Thermal processing of green coconut water (GCW) caused non-enzymic browning and development of rancidity. Effect of the addition of several combinations of ascorbic acid (AA) (0 to 100 ppm) and sodium metabisulfite (SMB) (0 to 30 ppm) on brown discolouration and rancidity of GCW during elevated thermal processing (121°C for 5 min at 15 psi) was investigated. Addition of AA and/or SMB significantly (P
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%).
Sugarcane juice deteriorates rapidly as enzymatic browning and microbial spoilage take place soon after juice extraction. Therefore, the effects of pasteurization treatment (70ºC for 10 minutes) and addition of calamansi juice at different concentrations (0, 1.0, 1.5, and 3.0%, v/v) on the physicochemical (color, pH, total soluble solid, titratable acidity, and peroxidase enzymatic activity), microbiological (total plate count, yeast and mold), and sensory characteristics of black stem sugarcane juice were investigated. Quantitative descriptive analysis (QDA®) was conducted by eight trained panelists to determine the intensity of the sensory attributes of sugarcane juice, such as brownish color, grassy aroma, citrus aroma, sweetness, sourness, sweet aftertaste, and overall acceptability using a 15 cm unstructured line scale. The results showed that the pasteurization treatment significantly (p ˂ 0.05) increased the L* value (lightness) of sugarcane juice at 0% (v/v) of calamansi juice. Increasing the concentration of calamansi juice decreased the pH value and increased the titratable acidity of the sugarcane juice at 5% significance level. The relative effectiveness in reducing the peroxidase (POD) enzymatic activity and microbial load of sugarcane juice was shown after the addition of calamansi juice and pasteurization treatment, respectively. Significant (p ˂ 0.05) changes in the intensity of brownish color, grassy aroma, and sweet aftertaste of sugarcane juice were observed after the pasteurization treatment at 0% (v/v) calamansi juice; whereas, the addition of calamansi juice at different concentrations increased the intensity of citrus aroma and sourness, but decreased the intensity of grassy aroma and sweet aftertaste of sugarcane juice at 5% significance level. Hence, pasteurization treatment coupled with the addition of 1.5% (v/v) calamansi juice was effective in achieving microbial stability and consumer’s acceptability on sugarcane juice.
Fifteen strains of Lactobacillus and Bifidobacterium were screened based on their ability to adhere to hydrocarbons via the determination of cellular hydrophobicity. Lactobacillus acidophilus ATCC 314, L. acidophilus FTCC 0291, Lactobacillus bulgaricus FTCC 0411, L. bulgaricus FTDC 1311, and L. casei ATCC 393 showed greater hydrophobicity and, thus, were selected for examination of cholesterol-removal properties. All selected strains showed changes in cellular fatty acid compositions, especially total fatty acids and saturated and unsaturated fatty acids in the presence of cholesterol compared with those grown in the absence of cholesterol. In addition, we found that cells grown in media containing cholesterol were more resistant to sonication and enzymatic lysis compared with those grown without cholesterol. We further evaluated the location of the incorporated cholesterol via the insertion of fluorescence probes into the cellular membrane. In general, enrichment of cholesterol was found in the regions of the phospholipid tails, upper phospholipids, and polar heads of the cellular membrane phospholipid bilayer. Our results also showed that lactobacilli were able to reduce cholesterol via conversion of cholesterol to coprostanol, aided by the ability of strains to produce cholesterol reductase. Our results provided experimental evidence to strengthen the hypothesis that probiotics could remove cholesterol via the incorporation of cholesterol into the cellular membrane and conversion of cholesterol to coprostanol. The strains studied may be potential health adjunct cultures in fermented dairy products with possible in vivo hypocholesterolemic effects.
Paralactobacillus selangorensis gen. nov., sp. nov. is described. This organism, isolated from a Malaysian food ingredient called chili bo, is an obligatory homofermentative, rod-shaped lactic acid bacterium. The G+C content is 46.1-46.2+/-0.3 mol%. Earlier 16S rRNA studies showed that this organism constitutes a new taxon distantly related to the Lactobacillus casei-Pediococcus group. A phenotypic description that distinguishes Paralactobacillus selangorensis from other genera of lactic acid bacteria is presented. The type strain of Paralactobacillus selangorensis is LMG 17710T.
Three strains of verotoxin-producing Escherichia coli isolated from patients with haemorrhagic colitis harboured plasmids ranging in size from 2.7 kb to 91.2 kb. Those plasmids ranging from 2.7 kb to 6.8 kb hybridized to Shiga-like toxin I and Shiga-like toxin II gene probes.
