Authors

Discussion

Introduction:

Parasitic lice are a significant problem in beef and dairy cattle, especially in colder climates. They can cause irritation, anemia, and decreased production, with economic losses estimated at $120 million annually in North America. Over the past decade, many anecdotal reports have reported increased hair loss, skin lesions and lice infestations in Western Canada and Northern USA. Current control relies heavily on endectocides such as macrocyclic lactones and insecticides such as pyrethroids, but their widespread, and often indiscriminate,  leads to concerns about drug resistance and environmental impacts. Urgent action is needed to assess changing lice populations, develop field-deployable diagnostic tools, support evidence-based control and reduce insecticide use.

Objectives:

1: To develop 18S rDNA and COI mtDNA Illumina and Oxford Nanopore Technologies (ONT) based metabarcoding to identify cattle lice species and characterize their genetic diversity.

 2: To develop species-specific and “pan-louse” PCR and Loop-Mediated Isothermal Amplification (LAMP) assay to detect louse DNA on cattle skin swabs as a rapid diagnostic and disease monitoring tool.

Methodology:

We are collecting lice and taking DNA skin swabs from beef and dairy cattle herds in western Canada and northern USA with suspected lice/pruritis problems.  Genomic DNA extraction is performed from the collected lice specimens, followed by PCR to develop the 18S rDNA and COX-1 mtDNA Illumina and Oxford Nanopore Technologies (ONT) metabarcoding assays. Amplified Sequence Variants (ASVs) will be mapped against insect 18s rDNA and COX-1 reference sequence databases that we are developing to identify lice species and assess their population genetic diversity.

For the detection of lice DNA on skin swabs, we are developing Loop-Mediated Isothermal Amplification (LAMP) diagnostic tool employing the WarmStart® LAMP Kit (NEB) with FAM dye and quantifying fluorescent signal on a Thermo Fisher QuantStudio 5. LAMP Primer sets are designed using Primer Explorer V5 and the NEB primer design tool. The LAMP protocols are first developed on adult lice DNA and then tested for their ability to detect louse DNA  from skin swabs taken from infected cattle.

Preliminary Results:

Our preliminary work has yielded promising results. The LAMP Protocol has been developed and optimized using DNA extracted from adult lice specimens. Species-specific primer sets (PE1) amplifying L. vituli DNA and PE3 amplifying both L. vituli and B. bovis DNA have been developed. Louse DNA can be detected directly from DNA skin swabs from infected cattle using both the LAMP and standard PCR. We successfully detected the presence of two major lice species: Linognathus vituli and Bovicola bovis. Lice DNA extracted from skin swabs allowed rapid detection via real-time LAMP within a 30-minute incubation period. 

Significance:

This study aims to develop new diagnostic methods for cattle lice detection and provide information about lice infestation in cattle herds including the most prevalent species and the genetic diversity of cattle lice populations. The LAMP test is being developed for rapid diagnosis and quantification of lice infestations from skin swabs using a fluorescence assay in the laboratory. We plan to transition this to colorimetric platforms to be used as a rapid pen-side test. This information and novel diagnostic tools should allow more targeted insecticide use to achieve more effective and sustainable ectoparasite control and improve animal health and welfare while reducing the environmental impacts of management practices.