Displaying publications 41 - 60 of 85 in total

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  1. Fulltext Genetic diversity of toxigenic Vibrio cholerae O1 from Sabah, Malaysia 2015
    Zaw MT, Emran NA, Ibrahim MY, Suleiman M, Awang Mohd TA, Yusuff AS, et al.
    J Microbiol Immunol Infect, 2019 Aug;52(4):563-570.
    PMID: 29428381 DOI: 10.1016/j.jmii.2018.01.003
    BACKGROUND: Cholera is an important health problem in Sabah, a Malaysian state in northern Borneo; however, Vibrio cholerae in Sabah have never been characterized. Since 2002, serogroup O1 strains having the traits of both classical and El Tor biotype, designated as atypical El Tor biotype, have been increasingly reported as the cause of cholera worldwide. These variants are believed to produce clinically more severe disease like classical strains.

    PURPOSE: The purpose of this study is to investigate the genetic diversity of V.cholerae in Sabah and whether V.cholerae in Sabah belong to atypical El Tor biotype.

    METHODS: ERIC-PCR, a DNA fingerprinting method for bacterial pathogens based on the enterobacterial repetitive intergenic consensus sequence, was used to study the genetic diversity of 65 clinical V.cholerae O1 isolates from 3 districts (Kudat, Beluran, Sandakan) in Sabah and one environmental isolate from coastal sea water in Kudat district. In addition, we studied the biotype-specific genetic traits in these isolates to establish their biotype.

    RESULTS: Different fingerprint patterns were seen in isolates from these three districts but one of the patterns was seen in more than one district. Clinical isolates and environmental isolate have different patterns. In addition, Sabah isolates harbor genetic traits specific to both classical biotype (ctxB-1, rstRCla) and El Tor biotype (rstRET, rstC, tcpAET, rtxC, VC2346).

    CONCLUSION: This study revealed that V.cholerae in Sabah were genetically diverse and were atypical El Tor strains. Fingerprint patterns of these isolates will be useful in tracing the origin of this pathogen in the future.

    Matched MeSH terms: Vibrio cholerae O1/genetics*; Vibrio cholerae O1/isolation & purification
  2. Comparative PCR-based fingerprinting of Vibrio cholerae isolated in Malaysia
    Shuan Ju Teh C, Thong KL, Osawa R, Heng Chua K
    J Gen Appl Microbiol, 2011;57(1):19-26.
    PMID: 21478644
    Vibrio cholerae, the causative agent of cholera, is endemic in many parts of the world, especially in countries poor in resources. Molecular subtyping of V. cholerae is useful to trace the regional spread of a clone or multidrug-resistant strains during outbreaks of cholera. Current available PCR-based fingerprinting methods such as Random Amplified Polymorphic DNA (RAPD)-PCR, Enterobacterial Repetitive Intergenic Consensus Sequence (ERIC)-PCR, and Repetitive Extragenic Palindromic (REP)-PCR were used to subtype V. cholerae. However, there are problems for inter-laboratory comparison as these PCR methods have their own limitations especially when different PCR methods have been used for molecular typing. In this study, a Vibrio cholerae Repeats-PCR (VCR-PCR) approach which targets the genetic polymorphism of the integron island of Vibrios was used and compared with other PCR-based fingerprinting methods in subtyping. Forty-three V. cholerae of different serogroups from various sources were tested. The PCR-fingerprinting approaches were evaluated on typeability, reproducibility, stability and discriminatory power. Overall, Malaysian non-O1/non-O139 V. cholerae were more diverse than O1 strains. Four non-O1/non-O139 strains were closely related with O1 strains. The O139 strain in this study shared similarity with strains of both O1 and non-O1/non-O139 serogroups. ERIC-PCR was the most discriminative approach (D value = 0.996). VCR-PCR was useful in discriminating non-O1/non-O139 strains. RAPD-PCR and REP-PCR were less suitable for efficient subtyping purposes as they were not reproducible and lacked stability. The combination of the ERIC-PCR and VCR-PCR may overcome the inadequacy of any one approach and hence provide more informative data.
