Spotty Liver Diagnostics at Charles Sturt University

Spotty liver disease (SLD) is widespread in the poultry industry in Australia, and its causative agent has only recently been identified to be Campylobacter hepaticus (C. hepaticus) (Crawshaw et al., 2015). Within Australia, the disease was first detected in the late 1980s, with reports of the disease in all eastern states (New South Wales, Queensland, South Australia and Victoria) (Scott, 2016). In USA and Canada, a similar disease was reported between the 1950s to the 1970s, which was referred to as avian vibrionic hepatitis (AVH) (Crawshaw, 2019). This was followed by subsequent reports of AVH in the United Kingdom, Austria, Estonia and Germany (Van et al., 2017). The causative agent of AVH was never fully defined, but the terms AVH and SLD are both frequently used to describe acute, random, focal, necrotic, hepatitis in poultry (Crawshaw, 2019).
SLD is recognised to have significant economic consequences for the Australian layer industry (Grimes, 2011) due to flock mortalities (up to 10%) and reduction in egg production (10-25%). The disease most commonly occurs in free range and barn layer flocks, broiler breeders housed in deep litter barns, and occasionally in caged layers (Van et al., 2016). The increasing incidence of the disease anecdotally coincides with the increasing demand for free-range eggs (Moore et al., 2019).
Given that C. hepaticus was only recently identified as the causative agent of SLD, there have been limited studies on this pathogen. However, two other common Campylobacter species of chicken, C. jejuni and C. coli, have been subjected to considerable research due to the human risk associated with infection from these pathogens. C. jejuni and C. coli are the most common causes of bacterial gastroenteritis in humans worldwide, and Campylobacter has been identified as the major source of food poisoning in Australia, the United States and Europe (Banowary et al., 2015). Both C. jejuni and C. coli are known to colonise the gastrointestinal tract (GIT) of chickens, and it is likely that C. hepaticus colonises the GIT in a similar fashion (Van et al., 2017). Furthermore, C. hepaticus bacteria are known to be transmitted via the faecal-oral route, as confirmed by field observations and a clinical infection model study (Van et al., 2017).
Laboratory diagnosis is important for both identification of SLD, and for routine monitoring for disease prevention. Laboratory diagnosis is also useful in cases where there are low mortalities but there is a decrease in egg production and SLD is the suspected cause. Isolation of C. hepaticus using bacterial culture methods is time consuming. Recently, an ELISA based diagnostic test has also been developed that can be used using chicken serum samples (Muralidharan et al., 2020). However, molecular diagnostics such as PCR have provided a significant improvement in both specificity and sensitivity of methods used to identify and differentiate Campylobacters and have been particularly useful to identify new species or genetic variants (Courtice et al., 2018).
Therefore, the overarching goal of this study was to test the efficacy of a developed Loop-Mediated Isothermal Amplification (LAMP) assay in order to collect evidence in support of LAMP being a viable inexpensive, rapid, robust, and sensitive ‘on-farm’ alternative to traditional lab-based diagnostics for spotty liver disease. This in turn, can create novel opportunities of broad disease surveillance and early intervention to reduce disease associated losses in the poultry industry.