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Parasites in Fresh Produce: a Brazilian Inter-laboratory Evaluation of a Standardized methodology for the Detection of Ascaris sp. in Leafy Vegetables

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FOOD ANALYTICAL methodS 2021年第5期14卷 989-996页

作者： Pineda, Carolina Ortiz Leal, Diego Averaldo Guiguet de Lima, Roberta Ribeiro, Patricia Padilha Rodrigues, Amanda Martini, Maria Helena Stancari, Regina Celia Arantes Matosinhos, Flaviane Cristina Lopes Mine, Julio Cesar Greinert-Goulart, Juliane Araujo Grott, Suelen Cristina Diefenbach, Lucia Maria Guedes Araujo, Beatriz Franco, Regina Maura Bueno Univ Estadual Campinas UNICAMP Lab Protozool Campinas SP Brazil Univ Fed Parana UFPR Lab Parasitol Ambiental Curitiba Parana Brazil Inst Adolfo Lutz IAL Campinas SP Brazil Inst Adolfo Lutz IAL Nucleo Ciencias Quim & Bromatol Bauru SP Brazil Fundacao Ezequiel Dias FUNED Belo Horizonte MG Brazil Univ Estadual Ponta Grossa UEPG Dept Anal Clin & Toxicol Ponta Grossa Parana Brazil Fundacao Univ Reg Blumenau FURB Lab Parasitol Blumenau SC Brazil Lab Cent Saude Publ LACEN Div Anal Prod Secao Microscopia & Triagem Porto Alegre RS Brazil

Vegetables are potential vehicles for the transmission of parasites. Helminth eggs have been detected in produce in various countries around the world, with high prevalence rates. However, there is no standard validated method or ISO to the isolation, concentration, and identification of helminth eggs in salad vegetables, which makes comparison difficult since each study utilized different analytical methods and food matrix. The purpose of the present study was to evaluate the effectiveness of a previously described methodology developed for the recovery of Ascaris sp. eggs in two different leaf greens under real conditions of use, through the simultaneous application of the same method by ten different laboratories. This is the first collaborative inter-laboratory trial by several laboratories across Brazil. Lettuce and arugula samples artificially contaminated with A. suum eggs were analyzed by various laboratories to determine the recovery efficiency of Ascaris sp. eggs. A final mean recovery efficiency of 57.10% for Ascaris sp. eggs in lettuce and 50.7% in arugula was identified. The methodology applied in this study can be used to evaluate Ascaris sp. eggs in vegetables. It is reasonably easy, fast, and efficient in comparison with other methods. The identification of helminth eggs in fresh vegetables using a reliable methodology is important if the microbiological safety of these foods is to be improved.

关键词： Ascaris sp. Foodborne pathogens Recovery rapid method Vegetables

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Genus-level identification of dermatophytes by MALDI-TOF MS after 2days of colony growth

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LETTERS IN APPLIED MICROBIOLOGY 2018年第2期67卷 136-143页

作者： Intra, J. Sarto, C. Tiberti, N. Besana, S. Savarino, C. Brambilla, P. Univ Milano Bicocca Desio Hosp Dept Lab Med Via Mazzini 1 I-20833 Desio MB Italy Univ Geneva Translat Biomarker Grp Fac Med Geneva Switzerland

Matrix-assisted laser desorption-ionization time-of-flight mass spectrometry (MALDI-TOF MS) is becoming a popular technology in clinical microbiology. It is a fast and highly specific method for the routine identification of micro-organisms. In this study, we evaluated the suitability of dermatophyte identification after only 2days of colony growth using MALDI-TOF MS. Two protein extraction protocols were also evaluated consisting of either formic acid alone or of ethanol-formic acid-acetonitrile to achieve a complete protein extraction. Morphology-based techniques were used as the diagnostic standard methods and MALDI-TOF MS results were obtained using the manufacturer's spectral library. Using the formic acid protein extraction protocol after 2days of colony growth, 70 and 46% of dermatophytes were properly identified at the genus and species-level respectively. The addition of ethanol-formic acid-acetonitrile extraction protocol increased the identification to 90 and 62%. Based on our observations, we propose a two-step workflow for the fast and reliable identification of dermatophytes after only 2days of colony growth. This flow chart consists of a first direct deposition procedure with the addition of formic acid, followed by a complete protein extraction when dermatophyte identification is not successful. Significance and Impact of the StudyIn this study, a two-step workflow for the identification of clinical dermatophytes using MALDI-TOF analysis and commercially available spectral library was developed. The workflow consists of an initial direct deposition of the sample on the MALDI plate and formic acid protein extraction at 2days of growth culture;if dermatophyte identification is not successful, a complete protein extraction using ethanol-formic acid-acetonitrile is subsequently performed. Using this workflow, the correct isolate identifications increase up to 90%;of these, 27% are identified at the genus-level, providing sufficient information to start a