    Matched MeSH terms: Vibrio cholerae/classification*; Vibrio cholerae/genetics*; Vibrio cholerae/isolation & purification
  3. Fulltext Host, pathogen and environment: a bacterial gbpA gene expression study in response to magnesium environment and presence of prawn carapace and commercial chitin
    Tiruvayipati S, Bhassu S
    Gut Pathog, 2016;8:23.
    PMID: 27231485 DOI: 10.1186/s13099-016-0105-5
    Vibrio parahaemolyticus is a Gram-negative halophilic bacterium which is found largely in estuarine and coastal waters. The bacteria has been a main focus in gastro-intestinal infections caused primarily due to the consumption of contaminated seafood. It was shown to survive in magnesium concentrations as high as 300 mM which are toxic to various other micro-organisms. Several genes of V. parahaemolyticus were studied, among which gbpA (N-acetyl glucosamine binding protein) was reported in Vibrio cholerae.
    Matched MeSH terms: Vibrio cholerae
  4. Fulltext Detection of Vibrio cholerae in raw cockles (Anadara granosa) by polymerase chain reaction
    International Food Research Journal, 2010;17(3):675-680.
    MyJurnal
    Aimed of this study was to determine the presence of Vibrio cholerae in cockles (Anadara granosa)
    from different coasts in Malaysia and to measure the biosafety of V. cholerae in raw cockles at wet market in Malaysia using the polymerase chain reaction (PCR) in combination with the most probable number (MPN) method. A total of 100 samples from 4 different wet markets in the West and East were examined for the presence of V. cholerae. The prevalence of V. cholerae between the two coasts was not significant different. In fact, the 74% of samples from West coast area was found positive while the 69% for samples collected in the East coast. West coast samples showed a prevalence of 60% for the wet market A=, 64% for B=, 88% for C= and 84% for the market D); East coast samples showed the same percentage with 72% for the wet markets E, F and H, followed by wet market G with 60%.With the MPN-PCR method, using 80 samples of raw cockles obtained from 4 wet markets, the occurrence of V. cholerae detected was of 95%. The frequency of V. cholerae in raw cockles obtained from wet market I and L was higher (100%) compared to other wet market (Wet market B=, 90%; Wet market C=, 95%).The density of V. cholerae detected in all samples ranged from 24000 MPN/g, but most of the samples (24 samples) were in category >24000 MPN/g concentration. V. cholerae was present in raw cockles in higher number. Hence, these results demonstrate the presence of pathogenic V.cholerae in cockles harvested and reveal the potential risk of illness associated with their consumption. This study will be the first biosafety assessment of V. choleare in raw cockles in Malaysia and it will provide significant insights about Malaysian scenario.
    Matched MeSH terms: Vibrio cholerae
  5. Fulltext Short communication : microbiological, histamine and 3-MCPD contents of Malaysian unprocessed ‘budu’
    Rosma, A., Afiza, T. S., Wan Nadiah, W. A., Liong, M. T., Gulam, R. R. A.
    International Food Research Journal, 2009;16(4):589-594.
    MyJurnal
    Unprocessed ‘budu’ is a mixture of anchovies and salt that has been fermented for a period of time, and has not been heat-treated nor formulated with additional ingredients. This study analyzed Malaysian
    unprocessed ‘budu’ from 12 producers for microbiological, salt, protein, histamine and 3-MCPD contents.
    The results demonstrated that Malaysian unprocessed ‘budu’ were free from pathogenic Coliform, E. coli,
    V. parahaemolyticus and V. cholerae contaminations. Carcinogenic 3-MCPD was below detection level of 2 ppb for all 12 samples tested. However, 58% of the unprocessed ‘budu’ had histamine content greater than the hazardous levels of 50 mg/100 g sample.
    Matched MeSH terms: Vibrio cholerae
  6. Fulltext Genomic diversity of cholera outbreak strains in East Malaysia
    Bilung, Lesley Maurice, Yong, Sy Fuh, Linang, Velnetti, Benjamin, Adam, Vincent, Micky, Apun, Kasing, et al.
    Malaysian Journal of Medicine and Health Sciences, 2014;10(2):19-26.
