A total of 106 beef samples which consisted of local (n = 59) and imported (n = 47) beef and 180 milk samples from cows (n = 86) and goats (n = 94) were collected from Selangor, Malaysia. Overall, 30.2% (32 of 106) of beef samples were found positive for Arcobacter species. Imported beef was significantly more contaminated (46.80%) than local beef (16.9%). Arcobacter butzleri was the species isolated most frequently from imported (81.8%) and local (60%) beef, followed by Arcobacter cryaerophilus in local (33.3%) and imported (18.2%) beef samples. Only one local beef sample (10%) yielded Arcobacter skirrowii. Arcobacter species were detected from cow's milk (5.8%), with A. butzleri as the dominant species (60%), followed by A. cryaerophilus (40%), whereas none of the goat's milk samples were found positive for Arcobacter. This is the first report of the detection of Arcobacter in milk and beef in Malaysia.
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.
Consecutive chloroform, ethanol, and ethyl acetate partitions of extracts from winged bean [Psophocarpus tetragonolobus (L.) DC] root, stem, leaf, and pod extracts were tested for their antimicrobial activity against 19 microbial species, including 11 bacterial pathogens, four yeasts, and four molds using the disk diffusion assay technique. The pod extract was found to be most effective against all of the tested organisms, followed by the stem, root, and leaf extracts, and the ethanol fraction showed the most significant (p < 0.05) antimicrobial activity against all of the tests among three soluble fractions of extract, followed by the ethyl acetate and chloroform fractions. The minimum inhibitory concentrations (MICs) of extracts determined by the broth dilution method ranged from 1.25 to 10.0 mg/mL. The MIC of ethanol fraction of pod extracts was the lowest by comparison with the other two extracts. The MIC for fungi was at or below 2.5 mg/mL and for bacteria was at or above 2.5 mg/mL.
Little work has been reported on the use of commercial antimicrobials against foodborne pathogens on duck meat. We investigated the effectiveness of trisodium phosphate (TSP) and sodium hypochlorite (SH) as antimicrobial treatments against Campylobacter and Salmonella on duck meat under simulated commercial water chilling conditions. The results were compared to the same treatments on well-studied chicken meat. A six strain Campylobacter or Salmonella cocktail was inoculated (5 ml) at two dilution levels (10(4) and 10(8) cfu/ml) onto 25 g duck or chicken meat with skin and allowed to attach for 10 min. The meat was exposed to three concentrations of pH adjusted TSP (8, 10 and 12% (w/v), pH 11.5) or SH (40, 50 and 60 ppm, pH 5.5) in 30 ml water under simulated spin chiller conditions (4 °C, agitation) for 10 min. In a parallel experiment the meat was placed in the antimicrobial treatments before inoculation and bacterial cocktails were added to the meat after the antimicrobial solution was removed while all other parameters were maintained. Untreated controls and controls using water were included in all experiments. Bacterial numbers were determined on Campylobacter blood-free selective agar and Mueller Hinton agar or xylose deoxycholate agar and tryptone soya agar using the thin agar layer method for Campylobacter and Salmonella, respectively. All TSP concentrations significantly (p<0.05) reduced numbers of Campylobacter (~1.2-6.4 log cfu/cm(2)) and Salmonella (~0.4-6.6 log cfu/cm(2)) on both duck and chicken meat. On duck meat, numbers of Campylobacter were less than the limit of detection at higher concentrations of TSP and numbers of Salmonella were less than the limit of detection at all concentrations of TSP except one. On chicken meat, numbers of Campylobacter and Salmonella were less than the limit of detection only at the lower inoculum level and higher TSP concentrations. By contrast only some of the concentrations of SH significantly (p<0.05) reduced numbers of Campylobacter and Salmonella (~0.2-1.5 log cfu/cm(2)) on both duck and chicken meats. None of the SH treatments resulted in numbers of either pathogen being less than limit of detection. Results indicate that chicken meat has the ability to effectively protect Campylobacter and Salmonella against the impact of trisodium phosphate and sodium hypochlorite while duck meat does not. This study suggests that trisodium phosphate has a strong potential for application in a commercial poultry processing to reduce Campylobacter and Salmonella specifically on duck meat.
