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Fulltext The distribution and characteristics of bacteria in recreational river water of a community resort in Baram, Sarawak, Malaysian Borneo

Lihan, S., Tian, P.K,, Chiew, T.S., Ching, C.L., Shahbudin, A., Hussain, H., et al.

International Food Research Journal, 2017;24(5):2238-2245.
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Enterobacteriaceae is a large family within the Gram-negative bacteria that primarily inhabits in the gastrointestinal tract of human and animals. The bacteria within this group are readily survived in the environment with some species found living free in the water where energy sources are scarce, making them ideal indicators for faecal contamination of the river water. Some species within the family have been used as indicator for the presence of pathogenic bacteria whilst on the other hand some species have been directly associated with various diseases in human and animals. The main aim of this research study was to determine the distribution and characteristics of the Enterobacteriaceae in water samples collected from river and waterfalls within a community resort. The health risk associated with the bacteria was analysed with regard to their susceptibility to antibiotics. Samples were collected from surface water and water falling down directly from waterfalls of river within the community resort. The samples collected were plated onto Eosine Methylene Blue agar (EMBA) for the isolation of the Enterobacteriaceae. Bacterial colonies growing on the agar were randomly picked, purified, stocked and then identified using API 20E identification kit. DNA fingerprinting using (GTG)5-PCR was utilised to determine their genetic profiles before the isolates were grouped into a dendrogram using RAPDistance software package. The level of antibiotic susceptibility of the bacteria isolates was analysed using disc diffusion technique. This study confirmed the presence of Enterobacter, Klebsiella, Citrobacter, Pantoea and Serratia in the water samples with their single and multiple antibiotic resistance and susceptible characteristics. The dendrogram presented in this study shows genetic similarities and differences among the strains, suggesting while there is a potential for single distribution of a clone, there is also possibility of the distribution of different strains within species in the water environment. Therefore, awareness on the potential risk associated with genetically diverse intermediate and resistant enteric bacteria in the recreational water should be communicated to the public especially communities within the study area.
Rapid isolation and detection of Escherichia coli O157:H7 by use of rainbow agar O157 and PCR assay

Radu S, Rusul G, Ling OW, Purwati E, Mustakim M, Lihan S

Southeast Asian J. Trop. Med. Public Health, 2000 Mar;31(1):77-9.
PMID: 11023069

This study has evaluated the use of a commercially available Rainbow agar O157 and polymerase chain reaction (PCR) assays for the detection of Shiga-like toxin producing Escherichia coli and to serotype E. coli O157:H7 from raw meat. The Rainbow agar O157 was found to be selective and sensitive for the screening of the E. coli O157 from artificially and naturally contaminated meat samples. Shiga-like toxin producing E. coli were identified with two primer pairs that amplified fragments of the SLT-I (384 bp) and SLT-II (584 bp). E. coli O157:H7 was serotyped with a primer pair specified for the H7 flagellar gene, which amplify specific DNA fragments (625 bp) from all E. coli O157:H7 strains. The use of Rainbow agar O157 described allows for the presumptive isolation of E. coli O157 in 24 hours. Identification and confirmation of the presumptive isolates as E. coli O157:H7 by PCR assays require additional 6-8 hours. The above-mentioned screening and identification procedures should prove to be a very useful method since it allows for the specific detection of E. coli O157:H7.
Characterization of Vibrio vulnificus isolated from cockles (Anadara granosa): antimicrobial resistance, plasmid profiles and random amplification of polymorphic DNA analysis

Radu S, Elhadi N, Hassan Z, Rusul G, Lihan S, Fifadara N, et al.

