Microbial Diversity Profiles Research Articles

Microbial Diversity Profiling of Polysaccharide (gum)-Producing Bacteria Isolated from a South African Sugarcane Processing Factory.

Polysaccharide (gum)-producing bacteria are responsible for severe economic losses in the sugarcane processing industry. Increased polysaccharide levels in raw sugar are normally an indication that biodeterioration occurred in the cane, soon after harvesting. Once in the sugar processing plant, the cell numbers of gum-producing bacteria escalate and may reach levels difficult to control. We have isolated 430 gum-producing bacteria from sugarcane and different sampling points in a South African sugarcane processing factory. As expected, high cell numbers of gum-producing bacteria were isolated from the factory during a time when sugar with a high dextran content was produced. What we did not expect was to find the same species in the factory at a time when sugar with a low dextran content was produced. Phylogenetic analyses of the 16S rRNA gene sequences differentiated the gum-producing bacteria into four genera and nine species. The majority of these isolates belonged to the genus Weissella (47%), followed by members of Bacillus (24%), Leuconostoc (19%) and Lactobacillus (10%). For the first time, we report on the isolation of Weissella confusa, Weissella cibaria and Bacillus amyloliquefaciens from a sugarcane processing factory.

Variations in Microbial Diversity and Metabolite Profiles of the Tropical Marine Sponge Xestospongia muta with Season and Depth.

Xestospongia muta is among the most emblematic sponge species inhabiting coral reefs of the Caribbean Sea. Besides being the largest sponge species growing in the Caribbean, it is also known to produce secondary metabolites. This study aimed to assess the effect of depth and season on the symbiotic bacterial dynamics and major metabolite profiles of specimens of X. muta thriving in a tropical marine biome (Portobelo Bay, Panamá), which allow us to determine whether variability patterns are similar to those reported for subtropical latitudes. The bacterial assemblages were characterized using Illumina deep-sequencing and metabolomic profiles using UHPLC-DAD-ELSD from five depths (ranging 9-28m) across two seasons (spring and autumn). Diverse symbiotic communities, representing 24 phyla with a predominance of Proteobacteria and Chloroflexi, were found. Although several thousands of OTUs were determined, most of them belong to the rare biosphere and only 23 to a core community. There was a significant difference between the structure of the microbial communities in respect to season (autumn to spring), with a further significant difference between depths only in autumn. This was partially mirrored in the metabolome profile, where the overall metabolite composition did not differ between seasons, but a significant depth gradient was observed in autumn. At the phyla level, Cyanobacteria, Firmicutes, Actinobacteria, and Spirochaete showed a mild-moderate correlation with the metabolome profile. The metabolomic profiles were mainly characterized by known brominated polyunsaturated fatty acids. This work presents findings about the composition and dynamics of the microbial assemblages of X. muta expanding and confirming current knowledge about its remarkable diversity and geographic variability as observed in this tropical marine biome.

Phylogenetic diversity and metabolic potential of microbiome of natural healing clay from Chamliyal (J&K).

Clay therapy for skin disease treatment is an ancient practice popular worldwide as a cheap alternative to pharmaceutical products. Effectiveness of clay against skin problems has been linked to its mineral composition and to microbial activity. The clay-water paste of a holy shrine Chamliyal in the Jammu region of J&K, India is used as an ointment to treat different skin disorders particularly psoriasis. Using the 16 SrDNA amplicon pyrosequencing and whole-metagenome direct shotgun Illumina sequencing, microbial phylogeny and potential metabolic functions were catalogued for Chamliyal's clay. Microbial diversity profile of the Chamliyal's clay is similar to other medicinal clays, particularly Dead Sea; there is some uniqueness as well. Although Proteobacteria, Actinomycetes and Firmicutes are common inhabitants of all the clay types, sulphur- and iron-reducing bacteria like Deferribacterales are particular to clays used for skin healing. In the present study it is proposed that healing properties of clay may be due to the microbes and microbial genes associated with metabolism of minerals like iron and sulphur, that lead to mineral acquisition in the Chamliyal's clay.

