Since the introduction of the molecularly imprinting technology (MIT) in 1970s, it becomes an emerging technology with the potential for wide-ranging applications in food manufacturing, processing, analysis and quality control. It has been successfully applied in food microbiology, removal of undesirable components
from food matrices, detection of hazardous residues or pollutants and sensors. Molecularly imprinted solid-phase extraction (MISPE) is the most common application so far. The review describes the methods of making the molecularly imprinted polymer systems, the application of the technology in food safety issues and the remaining challenges.
Salmonella remains to be a major foodborne pathogen for animals and humans and is the
leading cause of foodborne infections and outbreaks in various countries. Salmonella Enteritidis
is one of the most frequently isolated serotypes in poultry and poultry products from human
food poisoning cases. It can cause mild to acute gastroenterititis as well as other common
food poisoning symptoms when infection takes place in human. Nucleic acid amplification
technologies such as Polymerase Chain Reaction (PCR) is a tool that is rapid and sensitive
for detection of bacterial pathogen. We report the successful detection of S. Enteritidis by
PCR in raw chicken meat artificially-contaminated with serial concentration of S. Enteritidis
using crude DNA extracts as DNA template. PCR primers, ENT-F and ENT-R targeted on sdfI
gene were used to amplify DNA region unique to S. Enteritidis with crude DNA extract of the
samples, yielded product with the size of 303 bp. These primers were specific to S. Enteritidis
when tested by in-silico simulation against genome database of targeted bacterial species and
confirmed in PCR as amplification bands were observed with S. Typhimurium, S. Polarum and
S. Gallinarum. The established PCR can detect as few as 9.4 X 101
CFU/ml of inoculated S.
Enteritidis concentration and proved that pre-enrichment effect have significant effect on PCR
detection by increasing 1000-fold of the sensitivity limit compared to the non pre-enriched
samples. The PCR technique indicated that it can be successfully coupled with pre-enrichment
step to offer advantage in routine screening and surveillance of bacterial contamination in food
samples.
Application of surface plasmon resonance (SPR) biosensor in detection of genetically modified organism (GMO) is demonstrated. A total of four biotinylated probes namely Tnosb, P35Sb, LECb and TSQb were successfully immobilized onto the SA chip. Results analysis indicated that the SPR system with the sensor chip immobilized with the Tnosb, P35Sb, LECb and TSQb biotinylated probes potentially detect complementary standard fragments as low as 1 nM. Biospecific interaction analysis (BIA), employing surface plasmon resonance (SPR) and biosensor technologies provide easy, rapid and automatable approach in detection of GMOs. Short assay times, label free DNA hybridization reaction and no toxic compounds are required, i.e. ethidium bromide, and the reusability of the sensor surface are some of the factors that contribute to the general advantages of the surface plasmon resonance (SPR) biosensor system in detection of GMOs.
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.
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.
Salmonella has caused foodborne illnesses globally and it has been a rising threat on fresh produce. The objective of this study was to determine the prevalence and concentration of Salmonella spp., Salmonella Typhi and Salmonella Typhimurium in freshly prepared fruit juice sold at hawker stalls. Analysis was conducted by employing most probable number-polymerase chain reaction (MPN-PCR). A total of 50 freshly prepared fruit juices were examined and the prevalence of Salmonella spp., Salmonella Typhi and Salmonella Typhimurium in the fruit juices were 34%, 20% and 10%, respectively, with an estimated microbial load varying from 0 to 42 MPN/g. Of the five different fruits, carrot juice had the highest prevalence of Salmonella spp. (60%) and Salmonella Typhi (40%). However, Salmonella Typhimurium was detected in apple (30%), orange (10%) and starfruit juice (10%). Factors contributing to the presence of Salmonella were cross-contamination and poor sanitation practice. Besides, negligence on temperature and storage time also led to the growth of Salmonella. Proper monitoring and risk assessment are needed in order to establish control measures to ensure the quality and safety of fruit juices in Malaysia.
