scholarly journals Molecular Detection of Rotavirus in Mollusks from the Oued El Maleh Estuary of Mohammedia, Morocco

2021 ◽  
Author(s):  
Abderrahim Hatib ◽  
Najwa Hassou ◽  
Abdelouahab Benani ◽  
Jamal Eddine Hafid ◽  
Moulay Mustapha Ennaji
Keyword(s):  
Real Time ◽  
Enteric Bacteria ◽  
Human Populations ◽  
Rt Pcr ◽  
Wet Season ◽  
Rotavirus A ◽  
Viral Loads ◽  
Wide Range ◽  
Group A Rotaviruses ◽  
Group A

Viral outbreaks can result from the consumption of contaminated bivalve mollusks. However, despite the regulation related to enteric bacteria in food products, the consumption of raw and undercooked mollusks remains linked to viral epidemics in human populations. Real-time RT-PCR is a highly sensitive approach for detecting and quantifying enteric viruses, and after eliminating enzymatic amplification inhibitors from samples of interest, sensitive and specific tests, like real-time RT-PCR, can facilitate the detection and quantification of a wide range of viruses that are concentrated in mollusk digestive tissues. The aim of the present study was to evaluate the prevalence of Group-A rotaviruses in mussel (Mytilus edulis Linnaeus, 1758) specimens (n=576) collected downstream of the Oued El Maleh Estuary, which is along the coast of Mohammedia City in Morocco, using real-time RT-PCR. Rotavirus A RNA was detected in 37.5% (n=18) of the 48 sample batches, and viral loads ranged from 0.42×101 to 1.8603×104 genomic copies per g digestive tissue. Most (72.22%) of the positive samples were collected during the wet season (September-April), and the probability of detecting rotaviruses was significantly greater during the wet season than during the dry season (P<0.001). Monitoring Rotavirus A and similar viruses in shellfish may help prevent viral contamination and preserve public health.

scholarly journals Electropherotypes and G-Types of Group A Rotaviruses Detected in Children with Diarrhea in Lagos, Nigeria

ISRN Virology ◽  
2013 ◽  
Vol 2013 ◽  
pp. 1-5 ◽  
Author(s):  
Christianah Idowu Ayolabi ◽  
David Ajiboye Ojo ◽  
George Enyimah Armah
Keyword(s):  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Migration Pattern ◽  
Genomic Diversity ◽  
Rt Pcr ◽  
Group A Rotaviruses ◽  
Group A ◽  
Stool Samples ◽  
Health Care Centers ◽  
Virus Isolates

Approximately over 500,000 children die annually due to severe dehydrating diarrhea caused by rotaviruses. This work investigated rotavirus infection among children less than 5 years with diarrhea in Lagos and determined the circulating electropherotypes and genotypes of the virus isolates. Three hundred and two (n=302) stool samples from children below 60 months were collected from different hospitals and health care centers in Lagos and subjected to enzyme immunoassay (EIA) to determine the presence of Group A rotavirus, RT-PCR to determine the G-types, and polyacrylamide gel electrophoresis (PAGE) to determine the electropherotypes. The results show that 60.3% of the samples showed distinct rotavirus RNA migration pattern, having long electropherotypes (55.3%) of seven variations dominating over the short electropherotypes (44.5%). Six different G-types were detected (G1, G2, G3, G4, G9, and G12). Serotypes G1 and G12 showed long electropherotypic pattern while G2, G3, and G9 exhibited either short or long electropherotype. All G4 detected show short electropherotypic pattern. In conclusion, information on the genomic diversity and RNA electropherotypes of rotaviruses detected in children with diarrhea in Lagos is reported in this study.