FEMS Microbiol Lett, 1998 Aug 01;165(1):139-43.
PMID: 9711850

Antibiotic susceptibility, plasmid profiles and random amplification of polymorphic DNA (RAPD) were used to study strains of Vibrio vulnificus isolated from cockles (Anadara granosa). Thirty-six isolates were analyzed. The prevalent biotypes were 1 (72.2% of the isolates) and 2 (27.8%). Among these, 21 strains of biotype 1 and two strains of biotype 2 contained plasmid DNA bands ranging in size from 1.4 to 9.7 MDa. Thirty-one (83.3%) were found to be resistant to one or more of the antimicrobial agents tested, however no specific correlation between antimicrobial resistance patterns and a single biotype was found. In addition, no particular plasmid profile was predictive of a particular pattern of antibiotic susceptibility. Two primers produced polymorphisms in all strains tested, producing bands ranging from 0.25 to 2.7 kb, indicating a high variability among both biotype 1 and biotype 2 of the V. vulnificus strains investigated. RAPD identity across biotypes was also observed among Vibrio vulnificus strains.
Fulltext Genotypic and phenotypic relationship in Burkholderia pseudomallei indicates colonization with closely related isolates

Radu S, Lihan S, Idris A, Ling OW, Al-Haddawi MH, Rusul G

Southeast Asian J. Trop. Med. Public Health, 1999 Dec;30(4):760-3.
PMID: 10928372

Seven isolates of Burkholderia pseudomallei from cases of melioidosis in human (2 isolates) and animal (2 isolates), cat (one isolate) and from soil samples (2 isolates) were examined for in vitro sensitivity to 14 antimicrobial agents and for presence of plasmid DNA. Randomly amplified polymorphic DNA (RAPD) analysis was used to type the isolates, using two arbitrary primers. All isolates were sensitive to chloramphenicol, kanamycin, carbenicillin, rifampicin, enrofloxacin, tetracycline and sulfamethoxazole-trimethoprim. No plasmid was detected in all the isolates tested. RADP fingerprinting demonstrated genomic relationship between isolates, which provides an effective method to study the epidemiology of the isolates examined.
Molecular characterization of Vibrio cholerae O1 and non-O1 from human and environmental sources in Malaysia

Radu S, Ho YK, Lihan S, Yuherman, Rusul G, Yasin RM, et al.

Epidemiol Infect, 1999 Oct;123(2):225-32.
PMID: 10579441

A total of 31 strains of Vibrio cholerae O1 (10 from outbreak cases and 7 from surface water) and non-O1 (4 from clinical and 10 from surface water sources) isolated between 1993 and 1997 were examined with respect to presence of cholera enterotoxin (CT) gene by PCR-based assays, resistance to antibiotics, plasmid profiles and random amplified polymorphic DNA (RAPD) analysis. All were resistant to 9 or more of the 17 antibiotics tested. Identical antibiotic resistance patterns of the isolates may indicate that they share a common mode of developing antibiotic resistance. Furthermore, the multiple antibiotic resistance indexing showed that all strains tested originated from high risk contamination. Plasmid profile analysis by agarose gel electrophoresis showed the presence of small plasmids in 12 (7 non-O1 and 5 O1 serotypes) with sizes ranging 1.3-4.6 MDa. The CT gene was detected in all clinical isolates but was present in only 14 (6 O1 serotype and 8 non-O1 serotype) isolates from environmental waters. The genetic relatedness of the clinical and environmental Vibrio cholerae O1 and non-O1 strains was investigated by RAPD fingerprinting with four primers. The four primers generated polymorphisms in all 31 strains of Vibrio cholerae tested, producing bands ranging from < 250 to 4500 bp. The RAPD profiles revealed a wide variability and no correlation with the source of isolation. This study provides evidence that Vibrio cholerae O1 and non-O1 have significant public health implications.
In vitro antimicrobial efficacy of Cassia alata (Linn.) leaves, stem, and root extracts against cellulitis causative agent Staphylococcus aureus

Toh SC, Lihan S, Bunya SR, Leong SS

BMC Complement Med Ther, 2023 Mar 18;23(1):85.
PMID: 36934252 DOI: 10.1186/s12906-023-03914-z