The revolution of agriculture through biotechnology have produced large-scale of genetically
modified crops which brought up a controversy on the safety usage of genetically modified
organisms (GMOs). It has been implemented globally that all GMO products and its derived
ingredients should have regulations on the usage and labelling. Thus, it is necessary to develop
methods that allow rapid screening of GMO products to comply with the regulations. This
study employed a reliable and flexible multiplex polymerase chain reaction (PCR) method for
the rapid detection of transgenic elements in genetically modified soy and maize along with
the soybean LECTIN gene and maize ZEIN gene respectively. The selected four common
transgenic elements were 35S promoter (35S); Agrobacterium tumefaciens nopaline synthase
terminator (NOS); 5-enolypyruvylshikimate-3-phosphate synthase (epsps) gene; and Cry1Ab
delta-endotoxin (cry1Ab) gene. Optimization of the multiplex PCR methods were carried out
by using 1% Roundup ReadyTM Soybean (RRS) as the certified reference material for soybean
that produced fourplex PCR method detecting 35S promoter, NOS terminator, epsps gene and
soybean LECTIN gene and by using 1% MON810 as the certified reference material for maize
that produced triplex PCR method detecting 35S promoter, cry1Ab gene and maize ZEIN gene
prior to screening of the GMO traits in various food products and animal feeds. 1/9 (11.1%) of
the animal feed contained maize and 1/15 (6.7%) of the soybean food products showed positive
results for the detection of GMO transgenic gene. None of the maize food products showed
positive results for GMO transgenic gene. In total, approximately 4% of the food products
and animal feed were positive as GMO. This indicated GMOs have not widely entered the
food chain. However, it is necessary to have an appropriate screening method due to GMOs’
unknown potential risk to humans and to animals. This rapid screening method will provide
leverage in terms of being economically wise, time saving and reliable.
Ultra high temperature (UHT) treated milk products and formula milk are known to be
frequently contaminated with Bacillus cereus. Presence of B. cereus in these milk products is
of particular concern considering the majority of consumers are infants and children. Possible
sources of contamination are contaminated raw milk, cross-contamination during processing,
under-processing and mishandling of milk products. This study was conducted to detect the
presence of B. cereus in both formula milk (n=12) and UHT milk (n=20) sold in selected retail
markets. The approach consisted of enumerating by MPN/g followed by PCR assay aimed
at detecting gyrB gene in B. cereus, that encode for the subunit B protein of DNA gyrase
(topoisomerase type II). Contamination level of B. cereus in both types of samples examined
ranged from < 3 to > 1100 MPN/g. The contamination level of B. cereus was found to be
highest in full cream UHT milk (> 1100 MPN/g) and formula milk (> 1100 MPN/g). The PCR
analysis showed that 41.7% (5/12) formula milk and 30% (6/20) UHT milk samples were
detected with B. cereus, respectively. This is the first report of such study demonstrating the
presence of B. cereus in formula milk from Malaysia. Therefore, constant surveillance of these
milk products would reduce the potential risk of B. cereus-linked outbreaks.
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.
The safety level of microwaved foods remains at vague as this subject was less addressed
scientifically. A study was initiated to address the matter by investigating on the
survivability of Salmonella and Shiga-toxigenic Escherichia coli (STEC) O157 in
microwave heated ready-to-eat (RTE) foods using the Most Probable Number coupled
Polymerase Chain Reaction (MPN-PCR) technique. A total of 329 samples of various
ready-to-eat (RTE) convenience meals were collected around Wilayah Persekutuan Kuala
Lumpur and Selangor regions. Salmonella was positively identified in 66 samples (20.1%,
The effects of methanolic extract of Javanese turmeric (Curcuma xanthorrhiza Roxb.) at different level of concentrations on the inactivation of Bacillus cereus, Escherichia coli, Pseudomonas spp. and Staphylococcus aureus in oyster mushroom (Pleurotus sajor-caju) were investigated. This study was conducted principally for the achievement on the best combination between the
susceptibility of C. xanthorrhiza extract on natural microflora and foodborne pathogenic bacteria with the sensory acceptability of the soaked oyster mushroom. Three different concentrations (g/ml), 0.05%, 0.50% and 5.00%, of C. xanthorrhiza extract prepared with dilution method were designed as sanitizing agent in treating the oyster mushroom at 5 minutes and 10 minutes.
