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Cytotoxicity of Vibrio cholerae on Madin Darby Bovine Kidney cells

Haque QM, Mohamad NF, Helaluddin AB, Saeed M

Pak J Pharm Sci, 2010 Oct;23(4):393-7.
PMID: 20884452

The cytotoxicity of cell-free culture filtrates of 31 isolates of Vibrio cholerae O1 and O139, 5 reference strains and 26 clinical isolates, was tested on Madin Darby Bovine Kidney (MDBK) cells and Vero cells. The 3-[4,5-dimethylthiazol-2-y]-2, 5-diphenyltetrazolium bromide (MTT) test was used to detect the effect of the filtrates on the proliferation and viability of cultured cell populations. The filtrates were prepared from serial ten-fold dilutions of inoculated AKI and APW broth media with and without the addition of polymyxin B. The APW culture filtrates of both V. cholerae O1 and O139 with and without added polymyxin B showed greater toxicity to MDBK cells as compared to AKI filtrates. The cytotoxicity of AKI-grown V. cholerae O139 to MDBK cells was greater than that of V. cholerae O1 grown in the same medium. The cytotoxicity of APW filtrates on Vero cells was low and only noted when polymyxin was added to the medium.

Matched MeSH terms: Vibrio cholerae/drug effects; Vibrio cholerae/chemistry*
A signal-amplified electrochemical DNA biosensor incorporated with a colorimetric internal control for Vibrio cholerae detection using shelf-ready reagents

Low KF, Zain ZM, Yean CY

Biosens Bioelectron, 2017 Jan 15;87:256-263.
PMID: 27567251 DOI: 10.1016/j.bios.2016.08.064

A novel enzyme/nanoparticle-based DNA biosensing platform with dual colorimetric/electrochemical approach has been developed for the sequence-specific detection of the bacterium Vibrio cholerae, the causative agent of acute diarrheal disease in cholera. This assay platform exploits the use of shelf-stable and ready-to-use (shelf-ready) reagents to greatly simplify the bioanalysis procedures, allowing the assay platform to be more amenable to point-of-care applications. To assure maximum diagnosis reliability, an internal control (IC) capable of providing instant validation of results was incorporated into the assay. The microbial target, single-stranded DNA amplified with asymmetric PCR, was quantitatively detected via electrochemical stripping analysis of gold nanoparticle-loaded latex microspheres as a signal-amplified hybridization tag, while the incorporated IC was analyzed using a simplified horseradish peroxidase enzyme-based colorimetric scheme by simple visual observation of enzymatic color development. The platform showed excellent diagnostic sensitivity and specificity (100%) when challenged with 145 clinical isolate-spiked fecal specimens. The limits of detection were 0.5ng/ml of genomic DNA and 10 colony-forming units (CFU)/ml of bacterial cells with dynamic ranges of 0-100ng/ml (R(2)=0.992) and log10 (1-10(4) CFU/ml) (R(2)=0.9918), respectively. An accelerated stability test revealed that the assay reagents were stable at temperatures of 4-37°C, with an estimated ambient shelf life of 200 days. The versatility of the biosensing platform makes it easily adaptable for quantitative detection of other microbial pathogens.

Matched MeSH terms: Vibrio cholerae/genetics; Vibrio cholerae/isolation & purification*
Immunohistochemical, histological and ultrastructural evaluation of protection provided by cholera vaccine against V. cholerae O139

Murugaiah C, Nik Mohd Noor NZ, Al-Talib H, Mustafa S, Manickam R, Pattabhiraman L

Microb Pathog, 2020 Mar;140:103964.
PMID: 31904450 DOI: 10.1016/j.micpath.2020.103964

