Loop-Mediated Isothermal Amplification for the Detection of Pasteurella multocida, Mannheimia haemolytica, and Histophilus somni in Bovine Nasal Samples

This work develops a loop-mediated isothermal amplification (LAMP) assay that detects the presence of bacterial pathogens (Pasteurella multocida, Mannheimia haemolytica, and Histophilus somni) for bovine respiratory disease complex (BRD) in crude nasal samples. Diagnosing BRD involves a physical examination of cattle expressing physical symptoms linked to the disease. Unfortunately, these symptoms are not unique to BRD alone and do not identify specific pathogens to treat. Nucleic acid-based diagnostics, like polymerase chain reaction (PCR), identify BRD pathogens by amplifying species-specific genes present in samples. However, PCR-based approaches require (i) expensive equipment for successful operation and (ii) DNA extractions to remove PCR inhibitors in samples. LAMP offers an accurate, inhibitor-resistant approach to detecting BRD pathogens in a point-of-care format. This developed LAMP assay is 97% accurate in pure DNA samples, 99% sensitive, and 89% specific in DNA-spiked bovine nasal samples (with 104 DNA copies/reaction).