Street food is popular in Asia due to its availability, low price and good taste. The safety of
street food has been always questionable due to its poor handling which probably leads to
microbial contamination. The objective of this study was to determine the surviving quantities
of V. parahaemolyticus under various conditions in street-vended food, namely satar and otakotak
after anticipated cross-contamination to support policy and regulatory documents. The
satar and otak-otak were prepared from minced and unminced fish flesh, respectively, together
with other ingredients. Each satar and otak-otak were prepared with 0, 0.5, 1.5 and 3% of
sodium chloride (NaCl), respectively. V. parahaemolyticus inoculum at approximately 8.66 log
CFU/ml were inoculated into the samples and incubated for up to 6 h. Samples were taken at 0,
1, 3 and 6 h for enumeration of V. parahaemolyticus using spread plate method on Thiosulphate
Citrate Bile Salts Sucrose (TCBS) agar. For control samples, V. parahaemolyticus was not
immediately inactivated in distilled water even though significant better survivability was
observed in Phosphate Buffer Saline (PBS). The numbers of V. parahaemolyticus was found
to decrease by varying amounts based on the salt content and duration of holding. However,
significant amounts survived to indicate potential risk.
This study aimed to determine the biofilm formation ability by Salmonella Typhi on cucumber, mango and guava surface, as well as to determine the relationship between time contact and biofilm formation. Crystal violet assay was performed to quantify the biofilm formation based on the value of optical density at 570 nm of the destaining crystal violet at the specific interval time. The result showed that the attachment of the bacterial cells on the fresh produce surface increased with the contact time. The readings of OD570at time 12 h for cucumber, mango and guava surfaces were 0.824, 0.683 and 0.598, respectively, indicating that the biofilm formation by Salmonella Typhi on different fresh produce surface varied with time. Since the result showed that Salmonella Typhi formed biofilm on fresh produce surfaces, hygienic practice from farm to fork including handling, processing, distribution and storage of the fresh produce should be of concern.
Salmonellosis is an important public health problem and causes large economic losses in the poultry industry. The emergence of molecular technology has opened various possibilities for constructing tailor-made proteins, particularly protein E from bacteriophage PhiX174 for the
production of bacterial ghosts (BGs) applied in vaccines purposes. In the present study, the plamdaPRcI-Elysis plasmid carrying the PhiX174 lysis gene E and thermo-sensitive lamda PR-cl857 regulatory system was constructed. Two Salmonella Enteritidis (SE-2 and SE- 4) and one Salmonella Typhimurium (ST-4) isolates were able to uptake the lysis plasmid via electrotransformation. Generation of ghosts was enhanced by increasing the incubation temperature up to 42˚C. Cell viability of SE-2, SE-4 and ST-4 decreased ranging in log 2.7 to log 4.1 cycles after lysis induction. Moreover, SE-2 and SE-4 exhibited the earliest reduction of CFU after 3 h of incubation. Our results may provide a promising avenue for the development of Salmonella BGs vaccines.
Bacteriophages are the viruses of bacteria and are widely distributed in the biosphere, exhibiting
dramatic manifestations both in liquid cultures and on solid media. In this study, bacteriophages
were isolated from different types of food (beef, chicken meats, cucumber, lettuce, clam,
cockles and shrimp) and sewage samples using 6 reference pathogen strains (Salmonella
Enteritidis, Salmonella Typhimurium, Campylobacter jejuni, Vibrio parahaemolyticus, Listeria
monocytogenes and Escherichia coli). A total of 29 bacteriophage isolates were obtained and
further examined for titer via agar overlay assay. The titers were determined within the range
of 108
to 1011 PFU/mL. Our results showed that diverse of bacteriophages are naturally present
in a variety of foods.
Bacteriophages are ubiquitous in our world, mainly in the oceans, soil, the water and food
we consume. They can be used efficiently in modern biotechnology, as well as alternatives
to antibiotics for many antibiotic resistant bacterial strains. Phages can be used as vehicles
for vaccines both DNA and protein, for the detection of pathogenic bacterial strain, as biocontrol
agents in agriculture and food industry. This review outlines the properties as well
as the influence of different external physical and chemical factors like temperature and
acidity on phage persistence. A better understanding of the complex problem of phage
sensitivity to external factors may be useful for other researchers working with phages.
Furthermore, the applications of bacteriophages were described in this paper as well.
According to the World Health Organisation (WHO), globally 600 million people suffer
from food-borne diseases (FBD), and 420,000 people die as a result. The European Food
Safety Authority (EFSA) has stated that FBD are linked to the food industry, with the
most common means of transmission being due to poor food handling and hygiene by
food handlers working in the food industry. The aim of this research was to investigate the
effectiveness of mandatory food handler training programmes (FHTP) to prevent FBD in
Malaysia and Ireland. To do this, the FHTP existing in Malaysia and Ireland were
analysed, in addition to the legislation they fall under in each respective country.
