scholarly journals Phylogenetic Study of VP6 Gene of Bovine Rotavirus A and Molecular Survey of Bovine Rotaviruses B and C, and Human G and P Genotypes of Rotavirus A in Calves in Iran

2020 ◽  
Vol 8 (2) ◽  
pp. 66-72
Author(s):  
Ahmad Nazaktabar ◽  
Omid Madadgar
Keyword(s):  
Polyacrylamide Gel Electrophoresis ◽  
Phylogenetic Study ◽  
Rt Pcr ◽  
Japanese Strain ◽  
Rotavirus A ◽  
Bovine Rotavirus ◽  
Study Materials ◽  
Vp6 Gene ◽  
Rotavirus Diarrhea ◽  
Diarrheic Calves

Background: Rotavirus (RV) is one of the most important causes of diarrhea in the calf and human neonates. Rotaviruses are divided into nine different serogroups, of which group A is more important compared to other groups. Objective: This study was performed because of the lack of information about the importance and prevalence of bovine rotaviruses B (RVB) and C (RVC) and human genotypes of rotavirus A (RVA) in the bovine population in Iran. Phylogenetic analysis of VP6 of bovine RVA was the second aim of the present study. Materials and Methods: A total of 581 stool specimens were collected from diarrheic calves of 14 provinces and were analyzed by reverse transcription polymerase chain reaction (RT-PCR) and 485 of them were investigated by PAGE electrophoresis to determine the frequency of three rotaviruses A (RVA), B (RVB), and C (RVC). The presence of human G and P genotypes in Iranian bovine population was also evaluated using semi-nested multiplex RT-PCR. Results: RVA was detected by RT-PCR (VP6 gene detection) in 16.2% (94/581) and by PAGE in 22.16% (108/485) and no positive cases of RVB and RVC were confirmed by polyacrylamide gel electrophoresis (PAGE). This study showed that non-A RV groups (B and C) have little role in calf diarrhea in Iran. The results of the phylogenetic study of VP6 sequences of rotaviruses A identified in this study showed that they all belonged to genotype I2 and were classified into three different branches. Specimen isolated in Zanjan showed the highest difference (maximum identity of 94%) with other sequences and clustered along with the Japanese strain, R22. Human G and P genotypes were not found in the studied samples. Conclusion: The results showed that non-A rotaviruses and human genotypes of RVA are of little importance in calf rotavirus diarrhea in Iran. Also, there is the first phylogenetic study of rotavirus A VP6 protein in Iran.

Rotavirus A associated pathology of intestine and mesenteric lymph nodes and occurrence in bovine calves of Gwalior and Bareilly regions

2018 ◽  
Author(s):  
Gayatri Kashyap ◽  
R. Singh ◽  
Y. S. Malik ◽  
R. K. Agrawal ◽  
K. P. Singh ◽  
...  
Keyword(s):  
Small Intestine ◽  
Lymph Nodes ◽  
Mononuclear Cells ◽  
Dairy Farm ◽  
Rt Pcr ◽  
Mesenteric Lymph Nodes ◽  
Fecal Samples ◽  
Rotavirus A ◽  
Mesenteric Lymph ◽  
Vp6 Gene

To understand the pathology of natural cases of rotavirus (RVA) in bovine calves, a total of 40 cases below 6 months died due to diarrhoea were studied, out of which 7 cases (17.5%) turned positive for RVA by RT-PCR. Histopathology of small intestine showed loss of villous enterocytes, blunting and fusion of villi, elongation of crypts and mononuclear cells infiltration in the lamina-propria. The mesenteric lymph nodes were severely depleted of lymphocytes. These changes were corroborated with presence of RVA antigen in sections by dFAT and nucleic acid by RT-PCR. The fluorescent signals were more in mesenteric lymph nodes than in intestine. Besides, 115 rectal fecal samples were also collected from calves for RVA detection by RT-PCR using VP6 gene specific sets of primers. Dead carcasses of calves (n= 40) belonged to organized dairy farm of Bareilly, while rectal fecal samples belonged to both organized (n= 38) and unorganized farms (n= 77) of Bareilly and Gwalior. The overall occurrence of RVA was 19.3% (30/155), comprising 5/37 cases (13.5%) from Gwalior (MP) and 25/118 cases (21.1%) from Bareilly (UP). These findings suggest the infection of RVA widely prevalent in calves and have potential to escape from the intestinal site to mesenteric lymph nodes.