BACKGROUND: Cellulitis is a common skin disease encountered in medical emergencies in hospitals. It can be treated using a combination of antibiotics therapy; however, the causative agent Staphylococcus aureus has been reported to develop resistance towards the currently used antibiotics. Therefore, the search for more alternative herbal origin antimicrobial agents is critical.
AIM: In this study, maceration and Soxhlet extraction of the whole plant of Cassia alata Linn. (leaves, roots, and stem) were performed using four solvents with different polarities, namely n-hexane, ethyl acetate, ethanol and distilled water. The crude extracts were screened using agar well diffusion, colorimetric broth microdilution, grid culture and bacterial growth curve analysis against Staphylococcus aureus. The phytochemicals in the crude extracts were identified using Gas Chromatography-Mass Spectrometry (GC-MS).
RESULTS: Agar-well diffusion analysis revealed that extraction using ethyl acetate showed the largest inhibition zone with an average diameter of 15.30 mm (root Soxhlet extract) followed by 14.70 mm (leaf Soxhlet extract) and 13.70 mm (root maceration extract). The lowest minimum inhibitory and minimum bactericidal concentration in root Soxhlet extract using ethyl acetate was 0.313 and 0.625 µg µL-1, respectively. Our study proved that crude extract of the plant suppressed the growth of S. aureus as evidenced from a significant regression extension (p S. aureus in this study. The remaining 56 out of 88 phytochemicals of the plant should be intensively studied for more medicinal uses.
Characterization of Burkholderia pseudomallei isolated in Thailand and Malaysia

Radua S, Ling OW, Srimontree S, Lulitanond A, Hin WF, Yuherman, et al.

Diagn Microbiol Infect Dis, 2000 Nov;38(3):141-5.
PMID: 11109011

A total of 35 Burkholderia pseudomallei isolates from Thailand (16 clinical and eight soil isolates) and Malaysia (seven animal, two isolate each from clinical and soil) were investigated by their antimicrobial resistance, plasmid profiles and were typed by randomly amplified polymorphic DNA analysis. All isolates were found to be resistant to six or more of the 12 antimicrobial agents tested. Only two small plasmids of 1.8 and 2.4 megadalton were detected in two clinical isolates from Thailand. RAPD analysis with primer GEN2-60-09 resulted in the identification of 35 RAPD-types among the 35 isolates. The constructed dendrogram differentiated the 35 isolates into two main clusters and a single isolate. The wide genetic biodiversity among the 35 isolates indicate that RAPD-PCR can be a useful method to differentiate unrelated B. pseudomallei in epidemiological investigation.
Fulltext Prevalence of Staphylococcus aureus carriage by young Malaysian footballers during indoor training

William JL, Radu S, Aziz SA, Rahim RA, Cheah YK, Liwan A, et al.

