Piper nigrum, commonly known as black pepper, is one of the most important spice crops
with high demand by the world market. However, diseases like foot rot and stem blight
cause by Phytophthora capsici have become the important production constraints in black
pepper industry. The frequent application of toxic fungicides to counter the diseases in pepper
plantations has raised certain environmental issues. In order to mitigate the use of fungicides,
biological approach to control P. capsici has been suggested. In this study, endophytic bacteria
were isolated from six P. nigrum roots and screened for in vitro antagonistic activity against P.
capsici through dual culture, mycelial growth, spore germination and double plate assay. The
antagonism testing involved the secretion of volatile and diffusible bioactive compounds by
the endophytic bacteria. Out of 19 isolates tested, two isolates DB(2)7 and SB(2)6 produced
volatile bioactive compounds and these two isolates showed highest antagonism against P.
capsici mycelia with the percentage of inhibition up to 47.63% and 43.33%, respectively.
Diffusible compounds from isolates DB(2)7, DB(2)9 and SB(2)6 produced clear zones in spore
germination test with radii measurements of 10.0-17.0 mm. Three isolates with promising
antifungal activity were further characterised through 16S rDNA sequencing. The analysis
of their sequences via National Center for Biotechnology Information (NCBI) suggests close
identity towards Enterobacter cancerogenus, Enterobacter cloacae and Enterobacter asburiae.
This research study demonstrated that these endophytic bacteria isolates are potentially to be
used as biocontrol agent in pepper cultivation.
In Malaysia, the aquaculture industry, particularly the production of freshwater aquaculture fish, is growing rapidly. Nevertheless, the illegal use of banned antimicrobial agents such as chloramphenicol in aquaculture has become a major concern in relation to the safety of consumers and also the development of drug-resistant strains in bacteria. Driven by those factors, the main intention of this study was to determine the prevalence and types of chloramphenicol resistance genes in E. coli isolated from aquaculture and other environmental waters. The respective chloramphenicol-resistance genes in the isolates were detected by multiplex PCR with four sense primers C-1, C-2, C-3, C-4 and one antisense primer C-R for targeting cat I, cat II, cat III and cat IV genes, respectively. Out of 27 E. coli isolated, 19 were resistant to chloramphenicol. Cat I, cat II, cat III and cat IV genes were detected in 19, 13, 10, and 6 of the E. coli isolates, respectively. The results of this study revealed that chloramphenicol-resistance E. coli is present in aquaculture and environmental waters, in the study area. This finding suggested that although banned, there could be illegal usage of chloramphenicol antibiotic in local aquaculture. The bacteria in aquaculture may have spread to other environmental water through disposal of aquaculture waste water to other environments.
Antibiotic susceptibility and genetic diversity of E. coli isolated from cultured catfish and their surrounding environment were determined. The levels of resistance of the E. coli isolates towards six different antibiotics tested differed considerably. Though the isolates displayed resistance towards some of the antibiotics tested, none of the isolates showed resistant towards norfloxacin, sulphametoxazole/trimethoprim and chloramphenicol. RAPD-PCR analysis using single primer and primers combination clustered the E. coli isolates into 3 and 5 groups, respectively. The results of this study suggest that the E. coli isolates from the catfish and their surrounding environment derived from a mixture of sensitive and resistant strains with diverse genetic contents. The use of the RAPD analysis is sufficiently discriminatory for the typing of the E. coli isolates.
Bacillus cereus is a soil inhabitant gram positive bacterium, and is known to cause severe food poisoning. The objective of this study was to isolate and identify the presence of Bacillus cereus s.l. from selected ready to eat cereals purchased randomly from local supermarkets in Kuching and Kota Samarahan, Sarawak. The result showed that four of the 30 food samples were detected to be contaminated by B. cereus s.l. . Our findings suggested that it is important for the public to be aware of the safety of RTE cereals consumption, as it is possible that B. cereus s.l. may be present in high count number and pose hazardous health effects to the consumers.