This study was carried out to examine the efficacy of two biocontrol agents (Clonostachys rosea 016, BCA1; Gram-negative bacterium, BCA5) for control of FUM1 gene expression and fumonisin B1 (FB1) production by F. verticillioides FV1 on maize cobs of different ripening stages: R3, Milk (0.985 aw); R4, Dough (0.976 aw); R5, Dent (0.958 aw). Initially, temporal studies on FUM1 gene expression and FB1 production were performed on maize kernels for up to 14days. This revealed that day 10 was optimum for both parameters, and was used in the biocontrol studies. Maize cobs were inoculated with 50:50 mixtures of the pathogen:antagonist inoculum and incubated in environmental chambers to maintain the natural aw conditions for ten days at 25 and 30°C. The growth rates of F. verticillioides FV1, the relative expression of the FUM1 gene and FB1 production were quantified. It was found that, aw×temp had significant impacts on growth, FUM1 gene expression and FB1 production by F. verticillioides FV1 on maize cobs of different maturities. The fungal antagonist (BCA1) significantly reduced FB1 contamination on maize cobs by >70% at 25°C, and almost 60% at 30°C regardless of maize ripening stage. For the bacterial antagonist (BCA5) however, FB1 levels on maize cobs were significantly decreased only in some treatments. These results suggest that efficacy of antagonists to control mycotoxin production in ripening maize cobs needs to take account of the ecophysiology of the pathogen and the antagonists, as well as the physiological status of the maize during silking to ensure effective control.
Rice bran, a by-product of the rice milling process, has emerged as a functional food and being used in formulation of healthy food and drinks. However, rice bran is often contaminated with numerous mycotoxins. In this study, a method to simultaneous detection of aflatoxins (AFB1, AFB2, AFG1, and AFG2), ochratoxin A (OTA), deoxynivalenol (DON), fumonisins (FB1 and FB2), sterigmatocystin (STG), T-2 toxin, HT-2 toxin, diacetoxyscirpenol (DAS) and zearalenone (ZEA) in rice bran was developed, optimized and validated using dispersive liquid-liquid microextraction (DLLME) and liquid chromatography-tandem mass spectrometry (LC-MS/MS). In DLLME, using a solvent mixture of methanol/water (80:20, v/v) as the dispersive solvent and chloroform as the extraction solvent with the addition of 5% salt improved the extraction recoveries (63-120%). The developed method was further optimized using the response surface methodology (RSM) combined with Box-Behnken Design (BBD). Under the optimized experimental conditions, good linearity was obtained with a correlation coefficient (r2) ≥ 0.990 and a limit of detection (LOD) between 0.5 to 50 ng g-1. The recoveries ranged from 70.2% to 99.4% with an RSD below 1.28%. The proposed method was successfully applied to analyze multi-mycotoxin in 24 rice bran samples.
Heat exchanger performance degrades rapidly during operation due to formation of deposits on heat transfer surfaces which ultimately reduces service life of the equipment. Due to scaling, product deteriorates which causes lack of proper heating. Chemistry of milk scaling is qualitatively understood and the mathematical models for fouling at low temperatures have been produced but the behavior of systems at ultra high temperature processing has to be studied further to understand in depth. In diversified field, the effect of whey protein fouling along with pressure drop in heat exchangers were conducted by many researchers. Adding additives, treatment of heat exchanger surfaces and changing of heat exchanger configurations are notable areas of investigation in milk fouling. The present review highlighted information about previous work on fouling, influencing parameters of fouling and its mitigation approach and ends up with recommendations for retardation of milk fouling and necessary measures to perform the task.
The study assessed the effect of conscious halal slaughter and slaughter following minimal anesthesia on bleeding efficiency of goats and keeping quality of goat meat. Ten Boer cross bucks were divided into two groups and subjected to either halal slaughter without stunning (HS) or minimal anesthesia prior to slaughter (AS). The blood lost during exsanguination was measured. Residual blood was further quantified by determination of hemoglobin and myoglobin content in longissimus lumborum muscle. Storage stability of the meat was evaluated by microbiological analysis and lipid oxidation. Blood loss at exsanguination, residual hemoglobin and lipid oxidation were not significantly different (p>0.05) between HS and AS. Lactic acid bacteria was the only microbe that was significantly elevated after 24h of storage at 4°C in the AS group. In conclusion, slaughtering goats under minimal anesthesia or fully conscious did not affect bleeding efficiency and keeping quality of goat meat.
