Food Samples Of Animal Origin Research Articles

Boronate affinity molecularly imprinted electrochemical sensor based on electrospun carbon nanofibres for selective kanamycin detection

The presence of trace kanamycin (KANA) residues in animal-derived foods from its improper use present a critical public health issue, making the urgent need for the development of sensitive and selective KANA detection methods. In this study, we developed molecularly imprinted polymers (MIPs)-based electrochemical sensor for the detection of KANA, incorporating Co, Mo-doped carbon nanofibres (Co, Mo@CNFs) to modify a glassy carbon electrode (GCE). Then, the MIPs layer was synthesized via electropolymerization using KANA as a template molecule and 3-aminophenylboronic acid (3-APBA) as the functional monomer (as MIPs/Co, Mo@CNFs/GCE), facilitating selective recognition through boric acid affinity. The presence of Co, Mo@CNFs enhanced electron transport across the otherwise insulating MIPs layer, thereby improving the sensor’s selective recognition capabilities. Under optimal conditions, the MIPs/Co, Mo@CNFs/GCE sensor demonstrated precise KANA detection within a concentration range of 1 × 10−2 to 1 × 105 nmol·L−1, achieving a detection limit of 2.56 pmol·L−1. Furthermore, the sensor’s applicability was validated in complex matrices, successfully detecting KANA in serum and milk samples with recovery rates ranging from 95.05 % to 116.43 %. These results confirmed that the proposed sensor is highly effective for detecting KANA in both biological and animal-origin food samples.

Foraging animal origin food samples as passive indicators of dioxin-like POPs contamination in industry sites: Method development, characterisation and risk assessment

Titanium dioxide (TiO2) is an important industrial chemical, and studies suggest its major production route - the chloride process could lead to the generation of unintentional dl-POPs. However, no relevant studies assessed the occurrence of dl-POPs associated with TiO2 production in the industrial zones, which is mostly due to the ultra-trace level distribution of these compounds in environmental compartments. The present study explored the novel possibility of utilising foraging animal-origin foods as sensitive indicators for addressing this challenge and generated a globally beneficial dataset by assessing the background levels of dl-POPs in the vicinity of a TiO2 production house in Southern India. Systematic sampling of foraging cow's milk and free-ranging hen's eggs was carried out from the study site, and the dl-POPs assessments were conducted utilising an in-house developed cost-effective GC-MS/MS-based analytical methodology. The median dl-POPs levels in milk and egg samples were about 3 times higher than the control samples collected from farm-fed animals and retail markets. The contaminant loads in the foraging animal-origin food samples were further traced to their presence in environmental compartments of soil and sediment and admissible degree of correlations were observed in congener fingerprints. Elevated health risks were inferred for the population in the industrial zones with weekly intakes weighing about 0.15–17 times the European Food Safety Authority-assigned levels. The consumption of foraging cow's milk was observed to have a higher contribution towards the hazard indices and cancer risk estimates and were significantly higher (p < 0.05) for children. The study also presents a critical validation of the GC-MS/MS-based method for the purpose of regulatory monitoring of dl-POPs, which could be of practical significance in economies in transition.

Embedded ZnO nanorod in ZIF-8 as a high-efficiency core–shell structural electrocatalyst for subnanomole level sensing of diethylstilbestrol

In view of the core–shell nanocomposites can complement the functions of the core and shell components, we here demonstrate a facile self-template strategy for synthesizing a one-dimensional rod-like hierarchical ZnO@ZIF-8 core–shell structure composed of a stabilized core and an active shell. The ZnO nanorods not only served as the template, but also as source of Zn ions for the in situ formation of zeolitic imidazolate framework-8 (ZIF-8) crystal shell, and the well-stabilized ZnO core preserved its own nature during the growth of the ZIF-8 shell. Furthermore, a sensing platform using ZnO@ZIF-8 as electrocatalyst was developed, which demonstrated salient electrocatalytic activity for diethylstilbestrol (DES) oxidation. The concentration of DES was directly proportional to its anodic peak current in the range of 2 nmol/L to 2 µmol/L, and ultralow detection limit of 0.73 nmol/L ( S/ N = 3) was achieved. The proposed method also gave excellent selectivity, reproducibility, and stability, showing promising analytical performance for the measurement of DES in food samples of animal origin.

