Bovine Leukemia virus 11901 Bovine leukemia virus (BLV) is a bovine virus closely related to HTLV-I, a human tumour virus. BLV is a retrovirus which integrates a DNA intermediate as a provirus into the DNA of B-lymphocytes of blood and milk. It contains an oncogene coding for a protein called Tax. Nevertheless in its natural host the cattle leukemia is rare. Because the oncogenic properties of the virus were discovered early, a search for evidence of pathogenicity humans started soon after discovery. Mostly farm workers drinking raw milk were tested for disease, especially for leukemia. But neither leukemia nor other signs of infection could be detected. So many in many states it was not tried to get rid of this infection. Testing strategies have recently changed since the virus was first detected in Cows. Many potential routes of BLV transmission exist. Transmission through procedures that transmit blood between animals such as gouge dehorning, vaccination and ear tagging with instruments or needles that are not changed or disinfected between animals is a significant means of BLV spread. Rectal palpation with common sleeves poses a risk that is increased by inexperience and increased frequency of palpation. Transmission via colostrum, milk, and in utero exposure is generally considered to account for a relatively small proportion of infections. Embryo transfer and artificial insemination also account for a small number of new infections as long as common equipment and/or palpation sleeves are not used. While transmission has been documented via blood feeding insects, the significance of this risk is unclear. The bottom line appears to be that transmission relies primarily on the transfer of infected lymphocytes from one animal to the next and that BLV positive animals with lymphocytosis are more likely to provide a source for infection. In general BLV causes only a benign mononucleosis-like disease in cattle. Only some animals later develop a B-cell leukemia called enzootic bovine leukosis. Under natural conditions the disease is transmitted mainly by milk to the calf. Infected lymphocytes transmit the disease too. So for artificial infection infected cells are used or the more stable and even heat resistant DNA. Virus particles are difficult to detect and not used for transmission of infection. It is possible that a natural virus reservoir exists in the water buffalo (Wiki: Bovine leukemia virus). Baboon Papio cynocephalus 9556 Bank vole Clethrionomys glareolus 447135 Bear Ursus americanus 9643 Birds Passeroidea 175121 Brown Trout Salmo trutta 8032 Buffalo Bison bison 9901 Carnivores Vulpes 9625 Cat Felis catus 9685 Catfishes Siluriformes 7995 Cattle Bos taurus 9913 Chicken Gallus gallus 9031 Chimpanzee Pan troglodytes 9598 chinchillas Chinchillidae 10150 Copper Pheasant Syrmaticus soemmerringii 9067 Deer Cervus elaphus 9860 Deer mouse Peromyscus maniculatus 10042 Dog Canis familiaris 9615 Ducks Anas 8835 Ferret Mustela putorius furo 9669 Fish Hyperotreti 117565 Gerbil Gerbillina 10045 Goat Capra hircus 9925 Gray wolf Canis lupus 9612 Guinea pig Cavia porcellus 10141 Hamster Mesocricetus auratus 10036 Horse Equus caballus 9796 Human Homo sapiens 9606 Macaque Macaca fascicularis 9541 Mongolian Gerbil Meriones unguiculatus 10047 Monkey Platyrrhini 9479 Mouse Mus musculus 10090 None None