Microbial Contaminants in Drinking Water Collected from Different Regions of Kuwait

Water plays a major role in maintaining life on earth, but it can also serve as a matrix for pathogenic organisms, posing substantial health threats to humans. Although, outbreaks of diseases attributable to drinking water may not be common in industrialized countries, they still occur and can lead to serious acute, chronic, or sometimes fatal health consequences. The analysis of drinking water samples from different regions of Kuwait was performed in this study for bacterial and viral contaminations. Drinking tap water samples were collected from 15 different locations of the six Kuwait governorates. All samples were analyzed by confocal microscopy for the presence of bacteria. The samples were cultured in vitro to detect cultivable organisms. DNA was isolated from the cultured organisms and the identity of the bacteria was determined by sequencing the bacterial 16S rRNA genes, followed by BLAST analysis in the database of NCBI, USA. RNA was extracted from water samples and analyzed by real-time PCR for the detection of viruses with potential health risks, i.e. Astrovirus, Enterovirus, Norovirus, Rotavirus, and Hepatitis A. Confocal microscopy showed the presence of bacteria in some water samples. The 16S rRNA gene sequencing of culture grown organisms, followed by BLAST analysis, identified the presence of several non-pathogenic bacterial species. However, one sample had Acinetobacter baumannii, which often causes opportunistic infections in immunocompromised people, but none of the studied viruses could be detected in the drinking water samples analyzed. The results indicate that drinking water samples analyzed from various locations in Kuwait are relatively safe for drinking and do not contain many harmful pathogens.

Optimization of Lead Bioremediation by Marine Halomonas sp. ES015 Using Statistical Experimental Methods

Bioremediation technology is now used for treatment instead of traditional metal removal methods. A strain was isolated from Marsa Alam, Red sea, Egypt showed high resistance to high lead concentration and was identified by the 16S rRNA gene sequencing technique as Halomonas sp. ES015. Medium optimization was carried out using Plackett-Burman design, and the most significant factors were yeast extract, casamino acid and inoculums size. The optimized media obtained by the statistical design raised the removal efficiency from 84% to 99% from initial concentration 250 ppm of lead. Moreover, Box-Behnken experimental design was applied to study the relationship between yeast extract concentration, casamino acid concentration and inoculums size. The optimized medium increased removal efficiency to 97% from initial concentration 500 ppm of lead. Immobilized Halomonas sp. ES015 cells on sponge cubes, using optimized medium in loop bioremediation column, showed relatively constant lead removal efficiency when reused six successive cycles over the range of time interval. Also metal removal efficiency was not affected by flow rate changes. Finally, the results of this research refer to the possibility of lead bioremediation by free or immobilized cells of Halomonas sp. ES015. Also, bioremediation can be done in batch cultures and semicontinuous cultures using column technology.

Biodegradation of Malathion by Acinetobacter baumannii Strain AFA Isolated from Domestic Sewage in Egypt

Bacterial strains capable of degradation of malathion from the domestic sewage were isolated by an enrichment culture technique. Three bacterial strains were screened and identified as Acinetobacter baumannii (AFA), Pseudomonas aeruginosa (PS1), and Pseudomonas mendocina (PS2) based on morphological, biochemical identification and 16S rRNA sequence analysis. Acinetobacter baumannii AFA was the most efficient malathion degrading bacterium, so used for further biodegradation study. AFA was able to grow in mineral salt medium (MSM) supplemented with malathion (100 mg/l) as a sole carbon source, and within 14 days, 84% of the initial dose was degraded by the isolate measured by high performance liquid chromatography. Strain AFA could also degrade other organophosphorus compounds including diazinon, chlorpyrifos and fenitrothion. The effect of different culture conditions on the degradation of malathion like inoculum density, other carbon or nitrogen sources, temperature and shaking were examined. Degradation of malathion and bacterial cell growth were accelerated when culture media were supplemented with yeast extract, glucose and citrate. The optimum conditions for malathion degradation by strain AFA were; an inoculum density of 1.5x 10^12CFU/ml at 30°C with shaking. A specific polymerase chain reaction primers were designed manually using multiple sequence alignment of the corresponding carboxylesterase enzymes of Acinetobacter species. Sequencing result of amplified PCR product and phylogenetic analysis showed low degree of homology with the other carboxylesterase enzymes of Acinetobacter strains, so we suggested that this enzyme is a novel esterase enzyme. Isolated bacterial strains may have potential role for use in bioremediation of malathion contaminated.