Application of MALDI-TOF Mass Spectrometry for the Assessment of Prevalence of Listeria monocytogenes in Raw Milk, Dairy Products and Freshwater Fishes

Background: Foodborne infections like Listeriosis cover several disorders and are a worldwide public health emergency. Listeria monocytogenes has been isolated from various foodstuffs, including milk and fishes. MALDI-TOF mass spectrometry technique is known for its rapid and accurate identification of bacterial organisms. Methods: In the current research, a total of 360 samples comprising raw milk (130), milk products (125) and freshwater fishes (105) were screened for the detection of pathogenic Listeria species by using the USDA method. The recovered Listeria isolates were characterized using conventional set of biochemical analysis along with sugar fermentation tests and further confirmed by MALDI-TOF MS. The virulent nature of pathogenic Listeria isolates was also assessed by in vitro tests like hemolysis on blood agar, CAMP and PI-PLC assay. Result: In current study, on screening 360 animal origin food samples, three isolates were recovered from raw milk samples and identified as Listeria monocytogenes indicating an overall prevalence of Listeria monocytogenes to the tune of 0.83%. Excellent correlation was observed with identification of Listeria species using conventional phenotypic tests and advanced molecular tool Matrix Assisted Laser Desorption Ionization Time of Flight Mass Spectrometry (MALDI-TOF MS) technique. The results depict dependability of advanced technique for rapid and reliable identification of Listeria species.

Clostridioides difficile in food and food products of animal origin in Assam, India

ObjectiveConsidering the paucity of information about food-associated Clostridioides difficile from India, a study was undertaken to establish the prevalence of C. difficile in a variety of foods of animal origin, together with molecular strain characterization and antimicrobial resistance. MethodsA total of 235 samples comprising raw meat and meat products, fish products, and milk and milk products were screened for C. difficile. Toxin genes and other parts of PaLoc were amplified in isolated strains. The resistance pattern towards commonly used antimicrobial agents was studied by the Epsilometric test. ResultsC. difficile was isolated from 17(7.23%) different food samples of animal origin, including toxigenic (6) and non-toxigenic (11) isolates. In four toxigenic strains, the tcdA gene could not be detected under used conditions (tcdA-tcdB+). However, all strains had binary toxin-associated genes (cdtA and cdtB). The antimicrobial resistance was highest in non-toxigenic C. difficile isolates in food of animal origin. ConclusionMeat, meat products and dry fish, but not milk and milk products were contaminated with C. difficile. Contamination rates were low with diverse toxin profiles and antibiotic resistance patterns among the C. difficile strains.

Potential of simple, rapid, and non-target planar bioassay screening of veterinary drug residues

Veterinary drug residues in food samples of animal origin are currently analyzed by target analysis using high-performance liquid chromatography combined with sophisticated mass spectrometers. Since the results are only partially consistent with the microbiological results and positive findings occur rarely (in the per mil range in Germany), the potential of a simple planar bioassay screening was studied in the field of veterinary drug residue analysis. Using only a simple dilution of the milk for sample preparation, it was challenging to meet the maximum residue limits for antibiotic drug residues, exemplarily shown for the screening of two fluoroquinolones. However, the potential was evident for a simple, rapid, eco-friendly, and non-target screening without expensive instrumentation. Regardless of whether it is an active metabolite, contaminant, degradation product, or veterinary drug residue, the effect indicated on the planar surface due to bioassay detection will most likely also affect the human microbiome when consumed. The non-target screening of the milk samples revealed compounds with substantial antibacterial effects, which were not in the previous focus of interest. These antibacterial compounds will most likely also affect the human microbiome. Is it only the regulated antibiotic residues or generally all antibiotic compounds in a sample that count for consumer protection? The current prevailing understanding of food safety and antimicrobial resistance, based on the results of target (rather than effect) analyses, is being challenged. Non-target planar bioassay screening has been shown to fill a current gap by providing an understanding of inconsistencies and complementing routine target analysis of veterinary drug residues. As a highlight, it provides the full picture of the real levels of active compounds, regardless of the permitted limits of antibiotics.