Parrot Psittacidae 9224 Pig Sus scrofa 9823 Rabbit Oryctolagus cuniculus 9986 Rainbow trout Oncorhynchus mykiss 8022 Rat Rattus 10114 Raven Corvus corax 56781 sei whale Balaenoptera borealis 9768 Sheep Ovis aries 9940 Squirrel Spermophilus richardsonii 37591 Tree shrew Tupaiidae 9393 Trouts, salmons & chars Salmoninae 504568 Turkey Meleagris gallopavo 9103 Vole Microtus ochrogaster 79684 Water buffalo Bubalus bubalis 391902 Bovine leukemia virus DNA vaccine encoding Env VO_0011378 DNA vaccine Research pcDNA3 [Ref1047:Brillowska et al., 1999] Intramuscular injection (i.m.) Intramuscular injection (i.m.) Bovine leukemia virus envelope gene encoding extracellular glycoprotein gp51and transmembrane glycoprotein gp30 DNA vaccine construction The bovine leukemia virus (BLV) envelope gene encoding extracellular glycoprotein gp51 (encoded by env gene) and transmembrane glycoprotein gp30 was cloned into a vehicle expression vector under the human cytomegalovirus (CMV) intermediate early promoter (Brillowska et al., 1999). The intramuscular injection of this plasmid vector generated a cellular immune response. Seven out of ten cows vaccinated with the DNA construct resisted a drastic challenge (Brillowska et al., 1999). Bovine Leukemia Virus Vaccine pBLV6073DX Recombinant vector vaccine Licensed pSP64 vector (Promega) [Ref5906:Suárez et al., 2022] subcutaneous injection The proviral construct inserted in the pSP64 vector (Promega) was amplified in Escherichia coli C3040 (New England Biolabs). The standard protocol for production of the inoculum was based on transfection of a monolayer of CHOK1 cells in 25 cm2 cell culture flasks with 2.6 µg of pBLV6073DX and 5.2 µg of linear polyethylenimine (Polysciences) in the presence of 3 ml EMEM (Sigma-Aldrich). Four hours post-transfection, 3 ml of EMEM supplemented with 10% FBS (Internegocios SA) and 3% trehalose were added. Seventy-two hours post-transfection, lysate aliquots (6 ml/dose) were preserved at -80°C in the presence of 15% trehalose. (Suárez et al., 2022) subcutaneous injection Vaccine strain (pBLV6073DX) with a point mutation in the transmembrane protein gene (T>G at nucleotide 6073) and a partial deletion of the R3-G4 sequences (between positions 6614 and 6997). (Suárez et al., 2022) Env/Rex/Tax Bovine leukemia virus VO_0011051 2760849 9626230 BLVgp05 AF033818 NP_056898 11901 4614 7460 + p18 10.96 16183.28 156 >NC_001414.1:4614-7460 Bovine leukemia virus, complete genome AATGCCCAAAGAACGACGGTCCCGAAGACGCCCACAACCGATCATCAGATGGGTAAGTCTCACTCTTACT CTCCTCGCTCTCTGTCAGCCCATCCAGACTTGGAGATGCTCCCTGTCCCTAGGAAATCAACAATGGATGA CAACATATAACCAAGAGGCAAAATTTTCCATCTCCATTGACCAAATATTAGAGGCTCACAATCAATCACC TTTCTGTCCCAGGTCTCCCAGATACACCTTGGACTTCGTAAATGGTTATCCTAAGATCTATTGGCCCCCC CCACAAGGGCGGCGCCGGTTTGGAGCCAGGGCCATGGTCACATATGATTGCGAGCCCCGATGCCCTTATG TGGGGGCAGATCACTTCGACTGCCCCCACTGGGACAATGCCTCCCAGGCCGATCAAGGGTCCTTTTATGT CAATCATCAGATTTTATTCCTGCATCTCAAACAATGTCATGGAATTTTCACTCTAACCTGGGAAATATGG GGATATGATCCCCTGATCACCTTTTCTTTACATAAAATTCCTGATCCCCCTCAACCCGACTTCCCTCAGC TGAACAGTGACTGGGTTCCCTCTGTCAGATCATGGGCCCTGCTTTTAAATCAAACGGCACGGGCCTTCCC AGACTGTGCTATATGTTGGGAACCTTCCCCTCCCTGGGCTCCCGAAATATTAGTATATAACAAAACCATC TCCGGCTCTGGACCCGGTCTCGCCCTCCCGGACGCCCAAATCTTCTGGGTCAACACGTCCTTGTTTAACA CCACCCAAGGATGGCACCACCCTTCCCAGAGGTTGTTGTTCAATGTTTCTCAAGGCAACGCCTTATTATT ACCCCCTATCTCCCTGGTTAATCTCTCTACGGTTTCCTCCGCCCCTCCTACCCGGGTCAGACGCAGTCCC