In our previous study, complete protection was observed in rabbit immunized with 1 × 1010 CFU of live attenuated VCUSM21P vaccine against challenge with 1 × 109 CFU Vibrio cholerae O139. In the present study, we investigated whether the vaccines can effectively protect immunized animals from any pathologic changes using histological, immunohistochemical and ultrastructural techniques. Severe pathology is evident in wild type injected ileum in non-immunized, showing extensive villous destruction, edema, necrosis and inflammation with infiltration of large numbers of inflammatory cells, extensive damage to the villi and microvilli with pore formation. Histology of ileum injected with wild type in immunized rabbit shows no significant pathological changes except for a few inflammatory cells in lamina propria with mild edema in mucosa and submucosa. immunohistochemical staining revealed O139 antigens of wild type are seen in the lamina propria of edematous villi, muscularis mucosa and submucosa with weak presence in the muscle coat in non-immunized rabbit after challenged with wild type in non-immunized rabbits, but in immunized rabbit localisation of the O139 LPS antigen is seen at the tips of the intact villi, within lamina propria and muscularis mucosa only. These observations suggest that the vaccine can effectively protect animals from any pathologic changes and eliminate V. cholerae O139 from the immunized animals.

Matched MeSH terms: Vibrio cholerae O139/genetics; Vibrio cholerae O139/immunology*
A microdevice for rapid, monoplex and colorimetric detection of foodborne pathogens using a centrifugal microfluidic platform

Sayad A, Ibrahim F, Mukim Uddin S, Cho J, Madou M, Thong KL

Biosens Bioelectron, 2018 Feb 15;100:96-104.
PMID: 28869845 DOI: 10.1016/j.bios.2017.08.060

Outbreaks of foodborne diseases have become a global health concern; hence, many improvements and developments have been made to reduce the risk of food contamination. We developed a centrifugal microfluidic automatic wireless endpoint detection system integrated with loop mediated isothermal amplification (LAMP) for monoplex pathogen detection. Six identical sets were designed on the microfluidic compact disc (CD) to perform 30 genetic analyses of three different species of foodborne pathogens. The consecutive loading, mixing, and aliquoting of the LAMP primers/reagents and DNA sample solutions were accomplished using an optimized square-wave microchannel, metering chambers and revulsion per minute (RPM) control. We tested 24 strains of pathogenic bacteria (Escherichia coli, Salmonella spp and Vibrio cholerae), with 8 strains of each bacterium, and performed DNA amplification on the microfluidic CD for 60min. Then, the amplicons of the LAMP reaction were detected using the calcein colorimetric method and further analysed via the developed electronic system interfaced with Bluetooth wireless technology to transmit the results to a smartphone. The system showed a limit of detection (LOD) of 3 × 10-5ngμL-1 DNA by analysing the colour change when tested with chicken meat spiked with the three pathogenic bacteria. Since the entire process was performed in a fully automated way and was easy to use, our microdevice is suitable for point-of-care (POC) testing with high simplicity, providing affordability and accessibility even to poor, resource-limited settings.

Matched MeSH terms: Vibrio cholerae/genetics; Vibrio cholerae/isolation & purification*
Construction and evaluation of a O139 Vibrio cholerae vaccine candidate based on a hemA gene mutation

Ravichandran M, Ali SA, Rashid NH, Kurunathan S, Yean CY, Ting LC, et al.

Vaccine, 2006 May 1;24(18):3750-61.
PMID: 16102875

In this paper, we describe the development of VCUSM2, a live metabolic auxotroph of Vibrio cholerae O139. Auxotrophy was achieved by mutating a house keeping gene, hemA, that encodes for glutamyl-tRNA reductase, an important enzyme in the C5 pathway for delta-aminolevulenic acid (ALA) biosynthesis, which renders this strain dependent on exogenous ALA for survival. Experiments using the infant mouse and adult rabbit models show that VCUSM2 is a good colonizer of the small intestine and elicits greater than a four-fold rise in vibriocidal antibodies in vaccinated rabbits. Rabbits vaccinated with VCUSM2 were fully protected against subsequent challenge with 1 x 10(11) CFU of the virulent wild type (WT) strain. Experiments using ligated ileal loops of rabbits show that VCUSM2 is 2.5-fold less toxic at the dose of 1 x 10(6) CFU compared to the WT strain. Shedding of VCUSM2 in rabbits were found to occur for no longer than 4 days and its maximum survival rate in environmental waters is 8 days compared to the greater than 20 days for the WT strain. VCUSM2 is thus a potential vaccine candidate against infection by V. cholerae O139.