Antibiogram of Extended Beta and Metallo-beta-lactamase E. coli O157:H7 Isolated from Pigs and Periwinkles sold in Rivers State

The emergence of multi-drug resistant and Extended Beta-lactamase producing E. coli O157:H7 is a general public health problem. Food samples of animal origin are undoubtedly the reservoirs of MDR which are in most cases transmitted to humans. The presence of extended beta-lactamase and Metallo-beta-lactamase producing E. coli O157:H7 was investigated amongst food samples and pig dung. A total of 240 samples comprising pig dung, pork, and periwinkles were collected from three different locations in Port Harcourt, Rivers State, Nigeria. Samples were analyzed for the presence of ESBL, MBL, and AmpC E. coli O157:H7 using standard microbiological methods. The results of the antibiotics susceptibility test of E. coli O157:H7 isolated from various samples revealed that out of the 16 E. coli O157:H7 isolated from the samples, the percentage resistance to Tetracycline, Ciprofloxacin, Ampicillin, Cotrimoxazole, Chloramphenicol, Amikacin, Cefetrizole, Gentamycin, Streptomycin, and Nalidixic was 62.5, 18.7, 100, 43.7, 50, 87.5, 87.5, 31.2, 50 and 56.3 %, respectively. While the antibiotics susceptibility pattern for E. coli O157 revealed that out of the 35 E. coli O157 isolated from the samples, the percentage resistance to Tetracycline, Ciprofloxacin, Ampicillin, Cotrimoxazole, Chloramphenicol, Amikacin, Cefetrizole, Gentamycin, Streptomycin and Nalidixic was 60, 22.9, 57.1, 42.9, 34.3, 60, 71.4, 42.9, 68.6 and 45.7 %, respectively. The results of the E. coli O157:H7 with Extended \(\beta\) ─Lactamase, Metallo Beta-Lactamase, and Amp C \(\beta\) ─Lactamase Production revealed that only 25.5 % were ESBL positive, 62.5 % MBL positive while 12.5 % was AmpC positive. The food samples do not only habour enteropathogenic E. coli but are reservoirs of beta-lactamases resistant genes which conferred multidrug resistance to the isolates. These food samples and dungs could serve as vehicles for the transmission of these resistant genes in the food chain. Thus, proper cooking of food samples and avoidance of pig dung is recommended.

Prevalence of Listeria monocytogenes in Foods of Animal Origin: Study from Assam, A North-Eastern State of India

Listeriosis is an important emerging bacterial food-borne zoonotic disease caused by Listeria monocytogenes. During the last two decades, the organism has been implicated in several food-borne disease outbreaks making it a serious public health hazard. The pathogenic strains of Listeria can cause severe illness in man after the ingestion of contaminated food products, especially the refrigerated foods of animal origin. In India, very few studies have been carried out in respect to the occurrence and incidence of L. monocytogenes. Assam, one of the eight states of Northeastern India, where approximately 79.0 percent of the total population are non-vegetarians. Chicken meat, pork, chevon and beef are exclusively taken as regular food items throughout the state. The present study describes about isolation, identification and molecular characterization of L.monocytogens from different foods of animal origin from Assam, India. A total of 750 different meat (chicken, chevon, pork and beef) and milk and their products (curd and paneer) samples were collected and analyzed for presence of Listeria. Isolation of L. monocytogenes was done from different food samples of animal origin by using selective enrichment method. PCR was employed for confirmation of the suspected cultures of Listeria targeting prsand16SrRNAgene. Upon confirmation, the isolates were further characterized based on 16S rRNA gene and phylogenetic analysis was performed. Of 750 samples, 48 (6.4%) samples were found positive. The highest rate of isolation of Listeria spp. was recorded in raw milk (8.95%) followed by curd (6.58%), chevon (5.36%), chicken (5.74%), pork (5.98%) and beef (3.75%). Molecular characterization of the isolates based on 16S rRNA gene of L.monocytogens isolated from foods of animal origin from Assam with that of other sequences reported from different parts of the world revealed that all the isolates of the present study are clustered along with other L.monocytogens isolates reported from different parts of the world, however, showing more closely related to isolates reported from China.