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.

scholarly journals Comparison of Diagnostic Tests for Detection of Bovine Rotavirus a in Calf Feces

2021 ◽  
Vol 0 (0) ◽  
Author(s):  
Shama Ranjan Barua ◽  
Shariful Islam ◽  
А.М.А.М. Zonaed Siddiki ◽  
Md Masuduzzaman ◽  
Mohammad Alamgir Hossain ◽  
...  
Keyword(s):  
Diagnostic Tests ◽  
Rapid Test ◽  
Diagnostic Methods ◽  
Diagnostic Tools ◽  
Rt Pcr ◽  
Fecal Samples ◽  
Rotavirus A ◽  
Bovine Rotavirus ◽  
Test Elisa ◽  
Calf Mortality

AbstractBovine rotavirus A (BRVA) is a frequent causative agent of diarrhea in neonatal calves. Accurate and rapid diagnosis is crucial to prevent calf mortality from BRVA induced diarrhea. Currently, variety of diagnostic methods are being used to detect BRVA from calves’ feces: antibody-based rapid test and ELISA, and molecular-based RT-PCR and RT-qPCR. The aim of the study was to evaluate the accuracy (sensitivity and specificity) of the rapid test (Immunochromatography), ELISA, and RT-PCR assays, using RT-qPCR as the gold standard, in detection of BRVA in diarrheic calves’ fecal samples. One hundred (n=100) clinically diarrheic fecal samples were tested with four different diagnostic tools. The percent of samples positive by rapid test, ELISA, RT-PCR and RT-qPCR was 10%, 16%, 17%, and 33%, respectively. The agreement between different assays was 75% to 99%. The highest agreement was observed between ELISA and RT-PCR assay (99%). The lowest agreement was recorded (75%) between rapid test and RT-qPCR. The sensitivity of the rapid test, ELISA, and RT-PCR were 30%, 49%, and 52%, respectively when compared to the reference test (RT-qPCR), whereas specificity was 100% for all assays. In conclusion, none of the frequently used diagnostic tests showed a satisfactory level of sensitivity to identify BRVA in calves’ feces. Therefore, the use of a more sensitive rapid test should be used to identify infected calves in field conditions in order to prevent calf mortality from rotaviral diarrhea.

scholarly journals Is Porcine Kobuvirus 1 a Typical Diarrhoeic Pathogen of Piglets?

2017 ◽  
Vol 61 (2) ◽  
pp. 50-55
Author(s):  
H. McFall ◽  
Š. Vilček
Keyword(s):  
Enteric Virus ◽  
Enteric Pathogen ◽  
Porcine Kobuvirus ◽  
Rt Pcr ◽  
Rotavirus A ◽  
Pig Farms ◽  
Rectal Swabs ◽  
Vp6 Gene ◽  
3D Gene

AbstractThe objective of this study was to show if porcine kobuvirus 1 (PKV-1) participates in the development of diarrhoea in piglets. The experiments were focused on comparing the occurrence of PKV-1 with the occurrence of rotavirus A (RVA) infection in suckling pigs on Slovak pig farms. A total of 91 rectal swabs of piglets (age < 28 days) were collected from 8 pig farms. RT-PCR was employed to detect PKV-1 through amplification of the 495 bp fragment of the 3D gene using primers KoVF/ KoVR, and RVA was detected through amplification of the 309 bp fragment of the VP6 gene using primers rot3 and rot5. As expected, the detection of RVA in diarrhoeic piglets was 56.8 % (P < 0.01), while only 14.8 % in healthy animals. These results confirm that RVA is one of the main causes of diarrhoea in young piglets. Comparatively, PKV-1 was detected in approximately equal numbers in the same group of both healthy and diarrhoeic pigs, with 74.1 % in healthy animals and 81.1 % in diarrhoeic animals, which was not statistically significant (P < 0.05). The level of co-infection of both viruses was 11.1 % in healthy animals. A portion of 48.6 % (P < 0.01) of diarrhoeic animals were found with RVA and PKV-1 coinfections. The results of this study indicate that while RVA is an enteric virus, PKV-1 cannot confidently be confirmed as an enteric pathogen.

