Kasing Apun

Molecular characterization of Vibrio cholerae O1 outbreak strains in Miri, Sarawak (Malaysia)

Bacterial resistance to various antimicrobial agents is common in area with high usage of antibiotics. In this study, the data on antimicrobial susceptibility patterns of Vibrio cholerae O1 from patients during an outbreak period was found to be high but variable rates of multidrug resistance. Thirty-two of 33 V. cholerae isolates harboured the tcp, ctx, zot and ace genes, suggesting their possible roles in the outbreak cases. We analyzed the molecular diversity of a total of 33 strains of V. cholerae O1 isolated from 33 patients between November 1997 and April 1998 using random amplified polymorphic DNA (RAPD) analysis. The 30 typable isolates could be separated into four major clusters containing 5, 17, 2 and 6 isolates, respectively. However, no particular RAPD pattern was predictive of a particular pattern of antibiotic susceptibility. The findings of this study showed that multiple clones seemed to be responsible for cases in the outbreaks in the study area.

Quantifying Escherichia coli Release from Soil under High-Intensity Rainfall

Bacterial loading in surface runoff can only be reasonably assessed or predicted with quantitative knowledge of the release of bacteria from the soil under different rainfall conditions. Most studies of bacterial movement were conducted under rainfall intensities of less than 44 mm h-1. However, in the tropics, intensities higher than 44 mm h-1 are frequent. In this study, Escherichia coli release from the soil into surface runoff and its distribution in the soil under the impact of heavy rainfall (95 mm h-1) of different durations were investigated. Results of simulated heavy rainfall of different durations on gently sloping grass plots with spray-applied E. coli indicated that E. coli was released with relative ease, resulting in contaminated runoff. Runoff E. coli concentrations ranged from 2.09 log(CFU) mL-1 in 5 min simulated rainfall events to 4.45 log(CFU) mL-1 in 15 min simulated rainfall events. The first simulated rainfall events after spray applications produced the highest concentration of E. coli in the runoff. Runoff loss accounted for 0.001% of the total applied E. coli in 5 min rainfall events and 2.1% in 15 min rainfall events. Total solids explained 28% of the variation in the concentrations and 14% of the total loadings. E. coli concentration in the surface centimeter of the soil explained 80% to 89% of the variations in runoff concentrations and loadings with regression slope of less than unity. Such quantitative relationships have the potential to predict runoff E. coli concentrations under high-intensity rainfall events.

Diversity of Leptospira spp. in Rats and Environment from Urban Areas of Sarawak, Malaysia

Various prevalence studies on Leptospira in animals and humans, as well as environmental samples, had been conducted worldwide, including Malaysia. However, limited studies have been documented on the presence of pathogenic, intermediate, and saprophytic Leptospira in selected animals and environments. This study was therefore conducted to detect Leptospira spp. in rats, soil, and water from urban areas of Sarawak using the polymerase chain reaction (PCR) method. A total of 107 rats, 292 soil samples, and 324 water samples were collected from April 2014 to February 2015. Pathogenic Leptospira was present in 5.6% (6/107) of rats, 11.6% (34/292) of soil samples, and 1.9% (6/324) of water samples. Intermediate Leptospira was present in 2.7% (8/292) of soil samples and 1.9% (6/324) of water samples. Saprophytic Leptospira was present in 10.3% (11/107) of rats, 1.4% (4/292) of soil samples, and 0.3% (1/324) of water samples. From this study, 76 Leptospira spp. were isolated. Based on DNA sequencing, the dominant Leptospira spp. circulating in urban areas of Sarawak are pathogenic Leptospira noguchii, intermediate Leptospira wolffii serovar Khorat, and saprophytic Leptospira meyeri, respectively. Overall, this study provided important surveillance data on the prevalence of Leptospira spp. from rats and the environment, with dominant local serovars in urban areas of Sarawak.