Occurrence and Antimicrobial Susceptibility Profile of Salmonella Isolates from Animal Origin Food Items in Selected Areas of Arsi Zone, Southeastern Ethiopia, 2018/19.

The status of Salmonella and its antimicrobial susceptibility profile in animal origin food items from different catering establishments in Ethiopia is scarce. Hence, this study aimed to investigate the occurrence and antimicrobial susceptibility profile of Salmonella isolates from animal origin food items in the selected areas of Arsi Zone. One hundred ninety-two animal origin food samples were collected and processed for Salmonella isolation. Isolates were tested for their susceptibility to 13 antimicrobials using Kirby–Bauer disk diffusion assay. An overall prevalence of 9.4% (18/192) Salmonella spp. isolates were recovered from animal origin food samples collected from different catering establishments. Seven (21.9%) of “Dulet,” 4 (12.5%) of “Kitfo,” 3 (9.4%) of “Kurt,” 2 (6.3%) of raw milk, 1 (3.1%) of egg sandwich and 1 (3.1%) of cream cake samples were positive for Salmonella. Catering establishments, protective clothing, source of contamination, manner of hand washing, and money handling were among the putative risk factors that were significantly associated (p < 0.05) with Salmonella spp. occurrence. Ampicillin, nitrofurans, and sulphonamide resistance were significantly associated (p < 0.05) with Salmonella spp. occurrence in the selected food items. Three (16.7%), 5 (27.8%), 5 (27.8%), and 4 (22.2%) of the isolates were resistant to 3, 4, 5, and 6 antibiotics, respectively, whereas only a sole isolate was resistant to two antibiotics (viz. ampicillin and kanamycin). In conclusion, the general sanitary condition of the catering establishments, utensils used, and personnel hygienic practices were not to the recommended standards in the current study. Besides, detection of multidrug-resistant strains of Salmonella in animal origin food items from different catering establishments suggests the need for detailed epidemiological and molecular characterization of the pathogen so as to establish the sources of acquisition of resistant Salmonella strains. Hence, implementation of Salmonella prevention and control strategies from farm production to consumption of animal origin food items are crucial.

One-step icELISA developed with novel antibody for rapid and specific detection of diclazuril residue in animal-origin foods

ABSTRACT Diclazuril, a broad-spectrum anticoccidial drug, may be accumulated in edible tissues of animals through illegal use, which poses potential threats to human health through the food chain. In this study, an innovative hapten was designed and an immunogen of diclazuril was successfully synthesised with keyhole limpet haemocyanin as carrier protein; then a monoclonal antibody with high specificity was obtained. Furthermore, based on the novel antibody, a one-step indirect competitive enzyme-linked immunosorbent assay (icELISA) was established for rapid and specific detection of diclazuril residues. Compared with the traditional icELISA method, this method saves at least 0.5 hours and one washing step. Under the optimal conditions, the one-step icELISA for diclazuril exhibited good performance with a 50% inhibition concentration (IC50) value of 0.952 μg/kg. The average recoveries of the icELISA ranged from 73.1% to 115.5% with the coefficient of variation lower than 12.7%, which was evaluated by detecting spiked animal-origin food samples. Finally, the one-step icELISA shows a good correlation with an ultra-high liquid chromatography-tandem mass spectrometry method. Those results demonstrate that the one-step icELISA developed for diclazuril detection is time-saving, low-cost, specific, sensitive, and reliable. It shows good potential for social, environmental, and economic benefits in future use.

