A preliminary survey of viruses of bats of Canada (identification of viruses that might emerge as human pathogens)

Emerging infectious diseases (EID) pathogen infects host for the first time new strains of a pathogen with distinct properties increased incidence of preexisting disease new or reintroduction into area

Animals and emerging human disease most EID events emerge from animals most of these events (70%) involve wild animals nature 21 Feb 2008

Role of bats in EID Confirmed or suspected in: –Nipah –Hendra –SARS –Marburg –Ebola –Menangle –Tioman –Melaka –rabies (in some areas)

“Damn! what is it about bats?”* * David Quammen and Lynn Johnson, Deadly Contact, National Geographic, October 2007

Chiroptera “hand wing” Epomops franqueti -

Old order fossil record > 50 million years N.B. Simmons, et al nature 451: Onchonycteris finneyi

Simmons, 2005, Science 307:527

Diversity MegachiropteraMicrochiroptera Tim Menzies Indian flying fox 1.8 m wingspan 1.6 kg Thai bumblebee bat 15 cm wingspan 2 gm

Hibernation and torpor

Bats of Canada Little brown bat (Myotis lucifugus) Yuma myotis (M. yumanensis) Long-legged myotis (M. volans) Fringed myotis (M. thysanodes) Northern long-eared bat (Myotis septentrionalis), Eastern small-footed myotis (M. leibii) Keen’s myotis (M. keenii) Long eared myotis (M. evotis) Western small-footed myotis (M. ciliolabrum) California myotis (M. californicus) Big brown bat (Eptesicus fuscus) Silver haired bat (Lasionycteris noctivagans) Hoary bat (Lasiurus cinereus), Red bat (Lasiurus borealis) Western red bat (Lasiurus blossevillii) Eastern pipstrelle (Pipistrellus subflavus) Spotted bat (Euderma maculatum) Townsend’s big-eared bat (Corynorhinus townsendii) Pallid bat (Antrozous pallidus) Bat Conservation International

Bats in Manitoba Little brown bat (Myotis lucifugus) Big brown bat (Eptesicus fuscus) Northern long-eared bat (Myotis septentrionalis), Silver haired bat (Lasionycteris noctivagans), Hoary bat (Lasiurus cinereus), Red bat (Lasiurus borealis) Hibernating Migrating Derek Donald

Genesis of the project

How does one find bats? live-trapped little brown bats archived rabies-negative bats at the Ottawa and Lethbridge CFIA labs wind farms submission from local veterinarian

Craig Willis

remove liver, kidney, brain, spleen, lung, intestine fix in formaldehyde (pool tissues from same species of bats) homogenize (pooled tissues) freeze at -80 tissue culture (primate, rodent bats, avian cells) Purify nucleic acid QIAmp viral RNA mini kit (will purify RNA and DNA) cDNA synthesis PCR corona (Gr 1, pan) lyssa henipah morbilli WNV Hanta gel sequence observe daily for cpe + - amplify characterize 3 passages autoclave +- herpes polyoma flavi bunya alpha Cache Valley

Tissue Cultures Vero (monkey) NIH-3T3 (mouse) BHK-21 (hamster) TbrLu1 (bat) Quail Chick embryo

Live-trapped little brown bats bat noM/Fcoronapolyomabat noM/Fcoronapolyoma 1F12F + 2F13F 3F14M + 4?15F 5M16F 6M17M 7F18F 8M19F 9M20M 10F + 21F + 11F +

Archived rabies-negative BatNumBatNum Eptesicus fuscus (BBB) 55M. californicus2 M. lucifugus (LBB)10M. evotis (WLEB) 1 M. yumanensis6 1 + ve polyoma Lasionycteris noctivagans (SHB) 6 M. septentrionalis (NLEB) 3Lasiurus cinereus (HB) 1

Wind-farm BatNo Lasiurus cinereus (Hoary) 7* Lasionycteris noctivagans (Silver haired) 6 Lasiurus borealis (Red) 3 * Includes 1 local submission

Strategy for recovering entire bat polyomavirus genome product from broad-specificity PCR primers for amplifying genome

Phylogeny of bat (Myotis) polyomavirus ClustalX

WU KI Budge Finch Bat Mouse Pneumotropic Sq monkey Bovine Crow Goose Hamster Mouse AG monkey SV40 SV12 Baboon JC BK 0.1 VP1 Merkel

WU KI Budge Finch Mouse Pneumotropic Sq monkey Bovine Crow Hamster Mouse AG monkey SV40 SV12 Baboon JC BK 0.1 Bat VP2,3 Merkel

WU KI Budge Finch Mouse Pneumotropic Sq monkey Bovine Crow Hamster Mouse AG monkey SV40 SV12 Baboon JC BK 0.1 Bat T-Antigen Merkel

WU KI Budge Finch Mouse Pneumotropic Sq monkey Crow Hamster Mouse AG monkey SV40 SV12 Baboon JC BK 0.1 Bat Small t-Antigen Merkel

What do we know about bat coronaviruses? SARS Group 1 bat coronaviruses –asian –Rocky Mountain bat coronavirus –European Sero-conversion in bat biologists

“…bats are the natural reservoir of all currently known coronavirus lineages.” Vijaykrishna et al J. Virol 81:

What do we know about polyomaviruses? mouse polyomavirus s persistent, asymptomatic infections in immuno-competent individuals - disease in immuno-suppressed or young animals pathogenic avian viruses

Many Thanks! Tim Dumonceaux Harvey Artsob Robin Lindsay Mike Drebot Marta Sabara Antonia Dibernardo Jack Dubois Susan Nadin- Davis Craig Willis