ABSTRACT
Viral diseases like foot-and-mouth disease (FMD), calf scour (CS), bovine viral diarrhea (BVD), infectious bovine rhinotracheitis (IBR) etc. affect the growth and milk production of cattle (Bos taurus) causing severe economic loss. Epitope-based vaccine designing have been evolved to provide a new strategy for therapeutic application of pathogen-specific immunity in animals. Therefore, identification of major histocompatibility complex (MHC) binding peptides as potential T-cell epitopes is widely applied in peptide vaccine designing and immunotherapy. In this study, MetaMHCI tool was used with seven different algorithms to predict the potential T-cell epitopes for FMD, BVD, IBR and CS in cattle. A total of 54 protein sequences were filtered out from a total set of 6351 sequences of the pathogens causing the said diseases using bioinformatics approaches. These selected protein sequences were used as the key inputs for MetaMHCI tool to predict the epitopes for the BoLA-A11 MHC class I allele of B. taurus. Further, the epitopes were ranked based on a proposed principal component analysis based epitope score (PbES). The best epitope for each disease based on its predictability through maximum number of predictors and low PbES was modeled in PEP-FOLD server and docked with the BoLA-A11 protein for understanding the MHC-epitope interaction. Finally, a total of 78 epitopes were predicted, out of which 27 were for FMD, 25 for BVD, 12 for CS and 14 for IBR. These epitopes could be artificially synthesized and recommended to vaccinate the cattle for the considered diseases. Besides, the methodology adapted here could also be used to predict and analyze the epitopes for other microbial diseases of important animal species.
Subject(s)
Animals , Cattle , Computational Biology , Diarrhea Viruses, Bovine Viral/analysis , Diarrhea Viruses, Bovine Viral/genetics , Epitopes/analysis , Epitopes/genetics , Foot-and-Mouth Disease/diagnosis , Foot-and-Mouth Disease/genetics , Infectious Bovine Rhinotracheitis/diagnosis , Infectious Bovine Rhinotracheitis/geneticsSubject(s)
Animals , Cattle , Cattle Diseases/diagnosis , Foot-and-Mouth Disease Virus/isolation & purification , Foot-and-Mouth Disease/diagnosis , Real-Time Polymerase Chain Reaction/methods , Reverse Transcriptase Polymerase Chain Reaction/methods , Acute Disease , Body Fluids/virology , Cattle Diseases/virology , Esophagus/virology , Foot-and-Mouth Disease Virus/classification , Foot-and-Mouth Disease Virus/genetics , Foot-and-Mouth Disease Virus/immunology , Foot-and-Mouth Disease/virology , Pharynx/virology , Reproducibility of Results , RNA, Viral/analysis , Real-Time Polymerase Chain Reaction/instrumentation , Reverse Transcriptase Polymerase Chain Reaction/instrumentation , Sensitivity and Specificity , Specimen Handling , Vaccination/veterinary , Viral Vaccines/immunologyABSTRACT
A reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay was developed to rapidly detect foot-and-mouth disease virus serotype C (FMDV C). By testing 10-fold serial dilutions of FMDV C samples, sensitivity of the FMDV C RT-LAMP was found to be 10 times higher than that of conventional reverse transcription-PCR (RT-PCR). No cross-reactivity with A, Asia 1, or O FMDV or swine vesicular disease virus (SVDV) indicated that FMDV C RT-LAMP may be an exciting novel method for detecting FMDV C.
Subject(s)
Animals , Foot-and-Mouth Disease/diagnosis , Foot-and-Mouth Disease Virus/genetics , Nucleic Acid Amplification Techniques/methods , Reverse Transcriptase Polymerase Chain Reaction/veterinary , Reverse Transcription/genetics , Sensitivity and SpecificityABSTRACT
We compared genetic variations in the VP1 gene of foot-and-mouth disease viruses (FMDVs) isolated since 2000 from various region of the world. We analyzed relative synonymous codon usage (RSCU) and phylogenetic relationship between geographical regions, and calculated the genetic substitution patterns between Korean isolate and those from other countries. We calculated the ratios of synonymously substituted codons (SSC) to all observed substitutions and developed a new analytical parameter, EMC (the ratio of exact matching codons within each synonymous substitution group) to investigate more detailed substitution patterns within each synonymous codon group. We observed that FMDVs showed distinct RSCU patterns according to phylogenetic relationships in the same serotype (serotype O). Moreover, while the SSC and EMC values of FMDVs decreased according to phylogenetic distance, G + C composition at the third codon position was strictly conserved. Although there was little variation among the SSC values of 18 amino acids, more dynamic differences were observed in EMC values. The EMC values of 4- and 6-fold degenerate amino acids showed significantly lower values while most 2-fold degenerate amino acids showed no significant difference. Our findings suggest that different EMC patterns among the 18 amino acids might be an important factor in determining the direction of evolution in FMDV.
