Re-Emergence of BTV-4 in Sheep Farms in Kosovo, 2020: A Retrospective Study

Abstract

Kosovo has previously seen two bluetongue (BT) epizootics, each caused by a different serotype, BTV-9 in 2001 and BTV-4 in 2014. Since 2014, no clinical cases of BT have been reported in Kosovo. In September, 2020, clinical signs suggestive of BTV infection were observed in several sheep farms in Kosovo. Blood samples from sheep (n = 40) were collected and subjected to further molecular investigations. Molecular analyses confirmed BTV serotype 4 (BTV-4) infection in thirty-six sheep from five different farms across two different regions. Full genome sequence analyses indicated that the BTV-4 strains (KOS2020/01 and KOS2020/02) detected in Kosovo in 2020 had high sequence identity (99.9%-100%) with a strain responsible for an outbreak in North Macedonia in July, 2020, (MKD2020/06) and with previous isolates (≥99.3%) from Greece, Hungary, and France. The percent nucleotide sequence (nt%) identity and phylogenetic analyses suggest that the incursion of BTV-4 into Kosovo was a re-emergence of a previously seen strain and not a novel reassortant. This could be due to a reintroduction of the strain into the region or from subclinical circulation which had been ongoing and underreported for years. Surveillance across Kosovo and the Balkan region to monitor the circulation of BTV is crucial if outbreaks are to be brought under control.

Figure 1

Clinical signs observed in sheep affected by BTV on farms in Kosovo. (a) Nasal discharge, erosion of the nasal planum, and excessive salivation. (b) Scabby mouth and abrasions on the bare skin around the lips. (c) Erosions and ulceration of oral mucosa. Lips are swollen with greyish brown necrotic deposition.

Figure 2

Phylogenetic relationship of BTV-4 from outbreaks in Kosovo in 2014 and 2020 based on (a) VP2 coding region and (b) VP5 coding region. Sequences produced in this study are labelled with a blue circle. Maximum likelihood trees (1,000 bootstrap replicates) were constructed using MEGA X software. Bootstrap support values of ≥70% are shown.

Figure 3

Maps of countries in the Balkan Peninsula showing the spatial distribution of BTV activity between 1998 and 2013 (a) and 2014 and 2020 (b), and closeup of Kosovo (c) to provide additional details in the region. White areas indicate Balkan countries, whereas countries outside of the Balkan Peninsula are shown by grey areas. BTV-1, BTV-4, BTV-8, BTV-9, and BTV-16 positive cases are represented by green, blue, black, purple, and orange dotted circles, respectively. Red dotted circles represent BTV-4 positive cases from the outbreak in Kosovo in 2020. All data of BTV occurrences were retrieved from the literature and the World Organisation for Animal Health (WOAH) database.