    MyJurnal
    Thirty one Vibrio cholera isolates recovered from cholera outbreak in Bintulu, Sarawak (Malaysia) were detected with the presence of ctx gene by using specific PCR. These isolates were further characterized and differentiated by using the Enterobacterial Repetitive Intergenic Consensus PCR (ERIC-PCR) and BOX-PCR to determine their genomic fingerprints. The specific PCR result confirmed the identities of 27 isolates out of 31 as pathogenic V. cholerae. The ERIC-PCR generated several genetic profiles consisting of 4-6 bands with sizes in the range of 100 to 600 bp, while the BOX-PCR produced profiles numbering 2-7 bands in the sizes between 200 to 1000 bp. Based on the dendrogram generated from the DNA fingerprinting profiles (ERIC-PCR and BOX-PCR), all of the isolates can be divided into 2 main clusters that is further divided into 2 sub-clusters. The low genetic diversity of the isolates indicated the outbreak of V. cholerae in the study area was due to the contamination from a single or few sources of V. cholerae.
    Matched MeSH terms: Vibrio cholerae
  7. Fulltext Simultaneous multiplex Polymerase Chain Reaction detection of Salmonella spp., Escherichia coli O157, Vibrio parahaemolyticus, Vibrio cholerae, Listeria monocytogenes and Campylobacter spp.
    Ling, S., Noramirah, R., Abidatul, A.A., Nurfarhanah, N.M.J., Noor-Azira, A.M., Jambari, N.N., et al.
    Food Research, 2018;2(3):240-246.
    MyJurnal
    Foodborne illness is a global burden that impacts a country politically, economically and
    socio-economically. The severity of the burden can be unmeasurable as foodborne illness
    is often an underestimated problem. In order to enlighten the burden, appropriate food
    safety control measures should be taken. This study aimed to optimize a multiplex
    Polymerase Chain Reaction (mPCR) detection method to identify foodborne pathogens
    simultaneously. Six foodborne pathogens namely, Salmonella spp., Escherichia coli O157,
    Vibrio parahaemolyticus, Vibrio cholerae, Listeria monocytogenes and Campylobacter
    spp., were targeted in the mPCR detection method. Each mPCR parameter was tested and
    the outcome was analysed to obtain a successful mPCR protocol to detect the targeted
    foodborne pathogens. The amplified PCR products showed that the optimized mPCR
    protocol will be a potential rapid diagnostic tool in foodborne pathogen detection.
    Matched MeSH terms: Vibrio cholerae
  8. Fulltext Tetracycline-resistant vibrio cholerae El Tor
    Hassan H, Teh A
    Med J Malaysia, 1993 Mar;48(1):95-6.
    PMID: 8341181
    http://www.e-mjm.org/1993/v48n1/Vibrio_Cholerae_El_Tor.pdf
    Matched MeSH terms: Vibrio cholerae/isolation & purification*
  9. A new bacterial white spot syndrome (BWSS) in cultured tiger shrimp Penaeus monodon and its comparison with white spot syndrome (WSS) caused by virus
    Wang YG, Lee KL, Najiah M, Shariff M, Hassan MD
    Dis Aquat Organ, 2000 May 25;41(1):9-18.
    PMID: 10907134
    This paper describes a new bacterial white spot syndrome (BWSS) in cultured tiger shrimp Penaeus monodon. The affected shrimp showed white spots similar to those caused by white spot syndrome virus (WSSV), but the shrimp remained active and grew normally without significant mortalities. The study revealed no evidence of WSSV infection using electron microscopy, histopathology and nested polymerase chain reaction. Electron microscopy indicated bacteria associated with white spot formation, and with degeneration and discoloration of the cuticle as a result of erosion of the epicuticle and underlying cuticular layers. Grossly the white spots in BWSS and WSS look similar but showed different profiles under wet mount microscopy. The bacterial white spots were lichen-like, having perforated centers unlike the melanized dots in WSSV-induced white spots. Bacteriological examination showed that the dominant isolate in the lesions was Bacillus subtilis. The occurrence of BWSS may be associated with the regular use of probiotics containing B. subtilis in shrimp ponds. The externally induced white spot lesions were localized at the integumental tissues, i.e., cuticle and epidermis, and connective tissues. Damage to the deeper tissues was limited. The BWS lesions are non-fatal in the absence of other complications and are usually shed through molting.