Antibiotic susceptibility, plasmid profiles and random amplification of polymorphic DNA (RAPD) were used to study strains of Vibrio vulnificus isolated from cockles (Anadara granosa). Thirty-six isolates were analyzed. The prevalent biotypes were 1 (72.2% of the isolates) and 2 (27.8%). Among these, 21 strains of biotype 1 and two strains of biotype 2 contained plasmid DNA bands ranging in size from 1.4 to 9.7 MDa. Thirty-one (83.3%) were found to be resistant to one or more of the antimicrobial agents tested, however no specific correlation between antimicrobial resistance patterns and a single biotype was found. In addition, no particular plasmid profile was predictive of a particular pattern of antibiotic susceptibility. Two primers produced polymorphisms in all strains tested, producing bands ranging from 0.25 to 2.7 kb, indicating a high variability among both biotype 1 and biotype 2 of the V. vulnificus strains investigated. RAPD identity across biotypes was also observed among Vibrio vulnificus strains.
Nineteen strains of vancomycin-resistant Enterococcus faecium isolated from 10 of 75 (13.3%) tenderloin beef samples were examined for resistance to selected antibiotics, presence of plasmids, and genetic diversity by random amplification of polymorphic DNA analysis. All strains showed multiple resistant to the antibiotics tested. Multiple antibiotic indexing of the vancomycin-resistant E. faecium strains showed that all (100%) originated from high risk contamination environments where antibiotics were often used. Plasmids ranging in size from 1.5 to 36 megadalton were detected in 15 of 19 (79%) strains. Thus, three plasmid profiles and eight antibiotypes were observed among the E. faecium strains. A high degree of polymorphism was obtained by combining the results of the two primers used; with the 19 E. faecium strains being differentiated into 19 RAPD-types. These preliminary results suggest that RAPD-PCR has application for epidemiologic studies and that resistance patterns and plasmid profiling could be used as an adjunct to RAPD for the typing of E. faecium in the study area.
Salmonella enterica serovar Paratyphi B (S. Paratyphi B) is a major foodborne pathogen distributed all over the world. However, little is known about the antibiotic resistance, genetic relatedness and virulence profile of S. Paratyphi B isolated from leafy vegetables and the processing environment in Malaysia. In this study, 6 S. Paratyphi B isolates were recovered from different vegetables and drain water of processing areas obtained from fresh food markets in Malaysia. The isolates were characterized by antibiogram, Pulsed-field gel electrophoresis (PFGE) and virulence genes. Antibiotic susceptibility test showed that 3 of the isolates were resistant to the antibiotics. These include S. Paratyphi B SP251 isolate, which was resistant to chloramphenicol, ampicillin, sulfonamides and streptomycin; Isolate SP246 which was resistant to chloramphenicol, sulfonamides and streptomycin and Isolate SP235 showing resistance to nalidixic acid only. PFGE subtyped the 6 S. Paratyphi B isolates into 6 distinct XbaI-pulsotypes, with a wide range of genetic similarity (0.55 to 0.9). The isolates from different sources and fresh food markets location were genetically diverse. Thirteen (tolC, orgA, spaN, prgH, sipB, invA, pefA, sofB, msgA, cdtB, pagC, spiA and spvB) out of the 17 virulence genes tested were found in all of the S. Paratyphi B isolates. Another gene (lpfC), was found only in one isolate (SP051). None of the isolates possessed sifA, sitC and ironN genes. In summary, this study provides unique information on antibiotic resistance, genetic relatedness, and virulotyping of S. Paratyphi B isolated from leafy vegetables and processing environment.
The growth of Vibrio cholerae O139 inoculated into cendol (a mixture of coconut milk, brown sugar, and green jelly from rice flour), rojak (prawn paste, sugar, soy sauce, spices, garlic, and peanut gravy), gravy, tofu, fried tofu, and wheat-flour noodles (all except rojak gravy containing the natural microbial flora) was examined at four incubation temperatures (7, 15,25, and 35°C). V. cholerae O139 grew well in cendol incubated at 25 and 35°C but not at 15°C or below. No growth of V. cholerae O139 in rojak gravy was detected at any temperature except for very slow growth at 35°C. V. cholerae O139 inoculated into tofu exhibited slow growth at 25 and 35°C and growth was not detected at 7 and 15°C. However, in fried tofu, the organism entered the growth phase after 12 h of incubation at 25 and 35°C. Growth of V. cholerae O139 was not demonstrated in noodles at any incubation temperatures. Nutrient broth with 1% NaCl added supported the growth of V. cholerae O139 at 25 and 35°C. At both of these incubation temperatures mean generation time was longer at pH 5 than at pH 8. The high variation in growth of V. cholerae O139 in the distinct foods examined might have been due to differences in pH, fat content, and aw. Proper sanitary practices and storage of foods at refrigeration temperatures will help to reduce the possibility of growth by Vibrio cholerae O139 in foods to levels which do not imply a risk for food-poisoning.