A study was conducted to estimate the prevalence of Salmonella among broilers retailed at wet-markets and processing plants. Litter and feed samples obtained from both broiler and breeder farms were also examined for Salmonella. A total of 158 out of 445 (35.5%) and 52 out of 104 (50.0%) broiler carcasses obtained from wet-markets and processing plants were contaminated with Salmonella, respectively. Salmonella was isolated from 14 out of 98 (14.3%) samples of intestinal content. Litter samples from broiler and breeder farms were positive for Salmonella, 8/40 (20%) and 2/10 (20%), respectively. Salmonella isolates (230) belonging to 15 different serovars were isolated. Predominant serovars were S. enteritidis, S. muenchen, S. kentucky and S. blockley.
Enterotoxigenic Bacillus cereus was detected in cooked foods (17), rice noodles (3), wet wheat noodles (2), dry wheat noodles (10), spices (8), grains (4), legumes (11) and legume products (3). One hundred ninety-four (42.3%), 70 (15.3%) and 23 (5.2%) of the 459 presumptive B. cereus colonies isolated from PEMBA agar were identified as B. cereus, Bacillus thuringiensis and B. mycoides, respectively. B. cereus isolates were examined for growth temperature, pH profile and enterotoxin production using both TECRA-VIA and BCET-RPLA kits. One hundred seventy-eight (91.8%) and 164 (84%) of the strains were enterotoxigenic as determined using TECRA-VIA and BCET-RPLA, respectively. Eighty-two (50%) of the enterotoxigenic strains were capable of growing at 5 degrees C, and 142 (86.6%) grew at 7 degrees C within 7 days of incubation. The enterotoxigenic strains did not grow at pH 4.0 but 69 (42.0%) of the strains were able to grow at pH 4.5 within 7 days at 37 degrees C. The isolates were resistant to ampicillin (98.8%), cloxallin (100%) and tetracycline (61.0%), and susceptible to chloroamphenicol (87%), erythromycin (77.4%), gentamycin (100%) and streptomycin (98.7%).
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.
This study was conducted to determine the Campylobacter contamination rate of chicken carcasses and the processing lines of modern processing plants in Malaysia. Three hundred sixty samples were collected from 24 flocks of broiler chickens at 12 modern poultry processing plants in 6 states of Malaysia. Fresh fecal droppings were collected from crates in the arrival area. Neck skin samples were taken from processed chicken carcasses at 3 different processing stages: before inside-outside washing, after inside-outside washing and post chilling. Swab samples from the scalding tank, chilling tank and conveyer belt before chilling were also collected to determine contamination with Campylobacter in the slaughter house environment prior to slaughter. Isolation for Campylobacter was performed following ISO 10272-1:2006(E). The overall of contamination rate with Campylobacter at the 12 plants was 61.0% (220/360). Eighty point six percent of the samples from before the inside-outside wishing step were contaminated with Campylobacter, as were 62.5% of the samples after the inside washing and 38.9% after the post-chilling step. This study shows extensive contamination of chicken carcasses and slaughtering houses in Malaysia with Campylobacter.
A total of 360 samples including fresh fecal droppings, neck skins, and swab samples was collected from 24 broiler flocks and processed by 12 modern processing plants in 6 states in Malaysia. Ninety samples from 10 traditional wet markets located in the same states as modern processing plants were also collected. Microbiological isolation for Campylobacter was performed following ISO 10272-1:2006 (E). The overall rate of contamination for Campylobacter in modern processing plants and in traditional wet markets was 61.1% (220/360) and 85.6% (77/90), respectively. Campylobacter jejuni was detected as the majority with approximately 70% for both facilities. In the modern processing plants, the contamination rate for Campylobacter gradually declined from 80.6% before the inside-outside washing to 62.5% after inside-outside washing and to 38.9% after the post chilling step. The contamination rate for Campylobacter from processed chicken neck skin in traditional wet markets (93.3%) was significantly (P<0.01) higher than in modern processing plants (38.9%).