Subject(s)
Animals , Cattle , Capsid Proteins/genetics , Codon/genetics , Evolution, Molecular , Foot-and-Mouth Disease/diagnosis , Foot-and-Mouth Disease Virus/genetics , Gene Frequency , Geography , Korea , Phylogeography , Polymorphism, Genetic , RNA, Viral/analysis , Species SpecificityABSTRACT
We compared genetic variations in the VP1 gene of foot-and-mouth disease viruses (FMDVs) isolated since 2000 from various region of the world. We analyzed relative synonymous codon usage (RSCU) and phylogenetic relationship between geographical regions, and calculated the genetic substitution patterns between Korean isolate and those from other countries. We calculated the ratios of synonymously substituted codons (SSC) to all observed substitutions and developed a new analytical parameter, EMC (the ratio of exact matching codons within each synonymous substitution group) to investigate more detailed substitution patterns within each synonymous codon group. We observed that FMDVs showed distinct RSCU patterns according to phylogenetic relationships in the same serotype (serotype O). Moreover, while the SSC and EMC values of FMDVs decreased according to phylogenetic distance, G + C composition at the third codon position was strictly conserved. Although there was little variation among the SSC values of 18 amino acids, more dynamic differences were observed in EMC values. The EMC values of 4- and 6-fold degenerate amino acids showed significantly lower values while most 2-fold degenerate amino acids showed no significant difference. Our findings suggest that different EMC patterns among the 18 amino acids might be an important factor in determining the direction of evolution in FMDV.
Subject(s)
Animals , Cattle , Capsid Proteins/genetics , Codon/genetics , Evolution, Molecular , Foot-and-Mouth Disease/diagnosis , Foot-and-Mouth Disease Virus/genetics , Gene Frequency , Geography , Korea , Phylogeography , Polymorphism, Genetic , RNA, Viral/analysis , Species SpecificityABSTRACT
The aim pf the present study was to compare three different methods for detection of food and mouth disease virus antigens[A, Asia1, O1]from mouth epithelium samples and also to monitore the distribution of FMDV serotypes 338 cattle and 34 sheep of Iran during 2000-2001. The suspension fluids of processed tongue epithelium samples were clarified by centrifugation and tested for presence of FMDV antigens. using Complement Fixation Test[CFT], Enzyme-linked Immuno-sorbant assay [ELISA] and inoculating onto BHK-21 cell culture. 151 out of 372 [40%] epithelium samples were positive to one of the FMD viruses, using CFT. These samples were collected from most provinces of Iran during one year [200-2001]. The virus was isolated from 78 out of 100 mouth epithelium samples in BHK-21 cell culture. The CF and ELISAtests were positive in 25 [30%] and 50[64%], respectively. ELISAwas at least two times more sensitive than the CFTfor detection of FMDVin epithelial samples. The Northwest region showed the highest rate of infection in the country meanwhile, Tehran province had the highest rate of infection among provinces
Subject(s)
Animals , Foot-and-Mouth Disease/diagnosis , Sheep , Cattle , Enzyme-Linked Immunosorbent AssayABSTRACT
It was reported that the sera of convalescent animals contain antibodies to foot and mouth disease (FMD) virus (FMDV) 2C, highly conserved nonstructural protein (NSP), whereas the sera of vaccinated animals do not. But ELISA methods using this protein were not reported and developed until recently. In this study, NSP 2C peptides were synthesized within the amino acid sequence of the conserved 2C nonstructural region of FMDV according to the sequences from Genbank database and used for identifying antigenic determinants. One of the synthesized thirteen peptides gave strong positive reactivity with most of the sera from 13 FMD infected farms, but not with sera from vaccinated and non-infected animals. Moreover, with the sera collected through serial bleedings from four cattle and five goats infected with FMDV O/SKR/2000 experimentally, positive results were obtained in two species after 10 days post infection (DPI). Therefore, we tried to develop and evaluate this ELISA based on 2C peptides. In comparison with the commercial NSP ELISA, the 2C peptide based ELISA method showed good specificity and sensitivity. These results demonstrate that the synthetic 2C peptide ELISA can be a complementary marker to differentiate FMDV-infected from vaccinated on a herd basis.
