This study was undertaken to characterize the antibiotic resistance and randomly amplified polymorphic DNA (RAPD) profiles of Vibrio parahaemolyticus isolates from raw vegetable samples. A total of 46 isolates of V. parahaemolyticus recovered from raw vegetables samples and were confirmed by PCR were analyzed in this study. Most of the isolates were resistant to nalidixic acid (93.48%) and were the least resistant towards imipinem (4.35%). The MAR index results also demonstrated high individual and multiple resistances to antibiotics among the isolates. From the RAPD analysis, the size for RAPD fragments generated ranged from 250 bp to 1,500 bp, with most of the strains contained three major gene fragments of 350, 1,000 and 1,350 bp. The RAPD profiles revealed a high level of DNA sequence diversity within the isolates. Antibiotic resistance and RAPD proved to be effective tools in characterizing and differentiating the V. parahaemolyticus strains.
Matched MeSH terms: Random Amplified Polymorphic DNA Technique
This study described the antibiotic and heavy metal resistance pattern of 17 isolates of Edwardsiella tarda obtained from Asian seabass (Lates calcarifer). E.tarda isolates were resistant to oleandomycin, lincomycin, novobiocin and spiramycin. In contrast, most of the isolates showed high level of susceptibility to tetracycline, doxycycline, florfenicol, chloramplenicol, nitrofurantoin, fosfomycin, kanamycin, oxolinic acid and flumequine. MAR value was 0.35 which indicated that the cultured Asian seabass have received high exposure to those tested antibiotics. Besides, very high level of heavy metal resistance among these isolates was observed. Genotypic profile of DNA fingerprintings generated by RAPD-PCR using M13 universal primer and M13 wild type phage primer showed high degree of genetic diversity with percentages similarity and genetic distance among the isolates were ranging from 10.5% to 100% and 0 to 0.895, respectively. This result indicates that strains that belong to the same origin were not always closely related genetically.
Matched MeSH terms: Random Amplified Polymorphic DNA Technique
A total of 32 clinical strains of Vibrio cholerae, including members of the 01 and 0139 serogroup
were collected from Klang, Selangor; Penang Island; Samarahan, Sarawak and Miri, Sarawak in Malaysia. In general, all the isolates except the 0139 serotype expressed low resistance to all the antibiotics tested with their Multiple Antibiotic Resistance (MAR) indices ranged from 0.10 to 0.48. The presence of ctx gene that encoded the cholera toxin was confirmed in all these clinical isolates by polymerase chain reaction. The results from the RAPD-PCR were analyzed using the RAPDistance software (Version 1.04). From the dendrogram generated, two main groups were observed which were subdivided into two clusters each. The Selangor’s isolates and the 0139 Penang’s isolates formed one group whereas the Samarahan, Sarawak isolates and the Miri, Sarawak isolates made up the other group, thus delineating their different sources of origin based on their geographical location.
Matched MeSH terms: Random Amplified Polymorphic DNA Technique
In this study, 90 non-replicate imipenem-resistant Pseudomonas aeruginosa (IRPA) Malaysian isolates collected between October 2005 and March 2008 were subjected to a screening test for detection of the integron and the gene cassette. Class 1 integrons were detected in 54 IRPA clinical isolates, whilst three isolates contained class 2 integrons. Analysis of the gene cassettes associated with the class 1 integrons showed the detection of accC1 in isolates carrying bla(IMP-7) and aacA7 in isolates carrying bla(VIM-2). aadA6 was detected in two isolates carrying bla(IMP-4). Using random amplification of polymorphic DNA analysis, 14 PCR fingerprint patterns were generated from the 32 isolates carrying metallo-β-lactamase (MBL) genes (35.5 %), whilst 20 patterns were generated from the 58 non-MBL gene isolates (64.4 %). Based on the differences in the fingerprinting patterns, two clusters (A and B) were identified among the MBL-producing isolates. Cluster A comprised 18 isolates (56 %) carrying the bla(VIM) gene, whereas cluster B comprised 14 (44 %) isolates carrying the bla(IMP) gene. The non-MBL isolates were divided into clusters C and D. Cluster C comprised 22 non-MBL isolates harbouring class 1 integrons, whilst cluster D consisted of three isolates carrying class 2 integrons. These findings suggest that the class 1 integron is widespread among P. aeruginosa isolated in Malaysia and that characterization of cassette arrays of integrons will be a useful epidemiological tool to study the evolution of multidrug resistance and the dissemination of antibiotic resistance genes.