CLINICAL AND PARACLINICAL CHARACTERISTITICS OF ROTAVIRUS DIARRHEA IN CHILDREN LESS THAN 5 YEARS OLD IN PEDIATRIC DEPARTMENT OF BINH DINH GENERAL HOSPITAL

2013 ◽  
pp. 50-56
Author(s):  
Quoc Tinh Nguyen ◽  
Thi Cu Nguyen
Keyword(s):  
General Hospital ◽  
Acute Diarrhea ◽  
White Blood Cells ◽  
Blood Calcium ◽  
Pediatric Department ◽  
Rotavirus A ◽  
Initial Symptoms ◽  
Stool Specimens ◽  
The Mean ◽  
Rotavirus Diarrhea

Background: Diarrhea is a leading cause of morbidity and mortality in children in developing countries. Rotavirus is the most important factor cause of severe diarrhea and mortality in children, especially under 2 years. Objective: To study Clinical and paraclinical characteristitics of rotavirus diarrhea in children less than 5 years old in the Pediatric Department of Binh Dinh General Hospital. Methods: Cross observational study, 417 children from 2 months to 5 years old who were hospitalized for acute diarrhea without blood in stool in Pediatric Department of Binh Dinh general hospital, from 15/3/2012 to 14/3/2013. There were 228 patients with rotavirus - positive stool specimens and 189 patients with rotavirus - negative stool specimens. Tested for rotavirus A classification by agglutination techniques. Results: Percentage of Rotavirus diarrhea in children <5 years was 54.7%. The mean age of rotavirus diarrhea was 14.83 ± 9.08 tháng months. Ages with the highest cases of rotavirus diarrhea are 2 - 12 months: 48.2%. Rotavirrus diarrhea is experienced in every month but, the highest prevalence is experienced in the spring - summer) with 71,5%. Clinical characteristitics of rotavirus diarrhea: Vomit presents in 91,7% of cases. 32.9% of the initial symptoms of the disease is vomiting. Mean number of vomiting episodes per day: 7 ± 3.57 times, significantly higher compared with non-rotavirus acute diarrhea (p <0,01). Mean number of bowel movement per day: 10.61±4.18 times, mean duration of diarrhea: 6.66 ± 2.52 days. Younger children have higher frequency and duration of diarrhea compared to older children. Paraclinical characteristitics of rotavirus diarrhea: the mean number leukocyte: 9.74 ±3.61 (x 109/l). 2.2% with low blood sodium status; 8.8% with reduction in serum potassium concentration. 26.3% with low blood calcium and glucose status. Laboratory characteristics of stool: 8.8% with white blood cells in stool, 41.7% with carbohydrate in stool. Conclusion: Rotavirus accounting for 54.7% of the causes of diarrhea in children. Rotavirus diarrhea is experienced throughout the year but the highest prevalence is in the spring-summer. Prominent symptoms of the disease are vomiting and watery stool. Children <24 months experience higher frequency and duration of diarrhea compared to older age groups. Key words: Diarrhea, Rotavirus.

Environmental dissemination of group A rotavirus: P-type, G-type and subgroup characterization

2009 ◽  
Vol 60 (3) ◽  
pp. 633-642 ◽  
Author(s):  
F. F. M. Ferreira ◽  
F. R. Guimarães ◽  
T. M. Fumian ◽  
M. Victoria ◽  
C. B. Vieira ◽  
...  
Keyword(s):  
Environmental Samples ◽  
Health Hazard ◽  
Sewage Treatment ◽  
Sewage Treatment Plant ◽  
Treatment Plant ◽  
Northern Region ◽  
Rt Pcr ◽  
Vp6 Gene ◽  
Elution Method ◽  
P Type

Rotaviruses A (RV-A) infection is the most common cause of acute diarrheal diseases in infants and the dissemination of these viruses in the environment represents a public health hazard. The present study aims to evaluate reverse transcription-polymerase chain reaction (RT-PCR) based protocols for the detection of RV-A genes in different types of environmental samples. RV-A were concentrated by the adsorption-elution method using negatively charged membranes associated with a Centriprep Concentrator 50. The RV-A VP4, VP7 and VP6 genes were detected using RT-PCR in river water from the Amazon Hydrographic basin (Northern region) and from wastewater in a sewage treatment plant in Rio de Janeiro (Southeast region), Brazil. RV-A were successfully detected in water environmental samples by the methods used. The detection of the VP6 gene by RT-PCR was the most sensitive for detecting RV-A in environmental samples (44.0%), when compared to the detection of the VP4 (33.3%) and VP7 (25.3%) genes. Based on nucleotide sequence and phylogenetic analysis of the partial VP6 gene, 22 environmental samples were determined to be subgroup II (Wa-like). These results indicate that analysis of environmental samples could possibly make a valuable contribution to studies on the epidemiology of RV-A.