Detection of Cryptosporidium and Cyclospora Oocysts from Environmental Water for Drinking and Recreational Activities in Sarawak, Malaysia

Cryptosporidiosis and cyclosporiasis are caused by waterborne coccidian protozoan parasites of the genera Cryptosporidium and Cyclospora, respectively. This study was conducted to detect Cryptosporidium and Cyclospora oocysts from environmental water abstracted by drinking water treatment plants and recreational activities in Sarawak, Malaysia. Water samples (12 each) were collected from Sungai Sarawak Kanan in Bau and Sungai Sarawak Kiri in Batu Kitang, respectively. In addition, 6 water samples each were collected from Ranchan Recreational Park and UNIMAS Lake at Universiti Malaysia Sarawak, Kota Samarahan, respectively. Water physicochemical parameters were also recorded. All samples were concentrated by the iron sulfate flocculation method followed by the sucrose floatation technique. Cryptosporidium and Cyclospora were detected by modified Ziehl-Neelsen technique. Correlation of the parasites distribution with water physicochemical parameters was analysed using bivariate Pearson correlation. Based on the 24 total samples of environmental water abstracted by drinking water treatment plants, all the samples (24/24; 100%) were positive with Cryptosporidium, and only 2 samples (2/24; 8.33%) were positive with Cyclospora. Based on the 12 total samples of water for recreational activities, 4 samples (4/12; 33%) were positive with Cryptosporidium, while 2 samples (2/12; 17%) were positive with Cyclospora. Cryptosporidium oocysts were negatively correlated with dissolved oxygen (DO).

High Occurrence of Staphylococcus aureus Isolated from Fitness Equipment from Selected Gymnasiums

Introduction Staphylococcus aureus is a leading cause of cutaneous bacterial infection involving community. Methods In this study, a total of 42 swab samples were collected from the surface of various fitness equipment such as back machines, exercise mats, dip stations, dumbbells, and treadmills. Identification of the bacterial isolates was conducted using biochemical tests and further analysed molecularly using the PCR method targeting nuc gene (270 bp). The nuc gene encodes for the thermonuclease enzyme, a virulent factor of S. aureus. Results The findings showed 31 out of 42 swab samples (73.81%) were positive with S. aureus. Conclusion This study showed that gymnasium equipment is a potential reservoir for S. aureus and might play an important role in transmitting the pathogen to humans. Objective This study was undertaken to assess the presence of S. aureus on the surface of fitness equipment from selected gymnasiums in Kuching and Kota Samarahan, Sarawak (Malaysia).

Evaluation of BOX-PCR and ERIC-PCR as Molecular Typing Tools for Pathogenic Leptospira

In the last decades, leptospirosis had gained public health concern due to morbidity and mortality rates caused by pathogenic Leptospira. The need for rapid and robust molecular typing methods to differentiate this zoonotic pathogen is of utmost importance. Various studies had been conducted to determine the genetic relatedness of Leptospira isolates using molecular typing methods. In this study, 29 pathogenic Leptospira isolates from rat, soil, and water samples in Sarawak, Malaysia, were characterized using BOX-PCR and ERIC-PCR. The effectiveness of these two methods with regard to the ease of interpretation, reproducibility, typeability, and discriminatory power was also being evaluated. Using BOX-PCR, six clusters and 3 single isolates were defined at a genetic distance percentage of 11.2%. ERIC-PCR clustered the isolates into 6 clusters and 2 single isolates at a genetic distance percentage of 6.8%. Both BOX-PCR and ERIC-PCR produced comparable results though the discriminatory index for ERIC-PCR (0.826) was higher than that for BOX-PCR (0.809). From the constructed dendrogram, it could be summarized that the isolates in this study were highly heterogeneous and genetically diverse. The findings from this study indicated that there is no genetic relatedness among the pathogenic Leptospira isolates in relation to the locality, source, and identity, with some exceptions. Out of the 29 pathogenic Leptospira isolates studied, BOX-PCR and ERIC-PCR successfully discriminated 4 isolates (2 isolates each) into the same cluster in relation to sample sources, as well as 2 isolates into the same cluster in association with the sample locality. Future studies shall incorporate the use of other molecular typing methods to make a more thorough comparison on the genetic relatedness of pathogenic Leptospira.