GTCGCAGCCCTGACCTTAGGCCTAGCCCTGTCAGTGGGGCTCACTGGAATTAATGTAGCCGTGTCTGCCC TTAGCCATCAGAGACTCACCTCCCTGATCCACGTTCTGGAGCAAGATCAGCAACGCTTGATCACAGCAAT TAACCAGACCCATTATAATTTGCTTAATGTGGCCTCTGTGGTCGCCCAGAACCGACGGGGGCTTGATTGG TTGTACATCCGGCTGGGTTTTCAAAGCCTATGTCCCACAATCAATGAACCTTGCTGTTTCCTACGCATTC AAAATGACTCCATTATCCGCCTCGGTGATCTCCAGCCTCTCTCGCAAAGAGTCTCTACAGACTGGCAGTG GCCCTGGAATTGGGATCTGGGGCTCACCGCCTGGGTGCGAGAAACCATTCATTCTGTTCTAAGCCTATTC CTATTAGCCCTTTTTTTGCTCTTCTTGGCCCCCTGCCTGATAAAATGCTTGACCTCTCGCCTTTTAAAAC TCCTCCGGCAGGCTCCCCACTTCCCTGAAATCTCCTTCCCCCCTAAACCCGATTCTGATTATCAGGCCTT GCTACCGTCAGCACCAGAGATCTACTCTCACCTCTCCCCCACCAAACCCGATTACATCAACCTTCGACCC TGCCCTTGACACCCCCGTGTTTCACGCACCCTCAGGCTGTGGTGGGGCACTGGCTTAGTGGAATAGTCAG TGTACCATCACAAGCCTCTTCTTGCTGCCAGCACCGAGTTCGAACACAGCCCTACCCTGAGCCTCTCTGA GTACATGACTGAGTGTAGCGCAGAGAGGTTGTCGCTTCTGCGTGTCACTCAGTCATTTTTTATAGCCGAT TGGGGTTCGCGCCCTCCCATTGCCTGTGACACGGTTAAGACCTCTCTCACTTCTGCTTCACCATCCCCCT GCCAGCGTTGGTCTAGTGGAAAGAACTAACGCTGACGGGGGCGATTTCTTGCAGCTGTGCTAAGCGAGAG GCTCTGGTGCTGGGGATAAGATGCGGCCCCTAGCACCACAGTCTCTGCGCCTTTTGGGTTCGAATCTTCC CCATGCAGCTTCCGCTTTTTACGCCCTGTTGCACACCCTTTCTAGAGATACCTGAAAATCTCAGCTCGCA CCCCAAGGAAGGTTGTGGCTCAGAGGTTAAAATAGCTCGGACCGCAACCTCCCTTTCTTTTTATTCCACC CTCGCAAGGCCCCGGGTTCTGGGCCCCCTAACGGAGGTTCAGAATTTCCTCTACTAGGGGATGCTCAGGT CCAAGTGTGCACAATATCTCTTCCAAAAGGTCCTGATGAACATCTTCCCATGTAACAAGCCCCAGCAGAG ACATTCCAGCCACATCCAGCAGCATTTGGGCCGCCTTCTCTAACAGTGCCCATAAAGTCCCTTCCGTTTC CACAACGGCTGCCTCTGCATCTTCTATTTCCACCTCGGCACCGACTCCCCCGCCGAGCCCTTCAAGCTCT TCGGGATCCATTACCTGATAACGACAAAATTATTTCTTGTCTTTTAAGCAAGTGTTGTTGGTTGGGGGCC CCACTCTCTACATGCCTGCCCGGCCCTGGTTTTGTCCAATGATGTCACCATCGATGCCTGGTGCCCCCTC TGCGGGCCCCATGAACGACTCCAATTCGAAAGGATCGACACCACGCTCACCTGCGAGACCCACCGTATCA CCTGGACCGCCGATGGACGACCTTTTGGCCTCAATGGAACATTGTTCCCTCGACTGCATGTCTCCGAGAC CCGCCCCCAAGGGCCCCGACGACTCTGGATCAACTGCCCCCTTCCGGCCGTTCGCGCTCAGCCCGGCCCG GTTTCACTTTCCCCCTTCGAGCAGTCCCCCTTCCAGCCCTACCAATGCCAATTGCCCTCGGCCTCTAGCG ACGGTTGCCCCATCATCGGGCACGGCCTTCTTCCCTGGAACAACTTA >NP_056898.1 p18 [Bovine leukemia virus] MPKERRSRRRPQPIIRWQVLLVGGPTLYMPARPWFCPMMSPSMPGAPSAGPMNDSNSKGSTPRSPARPTV SPGPPMDDLLASMEHCSLDCMSPRPAPKGPDDSGSTAPFRPFALSPARFHFPPSSSPPSSPTNANCPRPL ATVAPSSGTAFFPGTT Protective antigen The bovine leukemia virus (BLV) envelope gene encoding extracellular glycoprotein gp51 (encoded by env gene) and transmembrane glycoprotein gp30 was cloned into a vehicle expression vector under the human cytomegalovirus (CMV) intermediate early promoter. The intramuscular injection of this plasmid vector generated a cellular immune response. Seven out of ten cows vaccinated with the DNA construct resisted a drastic challenge [Ref1047:Brillowska et al., 1999]. Brillowska et al., 1999 journal Brillowska A, Dabrowski S, RuÅ‚ka J, KubiÅ› P, BuzaÅ‚a E, Kur J 1999 46 4 971-976 Acta biochimica Polonica Kabeya et al., 1996 journal Kabeya H, Ohashi K, Ohishi K, Sugimoto C, Amanuma H, Onuma M 1996 14 12 1118-1122 Vaccine Larsen et al., 2013 journal Larsen A, Gonzalez ET, Serena MS, Echeverría MG, Mortola E 2013 54 2 475-483 Molecular biotechnology Usui et al., 2003 journal Usui T, Konnai S, Tajima S, Watarai S, Aida Y, Ohashi K, Onuma M 2003 65 11 1201-1205 The Journal of veterinary medical science / the Japanese Society of Veterinary Science Wiki: Bovine leukemia virus website http://en.wikipedia.org/wiki/Bovine_leukemia_virus