Evaluation of oral fluid as an aggregate sample for early detection of African swine fever virus using four independent pen‐based experimental studies

Abstract

The sustained spread of African swine fever (ASF) virus throughout much of the world has made ASF a global animal health priority, with an increased emphasis on enhancing preparedness to prevent, detect and respond to a potential outbreak of ASF virus (ASFV). In the event of ASFV entry to the North American swine population, enhanced surveillance and diagnostic testing strategies will be critical to facilitate progressive response and eradication of the disease. Compared to individual animal sampling, pen‐based oral fluid collection for active surveillance is a non‐invasive alternative that is less resource and time‐intensive. To evaluate the feasibility of using rope‐based oral fluid for early detection of ASFV, four independent animal experiments were conducted in weaned pigs housed in numbers that mimic the industry settings, utilising either highly virulent ASFV Georgia 2007/1 strain or moderately virulent ASFV Malta’78 strain. Pen‐based oral fluid and individual oropharyngeal swabs were collected daily and blood samples from each animal were collected every other day. All samples were subsequently tested for ASFV by real‐time PCR. ASFV genome was detected in individual blood samples as early as one day post‐infection and detected in oral fluids at low‐to‐moderate levels as early as 3–5 days post‐infection in all four independent experiments. These results suggest that pen‐based oral fluid samples may be used to supplement the use of traditional samples for rapid detection of ASFV during ASF surveillance.

Superficial Inguinal Lymph Nodes for Screening Dead Pigs for African Swine Fever

“…Experiment 6 (Table 1, pigs 26-44) was conducted as one of four experiments to assess the feasibility of using swine oral fluids as an aggregate sample type to detect ASF genome in commercial pens [24]. In this experiment, a pig (seeder pig) (Table 1, pig 35) was randomly selected from the group and infected with a 1 × 10 3 TCID 50 dose of ASFV Georgia 2007/01, 1 mL, intramuscularly in the left thigh.…”

“…Tissue homogenates (10% w/v) of the spleen and SILN tissues were prepared using Precellys lysing kits with the help of a Precellys tissue homogenizer (Bertin, France). Total nucleic acid extraction from whole blood and tissue homogenates was performed as described previously [24,25], using a MagMax™ Pathogen RNA/DNA kit (Thermofisher Scientific, Waltham, MA, USA), according to the low cell count protocol provided by the manufacturer on the MagMax™ Express-96 Deep Well Magnetic Particle Processor (Thermofisher Scientific, Waltham, MA, USA).…”

“…Virus isolation was conducted in porcine primary leukocytes (PPLs), following the NC-FAD standard operating protocol for African swine fever virus isolation [24]. Briefly, PPLs were isolated from fresh porcine blood, plated on 24-well plates, 1 mL/well (10 6 WBC/mL with 0.4% v/v RBC).…”

African Swine Fever, the forgotten pandemic

“…To control ASF, early detection of infection is critical. This is an active area of research; an example published in this Special Issue is penbased testing of oral fluid (Goonewardene et al, 2021). Also, the role of contaminated feed as a potential source of ASFV spread is being actively researched (Fischer et al, 2021).…”

Development Real-Time PCR Assays to Genetically Differentiate Vaccinated Pigs From Infected Pigs With the Eurasian Strain of African Swine Fever Virus

“…In this context, the use of alternative sample types, like nasal, oral swabs, and the collection of oral fluids may represent a good alternative to maintain optimal levels of diagnostic sensitivity in these tests ( 15 , 36 ). Further studies will involved the evaluation of different types of clinical samples, as oral, rectal and nasal swabs.…”