Prevalence, Genetic Heterogeneity, and Antibiotic Resistance Profile of Listeria spp. and Listeria monocytogenes at Farm Level: A Highlight of ERIC- and BOX-PCR to Reveal Genetic Diversity

This study aimed to identify Listeria spp. and L. monocytogenes, characterize the isolates, and determine the antibiotic resistance profiles of the isolates Listeria spp. and L. monocytogenes in fresh produce, fertilizer, and environmental samples from vegetable farms (organic and conventional farms). A total of 386 samples (vegetables, soil, water, and fertilizer with manure) were examined. The identification of bacterial isolates was performed using PCR and characterized using ERIC-PCR and BOX-PCR. The discriminating power of the typing method was analyzed using Simpsons Index of Diversity. Thirty-four (n=34) Listeria isolates were subjected to antimicrobial susceptibility test using the disc-diffusion technique. The PCR analysis revealed that Listeria spp. were present in 7.51% (29/386) of all the samples (vegetable, soil, fertilizer, and water). None of the samples examined were positive for the presence of L. monocytogenes. Percentages of 100% (15/15) and 73.30% (11/15) of the Listeria spp. isolated from vegetables, fertilizer, and soil from organic farm B had indistinguishable DNA fingerprints by using ERIC-PCR and BOX-PCR, respectively. Listeria spp. isolated from 86 samples of vegetable, fertilizer, and environment of organic farm A and conventional farm C had distinct DNA fingerprints. Simpsons Index of Diversity, D, of ERIC-PCR and BOX-PCR is 0.604 and 0.888, respectively. Antibiotic susceptibility test revealed that most of the Listeria spp. in this study were found to be resistant to ampicillin, rifampin, penicillin G, tetracycline, clindamycin, cephalothin, and ceftriaxone. The isolates had MAR index ranging between 0.31 and 0.85. In conclusion, hygienic measures at farm level are crucial to the reduction of Listeria transmission along the food chain.

Assessment of Listeria monocytogenes in pet food

BackgroundListeria monocytogenes is one of the commonly isolated foodborne pathogens which cause illness, and listeriosis is a disease caused by this pathogen in human beings. Pets that consume contaminated pet food diets can be colonized by L. monocytogenes without showing clinical signs making the pets a possible source of contamination in the household. This study aimed to detect and enumerate the presence of L. monocytogenes in pet food diets, namely cat and dog food.ResultA total of 32 samples consisting of wet food (25%), dry food (25%), treats (25%), and leftover household samples (25%) were examined for this study. The pet food diets were sampled from pet food shops, grocery stores, and households located in Kuching and Kota Samarahan. The analysis was conducted using the most probable number–polymerase chain reaction (MPN-PCR). According to the results obtained from MPN-PCR, none of the samples were contaminated by L. monocytogenes.ConclusionBeing the first biosafety assessment of L. monocytogenes in pet food in Malaysia, this study can contribute to the building of a database regarding the potential contamination of pet food diets by L. monocytogenes.

Enumeration and molecular detection of Bacillus cereus in local indigenous and imported rice grains

BackgroundBacillus cereus is frequently related to foodborne illness outbreak. The common food vehicles for transmission of the bacteria are rice, rice products and starchy foods. As rice is a staple food for some countries including Malaysia, knowledge about safety of B. cereus in rice is important. This study was conducted to enumerate and identify B. cereus in local indigenous and imported rice grains. As Malaysia depends on imported rice to complement the food demands, it is crucial to assess on the imported rice besides the locals.ResultsTwenty local indigenous and twenty imported rice grains were investigated in this study. All samples showed positive for the presence of B. cereus using polymerase chain reaction targeting the gryB gene (475 bp) which encodes for B protein subunit for DNA gyrase or also known as topoisomerase II. The microbial load of B. cereus in all samples was >1100MPN/g. However, PCR analysis revealed all the samples were contaminated with B. cereus except for three samples of local indigenous rice (LIR 3, LIR 9 and LIR 20).ConclusionsDue to the finding of high prevalence on the samples, it is therefore concluded that the local indigenous and imported rice grains can be one potential source of B. cereus transmission to the public.