Subject(s)
Animals , Cattle , Enzyme-Linked Immunosorbent Assay/veterinary , Foot-and-Mouth Disease/diagnosis , Foot-and-Mouth Disease Virus , Goats , Sensitivity and Specificity , Vaccination , Viral Nonstructural Proteins/chemical synthesis , Viral VaccinesABSTRACT
One step TaqMan real-time reverse transcription polymerase chain reaction (R/T RT-PCR)using a set of primers/probes was developed for the detection of foot-and-mouth disease (FMD)virus. The gene-specific probes labeled fluorogen for the internal ribosomal entry site, Leader sequence and 2B regions were used to detect FMD virus (FMDV). This assay specifically detected FMDV both in cell culture preparations and clinical samples, and was capable of distinguishing FMD from other viral diseases similar to clinical signs (swine vesicular disease, vesicular stomatitis and bovine viral diarrhea). This assay was shown to be 1000-fold more sensitive than the conventional RT-PCR method. The detection limits of this assay was 1 TCID 50 /ml of the FMDV RNA concentration. Quantification was obtained by a standard curves plotting threshold cycle values versus known infectivity titer. The assay was sensitive, specific and rapid enough to detect FMDV RNA genome in probang samples. As such, the described method is reliable and provides faster disease diagnostics than the conventional RT-PCR procedure to detect FMDV.
Subject(s)
Animals , Foot-and-Mouth Disease/diagnosis , Foot-and-Mouth Disease Virus , Reverse Transcriptase Polymerase Chain Reaction/methods , Sensitivity and Specificity , Taq PolymeraseABSTRACT
Sera from infected, vaccinated and negative cattle were tested using 3D ELISA to allow each category of animals to be clearly differentiated based on the presence of antibody to 3D antigen. The assay can recognize infected animals in countries where FMD outbreaks have occurred, specially if ring vaccination has been performed, also at the same time vaccinated animals can become infected without apparent clinical symptoms. 3D ELISA test is simple and rapid to discriminate between antibodies elicited by FMD infection or by vaccination
Subject(s)
Animals, Laboratory , Foot-and-Mouth Disease/diagnosis , Foot-and-Mouth Disease/veterinary , Cattle , Aphthovirus/isolation & purification , Enzyme-Linked Immunosorbent Assay , VaccinationABSTRACT
O método do "Sandwich indireto" do ELISA ("Enzyme-Linked Immunosorbent Assay"), foi nesse trabalho empregado com vantagens para detecçäo e tipificaçäo direta dos sorotipos O1, A24, AVenceslau e C3 do vírus da febre aftosa presentes em suspensöes antigênicas brutas provenientes de cultura celular, do epitélio de coxim plantar de cobaias, de carcaça de camundongos e de epitélio lingual bovino, infectados com essas estirpes virais. Os títulos obtidos no ELISA foram sempre superiores aos da reaçäo de fixaçäo de complemento, apresentando uma sensibilidade 3,8 a 80 vezes maior do que esta última reaçäo. Quanto à especificidade do ensaio imunoenzimático foi observado o desenvolvimento de reaçöes heterotípicas cruzadas principalmente da estirpe O1 com o anti-soro anti-C3, da estirpe A24 com o anti-O1, e anti-C3, da estirpe AVenceslau com anti-O1 e da estirpe C3 com o anti-O1, o que näo ocorreu na reaçäo de fixaçäo de complemento. No entanto, isto näo impediu que as 4 estirpes virais estudadas (O1, A24, AVenceslau e C3) fossem classificadas pelo ELISA em seus respectivos sorotipos
Subject(s)
Cattle , Mice , Animals , Enzyme-Linked Immunosorbent Assay , Aphthovirus/immunology , Foot-and-Mouth Disease/diagnosis , Antibodies, Viral/analysis , Antigens, Viral/analysisABSTRACT
La fiebre aftosa es una enfermedad altamente contagiosa, por lo que un diagnostico rapido es fundadamental para su control o erradicacion. Es causada por 7 tipos de virus inmunologicamente diferentes, com mas de 65 subtipos y varias cepas. De los siete tipos el que mas varia es el A, del que con frecuencia aparecen nuevas cepas en el campo, algunas de ellas son muy diferentes a las conocidas, por que su identificacion por fijacion del complemento, utilizando sueros hiperimunes convencionales es dificil y demorada. Para subsanar este problema se ha propuesto la utilizacion de diferentes mezclas de sueros hiperinmunes. En el presente trabajo se describe la produccion de un suero hiperinmunepolivalente por hiperinmunizacion de cobayos con diferentes cepas del tipo A. Elsuero hiperinmune obtenido tiene mayor amplitud antigenica que los sueros individuales o la mezcla de estos. Esta mayor amplitud antigenica hace de el un elemento de diagnostico valiosissimo para elevar el numero de resultados positivos obtenidos directamente de las muestras de campo, eliminando de esa manera los pasajes en cultivos celulares o en ratones para aumentar la concentracion de antigeno. Sueros asi elaborados estan siendo ampliamente utilizados en la mayoria de los laboratorios de diagnostico de las enfermedades vesiculares de America del Sur.