    Matched MeSH terms: Vibrio cholerae/isolation & purification
  10. Fulltext Immunogenicity and protective efficacy of a live, oral cholera vaccine formulation stored outside-the-cold-chain for 140 days
    Xian TH, Sinniah K, Yean CY, Krishnamoorthy V, Bahari MB, Ravichandran M, et al.
    BMC Immunol, 2020 05 25;21(1):29.
    PMID: 32450807 DOI: 10.1186/s12865-020-00360-1
    BACKGROUND: Cholera, an acute watery diarrhoeal disease caused by Vibrio cholerae serogroup O1 and O139 across the continents. Replacing the existing WHO licensed killed multiple-dose oral cholera vaccines that demand 'cold chain supply' at 2-8 °C with a live, single-dose and cold chain-free vaccine would relieve the significant bottlenecks and cost determinants in cholera vaccination campaigns. In this direction, a prototype cold chain-free live attenuated cholera vaccine formulation (LACV) was developed against the toxigenic wild-type (WT) V. cholerae O139 serogroup. LACV was found stable and retained its viability (5 × 106 CFU/mL), purity and potency at room temperature (25 °C ± 2 °C, and 60% ± 5% relative humidity) for 140 days in contrast to all the existing WHO licensed cold-chain supply (2-8 °C) dependent killed oral cholera vaccines.

    RESULTS: The LACV was evaluated for its colonization potential, reactogenicity, immunogenicity and protective efficacy in animal models after its storage at room temperature for 140 days. In suckling mice colonization assay, the LACV recorded the highest recovery of (7.2 × 107 CFU/mL) compared to those of unformulated VCUSM14P (5.6 × 107 CFU/mL) and the WT O139 strain (3.5 × 107 CFU/mL). The LACV showed no reactogenicity even at an inoculation dose of 104-106 CFU/mL in a rabbit ileal loop model. The rabbits vaccinated with the LACV or unformulated VCUSM14P survived a challenge with WT O139 and showed no signs of diarrhoea or death in the reversible intestinal tie adult rabbit diarrhoea (RITARD) model. Vaccinated rabbits recorded a 275-fold increase in anti-CT IgG and a 15-fold increase in anti-CT IgA antibodies compared to those of rabbits vaccinated with unformulated VCUSM14P. Vibriocidal antibodies were increased by 31-fold with the LACV and 14-fold with unformulated VCUSM14P.

    CONCLUSION: The vaccine formulation mimics a natural infection, is non-reactogenic and highly immunogenic in vivo and protects animals from lethal wild-type V. cholerae O139 challenge. The single dose LACV formulation was found to be stable at room temperature (25 ± 2 °C) for 140 days and it would result in significant cost savings during mass cholera vaccination campaigns.

    Matched MeSH terms: Vibrio cholerae/immunology
  11. Fulltext Wings of the common house fly (Musca domestica L.): importance in mechanical transmission of Vibrio cholerae
    Yap KL, Kalpana M, Lee HL
    Trop Biomed, 2008 Apr;25(1):1-8.
    PMID: 18600198
    The importance of house fly (Musca domestica L) wings in mechanical transmission of bacteria was studied. A droplet of phosphate-buffered saline containing Vibrio cholerae was rolled along one wing of each house fly. None adhered to the wings but small proportions of the bacterium were isolated from about half the wings. Vibrio cholerae was spread onto the ventral wing surfaces of each unconscious house fly which then was placed inside a bottle. When it regained consciousness, the types of activity it performed over five minutes were noted before the house fly was killed and the bacteria on its wings numerated. Control were house flies killed before inoculation. The proportion of house flies with bacteria on their wings and the mean number of bacteria remaining were significantly less on live house flies than killed controls. Among the live house flies, bacteria were detected on fewer house flies which flew (25%) than those which did not fly (81%). In addition, the mean number of bacteria on the former was significantly less than the latter (5 against 780 colonies). However, both these parameters were not significantly different between the group which performed and the group which did not perform wing grooming; takeoff and alighting over short distances, and somersaulting. Wings of unconscious house flies tethered by their thoraxes were inoculated with V. cholerae. After regaining consciousness, the house flies were allowed to move their wings in flight motions for up to 30 seconds. Small proportions of bacteria remained on all the house flies. House flies were placed in a chamber containing a liquid bait spiked with V. cholerae. After two hours, 10 were removed sequentially and cultured for V. cholerae. The bacterium was isolated from four house flies: two from the legs, and two others from their bodies minus legs and wings. In conclusion, house fly wings do not play an important role in mechanical transmission of bacteria suspended in a non-adhering liquid medium because of the low transfer rate of the bacteria to the wings and poor retention of bacteria on the wings during normal house fly activities.