Evaluation of dairy processing wastewater biotreatment in an IASBR system: Aeration rate impacts on performance and microbial ecology

Dairy processing generates large volumes of wastewater that require extensive nutrient remediation prior to discharge. Significant commercial opportunities exist therefore for cost-effective biotechnologies capable of achieving this requirement. In this study the authors evaluated the use of intermittently aerated sequencing batch reactors, (IASBRs), as a single-tank biotreatment system for co-removal of COD, nitrogen and phosphorus from synthetic dairy processing wastewater. Variation of the IASBR aeration rates, (0.8, 0.6 and 0.4 L/min), had significant impacts on the respective nutrient removal efficiencies and underlying microbial diversity profiles. Aeration at 0.6 L/min was most effective and resulted in >90% co-removal of orthophosphate and ammonium. 16S rRNA based pyrosequencing of biomass DNA samples revealed the family Comamonadaceae was notably enriched (>80% relative abundance) under these conditions. In silico predictive metabolic modelling also identified Comamonadaceae as the major contributor of several known genes for nitrogen and phosphorus assimilation (nirK, nosZ, norB, ppK, ppX and phbC).

Combined genotyping, microbial diversity and metabolite profiling studies on farmed Mytilus spp. from Kiel Fjord

The blue mussel Mytilus is a popular food source with high economical value. Species of the M. edulis complex (M. edulis, M. galloprovincialis and M. trossulus) hybridise whenever their geographic ranges overlap posing difficulties to species discrimination, which is important for blue mussel aquaculture. The aim of this study was to determine the genetic structure of farmed blue mussels in Kiel Fjord. Microbial and metabolic profile patterns were studied to investigate a possible dependency on the genotype of the bivalves. Genotyping confirmed the complex genetic structure of the Baltic Sea hybrid zone and revealed an unexpected dominance of M. trossulus alleles being in contrast to the predominance of M. edulis alleles described for wild Baltic blue mussels. Culture-dependent and -independent microbial community analyses indicated the presence of a diverse Mytilus-associated microbiota, while an LC-MS/MS-based metabolome study identified 76 major compounds dominated by pigments, alkaloids and polyketides in the whole tissue extracts. Analysis of mussel microbiota and metabolome did not indicate genotypic dependence, but demonstrated high intraspecific variability of farmed mussel individuals. We hypothesise that individual differences in microbial and metabolite patterns may be caused by high individual plasticity and might be enhanced by e.g. nutritional condition, age and gender.

Effects of moderate, voluntary ethanol consumption on the rat and human gut microbiome.

Many alcohol-induced health complications are directly attributable to the toxicity of alcohol or its metabolites, but another potential health impact of alcohol may be on the microbial communities of the human gut. Clear distinctions between healthy and diseased-state gut microbiota have been observed in subjects with metabolic diseases, and recent studies suggest that chronic alcoholism is linked to gut microbiome dysbiosis. Here, we investigated the effects of moderate levels of alcohol consumption on the gut microbiome in both rats and humans. The gut microbiota of rats voluntarily consuming a 20 percent ethanol solution, on alternate days, were compared with a non-exposed control group to identify differential taxonomic and functional profiles. Gut microbial diversity profiles were determined using culture-independent amplification, next-generation sequencing and bioinformatic analysis of bacterial 16S ribosomal RNA gene sequence libraries. Our results showed that, compared with controls, ethanol-consuming rats experienced a significant decline in the biodiversity of their gut microbiomes, a state generally associated with dysbiosis. We also observed significant shifts in the overall diversity of the gut microbial communities and a dramatic change in the relative abundance of particular microbes, such as the Lactobacilli. We also compared our results to human fecal microbiome data collected as part of the citizen science American Gut Project. In contrast to the rat data, human drinkers had significantly higher gut microbial biodiversity than non-drinkers. However, we also observed that microbes that differed among the human subjects displayed similar trends in the rat model, including bacteria implicated in metabolic disease.

