among South Africa’s top crops in terms of production figures. Over the past few years... more Soybean is among South Africa’s top crops in terms of production figures. Over the past few years there has been increasingly more damage caused to local soybean by plant-parasitic nematode infections. The presence of Meloidogyne (root-knot nematodes) and Pratylenchus spp. (root lesion nematodes) in soybean fields can cripple the country’s production, however, little is known about the soil microbial communities associated with soybean in relation to different levels of Meloidogyne and Pratylenchus infestations, as well as the interaction(s) between them. Therefore, this study aimed to identify the nematode population assemblages and endemic rhizosphere bacteria associated with soybean using Next Generation Sequencing (NGS). The abundance of bacterial genera that were then identified as being significant using linear discriminant analysis (LDA) Effect Size (LEfSe) was compared to the abundance of the most prevalent plant-parasitic nematode genera found across all sampled sites, viz....
PLOS ONE, 2020
Anthropogenic activities in catchments used for drinking water production largely contaminates so... more Anthropogenic activities in catchments used for drinking water production largely contaminates source waters, and this may impact the quality of the final drinking water product. These contaminants may also affect taxonomic and functional profiles of the bacterial communities in the drinking water. Here, we report an integrated insight into the microbiome and water quality of four water treatment plants (NWC, NWE, WCA and NWG) that supply portable water to communities in South Africa. A new scoring system based on combined significant changes of physicochemical parameters and microbial abundance from raw to treated water was used to evaluate the effectiveness of the treatment plants at water purification. Physicochemical parameters which include total soluble solids, turbidity, pH, nitrites and phosphorus among others, were measured in source, treated, and distributed water. There were general statistically significant (P � 0.05) differences between raw and treated water, demonstrating the effectiveness of the purification process. Illumina sequencing of the 16S rRNA gene was used for taxonomic profiling of the microbial communities and this data was used to infer functional attributes of the communities. Structure and composition of the bacterial communities differed significantly (P < 0.05) among the treatment plants, only NWE and NWG showed no significant differences (P > 0.05), this correlated with the predicted functional profile of the microbial communities obtained from Phylogenetic Investigation of Communities by Reconstruction of Observed States (PICRUSt), as well as the likely pollutants of source water. Bacteroidetes, Chlorobi and Fibrobacteres significantly differed (P < 0.05) between raw and distributed water. PICRUSt inferred a number of pathways involved in the degradation of xenobiotics such as Dichlorodiphenyltrichloroethane, atrazine and polycyclic aromatic hydrocarbons. More worryingly, was the presence of pathways involved in beta-lactam resistance, potential pathogenic Escherichia coli infection, Vibrio cholerae infection, and Shigellosis. Also present in drinking and treated water were OTUs associated with a number of opportunistic pathogens.
Chemosphere, 2021
South African surface waters are subject to various forms of pollution. Recent findings in aquati... more South African surface waters are subject to various forms of pollution. Recent findings in aquatic systems suggest an association exists between yeast diversity, chemical pollutants and land coverage, which are important water quality determinants. Yeast abundance and diversity, as well as antifungal agents in two river systems in South Africa, were investigated and related to the existing land coverage. Yeast abundance and diversity were determined from environmental DNA by quantitative polymerase chain reaction and next-generation sequencing, respectively, of the 26S ribosomal ribonucleic acid (rRNA) gene. Antifungal agents were qualitatively and/or quantitatively detected by ultra-high-pressure liquid chromatography-mass spectrometry. Analyses of 2 031 714 high-quality 26S rRNA sequences yielded 5554 amplicon sequence variants (ASVs)/species. ASV richness and Shannon-Wiener index of diversity reflected the southward flow of the river with higher values observed downstream compared to the upstream. Fluconazole concentrations were quantifiable in only two samples; 178 and 271 ng L-1. Taxonomically, at least 20 yeast species were detected, including the dominant Candida tropicalis, Cryptococcus spp. as well as the lesser dominant Bensingtonia bomiensis, Fereydounia khargensis, Hericium erinaceus, Kondoa changbaiensi, Pseudozyma spp. and Sphacelotheca pamparum. The two dominant species are known opportunistic pathogens which had antifungal resistant traits in previous studies from the same rivers and therefore is a public health threat. The present study provides further evidence that yeasts should be included as part of water quality parameters, especially in developing countries where much of the population are economically disadvantaged, and also immunocompromised due to age and disease.
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Papers by Charlotte Mienie