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.

scholarly journals Multiplex Detection of Rotavirus A, B, C, Norovirus GI and GII, Adenovirus, Astrovirus and Sapovirus in a Single PCR System in Stool Samples From Acute Diarrhea Children

2020 ◽  
Author(s):  
Wei Li ◽  
Weiwei Li ◽  
Ran Tao ◽  
Wenqing Xiang ◽  
Mingming Zhou ◽  
...  
Keyword(s):  
High Throughput ◽  
Acute Diarrhea ◽  
Cross Reaction ◽  
Melting Curve Analysis ◽  
Clinical Test ◽  
Rt Pcr ◽  
Rotavirus A ◽  
Diarrhea Virus ◽  
Intestinal Pathogens ◽  
Stool Samples

Abstract Background: Early and accurate identification of infection viruses among children can benefit the personalized medical treatment and management, and reduce the future occurrence of serious symptoms. Thus, it is critical to develop a high-throughput multiplex real-time RT-PCR method to improve the accuracy and efficiency in routine clinical lab tests. Here, we developed a RT-PCR combined with melting curve analysis (RRCMC) method for simultaneous detection of rotavirus A, B, C, norovirus GI and GII, adenovirus, astrovirus and sapovirus. Results: Stool samples were collected from 160 children with acute diarrhea and tested by RRCMC assay. A total of 71 patients were tested positive with norovirus, adenovirus or rotavirus. The RRCMC assay has high specificity. There is no internal cross-reaction through the 8 diarrhea viruses and no cross-reaction of other commonly intestinal pathogens and human genome. The detection limit was ranging from 1×102 to 1×105 copies/ml for each diarrhea virus. Conclusions: In conclusion, the RRCMC method is a suitable rapid clinical test for infection viruses, with the advantages of high-throughput, low cost, high sensitivity, and specificity.

scholarly journals Allamanda Mosaic Caused by Cucumber mosaic virus in Taiwan

Plant Disease ◽  
2005 ◽  
Vol 89 (5) ◽  
pp. 529-529 ◽  
Author(s):  
Y. K. Chen ◽  
C. C. Yang ◽  
H. T. Hsu
Keyword(s):  
Cucumber Mosaic Virus ◽  
Mosaic Virus ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Chenopodium Quinoa ◽  
Rabbit Antiserum ◽  
Nucleotide Sequence Analysis ◽  
Enzyme Linked Immunosorbent Assay ◽  
Rt Pcr ◽  
Indicator Plants

Allamanda (Allamanda cathartica L., family Apocynaceae) is native to Brazil and is a popular perennial shrub or vine ornamental in Taiwan. Plants showing severe mosaic, rugosity, and leaf distortion symptoms on leaves are common in commercial nurseries and private gardens. Examination of crude sap prepared from symptomatic leaves using an electron microscope revealed the presence of spherical virus particles with a diameter of approximately 28 nm. The virus was mechanically transmitted to indicator plants and induced symptoms similar to those incited by Cucumber mosaic virus (CMV). The virus caused local lesions on inoculated leaves of Chenopodium quinoa and C. amaranticolor and systemic mosaic in Cucumis sativus, Lycopersicon esculentum, Nicotiana benthamiana, N. glutinosa, N. rustica, and N. tabacum. On N. tabacum, necrotic ringspots developed on inoculated leaves followed by systemic mosaic. Tests of leaf sap extracted from naturally infected allamanda and inoculated indicator plants using enzyme-linked immunosorbent assay were positive to rabbit antiserum prepared to CMV. Viral coat protein on transblots of sodium dodecyl sulfate-polyacrylamide gel electrophoresis reacted with CMV subgroup I specific monoclonal antibodies (2). With primers specific to the 3′-half of RNA 3 (1), amplicons of an expected size (1,115 bp) were obtained in reverse transcription-polymerase chain reaction (RT-PCR) using total RNA extracted from infected allamanda and N. benthamiana. The amplified fragment (EMBL Accession No. AJ871492) was cloned and sequenced. It encompasses the 3′ part of the intergenic region of RNA 3 (158 nt), CP ORF (657 nt), and 3′ NTR (300 nt) showing 91.8–98.9% and 71.4–72.8% identities to those of CMV in subgroups I and II, respectively. Results of MspI-digested restriction fragment length polymorphism patterns of the RT-PCR fragment and the nucleotide sequence analysis indicate that the CMV isolate from allamanda belongs to subgroup IB, which is predominant on the island. To our knowledge, CMV is the only reported virus that infects allamanda and was first detected in Brazil (3), and this is the first report of CMV infection in allamanda plants occurring in Taiwan. References: (1) Y. K. Chen et al. Arch. Virol. 146:1631, 2001. (2) H. T. Hsu et al. Phytopathology 90:615, 2000. (3) E. W. Kitajima. Acta. Hortic. 234:451, 1988.

