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Universal mitochondrial PCR combined with species-specific dot-blot assay as a source-tracking method of human, bovine, chicken, ovine, and porcine in fecal-contaminated surface water

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WATER RESEARCH 2009年第7期43卷 2002-2010页

作者： Kortbaoui, Randa Locas, Annie Imbeau, Marianne Payment, Pierre Villemur, Richard INRS Inst Armand Frappier Laval PQ H7V 1B7 Canada

Mitochondrial (mt) DNA has the potential to be used as an animal-specific genetic marker for source-tracking of fecal contamination in surface waters and groundwaters. In this report, we describe the development of a method to detect in a single assay human, bovine, ovine, porcine and chicken mitochondrial (mt) DNA in water. Consensus nucleic sequences were found between human, bovine, porcine, ovine and chicken mtDNA to design three sets of PCR universal primers. Upon polymerase chain reaction (PCR) with a digoxigenin-labeled nucleotide and the universal primers, species determination was carried out by dot-blotting membranes containing specific oligonucleotides for these five animals. Our method was carried out with three river samples and three wastewater samples, and the results were compared with those obtained by multiple nested PCR with specific primers for these five species. Our results showed that the dot-blot assays were as specific and sensitive as the nested-PCR approach. The proposed method has the advantage that it requires the use of only one PCR per sample and very little amounts of DNA. Finally, it is an alternative to multiplex PCR approach which is less sensitive, and shows the way for the development of DNA arrays for source-tracking of many more animal species in fecal-contaminated water. (C) 2009 Elsevier Ltd. All rights reserved.

关键词： source-tracking Water Fecal contamination Mitochondrial DNA DNA array

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Mitochondrial genome sequence variation as a useful marker for assessing genetic heterogeneity among Cyclospora cayetanensis isolates and source-tracking

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PARASITES & VECTORS 2019年第1期12卷 47-47页

作者： Guo, Yaqiong Wang, Yuanfei Wang, Xiaolan Zhang, Longxian Ortega, Ynes Feng, Yaoyu South China Agr Univ Coll Vet Med Minist Agr Key Lab Zoonosis Guangzhou 510642 Guangdong Peoples R China Henan Agr Univ Coll Anim Sci & Vet Med Zhengzhou 450002 Henan Peoples R China Univ Georgia Coll Agr & Environm Sci Griffin GA 30223 USA

Background: Cyclospora cayetanensis is an important enteric pathogen, causing diarrhea and food-borne cyclosporiasis outbreaks. For effective outbreak identification and investigation, it is essential to rapidly assess the genetic heterogeneity of C. cayetanensis specimens from cluster cases and identify the likely occurrence of outbreaks. Methods: In this study, we developed a quantitative PCR (qPCR) targeting the polymorphic link region between copies of the mitochondrial genome of C. cayetanensis, and evaluated the genetic heterogeneity among 36 specimens from six countries using melt curve, gel electrophoresis, and sequence analyses of the qPCR products. Results: All specimens were amplified successfully in the qPCR and produced melt peaks with different Tm values in the melt curve analysis. In gel electrophoresis of the qPCR products, the specimens yielded bands of variable sizes. Nine genotypes were identified by DNA sequencing of the qPCR products. Geographical segregation of genotypes was observed among specimens analyzed, which could be useful in geographical source-tracking. Conclusions: The length and nucleotide sequence variations in the mitochondrial genome marker allow rapid assessment of the genetic heterogeneity among C. cayetanensis specimens by melt curve, gel electrophoresis, or DNA sequence analysis of qPCR products. The sequence data generated could be helpful in the initial source-tracking of the pathogen.

关键词： Cyclospora cayetanensis Mitochondrion qPCR Genotyping source-tracking

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Rapid identification and source-tracking of Listeria monocytogenes using MALDI-TOF mass spectrometry

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INTERNATIONAL JOURNAL OF FOOD MICROBIOLOGY 2015年 202卷 1-9页

作者： Jadhav, Snehal Gulati, Vandana Fox, Edward M. Karpe, Avinash Beale, David J. Sevior, Danielle Bhave, Mrinal Palombo, Enzo A. Swinburne Univ Technol Fac Sci Engn & Technol Dept Chem & Biotechnol Hawthorn Vic 3122 Australia CSIRO Food & Nutr Werribee Vic 3030 Australia CSIRO Land & Water Flagship Highett Vic 3190 Australia BioMerieux Australia Pty Ltd Parkview Business Ctr Unit 25 Baulkham Hills NSW 2153 Australia