Development of a real-time reverse transcription-PCR assay to detect and quantify group A rotavirus equine-like G3 strains

2021 ◽  
Author(s):  
Eric M. Katz ◽  
Mathew D. Esona ◽  
Rashi Gautam ◽  
Michael D. Bowen
Keyword(s):  
Viral Load ◽  
Real Time ◽  
Reverse Transcription ◽  
Dynamic Range ◽  
Limit Of Detection ◽  
Human Populations ◽  
Pcr Assay ◽  
Group A Rotavirus ◽  
Reverse Transcription Pcr ◽  
Group A

Since 2013, group A rotavirus strains characterized as novel DS-1-like inter-genogroup reassortant ‘equine-like G3’ strains have emerged and spread across five continents among human populations in at least 14 countries. Here we report a novel one-step TaqMan quantitative real-time reverse transcription-PCR assay developed to genotype and quantify the viral load for samples containing rotavirus equine-like G3 strains. Using a universal G forward primer and a newly designed reverse primer and TaqMan probe, we developed and validated an assay with a linear dynamic range of 2.3 × 10 9 – 227 copies per reaction and a limit of detection of 227 copies. The percent positive agreement, percent negative agreement, and precision of our assay were 100.00%, 99.63%, and 100.00%, respectively. This assay can simultaneously detect and quantify the viral load for samples containing DS-1-like inter-genogroup reassortant equine-like G3 strains with high sensitivity and specificity, faster turnaround time, and decreased cost and will be valuable for high-throughput screening of stool samples collected to monitor equine-like G3 strain prevalence and circulation among human populations throughout the world.

scholarly journals Sensitive SYBR Green—Real Time PCR for the Detection and Quantitation of Avian Rotavirus A

2018 ◽  
Vol 6 (1) ◽  
pp. 2 ◽  
Author(s):  
David De la Torre ◽  
Claudete Astolfi-Ferreira ◽  
Ruy Chacon ◽  
Antonio Piantino Ferreira
Keyword(s):  
Polymerase Chain Reaction ◽  
Real Time ◽  
Sybr Green ◽  
Molecular Techniques ◽  
Economic Losses ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Rotavirus A ◽  
Avian Nephritis Virus ◽  
Polymerase Chain

Avian rotavirus A (ARtV-A) is a virus that affects young birds, causing acute diarrhea and economic losses in the poultry industry worldwide. The techniques used for the diagnosis of ARtV-A include electron microscopy, isolation in cell culture, and serology, as well as molecular techniques, such as the reverse transcription-polymerase chain reaction (RT-PCR). The objective of this work was to standardize a real-time RT-polymerase chain reaction (RT-qPCR) using SYBR Green chemistry for the rapid detection and quantification of ARtV-A from bird tissues and materials fixed on FTA cards on the basis of the nucleotide sequence of segment 6 (S6), which codes for the structural VP6 protein of ARtV-A. The results show the efficient amplification of the proposed target, with a limit of detection (LoD) of one copy gene (CG) per microliter of cDNA and a limit of quantification (LoQ) of 10 CGs per microliter. The efficiency of the primers was determined to be 95.66% using a standard curve, with an R2 value of 0.999 and a slope of −3.43. The specificity was determined using samples coinfected with ARtV-A, the chicken parvovirus, the chicken astrovirus, and the avian nephritis virus as positive controls and commercially available vaccines of the infectious bronchitis virus, infectious bursa disease virus, avian reovirus and healthy organs as negative controls. This technique, which lacks nonspecific PCR products and dimers, demonstrated greater sensitivity and specificity than conventional RT-PCR, and it reduced the analysis time by more than 50%.

scholarly journals Diagnosing rotavirus A associated IID: Using ELISA to identify a cut-off for real time RT-PCR

2009 ◽  
Vol 44 (3) ◽  
pp. 242-245 ◽  
Author(s):  
Gemma Phillips ◽  
Ben Lopman ◽  
Clarence C. Tam ◽  
Miren Iturriza-Gomara ◽  
David Brown ◽  
...  
Keyword(s):  
Real Time ◽  
Rt Pcr ◽  
Rotavirus A

scholarly journals Evaluation of multiplex SYBR green real-time PCR assay for detection of pathogenic Escherichia coli

2020 ◽  
Vol 9 (2) ◽  
pp. 448
Author(s):  
Ema Komalasari ◽  
Winiati P. Rahayu ◽  
Siti Nurjanah
Keyword(s):  
Escherichia Coli ◽  
Real Time ◽  
Real Time Pcr ◽  
Sybr Green ◽  
Rt Pcr ◽  
E Coli ◽  
Melting Curves ◽  
Pathogenic Escherichia Coli ◽  
Wide Range ◽  
Pcr Method