Effectiveness was determined by investigating the level of food safety knowledge (FSK)
and food safety practices (FSP) of food handlers in Malaysia and Ireland. A systematic
literature review (SLR) and a narrative literature review (NLR) were conducted for this
research. The SLR was based on the PRISMA diagram, using the Confidence in the
Evidence from Reviews of Qualitative research (CERQual) approach to evaluate the
studies used for this research. A total of 8 Malaysian studies and 1 Irish study were used to
determine the level of FSK and FSP of food handlers in each respective country, to
examine the effectiveness of FHTP. The results of the studies used for this research have
depicted overall good FSP and FSK of food handlers in Malaysia and Ireland; yet trends
continue to show that food handlers are one of the biggest contributors to FBD,
demonstrating that FHTP are not effective in preventing FBD. The findings from this
research highlights that although these trainings can be an effective tool to prevent FBD, if
they are not executed correctly, food handlers will continue to contribute to FBD.
Recently, many cases related to viral gastroenteritis outbreaks have been reported all over the world. Noroviruses are found to be leading as the major cause of outbreaks of acute gastroenteritis. Patients with the acute gastroenteritis normally found to be positive with norovirus when stools and vomit were analyzed. This paper reviews various activities and previous reports that describe norovirus contaminated in various food matrixes and relationship between food handlers. Lately, a numbers of norovirus outbreaks have been reported which are involved fresh produce (such as vegetables, fruits), shellfish and prepared food. Food produces by infected food handlers may therefore easily contaminated. In addition, food that required much handling and have been eaten without heat treatment gave the high risk for getting foodborne illnesses. The standard method for detection of norovirus has already been available for stool samples. However, only few methods for detection of norovirus in food samples have been developed until now.
Presence of Norovirus in food can cause viral gasteroenteritis. Recently, lots of reports relating to Norovirus in food have been published. Special attention must be paid to the raw foods as they are not subjected to further heat treatment. In this study, pegaga, kesum, tauge and ulam raja (popular salad vegetables in Malaysia) were investigated for Norovirus. A total of 32 samples from each type of salad vegetables were purchased from local market and analyzed using One-step RT-PCR (Reverse Transcriptase-Polymerase Chain Reaction) for both genogroups namely Norovirus Genogroup I and Genogroup II. Results showed that tauge had the highest contamination with Norovirus Genogroup I (15.6%) comparing to pegaga (9.4%), kesum (12.5%)
and ulam raja (0%). Samples were free from Norovirus Genogroup II. The study showed that raw vegetables are high-risk foods and can be contaminated with Norovirus.
Food labeling in accordance with Novel Food Regulation has been enforced in the European Community since 1997 with a series of updated legislations namely, EC/258/97, EC/1139/98, EC/49/2000, EC/50/2000 and EC/1829/2003. Guidelines and labeling regulations for the use of GMOs materials in food and feed products has also been introduced in Malaysia and Vietnam. Therefore, the demand for the establishment and development of a robust and rapid operation procedure for GMO detection has increased recently in both countries. The procedure of GMO detection emphasizes not only on detection tests but also on confirmation assays. This study employed PCR technology for detection and direct DNA sequencing for confirmation procedures respectively. The results demonstrated for the first time the presence of GM plants with glyphosate-resistant trait led by the control of P35S promoter and NOS terminator in either Malaysian or Vietnamese feed with high frequency (20 positive samples out of 24 analyzed samples). The P35S promoter, EPSPS gene and NOS terminator sequences obtained showed some mutations on single-stranded and double-stranded targeted sequences caused by single nucleotide insertion or single nucleotide changes. These results reinforce the need for development of detection procedures to comply with food/feed labeling system.
The ability to detect the presence of transgenes in crop-derived foods depends on the quantity and quality of DNA obtained from a product to be analyzed. The efficiency of DNA extraction protocols differs due to the nature of each food product. In this paper, we described two main DNA extraction protocols and their modifications that have been applied and evaluated for DNA extraction from raw and processed food as well as animal feed. The yield and quality for five categories of food and feed samples namely, raw soybean, raw maize, animal feed, smooth tofu and soymilk are discussed. The statistical interaction analyses showed that the cetyltrimethyl ammonium bromide (CTAB) method was proven to be the best method to extract DNA from raw soybean, maize and animal feed samples which not only obtained high DNA yield of 32.7, 28.4 and 33.4 ng DNA/mg sample respectively, but also produced high quality DNA with the absorbance A260/A280 ratio of 1.9, 1.9 and 2.0, respectively. These DNA were suitable for PCR amplification which produced a 164 bp DNA fragment of the lectin gene from soybean, and a 277 bp DNA fragment of the zein gene from maize. In the processed food category, the Wizard isolation method was found to be the best for the extraction of DNA from smooth tofu and soymilk with the yield of 13.2 and 3.4 ng DNA/mg sample, and the quality of the DNA at the absorbance A260/A280 ratio ranged from 1.9 to 1.7. These DNA were successfully amplified using primers specific to the lectin gene of soybean.