Listeria monocytogenes is an important foodborne pathogen responsible for the sometimes fatal disease listeriosis. Public health concerns and stringent regulations associated with the presence of this pathogen in food and food processing environments underline the need for rapid and reliable detection and subtyping techniques. In the current study, the application of matrix assisted laser desorption/ionisation-time-of-flight mass spectrometry (MALDI-TOF MS) as a single identification and source-tracking tool for a collection of L. monocytogenes isolates, obtained predominantly from dairy sources within Australia, was explored. The isolates were cultured on different growth media and analysed using MALDI-TOF MS at two incubation times (24 and 48 h). Whilst reliable genus-level identification was achieved from most media, identification at the species level was found to be dependent-on culture conditions. Successful speciation was highest for isolates cultured on the chromogenic Agar Listeria Ottaviani Agosti agar (ALOA, 91% of isolates) and non-selective horse blood agar (HBA, 89%) for 24 h. Chemometric statistical analysis of the MALDI-TOF MS data enabled source-tracking of L. monocytogenes isolates obtained from four different dairy sources. Strain-level discrimination was also observed to be influenced by culture conditions. In addition, t-test/analysis of variance (ANOVA) was used to identify potential biomarker peaks that differentiated the isolates according to their source of isolation. source-tracking using MALDI-TOF MS was compared and correlated with the gold standard pulsed-field gel electrophoresis (PFGE) technique. The discriminatory index and the congruence between both techniques were compared using the Simpsons Diversity Index and adjusted Rand and Wallace coefficients. Overall, MALDI-TOF MS based source-tracking (using data obtained by culturing the isolates on HBA) and PFGE demonstrated good congruence with a Wallace coefficient of 0.71 and comparab

关键词： Mass spectrometry Food safety Chemometrics source-tracking PFGE

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Preliminary results in B-spline enhanced source-tracking

Preliminary results in B-spline enhanced source-tracking

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9th IEEE Signal Processing Workshop on Statistical Signal and Array Processing (SSAP98)

作者： Green, RA Pierre, JW Univ Wyoming Dept Elect Engn Laramie WY 82071 USA

ISBN: (纸本)0780350103

source-tracking algorithms typically require estimation of an array's unknown correlation matrix. Time-averaging forms the basis of estimating the correlation matrix in most applications. This paper provides a preliminary investigation in the use of B-splines rather than time-averages to estimate the correlation matrix. The smooth, continuous character of splines can accurately match the behaviour of an array's true correlation matrix. Preliminary results indicate tracking estimates based on the spline-fit correlation:matrix have improved smoothness over time-averaging methods and can have comparable or better accuracy. Furthermore, B-splines are computationally efficient and are easily adapted to near real-time processing.

关键词： sensor array processing source-tracking nonstationary signals and time-varying systems B-splines

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Secure Complaint-Enabled source-tracking for Encrypted Messaging 21

Secure Complaint-Enabled Source-Tracking for Encrypted Messa...

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ACM SIGSAC Conference on Computer and Communications Security (ACM CCS)

作者： Peale, Charlotte Eskandarian, Saba Boneh, Dan Stanford Univ Stanford CA 94305 USA Univ N Carolina Chapel Hill NC USA

ISBN: (纸本)9781450384544

While the end-to-end encryption properties of popular messaging schemes such as Whatsapp, Messenger, and Signal guarantee privacy for users, these properties also make it very difficult for messaging platforms to enforce any sort of content moderation. This can lead to the unchecked spread of malicious content such as misinformation on such platforms. In 2019, Tyagi et al. developed message traceback, which addresses this issue by allowing a messaging platform to recover the path of a forwarded message after a user reports it for malicious content. This paper presents an alternative to message traceback that offers more privacy to users and requires less platform-side storage. We term this approach source-tracking for encrypted messaging schemes. source-tracking enables messaging platforms to provide the privacy guarantees expected from standard end-to-end encryption, but also helps hold the sources of malicious messages accountable: if malicious content is reported by a user, the source can be identified. We formalize security goals for source-tracking schemes and design and implement two source-tracking schemes with different security and performance tradeoffs.

关键词： tracing abuse reporting message franking end-to-end encrypted messaging source-tracking

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Detecting early-warning biomarkers associated with heart-exosome genetic-signature for acute myocardial infarction: A source-tracking study of exosome

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JOURNAL OF CELLULAR AND MOLECULAR MEDICINE 2024年第8期28卷 e18334页

作者： Jin, Xiaojun Xu, Weifeng Wu, Qiaoping Huang, Chen Song, Yongfei Lian, Jiangfang Ningbo Univ Affiliated Lihuili Hosp Hlth Sci Ctr Ningbo Zhejiang Peoples R China Xi An Jiao Tong Univ Affiliated Hosp 1 Dept Genet Xian Shaanxi Peoples R China

