ABSTRACT
BACKGROUND: Lumpy skin disease (LSD) is caused by LSD virus which is a member of the Capripoxvirus (CaPV) genus. Although PCR provides for a rapid and sensitive diagnosis, it has limited use due to its complexity in terms of cost, time and equipment. Loop-mediated isothermal amplification (LAMP) is a simple, specific and cost-effective method with a diagnostic accuracy similar to PCR. OBJECTIVES/HYPOTHESIS: To compare the detection rate (DR) of two LAMP assays versus PCR for the detection of CaPV. ANIMALS: This study used 105 apparently health animals (AHA) and 59 clinically sick animals (CSA). METHODS AND MATERIALS: PCR and LAMP assays (LAMP1 and LAMP 2) were compared for detection of CaPV from AHA and CSA using blood and tissue samples. The detection was confirmed by sequencing of PCR positive samples. Analytical sensitivity and specificity of LAMP assays also were assessed. RESULTS: The DR in CSA was 13.6% for PCR whereas for LAMP it was 39.0% and 25.4% for LAMP 1 and 2 methods, respectively. In AHA, the LAMP assay DR was 14.3% and 1.9% for LAMP 1 and 2, respectively. Phylogenetic tree analysis confirmed the identity of CaPV. Analytic sensitivity showed a detection limit of 8 copies/µL. The analytic specificity test showed no cross detection with other infectious agents. CONCLUSION AND CLINICAL IMPORTANCE: Good sensitivity and specificity results for LAMP assay support its application in the routine diagnosis of LSD, whereas its ability to detect LSDV in apparently healthy animals shows its usefulness in identifying populations at risk of LSD.
Subject(s)
Lumpy Skin Disease/diagnosis , Animals , Capripoxvirus/genetics , Cattle , Lumpy Skin Disease/virology , Nucleic Acid Amplification Techniques/veterinary , Polymerase Chain Reaction/veterinary , Reproducibility of Results , Sensitivity and SpecificityABSTRACT
Phylogeography data are of paramount importance in studying the molecular epidemiology dynamics of foot-and-mouth disease virus (FMDV). In this study, epithelial samples and oesophageal-pharyngeal fluids were collected from 361 convalescent animals (cattle and buffaloes) in the field throughout Tanzania between 2009 and 2013. The single plex real-time RT-PCR (qRT-PCR) assay for rapid and accurate diagnosis of FMDV employing the Callahan 3DF-2, 3DF-R primers and Callahan 3DP-1 probe were used. Preparation of the samples was performed according to the OIE manual, with a Kenya O serotype obtained from the attenuated vaccine serving as a positive control and samples collected from healthy animals serving as true negatives. The results indicated that 53.49% of samples (n = 176) were positive for FMDV genome by qRT-PCR, with Ct values ranging from 14 to 32. In addition, molecular typing of the FMDV genome positive samples using serotype specific primers revealed the existence of several serotypes: serotype South Africa Territory 1 (SAT1) (34.25%, n = 60), serotype A (68.92%, n = 98), serotype O (59.20%, n = 98) and SAT2 (54.54%, n = 96). The virus protein 1 sequences analysis for 35 samples was performed and the collective results indicated: 54.28% serotype O, 25.71% serotype A, 14.28% serotype SAT1 and 2.85% serotype SAT2. Therefore in this study, both the phylogenetic trees and spatial distribution of serotypes elucidated the phylodynamics of multiple FMDV field strains in Tanzania and neighbouring countries.
