Expanding international pet trade has emerged as one of the main introduction pathways of aquatic invasive species, with ornamental crayfish species commonly available on the EU and global markets. Besides most frequently studied crayfish pathogens, such as Aphanomyces astaci and white spot syndrome virus (WSSV), ornamental crayfish carry associated microbial communities, which may potentially lead to the emergence of known or even novel diseases following intentional or unintentional release of animals into the wild. This is especially problematic in the case of viruses, which represent an important, yet considerably understudied, group of crayfish pathogens. Here we analyzed viromes of hepatopancreas tissue of four crayfish species acquired in the international pet trade in Europe (Procambarus clarkii, Procambarus alleni, Cherax holthuisi, and Cherax quadricarinatus) using a high throughput sequencing based metagenomic approach. Seven different known viruses were identified, which were previously either directly associated with crayfish (WSSV, Cherax quadricarinatus reovirus, chequa iflavirus, athtab bunya-like virus) or with hosts from subphylum Crustacea or invertebrates associated with freshwater environment (Shahe ispoda virus 5, Dicistroviridae sp.). Additional sequences represented 8 potential novel and divergent RNA viruses, most similar to sequences belonging to members of Picornavirales, Elliovirales, Reovirales, Hepelivirales, Tolivirales and Ghabrivirales orders. We discuss our findings in relation to their phylogenetic relationships, geographical origins, and putative pathogenicity implications. The results highlight the need for further research into the risks related to disease emergence associated with the pet trade.

Department of Forest Protection and Wildlife Management Faculty of Forestry and Wood Technology Mendel University in Brno Zemědělská 3 Brno Czechia

Faculty of Agrobiology Food and Natural Resources Czech University of Life Sciences Prague Prague Kamýcká 129 Czechia; Faculty of Science Humanities and Education Technical University of Liberec Studentská 1402 2 461 17 Liberec Czechia

Faculty of Food Technology and Biotechnology University of Zagreb Pierottijeva 6 10000 Zagreb Croatia

Faculty of Science University of Zagreb Horvatovac 102a Zagreb Croatia

National Institute of Biology Večna pot 121 1000 Ljubljana Slovenia

South Bohemian Research Centre of Aquaculture and Biodiversity of Hydrocenoses Faculty of Fisheries and Protection of Waters University of South Bohemia in České Budějovice Zátiší 728 2 389 25 Vodňany Czechia

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