    Matched MeSH terms: Vibrio cholerae/isolation & purification*
  12. Some interesting isolates from a diagnostic laboratory
    Lim PL
    J Clin Pathol, 1978 Mar;31(3):223-6.
    PMID: 641196
    Citrobacter koseri, Plesiomonas shigelloides, Edwardsiella tarda, Yersinia enterocolitica, Alkalescens dispar, Vibrio parahaemolyticus, and Vibrio alginolyticus were seven interesting microorganisms isolated recently in our diagnostic laboratory.
    Matched MeSH terms: Vibrio cholerae/isolation & purification
  13. Ultrasensitive pathogen detection with a rolling circle amplification-empowered multiplex electrochemical DNA sensor
    Yeap CSY, Chaibun T, Lee SY, Zhao B, Jan Y, La-O-Vorakiat C, et al.
    Chem Commun (Camb), 2021 Nov 16;57(91):12155-12158.
    PMID: 34726213 DOI: 10.1039/d1cc05181d
    We report a highly sensitive and selective multiplex assay by empowering an electrochemical DNA sensor with isothermal rolling circle amplification. The assay could simultaneously detect and discriminate three common entero-pathogens in a single reaction, with femtomolar sensitivity. It is useful for field- or resource-limited settings.
    Matched MeSH terms: Vibrio cholerae/isolation & purification*
  14. An ambient temperature stable and ready-to-use loop-mediated isothermal amplification assay for detection of toxigenic Vibrio cholerae in outbreak settings
    Engku Nur Syafirah EAR, Nurul Najian AB, Foo PC, Mohd Ali MR, Mohamed M, Yean CY
    Acta Trop, 2018 Jun;182:223-231.
    PMID: 29545156 DOI: 10.1016/j.actatropica.2018.03.004
    Cholera, caused by Vibrio cholerae is a foodborne disease that frequently reported in food and water related outbreak. Rapid diagnosis of cholera infection is important to avoid potential spread of disease. Among available diagnostic platforms, loop-mediated isothermal amplification (LAMP) is regarded as a potential diagnostic tool due to its rapidity, high sensitivity and specificity and independent of sophisticated thermalcycler. However, the current LAMP often requires multiple pipetting steps, hence is susceptible to cross contamination. Besides, the strict requirement of cold-chain during transportation and storage make its application in low resource settings to be inconvenient. To overcome these problems, the present study is aimed to develop an ambient-temperature-stable and ready-to-use LAMP assay for the detection of toxigenic Vibrio cholerae in low resource settings. A set of specific LAMP primers were designed and tested against 155 V. cholerae and non-V. cholerae strains. Analytical specifity showed that the developed LAMP assay detected 100% of pathogenic V. cholerae and did not amplified other tested bacterial strains. Upon testing against stool samples spiked with toxigenic V. cholerae outbreak isolates, the LAMP assay detected all of the spiked samples (n = 76/76, 100%), in contrast to the conventional PCR which amplified 77.6% (n = 59/76) of the tested specimens. In term of sensitivity, the LAMP assay was 100-fold more sensitive as compared to the conventional PCR method, with LOD of 10 fg per μL and 10 CFU per mL. Following lyophilisation with addition of lyoprotectants, the dry-reagent LAMP mix has an estimated shelf-life of 90.75 days at room temperature.