Malaysian population widely consumes the cereal-based foods, oilseeds, nuts, and spices in their daily diet. Mycotoxigenic fungi are well known to invade food products under storage conditions and produce mycotoxins that have threat to human and animal health. Therefore, determining toxigenic fungi and aflatoxin B(1) (AFB1) in foods used for human consumption is of prime importance to develop suitable management strategies and to minimize risk. Ninety-five food products marketed in Penang, Malaysia were randomly collected from different supermarkets and were analyzed for presence of Aspergillus spp. by agar plate assay and AFB1 by enzyme-linked immunosorbent assay (ELISA). A. flavus was the dominant fungi in all foods followed by A. niger. Fifty-five A. flavus strains were tested for their ability to produce aflatoxins on rice grain substrate. Thirty-six (65.4%) strains out of 55 produced AFB1 ranging from 1700 to 4400 μg/kg and 17 strains (31%) produced AFB2 ranging from 620 to 1670 μg/kg. Natural occurrence of AFB1 could be detected in 72.6% food products ranging from 0.54 to 15.33 μg/kg with a mean of 1.95 μg/kg. Maximum AFB1 levels were detected in peanut products ranging from 1.47 to 15.33 μg/kg. AFB1 levels detected in all food products were below the Malaysian permissible limits (<35 μg/kg). Aspergillus spp. and AFB1 was not detected in any cookies tested. Although this survey was not comprehensive, it provides valuable information on aflatoxin levels in foods marketed in Malaysia.
The aim of this study was to determine antibacterial activity of S. polyanthum L. (salam) leaves extract foodborne pathogens. All the foodborne pathogens were inhibited after treating with extract in disk diffusion test with range 6.67 ± 0.58-9.67 ± 0.58 mm of inhibition zone. The range of MIC values was between 0.63 and 1.25 mg/mL whereas MBC values were in the range 0.63 mg/mL to 2.50 mg/mL. In time-kill curve, L. monocytogenes and P. aeruginosa were found completely killed after exposing to extract in 1 h incubation at 4x MIC. Four hours had been taken to completely kill E. coli, S. aureus, V. cholerae, and V. parahaemolyticus at 4x MIC. However, the population of K. pneumoniae, P. mirabilis, and S. typhimurium only reduced to 3 log CFU/mL. The treated cell showed cell rupture and leakage of the cell cytoplasm in SEM observation. The significant reduction of natural microflora in grapes fruit was started at 0.50% of extract at 5 min and this concentration also was parallel to sensory attributes acceptability where application of extract was accepted by the panellists until 5%. In conclusion, S. polyanthum extract exhibits antimicrobial activities and thus might be developed as natural sanitizer for washing raw food materials.
A reversed-phase HPLC optimization strategy is presented for investigating the separation and retention behavior of aflatoxin B1, B2, G1, G2, ochratoxin A and zearalenone, simultaneously. A fractional factorial design (FFD) was used to screen the significance effect of seven independent variables on chromatographic responses. The independent variables used were: (X1) column oven temperature (20-40°C), (X2) flow rate (0.8-1.2 ml/min), (X3) acid concentration in aqueous phase (0-2%), (X4) organic solvent percentage at the beginning (40-50%), and (X5) at the end (50-60%) of the gradient mobile phase, as well as (X6) ratio of methanol/acetonitrile at the beginning (1-4) and (X7) at the end (0-1) of gradient mobile phase. Responses of chromatographic analysis were resolution of mycotoxin peaks and HPLC run time. A central composite design (CCD) using response surface methodology (RSM) was then carried out for optimization of the most significant factors by multiple regression models for response variables. The proposed optimal method using 40°C oven temperature, 1 ml/min flow rate, 0.1% acetic acid concentration in aqueous phase, 41% organic phase (beginning), 60% organic phase (end), 1.92 ratio of methanol to acetonitrile (beginning) and 0.2 ratio (end) for X1-X7, respectively, showed good prediction ability between the experimental data and predictive values throughout the studied parameter space. Finally, the optimized method was validated by measuring the linearity, sensitivity, accuracy and precision parameters, and has been applied successfully to the analysis of spiked cereal samples.