Matched MeSH terms: Vibrio cholerae O139/genetics; Vibrio cholerae O139/growth & development; Vibrio cholerae O139/immunology*
Fulltext Molecular quantitation and characterization of Vibrio cholerae from different seafood obtained from wetmarket and supermarket

Vengadesh, L., Son, R., Yoke-Kqueen, C.

International Food Research Journal, 2012;19(1):45-50.
MyJurnal

Vibrio cholerae still represents a significant threat to human health worldwide despite the advances in hygiene, consumer knowledge, food treatment and food processing. In Malaysia, statistics in year 2009 have shown that among the food and water borne diseases, food poisoning has the highest incidence rate of 36.17 per 100,000 populations and with a mortality rate of 0.01 per 100,000 populations. In this study, 22 seafood samples comprising of fish, squid, crustacean and mollusks purchased from wet market and supermarket were analyzed. The Most Probable Number (MPN) and real time PCR was used to enumerate the Vibrio cholerae in seafood sample. The results showed that MPN-real time PCR of the samples from wet market had a maximum of >1100 MPN/g compare to 93 MPN/g enumerated from the MPN plate. The MPN-real time PCR in the samples from supermarket indicated 290 MPN/g as compared to 240 MPN/g enumerated from the MPN plate. The standard curves showed that there was a good linear correlation between the Ct values. The minimum level of detection of Vibrio cholerae standard DNA at targeted gene was 3 x 10-5 ng/μl.

Matched MeSH terms: Vibrio cholerae
Fulltext Antibiotic resistance and biosafety of Vibrio cholerae and Vibrio parahaemolyticus from freshwater fish at retail level

International Food Research Journal, 2011;18(4):1523-1530.
MyJurnal

A total of 49 isolates of V. parahaemolyticus and 8 isolates of V. cholerae isolated from freshwater fish of patin (Pangasius hypopthalmus) and red tilapia (Oreochromis sp.) were purchased from different retail level in Selangor, Malaysia. All of the isolates showed a multiple resistances towards all 15 antibiotics tested. Some of the isolates show a high resistance to different antibiotics including bacitracin, vancomycin, tetracycline, furazididone, cephalothin and erythromycin. However, both species was susceptible towards imipenem. Overall antibiotics resistance patterns of all isolates were resistant from 2 to 14 resistance patterns with multiple antibiotic resistance (MAR) index ranging from 0.13 to 0.93 respectively. As the results obtained in the dendrogram produced from both species had indicates that these antibiotics were intensively used whether in the aquaculture farm through feeds during culture or at the hatchery production of seed. Thus, this study will provides an essential information of the MAR index and also the clustering analysis in order to determine the biosafety of Vibrio spp. in freshwater aquaculture fish sold at different retail level in Malaysia.

Matched MeSH terms: Vibrio cholerae
Fulltext Molecular characterization of clinical isolate of Vibrio cholerae isolated from outbreaks cases in Malaysia

Patrick, G. B., Nishibuchi, M., Tunung, R., Son, R.

International Food Research Journal, 2012;19(3):1267-1274.
MyJurnal

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: Vibrio cholerae
Fulltext Detection of Vibrio cholerae in street food (satar and otak-otak) by Loop-Mediated Isothermal Amplification (LAMP), multiplex Polymerase Chain Reaction (mPCR) and plating methods

Tang, J-Y-H., Farhana Sakinah, M.R., Nakaguchi, Y., Nishibuchi, M., Chai, L-C., New, C.Y., et al.

Food Research, 2018;2(5):447-452.
MyJurnal

This goal of this study was to investigate the presence of Vibrio cholerae in street food,
namely satar and otak-otak, using Loop-Mediated Isothermal Amplification (LAMP),
multiplex Polymerase Chain Reaction (mPCR) and conventional plating on Thiosulphate
Citrate Bile-Salt Sucrose (TCBS) agar methods. A total of 78 satar and 35 otak-otak were
purchased from different districts of Terengganu (Besut, Setiu, Kuala Terengganu and
Kemaman). V. cholerae was found in satar with LAMP (10.3%), mPCR (10.3%) and
plating (0%). No V. cholerae was found in otak-otak using the three methods. This might
be due to V. cholerae able to survive in satar after grilling due to its thickness which may
contribute to undercooking. This study concluded that low presence of V. cholerae in satar
and otak-otak can be detected by molecular methods but not the conventional plating
method. LAMP assay is a useful tool for rapid detection of pathogens in food due to its
simplicity, highly sensitive and visual interpretation capability. Though the prevalence of
V. cholerae was low in the samples, proper handling of this food will help in reducing the
risk of acquiring infection from V. cholerae in contaminated samples.