Fermented fish and meat samples were purchased from supermarket and wet market for microbiological analysis of Listeria species and Listeria monocytogenes contamination. Listeria species were isolated from 17 (73.9%) of 23 samples of imported frozen beef, 10 (43.5%) of the 23 samples of local beef and 14 (56%) of the 25 samples of fermented fish from wet market. Listeria monocytogenes occurred in 15 (75%) of the frozen beef samples, 6 (30.4%) of the 23 samples of local meat and 3 (12%) of the 25 samples from fermented fish. Listeria species was not isolated from any of the 23 samples of imported frozen beef from supermarket and from the 5 samples of buffalo meat examined. This highlights the possibility of Listeria spp or L. monocytogenes to persist in meat and fermented fish in wet market and raises the problem of illness due to the handling and consumption of Listeria-contaminated meat or fermented fish are likely as evidence by the high contamination rates of samples sold at the wet market.
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 57 Vibrio vulnificus isolates from coastal water were characterized for their antimicrobial resistance, plasmid profiles and were typed by the PCR-based techniques: a random amplification of polymorphic DNA (RAPD) method and the enterobacterial repetitive intergenic consensus sequence (ERIC) method. All isolates were susceptible to chloramphenicol, nalidixic acid, tetracycline and trimethoprim-sulfamethoxazole. Fifty-one isolates were resistant to one or more of the other antibiotics tested. Plasmid analysis indicated that only 18 isolates carried small plasmids of 1.6 to 16 megadaltons. Analysis of the RAPD and ERIC DNA fingerprints of the V. vulnificus isolates with Gel Compare and cluster analysis software revealed significant genetic heterogeneity among these isolates. The combination of RAPD and ERIC analysis allowed us to distinguish all isolates. Thus, the combination of the two techniques is recommended for epidemiological investigation.
The objective of the present study was to evaluate the effect of electroporation on the membrane properties of lactobacilli and their ability to remove cholesterol in vitro. The growth of lactobacilli cells treated at 7.5 kV/cm for 4 ms was increased by 0.89 to 1.96 log(10) cfu/mL upon fermentation at 37 °C for 20 h, the increase being attributed to the reversible and transient formation of pores and defragmentation of clumped cells. In addition, an increase of cholesterol assimilation as high as 127.2% was observed for most cells electroporated at a field strength of 7.5 kV/cm for 3.5 ms compared with a lower field strength of 2.5 kV/cm. Electroporation also increased the incorporation of cholesterol into the cellular membrane, as shown by an increased cholesterol:phospholipids ratio (50.0-59.6%) upon treatment at 7.5 kV/cm compared with treatment at 2.5 kV/cm. Saturation of cholesterol was observed in different regions of the membrane bilayer such as upper phospholipids, apolar tail, and polar heads, as indicated by fluorescence anisotropy using 3 fluorescent probes. Electroporation could be a useful technique to increase the ability of lactobacilli to remove cholesterol for possible use as cholesterol-lowering adjuncts in the future.
We report for the first time on the prevalence, antibiotic resistance and RAPD types of Campylobacter species in ducks and duck related environmental samples in Malaysia. Samples were examined by enrichment in Bolton Broth followed by plating onto modified Charcoal Cefoperazone Deoxycholate agar (mCCDA) and/or plating directly onto mCCDA. A total of 643 samples were screened, and the prevalence of Campylobacter spp. in samples from different sources ranged from 0% to 85%. The method of isolation had a significant (P<0.05) effect on the isolation rate. One hundred and sixteen Campylobacter isolates, comprising of 94 Campylobacter jejuni, 19 Campylobacter coli and three Campylobacter lari, were examined for their sensitivity to 13 antibiotics. Majority of the C. jejuni isolates were resistant to cephalothin (99%), tetracycline (96%), suphamethoxazole/trimethoprim (96%), and very few were resistant to gentamicin (5%), chloramphenicol (7%) and erythromycin (1%). All C. coli isolates were resistant to cephalothin, nalidixic acid, norfloxacin and tetracycline but susceptible to chloramphenicol, erythromycin and gentamicin. The three C. lari isolates were resistant to all the antibiotics tested except chloramphenicol and gentamicin (1/3 and 2/3 susceptible, respectively). Genetic diversity of Campylobacter isolates were determined using random amplification of polymorphic DNA (RAPD). C. jejuni and C. coli isolates belong to fifty-eight and twelve RAPD types, respectively.