The study compares the bacteriological quality on Asian seabass (Lates calcarifer) between ice and salt storage methods. The main objectives of the study were to identify different bacteria constituents and quantitative bacterial load in Asian seabass when preserved with ice and sea salt. For the purpose of this study, Asian seabass was stored in two different conditions of ice-chilled and salted for 2 days. All fish samples were analyzed by performing bacteriological analysis and the isolated bacteria were identified by using API identification system. In case of the quantity of bacteria in the flesh, Chilling and salting had no significant difference to the quantity of bacteria on fish flesh. As for the skin, salt-preserved fish showed higher quantity of bacteria than ice-preserved fish. Acinetobacter baumannii and Pseudomonas fluorescens had been identified from skin sample of ice-chilled fish. Besides P. fluorescens and A. baumannii other isolates identified include Vibrio and Myxobacteria. All bacteria were cocci-shaped except a few bacilli. In term of bacteria number and morphological characteristics, ice-chilled preserved fish was better than salt preserved fish. Overall, less number of bacteria was observed in both ice-chilled and sea salt preserved fish. The result of this study indicated that the quick preservation is a very important factor to control bacterial load in the preserved fish.
Twenty-five and three strains of Escherichia coli O157:H7 were identified from 25 tenderloin beef and three chicken meat burger samples, respectively. The bacteria were recovered using the immunomagnetic separation procedure followed by selective plating on sorbitol MacConkey agar and were identified as E. coli serotype O157:H7 with three primer pairs that amplified fragments of the SLT-I, SLT-II and H7 genes in PCR assays. Susceptibility testing to 14 antibiotics showed that all were resistant to two or more antibiotics tested. Although all 28 strains contained plasmid, there was very little variation in the plasmid sizes observed. The most common plasmid of 60 MDa was detected in all strains. We used DNA fingerprinting by randomly amplified polymorphic DNA (RAPD) and pulsed-field gel electrophoresis (PFGE) to compare the 28 E. coli O157:H7 strains. At a similarity level of 90%, the results of PFGE after restriction with XbaI separated the E. coli O157:H7 strains into 28 single isolates, whereas RAPD using a single 10-mer oligonucleotides separated the E. coli O157:H7 strains into two clusters and 22 single isolates. These typing methods should aid in the epidemiological clarification of the E. coli O157:H7 in the study area.
This study has evaluated the use of a commercially available Rainbow agar O157 and polymerase chain reaction (PCR) assays for the detection of Shiga-like toxin producing Escherichia coli and to serotype E. coli O157:H7 from raw meat. The Rainbow agar O157 was found to be selective and sensitive for the screening of the E. coli O157 from artificially and naturally contaminated meat samples. Shiga-like toxin producing E. coli were identified with two primer pairs that amplified fragments of the SLT-I (384 bp) and SLT-II (584 bp). E. coli O157:H7 was serotyped with a primer pair specified for the H7 flagellar gene, which amplify specific DNA fragments (625 bp) from all E. coli O157:H7 strains. The use of Rainbow agar O157 described allows for the presumptive isolation of E. coli O157 in 24 hours. Identification and confirmation of the presumptive isolates as E. coli O157:H7 by PCR assays require additional 6-8 hours. The above-mentioned screening and identification procedures should prove to be a very useful method since it allows for the specific detection of E. coli O157:H7.
A total of 87 market fish samples representing five types of fish were evaluated for the presence of Aeromonas spp. Of the samples examined, 69%, 55%, 11.5% and 2.3% harbored Aeromonas spp., A. veronii biovar sobria, A. hydrophila and A. caviae, respectively. The 60 isolated Aeromonas spp. strains were further examined for hemolytic activity, resistance to antimicrobial agents and presence of plasmids. Hemolytic activity varied widely among the isolated strains. Though all the isolates demonstrated resistance to three or more of the antibiotics tested, all were susceptible to ceptazidime. Thirty-four (56.7%) of the sixty isolates harbored plasmids, with sizes ranging from 2.3 to 15.7 kb. These results indicate that hemolytic, multiple antibiotic resistant and genetically diverse aeromonads are easily recovered from fish in this region.