    Matched MeSH terms: Vibrio cholerae/isolation & purification*
  15. Microbiological status of live eel and processed fish products for export to Japan
    Rohaya MA, Chuink BH, Aniran K
    Southeast Asian J Trop Med Public Health, 1997;28 Suppl 1:77-9.
    PMID: 9656354
    Live eels and processed fish products from Malaysia are routinely checked for microbial pathogens before export to Japan. The eels and water from the ponds are screened for Vibrio cholerae and Salmonella spp, whereas the processed fish products are tested for microbial contamination (aerobic plate count), coliforms, E. coil and Vibrio cholerae. Results showed that live eels and water samples were negative for Vibrio cholerae but Salmonella spp were isolated occasionally. Various types of processed fish products had counts below 1.0 x 10(5) whilst coliforms, E. coli and Vibrio cholerae were absent. Records available showed that procedures involved in the production and transportation of live eel, preparation and processing of fish products have resulted in relatively safe food products.
    Matched MeSH terms: Vibrio cholerae/isolation & purification
  16. Detection and molecular characterization of the zot gene in Vibrio cholerae and V. alginolyticus isolates
    Raja N, Shamsudin MN, Somarny W, Rosli R, Rahim RA, Radu S
    Southeast Asian J Trop Med Public Health, 2001 Mar;32(1):100-4.
    PMID: 11485069
    A total of 11 Vibrio cholerae isolates from 1996-1998 outbreaks in Malaysia and 4 V. alginolyticus were analyzed. Isolates were characterized by polymerase chain reaction (PCR) and Southern hybridization for the presence of the gene encoding zonula occludens toxin (zot). Screening of zot gene by PCR revealed the presence of this gene in V. cholerae and V. alginolyticus. The zot gene from one V. cholerae Ogawa isolate that was cloned in a pCR 2.1 TOPO vector was sequenced. The sequences obtained were 99% homologous to the zot gene sequence from the Gene Bank.
    Matched MeSH terms: Vibrio cholerae/genetics*
  17. Occurrence of Vibrio cholerae 01 strains in Southeast Asia resistant to vibriostatic compound 0/129
    Mahalingam S, Cheong YM, Kan SK, Pang T
    Southeast Asian J Trop Med Public Health, 1993 Dec;24(4):779-80.
    PMID: 7939959
    Matched MeSH terms: Vibrio cholerae/metabolism*
  18. Construction and characterization of an auxotrophic ctxA mutant of O139 Vibrio cholerae
    Chan M, Cheong TG, Kurunathan S, Chandrika M, Ledon T, Fando R, et al.
    Microb Pathog, 2010 Nov;49(5):211-6.
    PMID: 20558271 DOI: 10.1016/j.micpath.2010.06.001
    Cholera caused by the O139 serogroup still remains a public health concern in certain regions of the world and the existing O1 vaccines do not cross-protect cholera caused by this serogroup. An aminolevulinic acid (ALA) auxotroph vaccine candidate against the O139 serogroup, designated as VCUSM2, was recently developed. It was found to be immunogenic in animal model studies but showed mild reactogenic effects due to the presence of two intact copies of Vibrio cholerae toxin (CTX) genetic element. In the present study we have modified the ctx operon by systematic allelic replacement methodology to produce a mutant strain, designated as VCUSM14. This strain has two copies of chromosomally integrated and mutated ctxA gene, encoding immunogenic but not toxic cholera toxin A subunit (CT-A). The amino acids arginine and glutamic acid at position 7th and 112th, respectively, in CT-A of VCUSM14 were substituted with lysine (R7K) and glutamine (E112Q), respectively. Two copies of the ace and zot genes present in the ctx operon were also deleted. Cholera toxin-ELISA using GM1 ganglioside showed that the both wild type CT and mutated CT were recognized by anti-CT polyclonal antibodies. VCUSM14 produced comparatively less amount of antigenic cholera toxin when compared to the VCUSM2 and Bengal wild type strain. VCUSM14 did not elicit fluid accumulation when inoculated into rabbit ileal loops at doses of 10(6) and 10(8) CFU. The colonization efficiency of VCUSM14 was one log lower than the parent strain, VCUSM2, which can be attributed to the ALA auxotrophy and less invasive properties of VCUSM14. VCUSM14, thus a non-reactogenic auxotrophic vaccine candidate against infection by O139 V. cholerae.