关键词： dermatophyte identification incubation time MALDI-TOF MS rapid method

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A combination of direct viable count and fluorescence in situ hybridization for specific enumeration of viable Lactobacillus delbrueckii subsp bulgaricus and Streptococcus thermophilus

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LETTERS IN APPLIED MICROBIOLOGY 2012年第3期54卷 247-254页

作者： Garcia-Hernandez, J. Moreno, Y. Amorocho, C. M. Hernandez, M. Univ Politecn Valencia Dept Biotecnol Valencia 46022 Spain

Aims: We have developed a direct viable count (DVC)-FISH procedure for quickly and easily discriminating between viable and nonviable cells of Lactobacillus delbrueckii subsp. bulgaricus and Streptococcus thermophilus strains, the traditional yogurt bacteria. methods and Results: direct viable count method has been modified and adapted for Lact. delbrueckii subsp, bulgaricus and Strep. thermophilus analysis by testing different times of incubation and concentrations of DNA-gyrase inhibitors. DVC procedure has been combined with fluorescent in situ hybridization (FISH) for the specific detection of viable cells of both bacteria with specific rRNA oligonucleotide probes (DVC-FISH). Of the four antibiotics tested (novobiocin, nalidixic acid, pipemidic acid and ciprofloxacin), novobiocin was the most effective for DVC method and the optimum incubation time was 7 h for both bacteria. The number of viable cells was obtained by the enumeration of specific hybridized cells that were elongated at least twice their original length for Lactobacillus and twice their original size for Streptococcus. Conclusions: This technique was successfully applied to detect viable cells in inoculated faeces. Significance and Impact of the Study: Results showed that this DVC-FISH procedure is a quick and culture-independent useful method to specifically detect viable Lact, delbrueckii subsp, bulgaricus and Strep thermophilus in different samples, being applied for the first time to lactic acid bacteria.

关键词： direct viable count FISH Lactobacillus delbrueckii subsp bulgaricus rapid method Streptococcus thermophilus

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Novel perspectives to purify genomic DNA from high humic acid content and contaminated soils

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SEPARATION AND PURIFICATION TECHNOLOGY 2010年第1期75卷 81-86页

作者： Techer, Didier Martinez-Chois, Claudia D'Innocenzo, Marielle Laval-Gilly, Philippe Bennasroune, Amar Foucaud, Laurent Falla, Jairo Univ Metz Paul Verlaine IUT Thionville Yutz LIEBE CNRSUMR 7146 F-57970 Espace Cormontaigne Yutz France

Numerous soil nucleic acid isolation protocols published during the last decades reflect difficulties to remove all traces of DNA-coprecipitated contaminants such as humic acids and pollutants. The aim of this work was to optimize a rapid and innovative DNA isolation and purification procedure from high humic acid content and contaminated soil. Two types of vitamins, pyridoxal hydrochloride and thiamine hydrochloride, were used as PCR inhibitor removals. The combination of vitamins with a "salting-out" with potassium chloride (KCl) during the optimization experiments enabled the recovery of PCR-ready DNA extracts from polluted soils with high organic matter contents. To investigate the extent of this purification method, genomic nucleic acids were also extracted from soil samples artificially supplemented with 10% (w/w) of pure humic acids. Successful PCR amplifications confirmed the high purity of the DNA recovered from all samples. The use of cetyl trimethylammonium bromide (CTAB) with vitamins instead of KCI prevented DNA losses and also allowed the recovery of high quality genomic DNA from the different soils, depending on lysis buffer pH adjustment. The application of this protocol followed by bacterial diversity analysis by PCR-TGGE demonstrated that this method could be applied to a bacterial molecular diversity analysis from a wide variety of soils. The major advantage of the described procedure is the inherent short processing time with no preliminary soil washing requirement or additional purification step. (C) 2010 Elsevier B.V. All rights reserved.

关键词： DNA purification Soils Humic acid Vitamins rapid method

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Comparison between a cultural procedure using Rappaport-Vassiliadis broth and motility enrichments on modified semisolid Rappaport-Vassiliadis medium for Salmonella detection from food and feed

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INTERNATIONAL JOURNAL OF FOOD MICROBIOLOGY 1996年第2-3期29卷 353-360页

作者： Wiberg, C Norberg, P Biology Division National Food Administration Box 622 S-757 26 Uppsala Sweden

Motility enrichment on modified semisolid Rappaport-Vassiliadis (MSRV) medium was evaluated for a rapid Salmonella detection in naturally contaminated food- and feed samples. The results of this procedure were compared with a conventional cultural procedure using Rappaport-Vassiliadis (RV) broth as selective enrichment and modified brilliant green-, xylose lysine desoxycholate- and mannitol lysine crystal violet brilliant green-, agar for selective plating. The productivity of motility enrichment was 87% compared to a productivity of 99% for the cultural procedure. Statistical analysis showed that there was a significant difference at the 5% level. The present study shows that a combination of direct motility enrichment with the cultural procedure could be used as a very rapid and highly cost-efficient method for Salmonella detection.

