qPCR for the detection of foodborne Trypanosoma cruzi. 2017

Poliana Alves de Souza Godoi, and Claudio Adriano Piechnik, and Ana Caroline de Oliveira, and Michelle Zibetti Sfeir, and Emanuel Maltempi de Souza, and Hervé Rogez, and Vanete Thomaz Soccol
Unidade Integrada Sesi Senai, Niquelândia, Goiás, Brazil; Universidade Federal do Paraná, Department of Bioprocess Engineering and Biotechnology, Molecular Biology Laboratory, Curitiba, Paraná, Brazil.

Here we presented a potential real-time PCR (qPCR) method with public health importance and relevance for detection of Trypanosoma cruzi in açai pulp. There is not a current process to identify T. cruzi in açai, that ensures innocuity of this food concerning oral transmission. First, six new primers were designed using the DNA sequences of T. cruzi y152 and Emerald strains obtained from GenBank. For primers evaluation and titration they were validated regarding the amplification and not with the fluorophore chosen 1ngμL-1 of the T. cruzi DNA as target. For determination of the ideal concentration the titration of the primers drawn in this study showed T. cruzi DNA amplification in five primer pairs at concentrations 100, 200 and 300nM and DNA fixed concentrations at 1ngμL-1. For standardization all reactions were performed in triplicate with 5.0μL and positives and negatives controls were included in every run. As positive control DNA from two genotypes TcI and TcII were used. As negative control the reaction product without DNA of the parasite was used. The best primer concentration, for the expected fragments, was 300nM. From six primers improved the Ep1F/Ep1R primer detected 1×10-4ngμL-1 for both genotype of the parasite. The Bp1F/Bp1R showed amplification for 1.70.10-7ngμL-1 for TcI and 4.31.10-8ngμL-1 for TcII, based on the standard curve. The last step we tested the selected primers in qPCR for monitoring T. cruzi in açai pulp experimentally contaminated. The recovery rate for the TcII was 71%, whereas in açai samples contaminated with TcI it was 76%.

UI MeSH Term Description Entries
D011634 Public Health Branch of medicine concerned with the prevention and control of disease and disability, and the promotion of physical and mental health of the population on the international, national, state, or municipal level. Community Health,Environment, Preventive Medicine & Public Health,Environment, Preventive Medicine and Public Health,Health, Community,Health, Public
D005517 Foodborne Diseases Acute illnesses, usually affecting the GASTROINTESTINAL TRACT, brought on by consuming contaminated food or beverages. Most of these diseases are infectious, caused by a variety of bacteria, viruses, or parasites that can be foodborne. Sometimes the diseases are caused by harmful toxins from the microbes or other chemicals present in the food. Especially in the latter case, the condition is often called food poisoning. Food Poisoning,Food-borne Disease,Food-borne Illness,Foodborne Disease,Foodborne Illness,Food-borne Diseases,Food-borne Illnesses,Foodborne Illnesses,Poisoning, Food,Disease, Food-borne,Disease, Foodborne,Food Poisonings,Food borne Disease,Food borne Diseases,Food borne Illness,Food borne Illnesses,Illness, Food-borne,Illness, Foodborne,Illnesses, Foodborne
D005838 Genotype The genetic constitution of the individual, comprising the ALLELES present at each GENETIC LOCUS. Genogroup,Genogroups,Genotypes
D006801 Humans Members of the species Homo sapiens. Homo sapiens,Man (Taxonomy),Human,Man, Modern,Modern Man
D000818 Animals Unicellular or multicellular, heterotrophic organisms, that have sensation and the power of voluntary movement. Under the older five kingdom paradigm, Animalia was one of the kingdoms. Under the modern three domain model, Animalia represents one of the many groups in the domain EUKARYOTA. Animal,Metazoa,Animalia
D014349 Trypanosoma cruzi The agent of South American trypanosomiasis or CHAGAS DISEASE. Its vertebrate hosts are man and various domestic and wild animals. Insects of several species are vectors. Trypanosoma cruzus,cruzi, Trypanosoma
D014355 Chagas Disease Infection with the protozoan parasite TRYPANOSOMA CRUZI, a form of TRYPANOSOMIASIS endemic in Central and South America. It is named after the Brazilian physician Carlos Chagas, who discovered the parasite. Infection by the parasite (positive serologic result only) is distinguished from the clinical manifestations that develop years later, such as destruction of PARASYMPATHETIC GANGLIA; CHAGAS CARDIOMYOPATHY; and dysfunction of the ESOPHAGUS or COLON. Trypanosomiasis, South American,American Trypanosomiasis,Chagas' Disease,Trypanosoma cruzi Infection,Infection, Trypanosoma cruzi,Infections, Trypanosoma cruzi,South American Trypanosomiasis,Trypanosoma cruzi Infections,Trypanosomiasis, American
D016054 DNA, Protozoan Deoxyribonucleic acid that makes up the genetic material of protozoa. Protozoan DNA
D060888 Real-Time Polymerase Chain Reaction Methods used for detecting the amplified DNA products from the polymerase chain reaction as they accumulate instead of at the end of the reaction. Kinetic Polymerase Chain Reaction,Quantitative Real-Time PCR,Quantitative Real-Time Polymerase Chain Reaction,Real-Time PCR,PCR, Quantitative Real-Time,PCR, Real-Time,PCRs, Quantitative Real-Time,PCRs, Real-Time,Quantitative Real Time PCR,Quantitative Real Time Polymerase Chain Reaction,Quantitative Real-Time PCRs,Real Time PCR,Real Time Polymerase Chain Reaction,Real-Time PCR, Quantitative,Real-Time PCRs,Real-Time PCRs, Quantitative
D017931 DNA Primers Short sequences (generally about 10 base pairs) of DNA that are complementary to sequences of messenger RNA and allow reverse transcriptases to start copying the adjacent sequences of mRNA. Primers are used extensively in genetic and molecular biology techniques. DNA Primer,Oligodeoxyribonucleotide Primer,Oligodeoxyribonucleotide Primers,Oligonucleotide Primer,Oligonucleotide Primers,Primer, DNA,Primer, Oligodeoxyribonucleotide,Primer, Oligonucleotide,Primers, DNA,Primers, Oligodeoxyribonucleotide,Primers, Oligonucleotide

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