    Matched MeSH terms: Vibrio cholerae O139/genetics*; Vibrio cholerae O139/immunology*; Vibrio cholerae O139/metabolism; Vibrio cholerae O139/pathogenicity
  19. Fulltext Molecular evidence of cholera outbreak caused by a toxigenic Vibrio cholerae O1 El tor variant strain in Kelantan, Malaysia
    Ang GY, Yu CY, Balqis K, Elina HT, Azura H, Hani MH, et al.
    J Clin Microbiol, 2010 Nov;48(11):3963-9.
    PMID: 20826646 DOI: 10.1128/JCM.01086-10
    A total of 20 Vibrio cholerae isolates were recovered for investigation from a cholera outbreak in Kelantan, Malaysia, that occurred between November and December 2009. All isolates were biochemically characterized as V. cholerae serogroup O1 Ogawa of the El Tor biotype. They were found to be resistant to multiple antibiotics, including tetracycline, erythromycin, sulfamethoxazole-trimethoprim, streptomycin, penicillin G, and polymyxin B, with 35% of the isolates being resistant to ampicillin. All isolates were sensitive to ciprofloxacin, norfloxacin, chloramphenicol, gentamicin, and kanamycin. Multiplex PCR analysis confirmed the biochemical identification and revealed the presence of virulence genes, viz., ace, zot, and ctxA, in all of the isolates. Interestingly, the sequencing of the ctxB gene showed that the outbreak strain harbored the classical cholera toxin gene and therefore belongs to the newly assigned El Tor variant biotype. Clonal analysis by pulsed-field gel electrophoresis demonstrated that a single clone of a V. cholerae strain was responsible for this outbreak. Thus, we present the first molecular evidence that the toxigenic V. cholerae O1 El Tor variant has invaded Malaysia, highlighting the need for continuous monitoring to facilitate early interventions against any potential epidemic by this biotype.
    Matched MeSH terms: Vibrio cholerae O1/classification*; Vibrio cholerae O1/genetics; Vibrio cholerae O1/isolation & purification; Vibrio cholerae O1/pathogenicity*
  20. Ambient temperature detection of PCR amplicons with a novel sequence-specific nucleic acid lateral flow biosensor
    Ang GY, Yu CY, Yean CY
    Biosens Bioelectron, 2012 Oct-Dec;38(1):151-6.
    PMID: 22705404 DOI: 10.1016/j.bios.2012.05.019
    In the field of diagnostics, molecular amplification targeting unique genetic signature sequences has been widely used for rapid identification of infectious agents, which significantly aids physicians in determining the choice of treatment as well as providing important epidemiological data for surveillance and disease control assessment. We report the development of a rapid nucleic acid lateral flow biosensor (NALFB) in a dry-reagent strip format for the sequence-specific detection of single-stranded polymerase chain reaction (PCR) amplicons at ambient temperature (22-25°C). The NALFB was developed in combination with a linear-after-the-exponential PCR assay and the applicability of this biosensor was demonstrated through detection of the cholera toxin gene from diarrheal-causing toxigenic Vibrio cholerae. Amplification using the advanced asymmetric PCR boosts the production of fluorescein-labeled single-stranded amplicons, allowing capture probes immobilized on the NALFB to hybridize specifically with complementary targets in situ on the strip. Subsequent visual formation of red lines is achieved through the binding of conjugated gold nanoparticles to the fluorescein label of the captured amplicons. The visual detection limit observed with synthetic target DNA was 0.3 ng and 1 pg with pure genomic DNA. Evaluation of the NALFB with 164 strains of V. cholerae and non-V. cholerae bacteria recorded 100% for both sensitivity and specificity. The whole procedure of the low-cost NALFB, which is performed at ambient temperature, eliminates the need for preheated buffers or additional equipment, greatly simplifying the protocol for sequence-specific PCR amplicon analysis.
    Matched MeSH terms: Vibrio cholerae/genetics*; Vibrio cholerae/isolation & purification
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