Effects of Moderate Voluntary Ethanol Consumption on the Rat and Human Gut Microbiome

Many alcohol‐induced health complications are directly attributable to the toxicity of alcohol or its metabolites, but another potential health impact of alcohol may be on the microbial communities of the human gut. Clear distinctions between healthy and diseased‐state gut microbiota have been observed in subjects with metabolic diseases and recent studies suggest that chronic alcoholism is linked to gut microbiome dysbiosis. Here, we investigated the effects of alcohol consumption on the gut microbiome in both rats and humans. The gut microbiota of rats voluntarily consuming a 20% ethanol solution, on alternate days, were compared to a non‐exposed control group to identify differential taxonomic and functional profiles. Gut microbial diversity profiles were determined using culture‐independent amplification, next‐generation sequencing and bioinformatic analysis of bacterial 16S ribosomal RNA gene sequence libraries. Our results showed that, compared to controls, ethanol‐consuming rats experienced a significant decline in the biodiversity of their gut microbiomes, a state generally associated with dysbiosis. We also observed significant shifts in the overall diversity of the gut microbial communities and a dramatic change in the relative abundance of particular microbes, such as the Lactobacilli. We also compared our results to human fecal microbiome data collected as part of the citizen science American Gut Project. In contrast to the rat data, human drinkers had significantly higher gut microbial biodiversity than non‐drinkers. However, we also observed that microbes that differed among the human subjects displayed similar trends in the rat model, including the Lactobacilli as well as bacteria implicated in metabolic disease.Support or Funding InformationNIH AA023291This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Gut Microbial Diversity in Women With Polycystic Ovary Syndrome Correlates With Hyperandrogenism.

A majority of women with polycystic ovary syndrome (PCOS) have metabolic abnormalities that result in an increased risk of developing type 2 diabetes and heart disease. Correlative studies have shown an association between changes in the gut microbiome and metabolic disorders. Two recent studies reported a decrease in α diversity of the gut microbiome in women with PCOS compared with healthy women. We investigated whether changes in the gut microbiome correlated with specific clinical parameters in women with PCOS compared with healthy women. We also investigated whether there were changes in the gut microbiome in women with polycystic ovarian morphology (PCOM) who lacked the other diagnostic criteria of PCOS. Subjects were recruited at the Poznan University of Medical Sciences. Fecal microbial diversity profiles of healthy women (n = 48), women with PCOM (n = 42), and women diagnosed with PCOS using the Rotterdam criteria (n = 73) were analyzed using 16S ribosomal RNA gene sequencing. Lower α diversity was observed in women with PCOS compared with healthy women. Women with PCOM had a change in α diversity that was intermediate between that of the other two groups. Regression analyses showed that hyperandrogenism, total testosterone, and hirsutism were negatively correlated with α diversity. Permutational multivariate analysis of variance in UniFrac distances showed that hyperandrogenism was also correlated with β diversity. A random forest identified bacteria that discriminated between healthy women and women with PCOS. These results suggest that hyperandrogenism may play a critical role in altering the gut microbiome in women with PCOS.

Adaptation of Methanogenic Inocula to Anaerobic Digestion of Maize Silage.