The genetic information of plasma total-exosomes originating from tissues have already proven useful to assess the severity of coronary artery diseases (CAD). However, plasma total-exosomes include multiple sub-populations secreted by various tissues. Only analysing the genetic information of plasma total-exosomes is perturbed by exosomes derived from other organs except the heart. We aim to detect early-warning biomarkers associated with heart-exosome genetic-signatures for acute myocardial infarction (AMI) by a source-tracking analysis of plasma exosome. The source-tracking of AMI plasma total-exosomes was implemented by deconvolution algorithm. The final early-warning biomarkers associated with heart-exosome genetic-signatures for AMI was identified by integration with single-cell sequencing, weighted gene correction network and machine learning analyses. The correlation between biomarkers and clinical indicators was validated in impatient cohort. A nomogram was generated using early-warning biomarkers for predicting the CAD progression. The molecular subtypes landscape of AMI was detected by consensus clustering. A higher fraction of exosomes derived from spleen and blood cells was revealed in plasma exosomes, while a lower fraction of heart-exosomes was detected. The gene ontology revealed that heart-exosomes genetic-signatures was associated with the heart development, cardiac function and cardiac response to stress. We ultimately identified three genes associated with heart-exosomes defining early-warning biomarkers for AMI. The early-warning biomarkers mediated molecular clusters presented heterogeneous metabolism preference in AMI. Our study introduced three early-warning biomarkers associated with heart-exosome genetic-signatures, which reflected the genetic information of heart-exosomes carrying AMI signals and provided new insights for exosomes research in CAD progression and prevention.

关键词： clustering early-warning heart-exosomes prediction source-tracking

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Human norovirus infection: surveillance and source tracking

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FUTURE VIROLOGY 2011年第4期6卷 431-438页

作者： Maunula, Leena von Bonsdorff, Carl-Henrik Univ Helsinki Dept Food Hyg & Environm Hlth FI-00014 Helsinki Finland

The clinical importance of noroviruses has increased in recent years. It is challenging to control the annual gastroenteritis and vomiting outbreaks caused by noroviruses in hospital wards and also long-term care facilities. This problem is partly due to the repeated emergence of highly penetrant genotype GII.4 variants that cause global epidemics. Consequently, this has led to an increased awareness of norovirus receptor selection and the population immunity characteristics that drive the evolution of the virus. Noroviruses mainly transmit from person to person by the feco-oral route and also via food, water and environmental fomites. International norovirus surveillance helps in source-tracking and in being able to follow the dissemination of viruses with food products in addition to the early detection of emerging variants.

关键词： genotyping molecular epidemiology norovirus RT-PCR source-tracking surveillance

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Surveillance and source tracking of foodborne pathogens in the vegetable production systems of India

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FOOD CONTROL 2024年 162卷

作者： Mohanapriya, Rangasamy Paranidharan, Vaikuntavasan Karthikeyan, Subburamu Balachandar, Dananjeyan Tamil Nadu Agr Univ Dept Agr Microbiol Coimbatore 641003 India Tamil Nadu Agr Univ Dept Plant Pathol Coimbatore 641003 India

Fresh vegetables are prone to foodborne pathogen infection due to improper cultivation methods and poor postharvest handling practices. This paper describes the prevalence of foodborne pathogens in three vegetables (tomato, brinjal, and lablab) cultivated by small-scale producers (SSP) of India. We assessed the possible entry of pathogens into the vegetable production chain. Samples from soil, manure, irrigation water, harvesting bins, and workers ' hands were assessed as sources of possible pathogen entry. In addition, further sampling was done to determine prevalence of these pathogens in the produce from the agricultural fields and markets. Total plate counts, yeast and mold count, total coliforms like general microbial indices, and possible foodborne pathogens like Salmonella , Escherichia coli O157, Staphylococcus aureus , Pseudomonas aeruginosa , and norovirus were assessed in all the samples by standard microbiological, biochemical, and molecular methods. The results revealed that among the three vegetables we studied, brinjal had the highest load of general microbial indicators. Moreover, the total coliforms count of market vegetables was higher than that of on -farm vegetables, indicating poor hygiene in post -harvest operations. The pathogen Pseudomonas aeruginosa was found in the market tomatoes (0.35 log 10 cfu/g), and E. coli was detected in the market samples of brinjal (2.98 -3.89 log 10 cfu/g) and lablab (0.59 -2.21 log 10 cfu/g), thereby revealing fecal cross -contamination during pre or post -harvest operations. None of the vegetable samples were contaminated with E. coli O157:H7, Staphylococcus aureus , Salmonella sp., and norovirus. However, the microbiological source tracking of primary production farms showed that E. coli O157: H7 was present in manure (9/27), soil (6/27), harvest bins (6/27), and irrigation water samples (3/27) of SSP farms. In addition, soil (33%), manure (11%), and harvest bins (11%) had Pseudomonas aeruginosa. In conclusion

关键词： Fresh vegetables Foodborne pathogens Microbial risk assessment Pre-harvest contamination source-tracking

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Genotyping and source tracking of Cryptosporidium by real-time PCR assays

Genotyping and source tracking of Cryptosporidium by real-ti...