The purpose of this study was to investigate the biosafety of Vibrio parahaemolyticus in raw salad vegetables at wet market and supermarket in Malaysia. A combination of Most Probable Number - Polymerase Chain Reaction (MPN-PCR) method was applied to detect the presence of V. parahaemolyticus and to enumerate their density in the food samples. The study analyzed 276 samples of common vegetables eaten raw in Malaysia (Wild cosmos = 8; Japanese parsley = 21; Cabbage = 30; Lettuce = 16; Indian pennywort = 17; Carrot = 31; Sweet potato = 29; Tomato = 38; Cucumber = 28; Four winged bean = 26; Long bean = 32). The samples were purchased from two supermarkets (A and B) and two wet markets (C and D). The occurrence of V. parahaemolyticus detected was 20.65%, with higher frequency of V. parahaemolyticus in vegetables obtained from wet markets (Wet market C = 27.27%Wet Market D = 32.05%) compared to supermarkets (Supermarket A = 1.64%; Supermarket B = 16.67%). V. parahaemolyticus was most prevalent in Indian pennywort (41.18%). The density of V. parahaemolyticus in all the samples ranged from <3 up to >2400 MPN/g, mostly <3 MPN/g concentration. Raw vegetables from wet markets contained higher levels of V. parahaemolyticus compared to supermarkets. V. parahaemolyticus were present in raw vegetables although in low numbers. The results suggest that raw vegetables act as a transmission route for V. parahaemolyticus. This study will be the first biosafety assessment of V. parahaemolyticus in raw vegetables in Malaysia.
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.
Salmonella enterica is one of the major causes of bacterial foodborne infection. The aims of this study were to determine the antibiotic resistance and the genetic diversity of Salmonella enterica isolated from street foods and clinical samples and to understand the correlation between the prevalence of serovars and genotypes with their source (street food and clinical samples) and geographic origin (Negeri Sembilan, Malacca and Selangor in Peninsular Malaysia). The enterobacterial repetitive intergenic consensus (ERIC) PCR analysis distinguished the Salmonella isolates into 19 ERIC types, with one untypable isolate. Dendrograms were specifically constructed for the S. Biafra and S. Typhi isolates. Identical or very similar ERIC types among the S. Biafra isolates from street food samples indicate transmission of the S. Biafra among the street foods, as well as possible cross-contamination of the street foods. In addition, the identical or very similar ERIC types among the S. Typhi isolates from human samples examined suggest possible similarity in their source of infection. All the twenty four isolates were resistant to rifampin and none were resistant to cefuroxime. Most isolates displayed multiple resistances. Dendrogram of antibiotic resistances produced six clusters, with similarity levels between 18.8% and 100%. Generally, street food and clinical isolates tend to cluster apart. Dendrogram to cluster the antibiotic groups showed that they could be grouped according to classes based on mode of inhibition. The findings suggest that street food contaminated with drug-resistant Salmonella enterica can be an important factor in the continuous emergence of antibiotic resistant Salmonella enterica.
Broiler part samples (80 fresh and 80 chilled) were examined for the prevalence and numbers of C. jejuni and C. coli by employing most-probable-number (MPN) and polymerase chain reaction (PCR) techniques. The prevalence of the bacteria was high where C. jejuni was detected in 92.5% fresh and 53.8% chilled samples while C. coli in 80.0% fresh and 56.3% chilled. The number of these bacteria in the positive fresh and chilled samples was from 3 to more than 2400 MPN/g and from 3 to 290 MPN/g, respectively. Antibiotic resistance test (using Kirby-Bauer disc diffusion method) on 10 C. jejuni and 13 C. coli isolates toward ampicillin, tobramycin, enrofloxacin, ciprofloxacin, tetracycline, cephalothin, gentamicin and norfloxacin revealed high resistance toward all antibiotics (20.0% - 100.0%). All isolates were resistant to at least two antibiotics. This study highlights the potential of multidrug-resistant C. jejuni and C. coli transmission to humans through fresh and chilled broiler parts. Consecutive studies with bigger sample sizes and covering all over Malaysia are warranted in future.