Pathogenic Escherichia coli (E. coli) has been implicated in a wide range of disease causing infections. It is essential to generate a method for detecting and differentiating each pathotype of E. coli which is more quickly and efficiently by using less reagent. This study aimed to evaluate a SYBR Green multiplex real-time PCR method for detecting four types of pathogenic E. coli. Two of multiplex real-time PCR system, 6-plex and 3-plex, were set to detect six different virulence factors from ETEC, EPEC, EHEC, and EIEC and evaluate the melting curves and specificity compared to simplex method. The results showed that 3-plex rt-PCR method gave more reliable melting curves than 6-plex. The 3-plex rt-PCR also provided similar melting value (Tm) to simplex system. The results of this specificity assay supported the selection of 3-plex rt-PCR conditions for detection of pathogenic E. coli.

scholarly journals Isolation, immunochemical demonstration of field strains of porcine group A rotaviruses and electrophoretic analysis of RNA segments of group A and C rotaviruses

2012 ◽  
Vol 51 (No. 5) ◽  
pp. 288-295 ◽  
Author(s):  
R. Smitalova ◽  
L. Rodak ◽  
I. Psikal ◽  
B. Smid
Keyword(s):  
Cell Line ◽  
Electrophoretic Analysis ◽  
Elisa Test ◽  
Farm Animals ◽  
Economic Losses ◽  
Rt Pcr ◽  
Group C Rotavirus ◽  
Group A Rotaviruses ◽  
Group A ◽  
Virus Isolates

Rotaviruses are major cause of acute diarrhea in animals and humans which can result in huge economic losses in farm animals including pigs. We collected 195 samples of feces of diarrhoeic animals. Rotavirus was demonstrated by electron microscopy using the method of negative staining in 27 samples and by ELISA test using monoclonal antibodies to the group antigen VP6 in 44 samples. Nine samples were selected for virus isolation. Three virus isolates (P375/4, P410/4 and P646/1) were successfully adapted to growth in cell line MA-104. These isolates were allocated to group A rotaviruses based on ELISA, immunoperoxidase test and electropherotype analysis. Electropherotype analysis demonstrated changes during passage in cell line in two of the three isolates. The selected sample P543/1 proved negative in ELISA in a fecal sample. Electropherotype analysis of this sample revealed a &ldquo;longer&rdquo; electropherotype profile. The profile was suggestive of group C rotavirus. Rotavirus group C was confirmed by RT-PCR and by sequence analysis in this sample.

scholarly journals Histo-Blood Group Antigens in Children with Symptomatic Rotavirus Infection

Viruses ◽  
2019 ◽  
Vol 11 (4) ◽  
pp. 339 ◽  
Author(s):  
Raúl Pérez-Ortín ◽  
Susana Vila-Vicent ◽  
Noelia Carmona-Vicente ◽  
Cristina Santiso-Bellón ◽  
Jesús Rodríguez-Díaz ◽  
...  
Keyword(s):  
Blood Group ◽  
Control Group ◽  
Human Populations ◽  
Blood Group Antigens ◽  
Symptomatic Infection ◽  
Rotavirus Infection ◽  
Rotavirus Gastroenteritis ◽  
Group A Rotaviruses ◽  
Group A ◽  
Prevalent Strain

Group A rotaviruses are a major cause of acute gastroenteritis in children. The diversity and unequal geographical prevalence of rotavirus genotypes have been linked to histo-blood group antigens (HBGAs) in different human populations. In order to evaluate the role of HBGAs in rotavirus infections in our population, secretor status (FUT2+), ABO blood group, and Lewis antigens were determined in children attended for rotavirus gastroenteritis in Valencia, Spain. During three consecutive years (2013–2015), stool and saliva samples were collected from 133 children with rotavirus infection. Infecting viral genotypes and HBGAs were determined in patients and compared to a control group and data from blood donors. Rotavirus G9P[8] was the most prevalent strain (49.6%), followed by G1P[8] (20.3%) and G12P[8] (14.3%). Rotavirus infected predominantly secretor (99%) and Lewis b positive (91.7%) children. Children with blood group A and AB were significantly more prone to rotavirus gastroenteritis than those with blood group O. Our results confirm that a HBGA genetic background is linked to rotavirus P[8] susceptibility. Rotavirus P[8] symptomatic infection is manifestly more frequent in secretor-positive (FUT2+) than in non-secretor individuals, although no differences between rotavirus G genotypes were found.