Matched MeSH terms: Random Amplified Polymorphic DNA Technique
Haematococcus pluvialis (Flotow) is a unicellular green alga, which is considered to be the best astaxanthin-producing organism. Molecular markers are suitable tools for the purpose of finding out genetic variations in organisms; however there have been no studies conducted on ISSR or RAPD molecular markers for this organism. The DNA of 10 different strains of H. pluvialis (four strains from Iran, two strains from Finland, one strain from Switzerland and three strains from the USA) was extracted. A genetic similarity study was carried out using 14 ISSR and 12 RAPD primers. Moreover, the molecular weights of the bands produced ranged from 0.14 to 3.4 Kb. The PCA and dendrogram clustered the H. pluvialis strains into various groups according to their geographical origin. The lowest genetic similarity was between the Iran2 and USA2 strains (0.08) and the highest genetic similarity was between Finland1 and Finland2 (0.64). The maximum numbers of bands produced by the ISSR and RAPD primers were 35 and 6 bands, respectively. The results showed that ISSR and RAPD markers are useful for genetic diversity studies of Haematococcus as they showed geographical discrimination.
Matched MeSH terms: Random Amplified Polymorphic DNA Technique
Mackerel (Scombridae; Rastrelliger) are small commercially important pelagic fish found in tropical regions. They serve as a cheap source of animal protein and are commonly used as live bait. By using a truss morphometrics protocol and RAPD analysis, we examined morphological and genetic variation among 77 individual mackerel that were caught using long lines and gillnets at 11 locations along the west coast of Peninsular Malaysia. Nineteen morphometric traits were evaluated and genetic information was estimated using five 10-base RAPD random primers. Total DNA was extracted from muscle tissue. Morphometric discriminant function analysis revealed that two morphologically distinct groups of Rastrelliger kanagurta and a single group of R. brachysoma can be found along the west coast of Peninsular Malaysia. We also found that the head-related characters and those from the anterior part of the body of Rastrelliger spp significantly contribute to stock assessment of this population. RAPD analysis showed a trend similar to that of the morphometric analysis, suggesting a genetic component to the observed phenotypic differentiation. These data will be useful for developing conservation strategies for these species.
Matched MeSH terms: Random Amplified Polymorphic DNA Technique
We examined genetic variation in blood cockles in an effort to obtain information useful for the sustainability, management, and the stability of this species as a major commodity in the fisheries sector. Ten populations of cockles were sampled from the north to the south of the west coast of peninsular Malaysia. The cockles were collected in collaboration with the Fisheries Research Institute, Penang. The population genetic analysis of the cockles were studied via RAPD-PCR and mtDNA sequencing. Three hundred individuals were analyzed with RAPD-PCR experiments. High gene diversity over all loci was observed (Shannon index = 0.549 ± 0.056 and Nei's gene diversity = 0.4852 ± 0.0430 among 35 loci). The second method, mtDNA sequencing, was employed to complement the information obtained from RAPD-PCR. The gene selected for mtDNA sequencing was cytochrome c oxidase I (COI). One hundred and fifty individuals were sequenced, yielding a partial gene of 585 bp. Statistical analysis showed homogeneity in general but did reveal some degree of variability between the populations in Johor and the rest of the populations. The Mantel test showed a positive but nonsignificant correlation between geographic and genetic distances (r = 0.2710, P = 0.622), as in the RAPD analysis. We propose that the homogeneity between distant populations is caused by two factors: 1) the translocation of the spats; 2) larvae are carried by current movement from the north of the peninsula to the south. The different genetic composition found in Johor could be due to pollution, mutagenic substances or physical factors such as the depth of the water column. This population genetic study is the first for this species in peninsular Malaysia. The data from this study have important implications for fishery management, conservation of blood cockles and translocation policies for aquaculture and stock enhancement programs.
Matched MeSH terms: Random Amplified Polymorphic DNA Technique
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.