There was significance reduction in the survival of microbial load between the untreated fresh oyster mushroom and those soaked with 0.05%, 0.50% and 5.00% rhizome extract (P
Klebsiella pneumoniae is a foodborne pathogen associated with pneumoniae. Multiresistance to antibiotics of K. pneumoniae is a significant public health treat. Recently, the use of natural products such as herbs to inhibit the growth of pathogens is increasing. Java turmeric (Curcuma xanthorrhiza Roxb.) has been reported to possess antibacterial activity against foodborne pathogens. Unfortunately, the antibacterial activity of java turmeric extract against the resistance to multiantibiotics of K. pneumoniae has not been investigated. In this study, the antibacterial activity of Java turmeric extract was tested against 24 isolates of resistant K. pneumoniae that was isolated from several vegetables; lettuce, cucumber, tomato and carrot, using the methods recommended by the Clinical and Laboratory Standard Institute (CSLI), including disc diffusion method, minimal inhibitory concentration (MIC), minimal bactericidal concentration (MBC) and killing time at concentration 0× MIC, ½× MIC, 1× MIC, 2× MIC and 4× MIC with predetermined time of 0, 0.25 , 0.5, 1, 2 and 4 h. The results showed that Java turmeric extract is susceptible to all resistant K. pneumoniae with inhibition zones ranging from 8.67 ± 0.58 to 10.00 ± 0.00 mm. The MIC and MBC values for the K. pneumoniae isolates against all bacterial isolates was 1.25 and 2.5 mg/ml, respectively. The killing time curve shows the reduction of resistant K. pneumoniae cells is fast acting; > 3 log10 within less than 15 min at 4× MIC (5.0 mg/ml). Finally, the isolates were completely killed at 4× MIC for 15 min. In conclusion, the Java turmeric extracts can be developed as natural antimicrobial agent to inhibit the growth of K. pneumoniae in food system.
The prevalence of Campylobacter spp. in retailed sushi were examined using the techniques of polymerase chain reaction (PCR) in combination with most probable number (MPN) to quantify the bacteria in 150 samples obtained from three supermarkets. The average prevalence of Campylobacter spp. in retailed sushi was 26.6% with 32%, 16% and 32% from supermarket I, II and III, respectively. Campylobacter jejuni was found to be the predominant species in retailed sushi with 82.49% of all Campylobacter spp. positive samples. Campylobacter coli was not detected in all samples. The maximum MPN number of Campylobacter spp. in retailed sushi purchased from supermarket I, II and III ranged from 3.6-11.0 MPN/g, 9.4->1100 MPN/g and 27-1100 MPN/g, respectively. The isolation of C. jejuni from a variety of ready-to-eat retail sushi may indicate that these products can act as possible vehicles for the dissemination of food-borne campylobacteriosis.
Polymerase chain reaction (PCR) technique was used to assay for the detection of specific genes in the genomes of the Aeromonas spp. isolated from environmental and shellfish sources, particularly aero and hlyA genes, responsible for aerolysin and hemolysin toxins production in this genus. The results showed that: (i) the 1500 bp amplicon of the hlyA gene was detected in 20/38 of the Aeromonas hydrophila, 13/38 of the A. caviae and 6/9 of the A. veronii biovar sobria isolates; (ii) the 690 bp amplicon of the aero gene was detected in 20/38 of A. hydrophila, 17/38 of A. caviae and 6/9 of A. veronii biovar sobria isolates; (iii) the nucleotide blast results of aerolysin gene sequences of the representative strains of A. hydrophila, A. caviae and A. veronii biovar sobria revealed a high homology of 94%, 95% and 95% with published sequences, respectively and ; (iv) the protein blast showed 97%, 94% and 96% homology when compared to the published sequences, respectively. The finding of A. hydrophila virulence genes in other members of the genus Aeromonas, may give a new perspective to the significance of these results. The method described here may be a useful detection tool to assist in further investigation of aero and hlyA genes in the genus Aeromonas, especially for food microbiologist.