Detection of Enterotoxin type B gene on Methicillin Resistant Staphylococcus aureus (MRSA) isolated from raw milk in East java, Indonesia

Background and Aim: Methicillin resistant Staphylococcus aureus (MRSA) has an exponential rate of spread throughout the world and its occurrence rates have been detected in animal / food samples of animal origin. The methicillin resistance (MR) strain is a significant factor in the potential virulence of Staphylococcus aureus such as the presence of Stapylococcal enterotoxin B (SEB) which induces a super antigenic effect. This study aims to detect the presence of SEB gene in MRSA isolates that isolated from cow milk in East Java province. Materials and Methods: Raw cow's milk ingredients were inoculated in enrichment media and mannitol salt agar (MSA) which were then tested using oxacillin and cefoxitin disc diffusion combined with oxacillin screen agar (ORSA) test to detect MRSA strains. All MRSA strains were detected by the SEB gene by the polymerase chain reaction (PCR) method. Results: In this study confirmed 18 MRSA from 150 samples of cow's milk and detected 4 MRSA isolates having SEB gene with product size 478 bp. In Conclusion: Detection of MRSA carrying the SEB gene in milk can have an impact on the public sector / animal health (milk borne disease) and the economy on livestock. It is very necessary to further analyze the relationship between various virulence factors in MRSA isolates that have the potential to be transmitted through cow's milk in East Java Keywords: Methicillin resistant Staphylococcus aureus(MRSA), Stapylococcal enterotoxin B (SEB), food safety, milk borne disease

The Twice-Oxidized Graphene Oxide/Gold Nanoparticles Composite SERS Substrate for Sensitive Detection of Clenbuterol Residues in Animal-Origin Food Samples

The illegal use of clenbuterol (CL), as a growth promoter in animal feeding, is banned in the majority of countries across the world, such as the European Union and China. In this study, a rapid and sensitive method for detecting CL in animal-origin food samples based on liquid-liquid extraction and surface-enhanced Raman spectroscopy (SERS) has been developed for the first time. A twice-oxidized graphene oxide/gold nanoparticles composite substrate was innovatively synthesized and used to enhance the characteristic Raman signal of CL at △v = 1474 cm− 1 and 1608 cm− 1, and no more than 10 min of sample cleanup was required before SERS analysis. The optimized hybrid substrate demonstrated good stability over a period of 240 days, which is more stable than previous studies. Quantitation calculation was performed in the concentration range of 1–100 ng g− 1 with a limit of detection (LOD) as low as 1 ng g− 1. Furthermore, the novel detection method was verified for use in the identification of real samples, and the results were consistent with those of liquid chromatography-tandem mass spectrometry (LC-MS/MS). The simplicity and reliability of the proposed method demonstrate its great potential for practical applications in the field of quality control and food safety.

Trace level analyses of selected perfluoroalkyl acids in food: Method development and data generation

To comprise the future requirements to detect low levels of perfluoroalkyl acids (PFAAs) including branched and linear perfluorooctane sulfonic acid (PFOS), perfluorooctanoic acid (PFOA), and perfluorohexane sulfonic acid (PFHxS) in food items, analytical methods for their determination in five different food matrices (cow milk, butter, chicken meat, beef, and fish) were developed and validated. Analytical method for eggs was only validated for PFOS and PFOA because of interfering substance appeared in some samples. The method applied on foods of animal origin includes alkaline digestion, extraction, and clean-up with solid phase extraction and adsorption on granular carbon where necessary. The method was shown effective to eliminate taurodeoxycholic acid (TDC), a bile acid that is an endogenous interference compound in egg samples causing ionization suppression and false positive result for PFOS when 499 > 80 transition was used for quantification. The validation was performed and resulted in recoveries >70% for all three PFAAs, the limits of quantification (LOQs) in all matrices were 3.1 pg g−1, 3.4 pg g−1, and 4.9 pg g−1 for PFHxS, PFOA, and L-PFOS, respectively. The optimized method was successfully applied to 53 food samples from the Swedish market and from developing countries. PFOS and PFOA were detected in all samples. PFHxS was detected in 76% of the samples. Further method development on separating interfering substance from PFHxS in egg is warranted due to relatively high detection of this compound in other food items. With this method, concentrations in the low pg g−1 range in food samples of animal origin were quantified including the branched PFOS isomers. This method can be applied to enforce potential future limit values for PFOS and PFOA as a consequence of the recent European Food Safety Authority (EFSA) recommendation where the tolerable intakes have been drastically lowered. Further method development is needed for foods of plant origin such as vegetables, flour, nuts, or bread.

