Formal estimation of the seropositivity cut‐off of the hemagglutination inhibition assay in field diagnosis of influenza D virus in cattle and estimation of the associated true prevalence in Morocco

Abstract

The influenza D virus (IDV) was discovered less than ten years ago. Increased interest in this virus is due to its nature (RNA virus with high mutation rate), its worldwide circulation in livestock species, its probable role in bovine respiratory disease and its zoonotic potential. Until currently, the establishment of positivity cutoff of the hemagglutination inhibition (HI) assay was not formalized in field conditions for the detection of antibodies directed against IDV in cattle (i.e. the proposed reservoir). In this study, the positivity cutoff of the HI assays was formally established (titre = 10) using a receiver operating characteristic (ROC) curve. This information was used to estimate the sensitivity (68.04 to 73.20%) and the specificity (94.17 to 96.12%) of two different HI assays (HI 1 and HI 2 , with two different IDV antigens) relatively to virus micro-neutralization test (VNT) as reference test. Based on the above characteristics, the true prevalence of IDV was then estimated in Morocco using a stochastic approach. Irrespective of the HI assays used, the estimation of the true prevalence was statistically equivalent (between 48.44% and 48.73%). In addition, the Spearman rank correlation between HI titres and VNT titres was statistically good (0.76 and 0.81 for HA 1 and HA 2 , respectively). The positive (0.82 and 0.79 for HA 1 and HA 2 , respectively) and the negative (0.86 and 0.85 for HA 1 and HA 2 , respectively) agreement indices between results of HI assays and VNT were good and similar. This study allowed for a formal establishment of a positivity cutoff in HI assays for the detection of antibodies directed against IDV. This information is of prime importance to estimate the diagnostic sensitivity and specificity of the test relatively to the VNT (i.e. the reference test). Using these characteristics, the true prevalence of IDV should be determined in a country.

Serological Surveillance of Influenza D Virus in Ruminants and Swine in West and East Africa, 2017–2020

“…The HI titer was expressed as the reciprocal of the highest serum dilution showing complete inhibition of hemagglutination. A serum sample was determined as seropositive when the HI titer was ≥1: 10 [ 11 , 22 ].…”

Influenza A and D Viruses in Non-Human Mammalian Hosts in Africa: A Systematic Review and Meta-Analysis

“…Studies around the globe have reported the circulation of IDV in either healthy or sick cattle, small ruminants and swine from China, France and the USA [30,32,34,117,118]. This review and meta-analysis demonstrated the presence of IDV specific antibodies in cattle from Benin, Morocco and Togo [40,90,91], camels from Ethiopia and Kenya [40,92] and small ruminants from Ethiopia and Togo [40,91,92]. However, no viral RNA of IDV was detected, possibly due to the absence of active infection in the animals during the period of sampling, the limited number of samples collected in each study, and the limited number of studies conducted in Africa.…”

“…Exposure to IDV was reported in East Africa, North Africa and West Africa. IDV antibodies were detected in cattle from Benin [40], Morocco [90] and Togo [40,91], in dromedary camels from Kenya [40] and Ethiopia [92], and in small ruminants from Ethiopia and Togo (Table 2). Further, the review of the literature suggests that IDV has been circulating in Africa since 2012 as evidenced by the antibodies detected in Morocco [40].…”

First expert elicitation of knowledge on drivers of emergence of influenza D in Europe

“…Nevertheless, a proper estimation of relative importance of illegal movements of live animals and their introduction pathways is deemed essential to set up risk-based awareness, prevention and surveillance programmes that correspond to reality. While direct (isolation of the virus and real-time polymerase chain reaction) Ferguson et al, 2015;Hause et al, 2013) and indirect (seroneutralization, haemagglutination inhibition assays and monoclonal antibody-based competitive ELISA) (Moreno et al, 2018;Saegerman, Salem, et al, 2020) diagnostic tests have been set up for IDV, the absence of commercial assays limits the possibility to implement a proper testing strategy in order to control the trade of live animals and to certify the sanitary IDV status of the herd of origin.…”

“…The fifth most important driver was the vaccine availability (D3-2). The worldwide spread and inter-species transmission of IDV could be prevented and controlled either thanks to an effective vaccine, or thanks to antiviral strategies (Asha & Kumar, 2019;Saegerman, Salem, et al, 2020). The use of antivirals in animals is not advised due to high treatment costs and risk of emergence and spread of resistant influenza strains (Kesinger et al, 2018).…”