Matched MeSH terms: Vibrio cholerae
Pengesahan bagi mengesan bakteria patogen makanan menggunakan teknologi mikroaturan DNA

SAWEI J, NORRAKIAH ABDULLAH SANI, AMINAH ABDULLAH, SAHILAH ABD. MUTALIB

Sains Malaysiana, 2013;42:1715-1720.

Kajian ini dijalankan untuk mengesahkan kemampuan teknologi DNA mikroaturan cip gen OliproTM FoodPATH bagi mengesan bakteria patogen makanan. Sebanyak 9 jenis DNA bakteria patogen makanan telah digunakan iaitu Bacillus cereus, Escherichia coli O157:H7, Staphylococcus aureus, Vibrio cholerae, Vibrio parahaemolyticus, Listeria monocytogenes, Salmonella spp., Shigella spp. dan Campylobacter spp. Sebanyak 36 kombinasi templat DNA bakteria patogen makanan telah digunakan. Pengesahan bagi mengesan bakteria patogen makanan dilakukan dengan menggunakan kaedah reaksi berantai polimerase (PCR) dan penghibridan Southern-blotting di atas cip gen untuk mengesahkan kemampuannya. Keputusan daripada analisis hibridasi di atas cip gen telah dibandingkan dengan hasil gel elektroforesis 2.0% (w/v). Lima saringan diperlukan untuk menghabiskan 36 kombinasi templat DNA bakteria patogen makanan. Setiap saringan, satu cip gen telah digunakan sebagai kawalan negatif tidak diinokulasikan dengan sebarang kombinasi DNA bakteria patogen makanan. Daripada hasil kajian, didapati bahawa semua kombinasi templat DNA bakteria patogen makanan telah dapat dikesan. Cip yang digunakan sebagai kawalan negatif tidak menunjukkan kehadiran DNA. Oleh itu, daripada kajian ini cip gen OliproTM FoodPATH didapati memberikan keputusan yang lebih baik berbanding dengan 2.0% (w/v) gel elektroforesis.

Matched MeSH terms: Vibrio cholerae
Health problems of foreign workers - Microbiological investigations

Ngeow YF, Ng KP, Puthucheary SD, Lam SK

JUMMEC, 2002;7:67-69.

Foreign workers in Malaysia are screened for certain infectious diseases prior to their entry to the country but some escape medical screening and others acquire infection during their stay in the country. The Faculty of Medicine, University of Malaya was commissioned to study the impact of foreign labour on the local health system and, as part of the investigations, 584 foreign workers attending local outpatient clinics were examined for serological evidence of syphilis, HIV infection, viral hepatitis B, C and E, as well as for enteric infections by Salmonella, Shigella and Vibrio cholerae. The results showed that apart from viral hepatitis E, the prevalence rates of the infections looked for were not notably higher than those for the general Malaysian population. The seroprevalence rates obtained were 2.6% for syphilis, 0.2% HIV infection, 3.8% viral hepatitis B, 1.0% viral hepatitis C, 14.4% viral hepatitis E. The detection of HEV IgM in 7.7% of the workers screened indicates that these infections could have been acquired during their stay in Malaysia.

Matched MeSH terms: Vibrio cholerae
Electrochemical genosensor assay using lyophilized gold nanoparticles/latex microsphere label for detection of Vibrio cholerae

Liew PS, Lertanantawong B, Lee SY, Manickam R, Lee YH, Surareungchai W

Talanta, 2015 Jul 1;139:167-73.
PMID: 25882423 DOI: 10.1016/j.talanta.2015.02.054