Br J Sports Med, 2004 Feb;38(1):12-4.
PMID: 14751938

BACKGROUND: Research has shown that athletes are carriers of Staphylococcus aureus during physical activity.
OBJECTIVE: To estimate the mean total plate count of S aureus carried by footballers before and after training at an indoor venue.
METHODS: Forty Malay and 20 Indian students volunteered to participate. There was also a control group consisting of 40 Malay and 20 Indian students who were not active. The experimental group were active footballers who had played at school or club level. The subjects were healthy and free of skin infection. The experiment was divided into three sessions, with 20 subjects present at each. At each session, the subjects trained for one hour. Swabs were taken from the skin, nose, and ear before and after training. For the control group, swabs were taken only once from the skin, nose, and ear. The swabs were subjected to biochemical tests and then streaked and cultured aerobically in Baird Parker agar plates for 24 hours at 37 degrees C. Black colonies with a clear zone were presumed to be S aureus, and the mean total plate count of the colonies was estimated. Gram staining, catalase, coagulase slide, coagulase tube, acetoin production, o-nitrophenyl beta-D-galactopyranoside (ONPG), and mannitol fermentation tests were used to confirm the colonies as S aureus. A haemolysin test was conducted with human blood to confirm haemolytic activity.
RESULTS: All subjects in the experimental group were carrying S aureus both before and after training. The estimated mean total counts of colonies from the skin, ear, and nose for the Malays before training were 33, 71, and 312 respectively. Counts after training were 21, 44, and 452 respectively. The results for the Indians were 72, 80, and 309 respectively before training and 55, 200, and 466 respectively after training. The positive results for Gram staining, catalase, coagulase slide, coagulase tube, acetoin production, ONPG, and mannitol fermentation tests were 100%, 96%, 95%, 95%, 93%, 93%, and 90% respectively. All subjects in the control group were also carrying S aureus.
CONCLUSIONS: All of the players were carriers of S aureus during training. The decrease in total count from the skin for both races may be due to lysozyme activity lysing the bacterial cells. Contamination of the environment with these bacteria may have increased the estimated total plate count in the nose. The experimental group face a higher risk of infection because of lower immunity during training and higher rate of injuries compared with the control group.
Biosorption of copper by endophytic fungi isolated from Nepenthes ampullaria

Wong C, Tan LT, Mujahid A, Lihan S, Wee JLS, Ting LF, et al.

Lett Appl Microbiol, 2018 Oct;67(4):384-391.
PMID: 29998586 DOI: 10.1111/lam.13049

Copper (Cu) tolerance was observed by endophytic fungi isolated from the carnivorous plant Nepenthes ampullaria (collected at an anthropogenically affected site, Kuching city; and a pristine site; Heart of Borneo). The fungal isolates, capable of tolerating Cu up to 1000 ppm (11 isolates in total), were identified through molecular method [internal transcribed spacer 4+5 (ITS4+5); ITS1+NL4; β-tubulin region using Bt2a + Bt2b], and all of them grouped with Diaporthe, Nigrospora, and Xylaria. A Cu biosorption study was then carried out using live and dead biomass of the 11 fungal isolates. The highest biosorption capacity of using live biomass was achieved by fungal isolates Xylaria sp. NA40 (73·26 ± 1·61 mg Cu per g biomass) and Diaporthe sp. NA41 (72·65 ± 2·23 mg Cu per g biomass), NA27 (59·81 ± 1·15 mg Cu per g biomass) and NA28 (56·85 ± 4·23 mg Cu per g biomass). The fungal isolate Diaporthe sp. NA41 also achieved the highest biosorption capacity of 59·33 ± 0·15 mg g-1 using dead biomass. The living biomass possessed a better biosorption capacity than the dead biomass (P
Fulltext Lactic acid production by Enteroccocus faecium in liquefied sago starch

Nolasco-Hipolito C, Zarrabal OC, Kamaldin RM, Teck-Yee L, Lihan S, Bujang KB, et al.

AMB Express, 2012 Sep 28;2(1):53.
PMID: 23021076 DOI: 10.1186/2191-0855-2-53

Enterococcus faecium No. 78 (PNCM-BIOTECH 10375) isolated from puto, a type of fermented rice in the Philippines was used to produce lactic acid in repeated batch fermentation mode. Enzymatically liquefied sago starch was used as the sole carbon source, since sago (Metroxylon spp) is a sustainable crop for industrial exploitation. Liquefied sago starch was inoculated with E. faecium to perform the saccharification and fermentation processes simultaneously. Results demonstrated that E. faecium was reused for 11 fermentation cycles with an average lactic acid yield of 36.3 ± 4.71 g/l. The lactic acid production was superior to that of simple batch mode and continuous fermentation in terms of lactic acid concentration. An un-dissociated lactic acid concentration of 1.15 mM affected the productivity of the cells. Work is in progress to maintain and increase the usability of the cells over higher fermentation cycles.