关键词： Salmonella food rapid method Rappaport-Vassiliadis medium motility enrichment

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Authentication of sweetness of mango juice using Fourier transform infrared-attenuated total reflection spectroscopy

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JOURNAL OF FOOD ENGINEERING 2010年第3期101卷 337-342页

作者： Jha, S. N. Gunasekaran, S. Cent Inst Postharvest Engn & Technol Ludhiana 141004 Punjab India Univ Wisconsin Madison WI 53706 USA

Fourier transform infrared spectrometer equipped with attenuated total reflection and chemometrics were used to determine added sugar content (ASC), total soluble solids (TSS) and real juice content (RJC) in fresh and commercial mango juice. Sucrose solutions (0-27%), fresh mango juice adulterated with 0-27% sucrose, and two commercial brands were evaluated in wavenumber range of 4000-650 cm(-1). Partial least squares (PLS) discrimination and principal component analysis (PCA) were used to classify the samples with or without ASC. PLS and multiple linear regression (MLR) were carried out with and without data treatments. The detection limit for ASC was 3% for samples with low natural TSS, 5% for samples with natural TSS more than 10% and 3.6% for commercial samples. ASC, TSS, and RJC were predicted in the wavenumber range of 1476-912 cm(-1) using PLS (multiple correlation coefficient, R = 0.99) and three wavenumbers (1088, 1050, 991 cm(-1)) using MLR (R = 0.98). (c) 2010 Elsevier Ltd. All rights reserved.

关键词： Food quality Adulteration Fruit juice rapid method

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EVALUATION OF MOTILITY ENRICHMENT ON MODIFIED SEMISOLID RAPPAPORT-VASSILIADIS MEDIUM (MSRV) AND AUTOMATED CONDUCTANCE IN COMBINATION WITH RAMBACH AGAR FOR SALMONELLA DETECTION IN ENVIRONMENTAL-SAMPLES OF A MILK POWDER FACTORY

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INTERNATIONAL JOURNAL OF FOOD MICROBIOLOGY 1994年第2-3期22卷 201-206页

作者： JOOSTEN, HMLJ VANDIJCK, WGFM VANDERVELDE, F Coberco Research P.O. Box 87 NL 7400 AB Deventer The Netherlands

The efficacy of motility enrichment on modified semisolid Rappaport-Vassiliadis medium (MSRV) and an automated conductance method for the detection of Salmonella in environmental samples was evaluated. Two hundred and ten environmental samples from unrestricted areas of a milk powder factory, 49 of which were artificially contaminated with Salmonella infantis, were examined. From exactly 100 samples Salmonella could be isolated. With the conventional (ISO-DIS 6579) method a 100% score was obtained, whereas the MSRV method gave 82 positive results. The conductance method permitted the detection of Salmonella in only 66 samples. The use of Rambach agar improved isolation efficiency of Salmonella from enrichment broths.

关键词： SALMONELLA MILK POWDER rapid method RAMBACH AGAR

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Comparative studies of magnetic particle-based solid phase fluorogenic and electrochemiluminescent immunoassay

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JOURNAL OF IMMUNOLOGICAL methodS 1998年第1-2期218卷 1-8页

作者： Yu, H Veridian Corp ERDEC Aberdeen Proving Ground MD 21010 USA

Two solid phase immunoassays, an electrochemiluminescent immunoassay (ECLIA) and a magnetic particle fluorogenic immunoassay (MPFIA) were evaluated and compared for bacterial detection. Briefly, the ECLIA is based on a redox reaction between ruthenium (II)-trisbipyridyl Ru[(bpy)(3)](2+) labeled antibody and the excess of tripropylamine, which generates photons. The entire reaction is carried on the near surface area between the spherical magnetic beads and an anode electrode. The detectable bacterial spores are at a linear range from 5 x 10(3) to 5 x 10(5) colony forming units (cfu) of Bacillus subtilis var. niger spores, 10(2) to 10(4) cfu of Bacillus anthrax spores and 10(2) to 10(6) cells of Escherichia coli O157:H7 in ECLIA. The unique MPFIA technique employs antibody-coated magnetic beads as solid phase in suspension for bacterial capture and concentration in a 96-well microplate format. Primary capturing antibodies, bacteria form a sandwich with alkaline phosphatase (AP)-labeled antibodies as reporter followed by a reaction with the AP substrate, AttoPhos(TM) to generate fluorescence for detection. Immunomagnetic separation permits direct isolating and concentrating bacterial cells from the crude samples, such as blood and environmental water. The results of MPFIA for detecting bacteria showed less sensitivity compared with that of ECLIA, however it provides a means for direct, high throughput screening bacteria from crude biological samples. Both ECLIA and MPFIA are rapid (less than one hour) and easy to use. (C) 1998 Elsevier Science B.V. All rights reserved.