Vibrio cholerae is a Gram-negative bacterium that causes cholera, a diarrheal disease. Cholera is widespread in poor, under-developed or disaster-hit countries that have poor water sanitation. Hence, a rapid detection method for V. cholerae in the field under these resource-limited settings is required. In this paper, we describe the development of an electrochemical genosensor assay using lyophilized gold nanoparticles/latex microsphere (AuNPs-PSA) reporter label. The reporter label mixture was prepared by lyophilization of AuNPs-PSA-avidin conjugate with different types of stabilizers. The best stabilizer was 5% sorbitol, which was able to preserve the dried conjugate for up to 30 days. Three methods of DNA hybridization were compared and the one-step sandwich hybridization method was chosen as it was fastest and highly specific. The performance of the assay using the lyophilized reagents was comparable to the wet form for detection of 1aM to 1fM of linear target DNA. The assay was highly specific for V. cholerae, with a detection limit of 1fM of PCR products. The ability of the sensor is to detect LAMP products as low as 50ngµl(-1). The novel lyophilized AuNPs-PSA-avidin reporter label with electrochemical genosensor detection could facilitate the rapid on-site detection of V. cholerae.

Matched MeSH terms: Vibrio cholerae/genetics*
Fulltext Cholera outbreak by Sea Gypsies in Sabah, Malaysia: A challenge in North Borneo

Jikal M, Riduan T, Aarifin R, Jeffree MS, Ahmed K

Int J Infect Dis, 2019 Jun;83:83-85.
PMID: 30986543 DOI: 10.1016/j.ijid.2019.04.008

OBJECTIVES: In this study we investigated an outbreak of Vibrio cholera O1 Ogawa serotype, occurred during December 2014 in Kudat district, situated in Sabah state of the Malaysian part of Borneo.
METHODS: Active case detection and contact tracing were done at respective localities by house to house survey. Passive case detection was done among acute gastroenteritis patients attended at various health facilities. To determine the source, samples from food, water and environment were taken. A case control study was also done to determine the risk factors.
RESULTS: A total of 44 symptomatic and 34 asymptomatic cases from 19 localities were investigated. 39 cases were detected through passive case detection. Median age of cases was 23 years. All cases belonged to serogroup O1 and Ogawa serotype. The epidemiological investigation of time, place, and person identified that V. cholerae cross-transmission might have occurred in two fish markets and the fish-loading port. Circumstantial evidences indicated that cholera was possibly transmitted through contaminated sea foods.
CONCLUSIONS: We concluded that the life-style of Sea Gypsies is a challenge in cholera control; therefore vaccination might be an effective way to mitigate cholera in an outbreak prone area like Kudat.

Matched MeSH terms: Vibrio cholerae*
Laboratory study of Vibrio cholerae O1 survival on three types of boiled rice (Oryza sativa L.) held at room temperature

Huat JT, Leong YK, Lian HH

J Food Prot, 2008 Dec;71(12):2453-9.
PMID: 19244898

This study examined whether the survival of Vibrio cholerae O1 on contaminated cooked rice was influenced by the type of rice. Vibrios survived unchanged on clumps of glutinous white rice (wet, grains adhered) held at room temperature for 24 h. On nonglutinous white rice (slightly moist, grains separate), 30% viable vibrios remained at 24 h. On nonglutinous brown rice (moist, separate, covered with a mucus-like substance), the number of vibrios increased 2.7-fold at 24 h. Survival rates of vibrios on the surfaces of a row of five cooked rice grains after 2 h of exposure at room temperature were 86, 29, 12, and 4% for glutinous rice, white rice, and the endosperm and pericarp of brown rice, respectively. (Each boiled brown rice grain surface was partly pericarp and partly endosperm, which became exposed by a rupture of the pericarp.) Covering each inoculated grain with a similar cooked rice grain surface increased the corresponding figures to 93, 99, 60, and 94%. Scanning electron microscopy revealed that each type of cooked grain surface possessed a distinct microtopography. For example, the surfaces of glutinous rice grains consisted of separated overlapping strips with many holes, while the pericarps of brown rice were flat interspersed with small pits. In conclusion, each type of boiled rice produced a distinct survival pattern of V. cholerae O1 caused by both the distinct gross features and the fine surface characteristics of the rice. The significance of this finding is that the type of rice consumed can be a factor in cholera transmission by contaminated rice.