The Antibacterial and Anticancer Activity of Marine Actinomycete Strain HP411 Isolated in the Northern Coast of Vietnam

Since the marine environmental conditions are extremely different from the other ones, marine actinomycetes might produce novel bioactive compounds. Therefore, actinomycete strains were screened from marine water and sediment samples collected from the coastal areas of Northern Vietnam. Ninety-nine actinomycete strains were obtained on starch-casein agar media by dilution technique, only seven strains, named HP112, HP12, HP411, HPN11, HP 11, HPT13 and HPX12, showed significant antibacterial activity against both gram-positive and gram-negative bacteria (Bacillus subtilis ATCC 6633, Staphylococcus epidemidis ATCC 12228, Escherichia coli ATCC 11105). Further studies were carried out with the most active HP411 strain against Candida albicans ATCC 10231. This strain could grow rapidly on starch casein agar and other media with high salt containing 7-10% NaCl at 28-30oC. Spore-chain of HP411 showed an elongated and circular shape with 10 to 30 spores/chain. Identification of the strain was carried out by employing the taxonomical studies including the 16S rRNA sequence. Based on phylogenetic and phenotypic evidence it is proposed that HP411 to be belongs to species Streptomyces variabilis. The potent of the crude extract of fermentation broth of HP411 that are effective against wide range of pathogens: both grampositive, gram-negative and fungi. Further studies revealed that the crude extract HP411 could obtain the anticancer activity for cancer cell lines: Hep-G2 (liver cancer cell line); RD (cardiac and skeletal muscle letters cell line); FL (membrane of the uterus cancer cell line). However, the actinomycetes from marine ecosystem will be useful for the discovery of new drugs in the future.

Characterization of Novel Atrazine-Degrading Klebsiella sp. isolated from Thai Agricultural Soil

Atrazine, a herbicide widely used in sugarcane and corn production, is a frequently detected groundwater contaminant. An atrazine-degrading bacterium, strain KB02, was obtained from long-term atrazine-treated sugarcane field soils in Kanchanaburi province of Thailand. Strain KB02 had a rod-to-coccus morphological cycle during growth. Sequence analysis of the PCR product indicated that the 16S rRNA gene in strain KB02 was ranging from 97-98% identical to the same region in Klebsiella sp. Based on biochemical, physiological analysis and 16S rDNA sequence analysis of one representative isolate, strain KB02, the isolates belong to the genus Klebsiella in the family Enterobacteriaceae. Interestingly that the various primers for atzA, B and C failed to amplify genomic DNA of strain KB02. Whereas the expected PCR product of atzA, B and C were obtained from the reference strain, Arthrobacter sp. strain KU001.

Fractal Analysis of 16S rRNA Gene Sequences in Archaea Thermophiles

A nucleotide sequence can be expressed as a numerical sequence when each nucleotide is assigned its proton number. A resulting gene numerical sequence can be investigated for its fractal dimension in terms of evolution and chemical properties for comparative studies. We have investigated such nucleotide fluctuation in the 16S rRNA gene of archaea thermophiles. The studied archaea thermophiles were archaeoglobus fulgidus, methanothermobacter thermautotrophicus, methanocaldococcus jannaschii, pyrococcus horikoshii, and thermoplasma acidophilum. The studied five archaea-euryarchaeota thermophiles have fractal dimension values ranging from 1.93 to 1.97. Computer simulation shows that random sequences would have an average of about 2 with a standard deviation about 0.015. The fractal dimension was found to correlate (negative correlation) with the thermophile-s optimal growth temperature with R2 value of 0.90 (N =5). The inclusion of two aracheae-crenarchaeota thermophiles reduces the R2 value to 0.66 (N = 7). Further inclusion of two bacterial thermophiles reduces the R2 value to 0.50 (N =9). The fractal dimension is correlated (positive) to the sequence GC content with an R2 value of 0.89 for the five archaea-euryarchaeota thermophiles (and 0.74 for the entire set of N = 9), although computer simulation shows little correlation. The highest correlation (positive) was found to be between the fractal dimension and di-nucleotide Shannon entropy. However Shannon entropy and sequence GC content were observed to correlate with optimal growth temperature having an R2 of 0.8 (negative), and 0.88 (positive), respectively, for the entire set of 9 thermophiles; thus the correlation lacks species specificity. Together with another correlation study of bacterial radiation dosage with RecA repair gene sequence fractal dimension, it is postulated that fractal dimension analysis is a sensitive tool for studying the relationship between genotype and phenotype among closely related sequences.

Antibiotic Resistance Profile of Bacterial Isolates from Animal Farming Aquatic Environments and Meats in a Peri-Urban Community in South Korea

The increasing usage of antibiotics in the animal farming industry is an emerging worldwide problem contributing to the development of antibiotic resistance. The purpose of this work was to investigate the prevalence and antibiotic resistance profile of bacterial isolates collected from aquatic environments and meats in a peri-urban community in Daejeon, Korea. In an antibacterial susceptibility test, the bacterial isolates showed a high incidence of resistance (~ 26.04 %) to cefazolin, tetracycline, gentamycin, norfloxacin, erythromycin and vancomycin. The results from a test for multiple antibiotic resistance indicated that the isolates were displaying an approximately 5-fold increase in the incidence of multiple antibiotic resistance to combinations of two different antibiotics compared to combinations of three or more antibiotics. Most of the isolates showed multi-antibiotic resistance, and the resistance patterns were similar among the sampling groups. Sequencing data analysis of 16S rRNA showed that most of the resistant isolates appeared to be dominated by the classes Betaproteobacteria and Gammaproteobacteria in the phylum Proteobacteria.