Matched MeSH terms: Random Amplified Polymorphic DNA Technique
The present investigation was carried out in an attempt to study the phylogenetic analysis of different breeds of domestic chickens in Peninsular Malaysia inferred from partial cytochrome b gene information and random amplified polymorphic DNA (RAPD) markers. Phylogenetic analysis using both neighbor-joining (NJ) and maximum parsimony (MP) methods produced three clusters that encompassed Type-I village chickens, the red jungle fowl subspecies and the Japanese Chunky broilers. The phylogenetic analysis also revealed that majority of the Malaysian commercial chickens were randomly assembled with the Type-II village chickens. In RAPD assay, phylogenetic analysis using neighbor-joining produced six clusters that were completely distinguished based on the locality of chickens. High levels of genetic variations were observed among the village chickens, the commercial broilers, and between the commercial broilers and layer chickens. In this study, it was found that Type-I village chickens could be distinguished from the commercial chickens and Type-II village chickens at the position of the 27th nucleotide of the 351 bp cytochrome b gene. This study also revealed that RAPD markers were unable to differentiate the type of chickens, but it showed the effectiveness of RAPD in evaluating the genetic variation and the genetic relationships between chicken lines and populations.
Matched MeSH terms: Random Amplified Polymorphic DNA Technique
Forty three (n=43) genomic DNA of Escherichia coli (11 isolates from eggs and 32 isolates from imported beef meats) were characterized by shiga toxin 1 (stx1), enterobacterial repetitive intergenic consensus-PCR (ERIC-PCR) and random amplified polymorphic DNA-PCR (RAPD-PCR) analyses. In the shiga toxin 1 (stx1) gene detection with primer stx 1F (5’-TTCTTCGGTATCCTATTCCC-3’) and stx 1R (5’- CTGTCACAGTAACAACCGT-3’), 9 E. coli of beef meats isolates were positive toward sxt1 gene. The results of the ERIC-PCR and RAPD-PCR were analyzed using GelCompar II software. ERIC-PCR with primer ERIC1 (5’-CACTTAGGGGTCCTCGAATGTA -3’) and ERIC2 (5’-AAGTAAGTGACTGGGGTGAGCG-3’) discriminated the E. coli into 6 clusters and 10 single isolates at 80% similarity. RAPD-PCR with primer Gen8 and Gen9, produced 10 clusters and 15 single isolates and 12 clusters and 14 single isolates of 80%, respectively. These results demonstrated that both ERIC-PCR and RAPD-PCR are useful and suitable tools for molecular typing of those isolates examined.
Matched MeSH terms: Random Amplified Polymorphic DNA Technique
This study was conducted to determine whether there was any genetic heterogeneity among Helicobacter pylori strains isolated from the antrum and corpus of the same individual in a Malaysian population and to determine the presence of heterogeneous susceptibility of the isolates by comparing PCR-RAPD and antibiotic profiles. Forty-four H. pylori isolates cultured from the antrum and corpus of 22 patients were analyzed. Antibiotic susceptibility testing was carried out by minimum inhibitory concentration determination, using E-Test method strips. PCR-RAPD was carried out on all the strains and the profiles generated were analysed for cluster analysis. Twenty-nine different PCR-RAPD profiles were observed in the 44 isolates. Fifteen pairs of the isolates from the same patients had the same PCR-RAPD patterns while in 7 pairs, the profiles were different. The strains were clustered into 2 separate clusters at a low coefficient of similarity, where most of the strains were in cluster 1. The degree of similarity was very low among most of the isolates. Most of the patients (16 of 22) were infected with strains that have the same antibiotic susceptibility profiles. Out of these, only 10 pairs shared the same PCR-RAPD and antibiotic profiles. Five pairs of isolates with similar PCR-RAPD profiles differed in their antibiotic profiles due to metronidazole resistance in one of the sites. A large degree of genetic heterogeneity was observed among H. pylori strains circulating among Malaysian patients. An individual patient can be infected with multiple strains and the strains can be antibiotic resistant.
Matched MeSH terms: Random Amplified Polymorphic DNA Technique*
The genetic heterogeneity and antifungal susceptibility patterns of Candida parapsilosis isolated from blood cultures of patients were investigated in this study. Randomly amplified polymorphic DNA (RAPD) analysis generated 5 unique profiles from 42 isolates. Based on the major DNA fragments of the RAPD profiles, the isolates were identified as RAPD type P1 (29 isolates), P2 (6 isolates), P3 (4 isolates), P4 (2 isolates) and P5 (1 isolate). Sequence analysis of the internal transcribed spacer (ITS) gene of the isolates identified RAPD type P1 as C. parapsilosis, P2 and P3 as Candida orthopsilosis, P4 as Candida metapsilosis, and P5 as Lodderomyces elongisporus. Nucleotide variations in ITS gene sequences of C. orthopsilosis and C. metapsilosis were detected. Antifungal susceptibility testing using Etests showed that all isolates tested in this study were susceptible to amphotericin B, fluconazole, ketoconazole, itraconazole and voriconazole. C. parapsilosis isolates exhibited higher MIC(50) values than those of C. orthopsilosis for all of the drugs tested in this study; however, no significant difference in the MICs for these two Candida species was observed. The fact that C. orthopsilosis and C. metapsilosis were responsible for 23.8 and 4.8 % of the cases attributed to C. parapsilosis bloodstream infections, respectively, indicates the clinical relevance of these newly described yeasts. Further investigations of the ecological niche, mode of transmission and virulence of these species are thus essential.