The aim of this study was to evaluate the effect of ultrasound on the intestinal adherence ability, cell growth, and cholesterol removal ability of parent cells and subsequent passages of Lactobacillus fermentum FTDC 1311. Ultrasound significantly decreased the intestinal adherence ability of treated parent cells compared to that of the control by 11.32% (P<0.05), which may be due to the protein denaturation upon local heating. Growth of treated parent cells also decreased by 4.45% (P<0.05) immediately upon ultrasound (0-4h) and showed an increase (P<0.05) in the viability by 2.18-2.34% during the later stage of fermentation (12-20 h) compared to that of the control. In addition, an increase (P<0.05) in assimilation of cholesterol (>9.74%) was also observed for treated parent cells compared to that of the control, accompanied by increased (P<0.05) incorporation of cholesterol into the cellular membrane. This was supported by the increased ratio of membrane cholesterol:phospholipids (C:P), saturation of cholesterol in the apolar regions, upper phospholipids regions, and polar regions of membrane phospholipids of parent cells compared to that of the control (P<0.05). However, such traits were not inherited by the subsequent passages of treated cells (first, second, and third passages). Our data suggested that ultrasound treatment could be used to improve cholesterol removal ability of parent cells without inducing permanent damage/defects on treated cells of subsequent passages.
This study aimed to evaluate the effect of ultrasound treatment on the cholesterol removing ability of lactobacilli. Viability of lactobacilli cells was significantly increased (P < 0.05) immediately after treatment, but higher intensity of 100 W and longer duration of 3 min was detrimental on cellular viability (P < 0.05). This was attributed to the disruption of membrane lipid bilayer, cell lysis and membrane lipid peroxidation upon ultrasound treatment at higher intensity and duration. Nevertheless, the effect of ultrasound on membrane properties was reversible, as the viability of ultrasound-treated lactobacilli was increased (P < 0.05) after fermentation at 37 °C for 20 h. The removal of cholesterol by ultrasound-treated lactobacilli via assimilation and incorporation of cholesterol into the cellular membrane also increased significantly (P < 0.05) upon treatment, as observed from the increased ratio of membrane C:P. Results from fluorescence anisotropies showed that most of the incorporated cholesterol was saturated in the regions of phospholipids tails, upper phospholipids, and polar heads of the membrane bilayer.
Catfish is one of the most cultivated species worldwide. Antibiotics are usually used in catfish farming as therapeutic and prophylactic agents. In the USA, only oxytetracycline, a combination of sulfadimethoxine and ormetoprim, and florfenicol are approved by the Food Drug Administration for specific fish species (e.g., catfish and salmonids) and their specific diseases. Misuse of antibiotics as prophylactic agents in disease prevention, however, is common and contributes in the development of antibiotic resistance. Various studies had reported on antibiotic residues and/or resistance in farmed species, feral fish, water column, sediments, and, in a lesser content, among farm workers. Ninety percent of the world aquaculture production is carried out in developing countries, which lack regulations and enforcement on the use of antibiotics. Hence, efforts are needed to promote the development and enforcement of such a regulatory structure. Alternatives to antibiotics such as antibacterial vaccines, bacteriophages and their lysins, and probiotics have been applied to curtail the increasing emergence of antibiotic-resistant bacteria due to the imprudent application of antibiotics in aquaculture.
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.
Ten strains of Salmonella weltevreden isolated from poultry sources were examined and found to contain plasmid DNA ranging in size from 1.8 to 68.5 MD. All isolates were susceptible to carbenicillin, cephalothin, ceftriazone, gentamicin, kanamycin and nalidixic acid, but resistance to bacitracin (100%), penicillin G (100%), rifampicin (100%), sulphamethoxazole (100%), cefuroxime (80%) and tetracycline (60%) was recorded. The 55 MD plasmid of strain SW5 determined resistance to penicillin G and tetracycline, which was transmissible to the E. coli K12 recipient at a frequency of 3.52 x 10(-5) transconjugants per input donor cell. The results of arbitrarily primed polymerase chain reaction (AP-PCR), using two 10-mer oligonucleotides and PCR-ribotyping to differentiate between the ten strains of S. weltevreden were compared. The strains were separated into ten different genome types by AP-PCR but were indistinguishable by PCR-ribotyping. These results suggest that poultry may constitute a reservoir for disseminating antibiotic resistance and that AP-PCR may be a valuable tool for epidemiological studies.