Matched MeSH terms: Vibrio cholerae O1/growth & development*
Oral vaccination with Lactococcus lactis expressing the Vibrio cholerae Wzm protein to enhance mucosal and systemic immunity

Zamri HF, Shamsudin MN, Rahim RA, Neela V

Vaccine, 2012 May 2;30(21):3231-8.
PMID: 22426330 DOI: 10.1016/j.vaccine.2012.02.012

A gene associated with lipopolysaccharide (LPS) transport was cloned from a local clinical Vibrio cholerae O1 strain of the Ogawa serotype by using the Lactococcus lactis nisin-controlled expression (NICE) system. The V. cholerae wzm gene, which codes for an integral membrane transporter protein, was expressed and targeted to the cytoplasmic membrane, and was crudely isolated through simple centrifugation and SDS solubilization. To examine seroreactivity of this construct, rabbits were orally fed with 10(9) cfu/ml of live, recombinant L. lactis carrying the wzm gene, induced with nisin prior to administration. Recombinant plasmids were retrieved from L. lactis cultured directly from stool samples of inoculated rabbits. Reverse-transcriptase PCR of wzm using the retrieved plasmids confirmed transcription of this gene, indicating viability and stability of the recombinants in vivo. The L. lactis-Wzm construct elicited substantial levels of IgG and sIgA, and challenge with virulent V. cholerae O1 evoked severe diarrhoea in the naive, non-immunised control group, but not in those fed with either recombinant or non-recombinant L. lactis. Oral administration with recombinant L. lactis expressing the V. cholerae wzm gene increases both systemic and mucosal immunity, whereas L. lactis itself appears capable of protecting against the diarrhoeal symptoms caused by V. cholerae. Wzm is a conserved membrane protein associated with the LPS endotoxin, and together with the food-grade L. lactis, represent an attractive target for the development of a safer, live anti-infective therapy against V. cholerae.

Matched MeSH terms: Vibrio cholerae O1/genetics; Vibrio cholerae O1/immunology*
Antimicrobial activity of the crude extract of Piper sarmentosum against methicilin-resistant Staphylococcus aureus (MRSA), Escherichia coli, Vibrio cholera and Streptococcus pneumoniae

Fernandez L, Daruliza K, Sudhakaran S, Jegathambigai R

Eur Rev Med Pharmacol Sci, 2012 Jul;16 Suppl 3:105-11.
PMID: 22957424

The emergence of novel diseases caused by microbial pathogens and the undesirable side effects of certain antibiotics has been a recent dilemma in the medical arena. Consequently, it has stirred the discovery of many naturally occurring agents which could possibly provide important ramifications against various pharmacological targets and to combat various ailments. The main aim of the present study was to determine the antimicrobial activity of the crude methanolic extract of Piper (P.) sarmentosum against methicillin resistant Staphylococcus aureus (MRSA), Escherichia coli, Vibrio cholera and Streptococcus pneumoniae.

Matched MeSH terms: Vibrio cholerae/drug effects
Fulltext Antibacterial activities of a new brominated diterpene from Borneon Laurencia spp

Vairappan CS, Ishii T, Lee TK, Suzuki M, Zhaoqi Z

Mar Drugs, 2010;8(6):1743-9.
PMID: 20631866 DOI: 10.3390/md8061743

In our continuous interest to study the diversity of halogenated metabolites of Malaysian species of the red algal genus Laurencia, we examined the chemical composition of five populations of unrecorded Laurencia sp. A new brominated diterpene, 10-acetoxyangasiol (1), and four other known metabolites, aplysidiol (2), cupalaurenol (3), 1-methyl-2,3,5-tribromoindole (4), and chamigrane epoxide (5), were isolated and identified. Isolated metabolites exhibited potent antibacterial activities against clinical bacteria, Staphylococcus aureus, Staphylococcus sp., Streptococcus pyogenes, Salmonella sp. and Vibrio cholerae.

Matched MeSH terms: Vibrio cholerae/drug effects
Enzyme-linked amperometric electrochemical genosensor assay for the detection of PCR amplicons on a streptavidin-treated screen-printed carbon electrode

Yean CY, Kamarudin B, Ozkan DA, Yin LS, Lalitha P, Ismail A, et al.