Matched MeSH terms: Random Amplified Polymorphic DNA Technique
Background : Visual impairment among the rural community has rarely been studied in this country. The purpose of this study is to determine the prevalence of visual impairment and its related factors among the adult population of Kampung Kuala Pajam, Branang.
Methodology : A cross sectional study was carried out in Kampung Kuala Pajam, Mukim Beranang from 25th of December till 30th of December 2007. Universal sampling is adopted and the sample size consists of 321 respondents who is 18 years and above. Trained interviewer has been used to obtain the data through questionnaire and eye examinations using logMAR chart, Red Reflex eye test, Relative Afferent Pupillary Defect (RAPD) test and Pin-Hole test.
Result : A total of 261 respondents who fulfil the criteria were interviewed and had eyes examination during the study period. The prevalence of visual impairment in this population was 9.2% and factors that showed significant association were age (p
Matched MeSH terms: Random Amplified Polymorphic DNA Technique
Fifty-nine isolates of Salmonella enterica subsp. enterica (S. enterica) isolated from indigenous vegetables, ‘selom’ (Oenanthe stolonifera) associated with 13 different serovars were obtained from Chemistry Department of Malaysia. The isolates encompass the common serovar, Salmonella enterica subsp. enterica serovar Weltevreden (S. Weltevreden) (39%) and Salmonella enterica subsp. enterica serovar Agona (S. Agona) (8.5%). Frequencies of the other 11 Salmonella serovars were ranged from 1.7% to 5.1%. All isolates were characterized by Enterobacterial Repetitive Intergenic Consensus-Polymerase Chain Reaction (ERIC-PCR), random amplified polymorphic DNA (RAPD), plasmid profiling and antimicrobial susceptibility testing. The results demonstrated ERIC-PCR, RAPD and composite analysis of both are suitable typing methods for S. enterica by demonstrating good discriminative ability and can be utilize as a rapid approach of comparing S. enterica isolates for epidemiological investigation. From this study, ERIC-PCR is exhibited lower discriminatory power when compare with RAPD. On the other hand, plasmid profiles yielded 32 profiles with molecular size ranging from 1129 bp to 17911 bp. Thirteen antimicrobial agents were included in this study and all isolates showed 100% (59/59) resistant to erythromycin and showed Multiple Antimicrobial Resistance (MAR) indexes ranging from 0.08 to 0.68. Dendrogram generated from antimicrobial resistance profiling exhibited poor discriminatory capability at serovar level. Although poultry still remain as the common reservoir for multidrug resistant (MDR) Salmonella. The isolation of 13 Salmonella serovars from selom that showed high MDR in this study is alarming. These results supported the notion that indigenous vegetable (selom) are gaining more antimicrobial resistance and could be potential health hazards.
Matched MeSH terms: Random Amplified Polymorphic DNA Technique
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.
Matched MeSH terms: Random Amplified Polymorphic DNA Technique
A study to determine the antibiotic sensitivity pattern and genotyping using RAPD-PCR was performed on 50 C. jejuni isolated from sushi retailed in different supermarkets. With less than half of the isolates susceptible to the antibiotics tested, resistant to two or more antibiotics were observed in most of the isolates. The banding patterns obtained from RAPD-PCR revealed that no predominant clone exists and the bacterial population is rather diverse. Hence, the resistance of the C. jejuni to different classes of antibiotic as well as their diverse genotypes suggests that these C. jejuni isolates were generated from different sources in the contaminated supermarkets where sushi were retailed. Our data showed that C. jejuni can be an important reservoir for resistance genes and that study with comprehensive collections of samples are urgently required to establish better measures to reduce or eliminate the risk from antibiotic resistant and pathogenic bacteria originating from minimally processed ready-to-eat food.