Studies indicate that bacterial cross-contamination occurs during food preparation where bacteria can retent on the food contact surfaces and cause illness. The study evaluated the adherence of Campylobacter spp. to cutting boards, blades of knives and hands after cutting chilled, raw broiler parts (thighs + drumsticks, wings and livers). The adherence to cucumber cuts that were cut using the unwashed boards and knives was also analyzed. Generally, utensils have higher mean of Campylobacter spp. retained to them (1.4-223.3 MPN/ml rinse) than hands (0.7-43.4 MPN/ml rinse); however, Mann-Whitney U test showed no significant differences in the bacterial numbers found among the different surfaces. The transfer rates of Campylobacter spp. from utensils to cucumber cuts varied from 0% to more than 100%. The bacteria detected could be from the utensils and cucumber contamination before purchase or due to other factors where further investigation is required. The possibility is there for Campylobacter to spread to contact surfaces during chilled broiler handling; therefore, utensils and hands involved should be washed thoroughly especially before ready-to-eat food preparation.
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.
Nowadays, the incidence rate of foodborne disease has increased and become one of the global burdens affecting all individual ages in South East Asia region. Foodborne disease is responsible for mortality and morbidity worldwide thus affecting socio-economic and quality of life. Major causes of foodborne hazards diseases include diarrheal and invasive infectious disease agent, helminthes and chemicals. However, in developing countries, data and record is insufficient with poor surveillance systems leading to incomplete information on the real burden of foodborne disease. The introduction of Actor Network Theory (ANT) as tools for assessing and analyzing the food safety issues has drawn attention from various researcher as it is proven to be able to point out and identify the human and non human actors which is directly and contingently involved. The interaction between the actors such as a worker in an organization, student in school, and peoples in public provide information that can be used to minimize the risk of foodborne disease. The ultimate use of ANT is it helps the researcher to draw a framework of the source of contamination, agent responsible, factors involved, and idea to control the spread.
A total of 112 burger patties (35 beef burger patties, 39 chicken burger patties and 38 fish burger patties) which are commercially available at retail level were investigated for the presence and number of Listeria monocytogenes. These samples were analyzed using MPN-PCR method and conventional culturing methods. L. monocytogenes was detected in 33.3% of chicken burger patties, 22.9% of beef patties, and 10.5% of fish patty samples. From all contaminated raw burger patties, the estimated count of L. monocytogenes was ranged from 3 to 75 MPN/g. The results suggest that burger act as a potential source of listeriosis if the contaminated burger patty is consumed without adequate cooking. The risk associated with consumption of these samples was found to be high particularly for processed food at retail level in Malaysia. Therefore, food manufacturers play an important role in monitoring the manufacturing process and conduct a periodical surveillance on microbiological quality assessment on the processing plants. Besides, there is a need to increase awareness of consumers and food handlers to practice proper cooking of the burger patties before the point of consumption, to reduce the risk of listeria infection.
The aims of this communication were to study characterization of serogroups among Salmonella isolates and the relationship of antimicrobial resistance to serogroups. Multiple antimicrobial resistance (MAR) was performed on 189 Salmonella enterica isolates associated with 38 different serovars that were recovered from poultry and four types of indigenous vegetables.
Molecular typing methods have been widely applied for many purposes. In this study, such methods were adopted as DNA fingerprinting tools to determine the origin and divergence of virulent Vibrio parahaemolyticus strains found in local seafood. Although not all strain carry virulent tdh and trh gene, increasing prevalence demands an effective fingerprinting scheme which can constantly monitor and trace the sources of such emerging food pathogens. By using ERIC-, RAPD-, and BOX-PCR methods, 33 Vibrio parahaemolyticus isolates from local Malaysia bloody clam (Anadara granosa) and Lala (Orbicularia orbiculata) with confirmed presence of tdh and trh gene were characterised, followed by determination of clonal relatedness among virulent strains using cluster analysis and discriminatory index. This study also involved application of Immunomagnetic Separation (IMS) Method which significantly improved the specificity of strain isolation. Cluster analysis using Unweighted Pair Group Mathematical Averaging (UPGMA) and Dice Coefficient shown clustering according to isolation food source, IMS level and haemolysin gene possessed. Nevertheless, different DNA fingerprinting methods generated different clustering at different similarity cut-off percentage, regardless as individual or as composite dendrograms. ERIC- and RAPD-PCR composite fingerprinting relatively shown the highest discriminatory index at following similarity cutoff percentage: 0.68 at 50%; 0.83 at 65%; and 0.93 at 75%. Discriminatory power increased with similarity cut-off percentage. However, result also suggested that BOX-PCR might be an effective fingerprinting tool, as it generated three clusters with no single-colony isolate at 70% similarity cut-off. This study not only achieved its objective to determine clonal relatedness among virulent strains from local seafood via characterisation, but also speculated the best possible combination of molecular typing methods to effectively do so.