Real-time PCR Detection of Rhodococcus fascians and Discovery of New Plants Associated with R. fascians in Pennsylvania

2012 ◽  
Vol 13 (1) ◽  
pp. 24 ◽  
Author(s):  
Ekaterina V. Nikolaeva ◽  
Seogchan Kang ◽  
Tracey N. Olson ◽  
SeongHwan Kim
Keyword(s):  
Real Time ◽  
Direct Detection ◽  
Infected Plant ◽  
Clinical Samples ◽  
Specific Method ◽  
Rt Pcr ◽  
Plant Materials ◽  
Rhodococcus Fascians ◽  
Wide Range ◽  
Positive Direct

Rhodococcus fascians is a gram-positive bacterium that causes bacterial fasciation on a wide range of ornamental plants. To address the need for a reliable, sensitive, and specific method for detecting R. fascians in infected plant materials, a real-time (RT) PCR assay was developed. The target for detection was fas-1, a plasmid-borne gene that is essential for virulence. DNAs from all confirmed pathogenic strains of R. fascians consistently tested positive, with detection limit of 30 fg of R. fascians DNA. In repeated PCR experiments with R. fascians pure culture, as few as 2.5 CFU were tested positive. Direct detection of R. fascians from clinical samples of Coreopsis was successful, whereas detection of R. fascians in Chrysanthemum, Pelargonium, Phlox, and Veronica required enrichment on modified D2 (mD2) medium. In the case of geraniums, as few as 102 CFU/100 mg plant tissues were successfully detected after 72 h enrichment on mD2. In total, 115 strains, isolated from 41 different kinds of flowering crops in Pennsylvania greenhouses during 1984-2010, were confirmed to be R. fascians by RT PCR. Geraniums and speedwell were the most frequently submitted clinical samples to Pennsylvania Department of Agriculture. The following are the first report of plants associated with R. fascians in PA: Ajania pacifica, Anemone sp., Aruncus sp., Baptisia sp., Eutrochium maculatum, Helianthemum sp., Lewisia sp., Monarda sp., Osteospermum ecklonis, Rudbeckia nitida, and Saponaria ocymoides. Accepted for publication 30 November 2011. Published 27 February 2012.

scholarly journals Rotavirus genotyping in gastroenteritis cases of an infantile population from Western Brazilian Amazonia

2012 ◽  
Vol 45 (4) ◽  
pp. 520-522 ◽  
Author(s):  
Maria Sandra Moura Costa ◽  
Paulo Afonso Nogueira ◽  
Gleicienne Félix Magalhães ◽  
Paula Taquita ◽  
Luis André Mariúba ◽  
...  
Keyword(s):  
Molecular Characterization ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Brazilian Amazon ◽  
Rt Pcr ◽  
Rotavirus A ◽  
Positive Stool ◽  
Group A ◽  
Stool Samples

INTRODUCTION: During the period from 2000 to 2002, 79 rotavirus-positive stool samples were collected from children presenting diarrhea in the Western Brazilian Amazon. METHODS: Molecular characterization of the G and P genotypes was performed using RT-PCR and electropherotyping analysis by polyacrylamide gel electrophoresis. RESULTS: A total of 59 samples were confirmed as group A rotavirus. A long electrophoretic profile was exhibited by the G1P[8], G3P[8], and G4P[8] genotypes. The G1P[8] genotype was found in greater proportion. The short electropherotype was exhibited only by G2 genotype strains. CONCLUSIONS: The proportion of the rotavirus genotypes observed was not different from that in other areas of Brazil. This study is the first genotyping of rotavirus in the Western Brazilian Amazon.

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