In this study, RAPD-PCR and ERIC-PCR were used to study the epidemiology of V. parahaemolyticus isolated from cockles in Padang, Indonesia. The Gold Oligo OPAR3 primer produced bands ranged from 1-8 with sizes from 0.2 – 5.0 kb and the Gold Oligo OPAR8 primer produced 1-7 bands with sizes 0.7 – 1.5 kb. Both primers produced twenty five RAPD patterns with a few isolates failed to produce any products. Based on phylogenetic dendrogram, all the isolates can be divided into 6 major clusters with similarity between 0 to 52%. For the ERIC primer, it produced bands ranged from 3-15 with sizes from 0.1 – 5.0 kb and twenty seven different ERIC patterns. Construction of the phylogenetic dendogram showed the isolates can be divided into 4 major clusters with similarity between 56 to 86%. The high diversity of both processes may be due to the multiple contamination sources of V. parahaemolyticus.
Fish can live healthier in aquarium with good water quality than they do in the wild. Maintaining
the quality of the water in fish facility is needed to avoid fluctuation of physicochemical
parameter values and contamination with pathogenic microorganisms that may cause serious
illness or even death among the fish. Contamination of the water, especially with animal
pathogens which are also pathogenic to human may pose health risk to those who are handling
or in direct contact with the water and fish in the facility. Therefore, there is a need to assess the
water quality and the risk associated with microorganisms in the water and the cultured animals.
The aim of this study was to determine the water quality with regard to the physicochemical
and microbiological parameters as well as the risk associated with bacteria in the water of the
fish facility. Samples of water from the water source and also from aquariums in the fish facility
were collected and analyzed. The water samples were plated on nutrient agar for bacterial
enumeration then bacterial colonies growing on the agar plates were randomly picked and
purified. (GTG)5
-PCR analysis was carried out to analyse the heterogeneity of the genome of the
bacterial isolated and a dendrogram was constructed from the (GTG)5
-PCR profile to determine
the genotypic group of the bacterial isolates. The risk associated with the bacteria from the
water was analyzed with respect to their antibiotic resitance. The result of this study revealed
that the (GTG)5
-PCR analysis was able to group the bacteria into 2 main genotypic clusters
which were further grouped into several sub-clusters. From the dengrogram, 12 representative
isolates were selected and identified using 16S rRNA sequencing. The identification confirmed
the presence of Aeromonas veronii (8 isolates), Aeromonas jandaei (2 isolates), Plesiomonas
shigelloides (1 isolate) and Pseudomonas alcaligene (1 isolate) from the water samples. All
of the isolates exhibited resistant towards ampicillin, penicillin and gentamicin. This study
revealed that the water from the fish facility harboured genetically diverse antibiotic resistance
bacteria which may pose health risk to the fish and also to those who are in direct contact
with the contaminated water and fish in the facility. Therefore, water in fish facility should be
monitored regularly and handled with caution.
This study was conducted to detect the presence of Listeria monocytogenes (L. monocytogenes)
and screen for its antibiotic susceptibility characteristic from wildlife and water samples at
Kubah National Park, Sarawak, Malaysia. Samples collected were incubated and streaked on
selective medium PALCAM agar to confirm the presence of Listeria spp. before they were
further tested using molecular analysis. Specific Polymerase Chain Reaction (PCR) assay were
performed to target specific virulence gene, haemolysin gene, hlyA to further distinguish the
presence of this pathogenic bacteria in the samples. Overall, out of the 30 samples tested, 10
samples were confirmed as to contain L. monocytogenes strains and selected to subsequent
antibiotic susceptibility test. Susceptibility patterns to 10 antibiotics were investigated
among the L. monocytogenes strains. All strains were uniformly resistant to tetracycline and
erythromycin. On the other hand, all strains were sensitive to gentamycin and tobramycin. The
multiple antibiotic resistance shown by the strains in this study indicate the potential health
hazard associated with the possible transmission between wildlife and water to its surrounding
environment especially visitors and workers of Kubah National Park, Sarawak, Malaysia.