Matched MeSH terms: Random Amplified Polymorphic DNA Technique
Forty-eight strains of Salmonella enterica subsp. enterica serovar Agona and 33 strains of Salmonella enterica subsp. enterica serovar Weltevreden were characterized by random amplified polymorphic DNA (RAPD) fingerprinting using 3 different arbitrary primer, Enterobacterial Repetitive Intergenic Consensus-Polymerase Chain Reaction (ERIC-PCR) and antimicrobial susceptibility testing. By using RAPD, 81 strains (44 strains of S. Agona and 33 strains of S. Weltevreden) can be clustered into 14 groups and 6 single isolates whereas ERIC-PCR produced 7 clusters and 3 single isolates. Thirteen antimicrobial agents were used and all the isolates were resistant to erythromycin and showed Multiple Antimicrobial Resistance indexes, ranging from 0.08 to 0.62. Poultry still remain as the common reservoir for multi-drug-resistant Salmonella. On the other hand, vegetables contaminated with S. Weltevreden showed a gain in antimicrobial resistance. Besides that, consistent antibiograms were observed from S. Weltevreden isolated at Kajang wet market on 2000/08/02.
Matched MeSH terms: Random Amplified Polymorphic DNA Technique
The inheritance of 31 amplicons from short and long primer RAPD was tested for segregating ratios in two families of the brown planthopper, Nilaparvata lugens, and they were found to be inherited in a simple Mendelian fashion. These markers could now be used in population genetics studies of N. lugens. Ten populations of N. lugens were collected from five locations in Malaysia. Each location had two sympatric populations. Cluster and principal coordinate analyses based on genetic distance along with AMOVA revealed that the rice-infesting populations (with high esterase activity) at five localities clustered together as a group, and Leersia-infesting populations (with low esterase activity) at the same localities formed another distinct cluster. Two amplicons from primers OPD03 (0.65 kb) and peh#6 (1.0 kb) could be considered diagnostic bands, which were fixed in the Leersia-infesting populations. These results represent evidence of a sibling species in the N. lugens complex.
Matched MeSH terms: Random Amplified Polymorphic DNA Technique
A total of 78 samples comprising different types of street foods, sold in different locations in Malaysia, were examined for the presence of Enterobacter cloacae. E. cloacae contamination was recorded in 9% of the samples examined. Tests for susceptibility to 12 different antibiotics showed that all were resistant to six or more antibiotics, but susceptible to chloramphenicol and gentamicin. Plasmids of four different sizes were detected from the three plasmid positive isolates. RAPD analysis using four primers yielded completely different banding patterns for all E. cloacae studied. In Malaysia, no published information on street foods in the epidemiological investigation of E.cloacae related disease is available. However, their occurrences have provided compelling evidence that the risk of disease transmission caused by E. cloacae through street foods is moderate.
Matched MeSH terms: Random Amplified Polymorphic DNA Technique
Recurrent vulvovaginal candidiasis affects women worldwide and the resistance to azole drugs may be an important factor. The extent of strain-to-strain variation within a species and its relationship to the ability of the organism to colonize the vulvovaginal mucosa is not well established. The aims of this study were to compare: (i) the genotypes of Candida strains in sequential infections in patients with recurrent vaginitis, (ii) the genotypes of strains in patients with only one episode of infection in a period of 1 year and (iii) determine the in vitro antifungal susceptibilities of strains that cause recurrent vaginitis. Fifty-one cultured specimens from six distinct Candida species were genotyped via random amplified polymorphic DNA-polymerase chain reaction (RAPD-PCR) method using the ERIC1 and ERIC2 primers (ERIC, enterobacterial repetitive intergenic consensus). Statistical analyses allowed three different scenarios to be discerned for recurrent cases: (i) strain maintenance without genetic variation, (ii) strain maintenance with minor genetic variation and (iii) outright strain replacement. The genetic relatedness between strains from patients with recurrent vaginitis and patients with single episode of vaginitis were demonstrated by the dendogramme and the mean pairwise similarity coefficient S(AB) for the intergroup comparison was 0.223. However, intragroup genetic relatedness was slightly higher than intergroup comparison, with mean S(AB) of 0.261 and 0.331 for Groups I and II, respectively. A high proportion of Group I isolates (87.5%) causing recurrent infections were resistant to ketoconazole, whereas 41.7% of these isolates were cross-resistant to both clotrimazole and ketoconazole as shown by the in vitro antifungal susceptibility test, especially for C. glabrata isolates. Pregnancy status of patients displayed a highly significant association with C. albicans species whereas non-albicans species had a markedly higher prevalence in non-pregnant patients (p<0.001). These results may have a profound impact on the management of vaginal candidiasis, especially in recurrent cases.
Matched MeSH terms: Random Amplified Polymorphic DNA Technique