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中国动物学会寄生虫学专业委员会第十届全国寄生虫学青年工作者学术研讨会

作者： Na Li Norman F.Neumann Norma Ruecker Yaoyu Feng Lihua Xiao School of Resources and Environmental Engineering East China University of Science and Technology Division of Foodborne Waterborne and Environmental Diseases Centers for Disease Control and Prevention School of Public Health University of Alberta

Objectives: The occurrence of Cryptosporidium oocysts in drinking source water can present a serious public health risk. To rapidly and effectively assess the source and human-infective potential of Cryptosporidium oocysts in water, sensitive detection and correct identification of oocysts to the species level(genotyping) are essential. Thus we developed three real-time PCR genotyping assays for rapid tracking of Cryptosporidium contamination. Methods: Light Cycler PCR and melt curve analysis using fluorescence resonance energy transfer(FRET) probes were used in the development of three real-time PCR assays, two targeting the small subunit(SSU) rRNA gene(18S-LC1 and 18S-LC2 assays) and one targeting the 90 k Da heat shock protein(hsp90) gene(hsp90 assay). We evaluated the sensitivity and Cryptosporidium species detection range of these assays. Results: The 18S-LC1 and hsp90 assay could differentiate common human-pathogenic species(C. parvum, C. hominis, and C. meleagridis), while the 18S-LC2 assay was able to differentiate non-pathogenic species(such as C. andersoni) from human-pathogenic ones commonly found in source water. In sensitivity evaluations, the 18S-LC2 and hsp90 genotyping assays could detect as few as one Cryptosporidium oocyst per sample, indicating it is possible for these two assays to detect low levels of oocysts in water samples. Conclusion: The 18S-LC2 and hsp90 genotyping assays might be used in environmental monitoring, whereas the 18S-LC1 genotyping assay could be useful for genotyping Cryptosporidium spp. in clinical specimens or wastewater samples.

关键词： Cryptosporidium Genotyping Real-time PCR Water source-tracking

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Overlooked dissemination risks of antimicrobial resistance through green tide proliferation

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WATER RESEARCH 2025年第PT B期268卷 122714页

作者： Zhao, Yanhui Zhang, Jian Zheng, Yu Shi, Jingliang Hu, Zhen Xie, Huijun Guo, Zizhang Liang, Shuang Wu, Haiming Shandong Univ Sch Environm Sci & Engn Shandong Key Lab Water Pollut Control & Resource R Qingdao 266237 Peoples R China Shandong Normal Univ Chem Engn & Mat Sci Jinan 250014 Peoples R China RIKEN Ctr Sustainable Resource Sci Saitama 3510198 Japan Nankai Univ Coll Environm Sci & Engn Tianjin 300071 Peoples R China Shandong Univ Environm Res Inst Qingdao 266237 Peoples R China

Green tides, particularly those induced by Enteromorpha, pose significant environmental challenges, exacerbated by climate change, coastal eutrophication, and other anthropogenic impacts. More concerningly, these blooms may influence the spread of antibiotic-resistant bacteria (ARB) and antibiotic resistance genes (ARGs) within ecosystems. However, the manner in which Enteromorpha blooms affect the distribution and spread of antimicrobial resistance (AMR) remains uncertain. This study investigated ARG profiles, dynamic composition, and associated health risks within the Enteromorpha phycosphere and surrounding seawater in typical bays (Jiaozhou, Aoshan, and Lingshan) in the South Yellow Sea. The Enteromorpha phycosphere exhibited significantly higher ARG abundance (p < 0.05) but lower diversity compared to the surrounding seawater. source-tracking and metagenomic analyses revealed that the phycosphere was the main contributor to the resistome of surrounding seawater. Moreover, resistant pathogens, especially ESKAPE pathogens, with horizontal gene transfer (HGT) potential, were more abundant in the phycosphere than in the surrounding seawater. The phycosphere released high-risk ARGs to the surrounding seawater during Enteromorpha blooms, posing serious health and ecological AMR risks in marine environments. This study highlights the significant role of Enteromorpha blooms in ARG spread and associated risks, urging a reassessment of AMR burden from a public health perspective.

关键词： Enteromorpha blooms Antibiotic resistance genes (ARGs) Metagenomics source-tracking Health risks

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