Anal Chem, 2008 Apr 15;80(8):2774-9.
PMID: 18311943 DOI: 10.1021/ac702333x

A general purpose enzyme-based amperometric electrochemical genosensor assay was developed wherein polymerase chain reaction (PCR) amplicons labeled with both biotin and fluorescein were detected with peroxidase-conjugated antifluorescein antibody on a screen-printed carbon electrode (SPCE). As a proof of principle, the response selectivity of the genosensor was evaluated using PCR amplicons derived from lolB gene of Vibrio cholerae. Factors affecting immobilization, hybridization, and nonspecific binding were optimized to maximize sensitivity and reduce assay time. On the basis of the background amperometry signals obtained from nonspecific organisms and positive signals obtained from known V. cholerae, a threshold point of 4.20 microA signal was determined as positive. Under the optimum conditions, the limit of detection (LOD) of the assay was 10 CFU/mL of V. cholerae. The overall precision of this assay was good, with the coefficient of variation (CV) being 3.7% using SPCE and intermittent pulse amperometry (IPA) as an electrochemical technique. The assay is sensitive, safe, and cost-effective when compared to conventional agarose gel electrophoresis, real-time PCR, and other enzyme-linked assays for the detection of PCR amplicons. Furthermore, the use of a hand-held portable reader makes it suitable for use in the field.

Matched MeSH terms: Vibrio cholerae/genetics
New anti-bacterial halogenated tricyclic sesquiterpenes from Bornean Laurencia majuscula (Harvey) Lucas

Kamada T, Phan CS, Vairappan CS

Nat Prod Res, 2019 Feb;33(4):464-471.
PMID: 29092618 DOI: 10.1080/14786419.2017.1396593

Three new halogenated tricyclic sesquiterpenes, omphalaurediol (1), rhodolaurenones B (2) and C (3) were isolated together with nine known haloganated sesquiterpenes such as rhodolaurenone A (4), rhodolaureol (5), isorhodolaureol (6), (-)-laurencenone D (7), elatol (8), (+)-deschloroelatol (9), cartilagineol (10), (+)-laurencenone B (11) and 2-chloro-3-hydroxy-α-chamigren-9-one (12) from a population of Bornean red algae Laurencia majuscula. The structures of three new metabolites were determined based on their spectroscopic data (IR, 1D and 2D NMR, and MS). These compounds showed antibacterial activity against three human pathogenic bacteria (Escherichia coli, Salmonella typhi and Vibrio cholera).

Matched MeSH terms: Vibrio cholerae/drug effects
Development of a dry-reagent-based nucleic acid-sensing platform by coupling thermostabilised LATE-PCR assay to an oligonucleotide-modified lateral flow biosensor

Ang GY, Yu CY, Chan KG, Singh KK, Chan Yean Y

J Microbiol Methods, 2015 Nov;118:99-105.
PMID: 26342435 DOI: 10.1016/j.mimet.2015.08.024

In this study, we report for the first time the development of a dry-reagent-based nucleic acid-sensing platform by combining a thermostabilised linear-after-the-exponential (LATE)-PCR assay with a one-step, hybridisation-based nucleic acid lateral flow biosensor. The nucleic acid-sensing platform was designed to overcome the need for stringent temperature control during transportation or storage of reagents and reduces the dependency on skilled personnel by decreasing the overall assay complexity and hands-on time. The platform was developed using toxigenic Vibrio cholerae as the model organism due to the bacterium's propensity to cause epidemic and pandemic cholera. The biosensor generates result which can be visualised with the naked eyes and the limit of detection was found to be 1pg of pure genomic DNA and 10CFU/ml of toxigenic V. cholerae. The dry-reagent-based nucleic acid-sensing platform was challenged with 95 toxigenic V. cholerae, 7 non-toxigenic V. cholerae and 66 other bacterial strains in spiked stool sample and complete agreement was observed when the results were compared to that of monosialoganglioside (GM1)-ELISA. Heat-stability of the thermostabilised LATE-PCR reaction mixes at different storage temperatures (4-56°C) was investigated for up to 90days. The dry-reagent-based genosensing platform with ready-to-use assay components provides an alternative method for sequence-specific detection of nucleic acid without any cold chain restriction that is associated with conventional molecular amplification techniques.

Matched MeSH terms: Vibrio cholerae

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