Dietary exposure and risk assessment of short-chain chlorinated paraffins in supermarket fresh products in Jinan, China.

Dietary intake is the major route for the exposure of residents to short-chain chlorinated paraffins (SCCPs). SCCPs are classified as persistent organic pollutants (POPs) by the Stockholm Convention since May 2017. This study assessed the general population's dietary exposure to SCCPs through supermarket products. Five food categories, which included 40 food species, were collected from five supermarkets in April 2019. The SCCP concentrations in all food matrices varied between 58.6 and 1977ngg-1 dry weight (dw), with the average and standard deviation (SD) value of 301±379ngg-1 dw. Generally, the SCCP concentrations in animal-origin food matrices were higher than those in plant-origin food matrices. The C10Cl7 congeners were predominant among the congener groups of SCCPs. The proportion of C10 congeners in the animal-origin food samples (29.8%) was lower than that in the plant-origin food samples (39.7%), and the C13 congeners showed a contrasting result. The estimated daily intake (EDI) of SCCPs through dietary intake for the residents was 3109ngkg-1 day-1, which is much lower than the standards of European Food Safety Authority (10mgkg-1 day-1) for SCCPs. Risk assessment based on the supermarket foods indicated that SCCP exposure through dietary intake does not cause adverse effects to human health according to the margin of exposure (MOE).

Multi-analyte method for the analysis of legacy and alternative brominated and chlorinated flame retardants in food products of animal origin using gas chromatography - magnetic sector high resolution mass spectrometry

An analytical method was developed and validated for the analysis of 32 halogenated flame retardants (HFRs), including 13 polybrominated diphenyl ethers (PBDEs), 9 dechlorane-related compounds (DRCs) and 10 of the so-called alternative BFRs in food samples of animal origin. Gas chromatography (GC) coupled with magnetic sector high resolution mass spectrometry (HRMS) was used for the instrumental analysis. Intralaboratory validation of the developed method was performed in terms of recovery, repeatability, linear calibration ranges, instrumental and method limits of quantitation (i-LOQ and m-LOQ). Where possible, trueness was verified by the analysis of reference materials (RMs). For analytes with available isotopically labeled surrogates, recoveries in the range of 70–120% and repeatability rates below 20% were observed, while for compounds quantitated using absolute calibration the values of recovery ranged from 59 to 153% and the repeatability in terms of RSDs was in the range of 1–26%. The values measured for the four analyzed RMs agreed with the provided consensus values, revealing that the recovery of reference concentrations for compounds with assigned consensus values was in 72–119% range. The elaborated method met the sensitivity criteria according to the Commission Recommendation, 2014/118/EU on the monitoring of BFRs in food products. The developed method was successfully applied for the analysis of numerous food samples. The analyses revealed ubiquitous presence of PBDEs and the majority of DRCs in the analyzed samples, while alternative BFRs were detected only occasionally.

Polybrominated diphenyl ethers (PBDEs) in chicken eggs and cow milk around municipal dumpsites in Abuja, Nigeria