A well-balanced microbial consortium is crucial for efficient biogas production. In turn, one of a major factor that influence on the structure of anaerobic digestion (AD) consortium is a source of microorganisms which are used as an inoculum. This study evaluated the influence of inoculum sources (with various origin) on adaptation of a biogas community and the efficiency of the biomethanization of maize silage. As initial inocula for AD of maize silage the samples from: (i) an agricultural biogas plant (ABP) which utilizes maize silage as a main substrate, (ii) cattle slurry (CS), which contain elevated levels of lignocelluloses materials, and (iii) raw sewage sludge (RSS) with low content of plant origin materials were used. The adaptation of methanogenic consortia was monitored during a series of passages, and the functionality of the adapted consortia was verified through start-up operation of AD in two-stage reactors. During the first stages of the adaptation phase, methanogenic consortia occurred very slowly, and only after several passages did the microbial community adapts to allow production of biogas with high methane content. The ABP consortium revealed highest biogas production in the adaptation and in the start-up process. The biodiversity dynamics monitored during adaptation and start-up process showed that community profile changed in a similar direction in three studied consortia. Native communities were very distinct to each other, while at the end of the Phase II of the start-up process microbial diversity profile was similar in all consortia. All adopted bacterial communities were dominated by representatives of Porphyromonadaceae, Rikenellaceae, Ruminococcaceae, and Synergistaceae. A shift from low acetate-preferring acetoclastic Methanosaetaceae (ABP and RSS) and/or hydrogenotrophic Archaea, e.g., Methanomicrobiaceae (CS) prevailing in the inoculum samples to larger populations of high acetate-preferring acetoclastic Methanosarcinaceae was observed by the end of the experiment. As a result, three independent, functional communities that syntrophically produced methane from acetate (primarily) and H2/CO2, methanol and methylamines were adapted. This study provides new insights into the specific process by which different inocula sampled from typical methanogenic environments that are commonly used to initiate industrial installations gradually adapted to allow biogas production from maize silage.

Dramatic Differences in Gut Bacterial Densities Correlate with Diet and Habitat in Rainforest Ants.

Abundance is a key parameter in microbial ecology, and important to estimates of potential metabolite flux, impacts of dispersal, and sensitivity of samples to technical biases such as laboratory contamination. However, modern amplicon-based sequencing techniques by themselves typically provide no information about the absolute abundance of microbes. Here, we use fluorescence microscopy and quantitative polymerase chain reaction as independent estimates of microbial abundance to test the hypothesis that microbial symbionts have enabled ants to dominate tropical rainforest canopies by facilitating herbivorous diets, and compare these methods to microbial diversity profiles from 16S rRNA amplicon sequencing. Through a systematic survey of ants from a lowland tropical forest, we show that the density of gut microbiota varies across several orders of magnitude among ant lineages, with median individuals from many genera only marginally above detection limits. Supporting the hypothesis that microbial symbiosis is important to dominance in the canopy, we find that the abundance of gut bacteria is positively correlated with stable isotope proxies of herbivory among canopy-dwelling ants, but not among ground-dwelling ants. Notably, these broad findings are much more evident in the quantitative data than in the 16S rRNA sequencing data. Our results provide quantitative context to the potential role of bacteria in facilitating the ants' dominance of the tropical rainforest canopy, and have broad implications for the interpretation of sequence-based surveys of microbial diversity.

Microbial diversity and biochemical profile of aguamiel collected from Agave salmiana and A. atrovirens during different seasons of year.

Aguamiel is a beverage produced by some Agave species that is consumed in its fresh or fermented form. Despite its uses and popularity, seasonal effects on its microbial and chemical profiles are unknown. In this study, using aguamiel collected from A. salmiana and A. atrovirens during different seasons, we identified microorganisms by sequencing the 16S and 18S rDNA genes and determined their chemical profiles. In total, 49 microbial strains were identified (38 bacteria and 11 yeasts). The highest richness and biodiversity were observed during winter and summer. Different lactic acid bacteria and yeast genera with potential industrial applications were identified, such as Acetobacter, Lactobacillus, Leuconostoc, and Clavispora. The analysis of the chemical profiles indicated the presence of maltooligosaccharides and fructooligosaccharides, which are associated with human health improvements, during spring in Agave aguamiel. Aguamiel can be used in the food industry due to its microbiological and chemical profiles.