关键词： solid phase immunoassay microplate magnetic beads bacteria detection rapid method

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Development of a multiplex real-time recombinase polymerase amplification (RPA) assay for rapid quantitative detection of Campylobacter coli and jejuni from eggs and chicken products

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FOOD CONTROL 2017年 73卷 1247-1255页

作者： Kim, Ji Yeun Lee, Jung-Lim Delaware State Univ Coll Agr & Related Sci Food Sci & Biotechnol Program Dept Human Ecol Dover DE 19901 USA

Chicken products are known as main vehicles for human infections and foodborne illness with campylobacteriosis. This study describes a rapid and sensitive method for the detection of C. coli and C. jejuni from eggs, chicken meat, and chicken broth using the multiplex real-time recombinase polymerase amplification (Rti-RPA) and its comparison to a culture dependent method. The optimal procedure for the multiplex Rti-RPA assay was set in isothermal condition at 45 degrees C for 20 min and the detection sensitivity was at 1 CFU/reaction in pure culture. In the case of food applications, the detection limit of C. coli and C. jejuni using the RPA assay was 1 CFU/mL from chicken broth, 10(3) CFU/g from egg and chicken meat samples without a pre-enrichment procedure, and 1 CFU/g from 24 h enriched egg and chicken meat samples. Quantifications of Campylobacter spp. in spiked food samples by the RPA assay were not significantly different with the results by the culture method. This multiplex Rti-RPA assay is considerably faster than other DNA amplification methods and exhibits no losses in detection sensitivity and specificity. This multiplex Rti-RPA is a simple, rapid detection and quantification assay that is completed within a 20 min runtime. The assay is applicable in the surveillance of C. coli and C. jejuni contamination in eggs and chicken products. (C) 2016 Elsevier Ltd. All rights reserved.

关键词： Campylobacter RPA rapid method Food safety Poultry products

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A head space gas chromatographic approach for the monitoring of the microbial cell activity and the cell viability evaluation

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JOURNAL OF MICROBIOLOGICAL methodS 1997年第2期29卷 103-114页

作者： Gardini, F Lanciotti, R Sinigaglia, M Guerzoni, ME UNIV BOLOGNA DIPARTIMENTO PROT & VALORIZZONE AGROALIMENTAREI-40126 BOLOGNAITALY UNIV BARI IST PROD & PREPARAZ ALIMENTARII-71100 FOGGIAITALY

A gas chromatographic method, based on the analysis of volatile metabolites in the head space of cultures or food systems has been developed and is widely used for the modelling and prediction of microbial behaviour in food systems and fermentation processes. In this paper, different applications based on two experimental approaches are described: (1) the detection of contamination levels of specific organisms or groups on the basis of previously prepared calibration curves;(2) the monitoring of the activity of microorganisms actually present in fermentation medium or in food model systems, by means of periodical gas chromatographic analysis of the vapours of the head space. The sensitivity and linearity within a value range from 10(1) to 10(8) g CFU/ml are the principal advantages offered by the head space gas chromatographic approach for detecting the contamination level of a system, based on previously prepared calibration curves. Moreover the gas chromatographic analysis of the head space presents the advantages of being inexpensive, relatively rapid and giving linear responses within wide intervals of cell loads. Preliminary research demonstrated that good selectivity could be obtained for Salmonella by inoculating samples in duplicate in a medium containing dulcitol as the only carbon source and with and without the presence of a polyvalent H antiserum as already used for radiometric Salmonella detection. The monitoring of the metabolic activity of microorganisms, in real or model systems, by means of an automatic GC analysis of the vapour in equilibrium in the head space, provided data suitable for analysis with equations, previously developed for data expressed as lop. CFU, The possibility of fitting CO2 data by means of equations, in which parameters with biological significance are calculated, is an important instrument for modelling the growth of microorganisms as a function of the environmental variables. (C) 1997 Elsevier Science Ireland Ltd.

关键词： head space gas chromatography growth modelling carbon dioxide rapid method indirect method

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