Polybrominated diphenyl ethers (PBDEs) are used as flame retardants in electronic equipment, as polymers in vehicles or construction, and in textiles. These additive flame retardants are emerging pollutants in Africa, released by the non-environmentally sound disposal of consumer products, often imported as secondhand, that have increasingly reached their end-of-life in the last decade. In Nigeria, which is a major receiver of e-waste and secondhand cars, there is a dearth of information regarding the levels of PBDEs in the environment, biota, and food. Thus, this study was designed to investigate the PBDE contamination of food samples of animal origin (chicken eggs and cow milk) around municipal waste dumpsites and background areas in Nigeria, to elucidate the role of dumpsites as potential sources of PBDE pollution and exposure in the country. Biological samples were collected over two years from two municipal waste dumpsites in Abuja.Fifty-six samples each of free-range chicken eggs and cow milk were collected. Control samples were collected approximately 5 km away from the dumpsites. After extraction and clean-up, the levels of POP-PBDEs listed in 2009 (major congeners of tetraBDE to heptaBDE), plus BDE-28 (Σ7PBDEs) were determined using GC-ECD. Data were analysed using descriptive statistics, t-test at α0.05. Levels of Σ7PBDEs (ng/g lipid weight (lw)) in chicken eggs at the two study sites ranged from 262.3 to 313.4 (ng/g lw), more than one order of magnitude higher than those at the control site in a village near the dumpsites (14.9 ± 3.73 ng/g lw), and two and more orders of magnitude higher compared to PBDE levels in these products in industrial countries and Nigerian supermarkets. Median PBDE levels in cow milk from the two dumpsites were 49.1 and 81.5 ng/g lw, respectively, considerably higher than levels in other studies. Proper disposal methods of waste polymers containing PBDEs such as co-incineration in BAT cement kilns and plastic recycling with separation is urgently needed in Nigeria and other parts of Africa, to prevent open burning as well as crude recycle to reduce PBDE levels in the environment and human food.

Rapid Detection of Shiga toxin-Producing E. Coli in Animal Origin Foods Using Loop-Mediated Isothermal Amplification (LAMP) Assay

The aim of this study was comparative evaluation of loop-mediated isothermal amplification (LAMP) and polymerase chain reaction (PCR) assay for rapid and inexpensive detection of shiga toxin-producing E. coli in animal origin foods by targeting stx1 and stx2 genes. The LAMP assay was performed using a water bath. The standardized LAMP assay was evaluated on 122 E. coli field isolates obtained from various animal origin food samples to ensure its reliability and usefulness. The result showed that conventional PCR could detect 68 (55.73%) and 75 (61.47%) positive E. coli isolates for stx1 and stx2 genes. Whereas, LAMP showed higher sensitivity by detecting 79 (64.75%) and 87 (71.31%) positive isolates of E. coli for stx1 and stx2 genes, respectively. LAMP assay was found to be highly specific and 10 times more sensitive as it could detect 1.11 × 102 cfu/ml for both stx1 and stx2 genes of E. coli isolates, whereas conventional PCR could detect 1.85 x 103 cfu/ml for both stx1 and stx2 genes of E. coli isolates. The rapidness, sensitivity, specificity, easiness and cost-effectiveness of LAMP assays will be very useful for the detection of foodborne pathogens for improving food sanitation and maintaining food safety.

Genotyping and distribution of putative virulence factors and antibiotic resistance genes of Acinetobacter baumannii strains isolated from raw meat

BackgroundAcinetobacter baumannii strains with multiple antimicrobial resistance are primarily known as opportunistic nosocomial bacteria but they may also be regarded as emerging bacterial contaminants of food samples of animal origin. Here we aimed to study the molecular characteristics of the A. baumanni strains isolated from raw meat samples.MethodsA total of 22 A. baumanni strains were isolated from 126 animal meat samples and were genotyped by ERIC-PCR method and by PCR detection of their virulence and antimicrobial resistance determinants. A. baumannii strains with 80% and more similarities were considered as one cluster.ResultsSixteen different genetic clusters were found amongst the 22 A. baumanni strains. Of the 22 strains, 12 (54.54%) had similar genetic cluster. A. baumannii strains exhibited the highest percentage of resistance against tetracycline (90.90%), trimethoprim (59.09%), cotrimoxazole (54.54%) and gentamicin (50.00%). TetA (81.81%), tetB (72.72%), dfrA1 (63.63%), aac(3)-IV (63.63%), sul1 (63.63%) and aadA1 (45.45%) were the most commonly detected antibiotic resistance genes. FimH (81.81%), afa/draBC (63.63%), csgA (63.63%), cnf1 (59.09%), cnf2 (54.54%) and iutA (50.00%) were the most commonly detected virulence factors. A. baumannii strains isolated from the chicken meat samples had the highest similarities in the genetic cluster.ConclusionsA. baumannii strains with similar genetic cluster (ERIC-Type) had the same prevalence of antibiotic resistance, antibiotic resistance genes and virulence factors. Genetic cluster of the A. baumannii strains is the main factor affected the similarities in the genotypic and phenotypic properties of the A. baumannii strains.