Impact of environmental conditions and process modifications on microbial diversity, fermentation efficiency and chemical profile during the fermentation of Mezcal in Oaxaca

Mezcal, a distilled alcoholic beverage derived from cooked agave plants, is manufactured traditionally on a small scale where raw materials and production practices differ noticeably between production regions, localities and even factories resulting in a set of highly distinctive products. The present study aimed to draw a comprehensive picture of the mezcal production in Oaxaca State, relating the microbial consortia with the chemical compounds detected during mezcal fermentation, and to elucidate the impact of process modifications made in order to improve fermentation performance. In two mezcal distilleries, fermentation processes were monitored in two consecutive years. Isolated yeasts (553) and bacteria (527) were identified combining traditional and molecular techniques, detecting up to 21 different yeast species and 27 different bacterial species during a single process. Fermentation kinetics and volatile compound generation varied strongly between the studied processes. A highly diverse microbiota associated to these traditional alcoholic fermentations was detected wherein several yeast species and numerous bacterial groups built up the microbial consortia. Modifications introduced between two production seasons showed a clear impact on microbial diversity. The employment of wild inoculums increased alcoholic fermentation efficiency, nevertheless, prominent changes in the volatile compound profile should be taken into account.

Metagenomic Analysis of a Complex Community Present in Pond Sediment.

The metagenomic profiling of complex communities is gaining immense interest across the scientific community. A complex community present in the pond sediment of a water body located close to a hexachlorocyclohexane (HCH) production site of the Indian Pesticide Limited (IPL) (Chinhat, Lucknow) was selected in an attempt to identify and analyze the unique microbial diversity and functional profile of the site. In this study, we supplement the metagenomic study of pond sediment with a variety of binning approaches along with an in depth functional analysis. Our results improve the understanding of ecology, in terms of community dynamics. The findings are crucial with respect to the mechanisms such as those involving the lin group of genes that are known to be implicated in the HCH degradation pathway or the Type VI secretory system (T6SS) and its effector molecules. Metagenomic studies using the comparative genomics approach involving the isolates from adjacent HCH contaminated soils have contributed significantly towards improving our understanding of unexplored concepts, while simultaneously uncovering the novel mechanisms of microbial ecology.

Distinctive Feature of Microbial Communities and Bacterial Functional Profiles in Tricholoma matsutake Dominant Soil.

Tricholoma matsutake, the pine mushroom, is a valuable forest product with high economic value in Asia, and plays an important ecological role as an ectomycorrhizal fungus. Around the host tree, T. matsutake hyphae generate a distinctive soil aggregating environment called a fairy ring, where fruiting bodies form. Because T. matsutake hyphae dominate the soil near the fairy ring, this species has the potential to influence the microbial community. To explore the influence of T. matsutake on the microbial communities, we compared the microbial community and predicted bacterial function between two different soil types—T. matsutake dominant and T. matsutake minor. DNA sequence analyses showed that fungal and bacterial diversity were lower in the T. matsutake dominant soil compared to T. matsutake minor soil. Some microbial taxa were significantly more common in the T. matsutake dominant soil across geographic locations, many of which were previously identified as mycophillic or mycorrhiza helper bacteria. Between the two soil types, the predicted bacterial functional profiles (using PICRUSt) had significantly distinct KEGG modules. Modules for amino acid uptake, carbohydrate metabolism, and the type III secretion system were higher in the T. matsutake dominant soil than in the T. matsutake minor soil. Overall, similar microbial diversity, community structure, and bacterial functional profiles of the T. matsutake dominant soil across geographic locations suggest that T. matsutake may generate a dominance effect.

Comparative evaluation of microbial diversity and metabolite profiles in doenjang, a fermented soybean paste, during the two different industrial manufacturing processes.