(GTG)5 PCR is a type of repetitive extragenic palindromic (rep)-PCR which amplifies the (GTG)5 repetitive element that lays throughout the bacterial genome. In this study, fifty, thirty-nine and forty-nine unknown bacteria were isolated from aquaculture farms in Miri, Limbang and Lundu, respectively. (GTG)5 PCR was used to screen for clonal diversity among the isolates according to sampling sites. Banding profiles obtained from electrophoresed (GTG)5 PCR products were analyzed by RAPDistance Software to generate a dendrogram of neighbor joining tree (NJT) format. Based on the constructed dendrogram, representative isolates were selected for further identification. Conserved 16S rRNA region of the selected bacteria isolates were amplified and purified DNA products were sequenced. (GTG)5 PCR is useful in differentiation of unknown bacterial isolates and 16S rRNA analysis species identity of the bacteria in Sarawak aquaculture environment. The high diversity of bacteria in aquaculture environment may be caused by contamination from various sources.
Nineteen strains of vancomycin-resistant Enterococcus faecium isolated from 10 of 75 (13.3%) tenderloin beef samples were examined for resistance to selected antibiotics, presence of plasmids, and genetic diversity by random amplification of polymorphic DNA analysis. All strains showed multiple resistant to the antibiotics tested. Multiple antibiotic indexing of the vancomycin-resistant E. faecium strains showed that all (100%) originated from high risk contamination environments where antibiotics were often used. Plasmids ranging in size from 1.5 to 36 megadalton were detected in 15 of 19 (79%) strains. Thus, three plasmid profiles and eight antibiotypes were observed among the E. faecium strains. A high degree of polymorphism was obtained by combining the results of the two primers used; with the 19 E. faecium strains being differentiated into 19 RAPD-types. These preliminary results suggest that RAPD-PCR has application for epidemiologic studies and that resistance patterns and plasmid profiling could be used as an adjunct to RAPD for the typing of E. faecium in the study area.
Four Vibrio cholerae O139 Bengal strains isolated from surface water were characterized by antibiotic resistance, plasmid profile, presence of cholera toxin gene and random amplification of polymorphic DNA (RAPD) analysis. All four strains exhibit multiple resistance towards the antibiotics tested with a multiple antibiotic resistance index of 0.5-0.66, and harboured a 2.0 MDa non-conjugative plasmid. The Vibrio cholerae O139 Bengal were positive for the cholera toxin gene. Antibiotyping and random amplification of polymorphic DNA analysis with four primers proved to be useful in discriminating the isolates. RAPD proved to be more sensitive. These results reveal that there is significant genetic diversity among the Vibrio cholerae O139 Bengal strains studied.
The administration of antimicrobials in aquaculture provides a selective pressure creating a reservoir of multiple resistant bacteria in the cultured fish and shrimps as well as the aquaculture environment. The objective of this study was to determine the extent of antibiotic resistance in aquaculture products and aquaculture's surrounding environment in Sarawak, Malaysian Borneo. Ninety-four identified bacterial isolates constituted of 17 genera were isolated from sediment, water, and cultured organisms (fish and shrimp) in selected aquaculture farms. These isolates were tested for their antibiotic resistance against 22 antibiotics from several groups using the disk diffusion method. The results show that the highest resistance was observed towards streptomycin (85%, n = 20), while the lowest resistance was towards gentamicin (1.1%, n = 90). The multiple antibiotic resistant (MAR) index of the isolates tested ranged between 0 and 0.63. It was suggested that isolates with MAR index > 0.2 were recovered from sources with high risk of antibiotic resistant contamination. This study revealed low level of antibiotic resistance in the aquaculture bacterial isolates except for streptomycin and ampicillin (>50% resistance, n = 94) which have been used in the aquaculture industry for several decades. Antibiotic resistant patterns should be continuously monitored to predict the emergence and widespread of MAR. Effective action is needed to keep the new resistance from further developing and spreading.