scholarly journals Detection of Multiple Infections of Citrus exocortis viroid, Citrus viroid III, and Hop stunt viroid Variants in Hunan Province, China

Plant Disease ◽  
2007 ◽  
Vol 91 (9) ◽  
pp. 1205-1205 ◽  
Author(s):  
S. Rizza ◽  
A. Catara ◽  
X. F. Ma ◽  
Z. Deng
Keyword(s):  
Polyacrylamide Gel Electrophoresis ◽  
Leaf Roll ◽  
Poncirus Trifoliata ◽  
Hunan Province ◽  
Variable Degree ◽  
Rt Pcr ◽  
Stunt Viroid ◽  
Citrus Exocortis Viroid ◽  
Hop Stunt Viroid ◽  
Citrus Viroids

Citrus cultivation in China has increased since the late 1970s, with China now having the largest area of citrus in culture in the world that is spread in 22 provinces and municipalities. Hunan Province has undergone a program to become one of the major citrus producers in China. Poncirus trifoliata is the main rootstock, so citrus viroids are a limiting factor for further citriculture development. In mainland China, only the presence of Citrus exocortis viroid (CEVd) has been reported from Etrog citron indexing, sPAGE (sequential polyacrylamide gel electrophoresis) analysis (2), and reverse transcription (RT)-PCR (3). Three viroid-like RNAs, a1, b1, and d, based on sPAGE patterns were detected years ago in our laboratory in budsticks received from Sichuan Province. To identify different viroids and determine their distribution, a survey has been undertaken. Field trees showing stunting, bark scaling and cracking of the rootstock, and poor yield were tested using biological indexing and PCR for the most frequent citrus viroids. Samples from six trees of a local sweet orange variety and three of a Clementine variety introduced from abroad, both grafted on P. trifoliata and showing a variable degree of bark scaling and cracking, were collected near Changsha and in the County of Xin Ning at the end of summer 2006. Small pieces of bark were inserted in stems of young E. citron budwood grafted on rough lemon and maintained in a warm greenhouse (24 to 32°C). Indexing on E. citron showed mild epinasty and leaf roll typical of citrus viroid infections. To identify specific viroids, bark was ground to a fine powder with liquid nitrogen and total RNA was extracted with TRIZOL Reagent (Invitrogen, San Diego, CA) and tested by RT-PCR to detect CEVd, Hop Stunt viroid (HSVd), and Citrus viroid III (CVd-III), as well as to identify the cachexia variants of HSVd. Four primer pairs were used to test the RNA extracts by RT-PCR (1). All samples were infected by HSVd, eight with CVd-III, and six with CEVd. The cachexia variants of HSVd were detected in four of nine samples. Mixed infections were as follows: one sample had CEVd and HSVd, eight had HSVd and CVd-III, and five were infected by the three viroids. A second sampling 3 months after inoculation gave the same amplification patterns. The results show that at least three viroids are present in citrus orchards in Hunan Province. To our knowledge, this is the first report of cachexia variants of HSVd and CVd-III in China. The common occurrence of these viroids supports the need for proper indexing of mother trees and a specific shoot tip grafting program to create healthy budwood sources to provide healthy plants. References: (1) L. Bernard and N. Duran-Vila. Mol. Cell. Probes, 20:105, 2006. (2) L. Han et al. Viroids. CSIRO Publishing, Melbourne, 283, 2003. (3). Q. Hu et al. Acta Bot. Sin. 39:613, 1997.

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