Two different doenjang manufacturing processes, the industrial process (IP) and the modified industrial process (mIP) with specific microbial assortments, were subjected to metabolite profiling using liquid chromatography-mass spectrometry (LC-MS) and gas chromatography time-of-flight mass spectrometry (GC-TOF-MS). The multivariate analyses indicated that both primary and secondary metabolites exhibited distinct patterns according to the fermentation processes (IP and mIP). Microbial community analysis for doenjang using denaturing gradient gel electrophoresis (DGGE), exhibited that both bacteria and fungi contributed proportionally for each step in the process viz., soybean, steaming, drying, meju fermentation, cooling, brining, and aging. Further, correlation analysis indicated that Aspergillus population was linked to sugar metabolism, Bacillus spp. with that of fatty acids, whereas Tetragenococcus and Zygosaccharomyces were found associated with amino acids. These results suggest that the components and quality of doenjang are critically influenced by the microbial assortments in each process.

A preliminary insight of correlation between human fecal microbial diversity and blood lipid profile

The study aimed to evaluate the effect of human gut-derived lactic acid bacteria and yeast on cholesterol levels. Fecal samples from five healthy volunteers were examined for the level and diversity of dominant microbiota. Pichia kudriavzevii (QAUPK01, QAUPK02, QAUPK03, QAUPK04 and QAUPK05) and Candida tropicalis (QAUCT06) were identified by phenotypic methods and DNA sequencing and tested for in vitro cholesterol assimilation ability. Significant correlations (p < 0.05) between fecal microbial diversity, volunteers’ age, body mass index (BMI) and serum lipid profile were established. From biochemical tests, eight strains of lactic acid bacteria (M1.1, M1.2, M2.1, M3.1, M3.2, M4.1, M5.1 and M5.2) were identified but no bsh activity was found in them. However, all yeast strains were able to assimilate cholesterol and maximum assimilation ability was shown by QAUPK03 (83.6%) and QAUPK05 (85.2%) after 72 h of growth at 37 °C.

Effect of bio-fertilizers application on microbial diversity and physiological profiling of microorganisms in arable soil

In laboratory assay, the diversity of bacteria and microscopic fungi and the community-level physiological profiling (CLPP) of microorganisms were observed after the addition of bio-sludge (40 t.ha -1 ) from a biogas station and addition of bio-fertlizers - AZOTER (10 dm -3 .ha -1 ) to the arable soil with PCR-DGGE and BIOLOG ® method (Eco Plates). The differences were recorded in the microbial diversity (bacteria and microscopic fungi) among variants according to the Shannon index. The differences in community of microscopic fungi were markedly higher among the soil samples with the additions of both bio-fertilizers compared to control soil samples. The occurrence of individual OTUs (operational taxonomic units) bacteria and microscopic fungi were different after 105 days of incubation from the status after the 1 st day of incubation. The community metabolic diversity (CMD) was influenced by the incubation time (105 days) as well, but not by application of bio-fertilizers. We observed a significant decrease (LSD test, P <0.05) in community metabolic diversity (CMD) and average metabolic response (AMR) of microorganisms in samples collected on the 105 th day of the experiment compared to samples collected on the 1 st day of the experiment in all tested samples.

Dynamics of microbial diversity profiles in waters of different qualities. Approximation to an ecological quality indicator

Over the past two decades, the amount of reclaimed water has increased throughout the world to face the current water shortage, and as a consequence there is an increasing interest to develop good indicators of water quality, beyond the traditional fecal indicators. In order to meet this need, in this work the microbial profiles of different wastewater treatment plant effluents, both secondary and tertiary, were studied and compared with water samples from an uncontaminated natural aquifer. Taking into account the most abundant phylogenetic groups found in these water samples, we calculated the Bacteroidetes, Gammaproteobacteria and Nitrospira/Betaproteobacteria (BGN:β) ratio and found significant differences between the mean ratios of the four water qualities. The secondary effluent ratios were never below 1.3 and the tertiary effluent and groundwater ratios were never over 0.85. Furthermore, calculation of this index with previous published data supports our results and indicates that the BGN:β ratio is a possible alternative indicator of water quality.