Phytochemicals belonging to the group’s phenols, terpenes, betalains, organosulfides, indoles and protein inhibitors are important components in fruits, vegetables, legumes, whole grains and nuts that have health promoting benefits and a variety of applications in food and pharmaceutical industries. Initially only a few of these important phytochemicals are produced commercially by chemical synthesis. However, recent developments in the field of biotechnology have provided metabolic engineering strategies that use microorganisms as cell factories for high production of these products. This review will discuss the general biosynthetic pathways, metabolic engineering and optimization strategies of functional phytochemicals that have received a lot of attention from investigators.
Ninety one leaf samples of Josapine pineapple cultivar (Kelantan, n=8; Pahang, n=20; Perak, n=11; Sabah, n=15; Johor, n=37) showing symptoms of heart rot disease were collected to determine the incidence of Erwinia chrysanthemi. Sixteen strains of E. chrysanthemi were isolated from 13 leaf samples from Pahang (n=4), Sabah (n=2) and Johor (n=7). All of the E. chrysanthemi strains displayed resistance to bacitracin with two strains showing resistance to sulfamethoxazole. None of the E. chrysanthemi strains were resistant toward ampicillin, carbenicillin, cephalothin, ceftriaxone, cefuroxime, gentamicin, kanamycin, nalidixic acid, penicillin G, streptomycin and tetracycline. All of the E. chrysanthemi strains were plasmidless. The dendrogram generated from the ERIC-PCR fingerprinting showed that the E. chrysanthemi strains formed 4 clusters and 7 single isolates at 80% similarity level. The restriction fragment length polymorphism (RFLP) analysis for 16 strains of E. chrysanthemi with HinfI and HaeIII endonuclease, 2 and 4 restriction profiles were obtained, respectively. The combinations of the four techniques were able to differentiate the 16 E. chrysanthemi strains into 14 genome types, suggesting a wide diversity of strains examined. ERICPCR fingerprinting method is found to be more discriminating and useful for the determination of the E. chrysanthemi strains relatedness.
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.
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.
The main source of E. coli 0157:H7 is cattle, but recent studies showed high percentage of outbreaks
contributed by contaminated water. The occurrence of E. coli O157:H7 in environmental water samples poses a potential threat to human health. The aim of this study was to establish a protocol for the detection of the pathogen E. coli O157:H7 and E. coli virulence genes (eaeA, rfbE, hly, stx1, and stx2) in a multiplex PCR protocol using six specific primer pairs. The target genes produced species-specific amplicons at 625 bp, 397 bp, 296 bp, 166 bp, 210 bp and 484 bp for E. coli O157:H7 (fliCh7 gene) and virulence genes (eaeA, rfbE, hly, stx1, and stx2) respectively. The results obtained show that the established PCR protocol is suitable for a rapid and specific analysis of the pathogenic E. coli O157:H7 in environmental water samples for the assessment of microbiological risks.
Legislation concerning the safety assessment and labelling of foodstuffs has been implemented in many countries. Consequential to a number of cases of food adulteration reported globally, a fast and reliable detection method for the food traceability is required in ensuring effective implementation of food legislation in a country. In this study, PCR-RFLP technique based on cyt b gene has been tested for its suitability for these purposes. This method combines the use of a pair of universal primer that amplifies a 359 bp fragment on the cyt b gene from meat muscle DNA and restriction enzyme analysis. Analysis of experimental beef frankfurter, minced beef, pork frankfurter and pork cocktail samples demonstrated the suitability of the assay for the detection of the beef (Bos taurus) and pork (Sus scrofa), but not applicable for some processed food, particularly detection of mackerel (Rasterelliger brachysoma), sardine (Saedinella Fimbriata) and tuna (Thunnus tonggol) origin in canned food. Commercial frauds through species mislabelling or misdescribed were not detected. The assay is demonstrated applicable for routine analysis of meat traceability of foodstuffs and legislation purposes, if sufficient availability of detectable mtDNA in the foodstuffs is ensured.