Poly[(2-(acryloyloxy) ethyl]trimethylammonium chloride-co-ethylene dimethacrylate monolith on-line solid-phase extraction coupled with liquid chromatography and tandem mass spectrometry for the fast determination of salicylic acid in foodstuffs.

We report the fabrication of an anion-exchange monolithic column in a stainless-steel chromatographic column (10mm×2.1mm i.d.) using [2-(acryloyloxy) ethyl]trimethylammonium chloride as the monomer and ethylene dimethacrylate as the crosslinker. The prepared monolith was developed as the adsorbent for the on-line solid-phase extraction of salicylic acid in various animal-origin foodstuffs combined with liquid chromatography and tandem mass spectrometry. The monolith was characterized by using Fourier transform infrared spectroscopy, scanning electron microscopy, nitrogen adsorption analysis, and elemental analysis. Potential factors affecting the on-line solid-phase extraction and liquid chromatography with tandem mass spectrometry analysis were studied in detail. Under the optimized conditions, the total analysis time including cleanup and liquid chromatography with tandem mass spectrometry separation was 17min. The developed method gave the linear range of 15-750μg/kg, detection limits (S/N=3) of 5μg/kg, and quantification limits (S/N=10) of 15μg/kg. The recoveries obtained by spiking 10, 20, and 100μg/kg of salicylic acid in the animal-origin food samples were in the range of 85.2-98.4%. In addition, the monolith was stable enough for 550 extraction cycles with the precision of peak area ≤11.6%.

Diversity of CTX-M-1-producing E. coli from German food samples and genetic diversity of the blaCTX-M-1 region on IncI1 ST3 plasmids

Antimicrobial resistance to cephalosporins is commonly mediated by extended-spectrum β-lactamases (ESBL) or plasmidic AmpC β-lactamases (pAmpC). In livestock blaCTX-M-1 is the most frequently detected ESBL-encoding gene. As transmission to consumers through contaminated food is often proposed, this study characterized ESBL/pAmpC-producing E. coli collected from food samples. Therefore, samples from food products of animal origin and vegetables were screened for phenotypically resistant E. coli by selective cultivation. The ESBL genotype was confirmed for 404 isolates with the majority of them (n = 212) harboring the blaCTX-M-1 gene. PFGE and MLST analyses as well as plasmid characterization were carried out for 89 isolates, selected under epidemiological aspects. In addition, 44 isolates were investigated by whole genome sequencing and/or sequencing of their plasmids on an Illumina Miseq platform.MLST and PFGE indicated a diverse population of CTX-M-1-producing E. coli in German food samples with no spread of single clonal lineages. The majority of the isolates harbored the blaCTX-M-1 gene on IncI1 plasmids. Frequently, the gene was associated with the ISEcp1 element and located on a ∼100 kb IncI1 plasmid depicting the plasmid multilocus sequence type (ST) 3. The blaCTX-M-1 gene and its flanking sequences were located within the shufflon of the type IV pilus region in diverse orientations.In conclusion, dissemination of the CTX-M-1 β-lactamase within food samples of animal origin is driven by the transmission of a ∼100 kb large IncI1 ST3 plasmid. Apart from conjugal transfer of IncI1 ST3 plasmids the transmission of the blaCTX-M-1 gene might be further promoted through mobilization due to its location within a recombination hot-spot of IncI1 plasmids.