Geomicrobiological investigations in subsaline maar lake sediments over the last 1500 years

Living microorganisms inhabit every environment of the biosphere but only in the last decades their importance governing biochemical cycles in deep sediments has been widely recognized. Most investigations have been accomplished in the marine realm whereas there is a clear paucity of comparable studies in lacustrine sediments. One of the main challenges is to define geomicrobiological proxies that can be used to identify different microbial signals in the sediments. Laguna Potrok Aike, a maar lake located in Southeastern Patagonia, has an annually not stratifying cold water column with temperatures ranging between 4 and 10 °C, and most probably an anoxic water/sediment interface. These unusual features make it a peculiar and interesting site for geomicrobiological studies. Living microbial activity within the sediments was inspected by the first time in a sedimentary core retrieved during an ICDP-sponsored drilling operation. The main goals to study this cold subsaline environment were to characterize the living microbial consortium; to detect early diagenetic signals triggered by active microbes; and to investigate plausible links between climate and microbial populations. Results from a meter long gravity core suggest that microbial activity in lacustrine sediments can be sustained deeper than previously thought due to their adaptation to both changing temperature and oxygen availability. A multi-proxy study of the same core allowed defining past water column conditions and further microbial reworking of the organic fraction within the sediments. Methane content shows a gradual increase with depth as a result of the fermentation of methylated substrates, first methanogenic pathway to take place in the shallow subsurface of freshwater and subsaline environments. Statistical analyses of DGGE microbial diversity profiles indicate four clusters for Bacteria reflecting layered communities linked to the oxidant type whereas three clusters characterize Archaea communities that can be linked to both denitrifiers and methanogens. Independent sedimentary and biological proxies suggest that organic matter production and/or preservation have been lower during the Medieval Climate Anomaly (MCA) coinciding with a low microbial colonization of the sediments. Conversely, a reversed trend with higher organic matter content and substantial microbial activity characterizes the sediments deposited during the Little Ice Age (LIA). Thus, the initial sediments deposited during distinctive time intervals under contrasting environmental conditions have to be taken into account to understand their impact on the development of microbial communities throughout the sediments and their further imprint on early diagenetic signals.

Microbial quality, diversity and antibiotic susceptibility profiles of bacterial isolates from borehole water used by schools in Greater Giyani Municipality, Mopani District, South Africa

In the present study, the microbial quality of several boreholes, used by rural schools in Greater Giyani Municipality, was assessed over a six months period to determine their safety for human consumption and to highlight the potential occurrence of water-borne diseases. The microbiological quality of the water sources was performed using the membrane filtration technique, while standard culture methods were used for bacteria isolation. The MicroScan was used for identification and antibiotic susceptibility profiles of isolated potentially pathogenic bacteria. The results obtained indicated that the water quality of the boreholes was poor over the study period (June to October, 2009). Indicator organisms were higher than the acceptable maximum limits prescribed by the South African Department of Water Affairs and Forestry (DWAF) and the World Health Organisation. Numerous organisms that are potential enteric pathogens such as Vibriospp., Shigella species and Klebsiella spp, were isolated throughout the study period. There was high resistance to many antibiotics particularly ampicillin (92%), cefazolin (95.9%), cefuroxime (91.7%) and cefoxitin (92.6%). The most active antibiotics were tobramycin with resistance level of 8.7% and levofloxacin (23.0%). The study indicates that water from the studied boreholes was not suitable for human consumption and may pose a serious threat to the health of consumers and therefore calls for urgent intervention. High level of antibiotic resistance is of concern in the management of infections caused by these organisms. Further studies are needed to identify the sources of contamination in order to curb the negative effect of contaminated water particularly in children. Key words: Antibiotic resistance, bacterial indicators, children, water quality, borehole, schools, South Africa.