A novel multiplex PCR-electronic microarray assay for rapid and simultaneous detection of bovine respiratory and enteric pathogens
| العنوان: | A novel multiplex PCR-electronic microarray assay for rapid and simultaneous detection of bovine respiratory and enteric pathogens |
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| المؤلفون: | Oliver Lung, Tara Furukawa-Stoffer, Cody Buchanan, Niroshan Thanthrige-Don, Dale L. Godson, Tomy Joseph, John S. Gilleard, Aruna Ambagala, Trevor W. Alexander |
| المصدر: | Journal of Virological Methods |
| بيانات النشر: | Elsevier BV, 2018. |
| سنة النشر: | 2018 |
| مصطلحات موضوعية: | 0301 basic medicine, BED, bovine enteric diseases, Time Factors, Microarray, Gastrointestinal Diseases, BCoV, bovine corona virus, DENV, dengue virus, Lt, heat labile toxin, ETEC, enterotoxigenicE. coli, law.invention, 0403 veterinary science, BRSV, bovine respiratory syncytial virus, PCR, polymerase chain reaction, law, EPEC, enteropathogenic E. coli, Multiplex, EHEC, enterohemorrhagic E. coli, Pasteurella multocida, Respiratory Tract Infections, Polymerase chain reaction, BToV, bovine torovirus, biology, LNA, lock nucleic acid, lktA, leukotoxin-A, stx-1, shiga-like toxin 1, Bovine enteric diseases, 04 agricultural and veterinary sciences, E.coli, Escherichia coli, Molecular Diagnostic Techniques, Salmonella enterica, Viruses, BoHV-1, Bovine herpesvirus 1, Veterinary Medicine, Bovine respiratory disease complex, 040301 veterinary sciences, 030106 microbiology, Cattle Diseases, BPI-3, bovine parainfluenza virus 3, Sensitivity and Specificity, Article, 03 medical and health sciences, RT-PCR, reverse transcription polymerase chain reaction, Virology, Multiplex polymerase chain reaction, Animals, 5′UTR, 5′ untranslated region, FI, fluorescence intensity, Bovine coronavirus, LOD, limit of detection, Bacteria, BRoV, bovine rotavirus, DNA microarray, Multiplex PCR, Microarray Analysis, biology.organism_classification, Bovine Respiratory Disease Complex, Coccidia, BRDC, bovine respiratory disease complex, LSB, low salt buffer, Cattle, BVDV, bovine viral diarrhea virus, nmaA, UDP-N-acetyle-D-glucosamine-2-epimerase gene, tbpB, transferring-binding protein B gene, Multiplex Polymerase Chain Reaction, GI, Geninfo identifier, NTC, no-template control |
| الوصف: | Highlights • Two Multiplex PCR-DNA microarray assays (BRDC and BED) were developed and validated. • The BRDC assay simultaneously detects 4 bacteria and 5 viruses. • The BED assay simultaneously detects 4 bacteria, 3 protozoa and 4 viruses. • Both assays were validated using laboratory and clinical samples. Respiratory and enteric diseases continue to be two major causes of economic losses to the cattle industry worldwide. Despite their multifactorial etiology, the currently available diagnostic tests for bovine respiratory disease complex (BRDC) and bovine enteric disease (BED) are single-pathogen-tests. DNA microarray when combined with multiplex polymerase chain reaction (PCR) is a powerful tool in detection and differentiation of multiple pathogens in a single sample. This study reports development and initial validation of two independent highly sensitive and specific multiplex PCR-electronic microarray assays, one for the detection and differentiation of pathogens of the BRDC and the other for detection and differentiation of pathogens of the BED. The BRDC multiplex PCR-microarray assay was able to detect and differentiate four bacteria (Mannheimia haemolytica, Histophilus somni, Pasteurella multocida, and Mycoplasma bovis) and five viruses [bovine parainfluenza virus-3, bovine respiratory syncytial virus, bovine herpesvirus-1, bovine coronavirus (BCoV), and bovine viral diarrhea virus (BVDV)] associated with BRDC. The BED multiplex PCR- microarray- assay was able to detect and differentiate four bacteria (Clostridium perfringens, Escherichia coli, Salmonella enterica Dublin, and Salmonella enterica Typhimurium), three protozoa (Eimeria zuernii, Eimeria bovis, and Cryptosporidium parvum), and four viruses (bovine torovirus, bovine rotavirus, BCoV, and BVDV) associated with the BED. Both assays detected their respective targets individually or in combination when present. The limit-of-detection of each assay at the PCR amplification and DNA microarray levels was determined using previously titrated laboratory amplified target pathogens or using quantified synthetic nucleotides. Both assays showed very high analytical sensitivity and specificity, and were validated using a limited number of clinical samples. The BRDC and BED multiplex PCR- microarray-assays developed in this study, with further clinical validation, could be used in veterinary diagnostic laboratories for the rapid and simultaneous identification of pathogens to facilitate quick and accurate decision making for the control and treatment of these two economically important disease complexes. Furthermore, these assays could be very effective tools in epidemiological studies as well as for screening of healthy animals to identify carriers that may potentially develop BRDC or BED. |
| تدمد: | 0166-0934 |
| الوصول الحر: | https://explore.openaire.eu/search/publication?articleId=doi_dedup___::2901a544b92ed955f6421ca506ebf855Test https://doi.org/10.1016/j.jviromet.2018.08.010Test |
| حقوق: | OPEN |
| رقم الانضمام: | edsair.doi.dedup.....2901a544b92ed955f6421ca506ebf855 |
| قاعدة البيانات: | OpenAIRE |
| تدمد: | 01660934 |
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