Shared Human/Rabbit Ligands for Rabbit Hemorrhagic Disease Virus

LETTERS

DOI: http://dx.doi.org/10.3201/eid1803.111486 Shared Human/ saline, RHDV strains isolated from whole liver extracts of infected References Rabbit Ligands for animals were incubated on coated Rabbit Hemorrhagic plates at dilutions corresponding to 1 × 1. Laemmert HW, Hughes T. The virus of 9 Ilheus encephalitis: isolation, serological Disease Virus 10 genome copies (0.2 μg/mL capsid specifi city and transmission. J Immunol. protein equivalent) as determined 1947;55:61–7. To the Editor: Rabbit by Nyström et al. (2). Monoclonal 2. Shope RE. Epidemiology of other arthro- hemorrhagic disease virus (RHDV) antibody 2G3, biotinylated anti-mouse pod-borne fl aviviruses infecting humans. In: Chambers T, Monath T, editors. The is a calicivirus of the genus Lagovirus IgG, and peroxidase-conjugated fl aviviruses: detection, diagnosis and vac- that causes epidemics of an acute avidin were used for RHDV detection; cination development. Vol. 61. Amster- disease and mortality rates of 50%– 3,3′,5,5′-tetramethylbenzidine was dam: Elsevier Academic; 2003. p. 386–7. 90% among rabbits. The disease, used as a substrate; and optical density 3. Nassar ES, Coimbra TL, Rocco IM, Pereira LE, Ferreira IB, de Souza LT, et which was fi rst described in 1984, is values at 450 nm were measured (2). al. Human disease caused by an arbovirus characterized by hemorrhagic lesions, Binding to the B type 2 epitope closely related to Ilheus virus: report of mainly affecting the liver and lungs was observed for all 6 strains (online fi ve cases. Intervirology. 1997;40:247–52. 24–72 h after infection (1). Technical Appendix Figure, panel A, http://dx.doi.org/10.1159/000150554 4. Spence L, Anderson CR, Downs WG. Iso- Similar to human caliciviruses wwwnc.cdc/gov/EID/pdfs/11-1402- lation of Ilheus virus from human beings of the genus Norovirus, RHDV Techapp.pdf). Human saliva samples in Trinidad, West Indies. Trans R Soc Trop binds to histo-blood group antigens were recognized by 5 of 6 RHDV Med Hyg. 1962;56:504–9. http://dx.doi. (HBGAs), and we recently showed strains. Only G6, an RHDV antigenic org/10.1016/0035-9203(62)90074-3 5. Srihongse S, Johnson CM. Isolation of that HBGAs serve as attachment variant also known as RHDVa (6), did Ilheus virus from man in Panama. Am J factors (ligands) that facilitate not show binding to saliva. Strains G1 Trop Med Hyg. 1967;16:516–8. RHDV infection (2). HBGAs are and G2 showed preferential binding 6. Johnson BW, Cruz C, Felices V, Espinoza polymorphic carbohydrate structures to saliva from B secretors over that WR, Manock SR, Guevara C, et al. Il- heus virus isolate from a human, Ecuador. representing terminally exposed from O secretors, and A secretors were Emerg Infect Dis. 2007;13:956–8. portions of larger glycans linked poorly recognized. Better recognition 7. Forshey BM, Guevara C, Laguna-Torres to proteins or glycolipids. In many of A secretor saliva was obtained VA, Cespedes M, Vargas J, Gianella A, vertebrate species, they are mainly with the G3 strain. The G4 and G5 et al. Arboviral etiologies of acute fe- brile illnesses in Western South Amer- expressed on epithelial surfaces. strains showed a clear preference for ica, 2000–2007. PLoS Negl Trop Dis. Because phylogenetic conservation A secretors over B and O secretors, 2010;4:e787. http://dx.doi.org/10.1371/ of receptors is a major risk factor for which indicated a shift in specifi city journal.pntd.0000787 cross-species transmission (3), we toward recognition of the A antigen 8. Martin DA, Muth D, Brown T, Johnson A, Karabatsos N, Roehrig J. Standardization analyzed the ability of RHDV strains from the H and B antigens, as reported of immunoglobulin M capture enzyme- to recognize human HBGAs expressed (2). None of the strains recognized linked immunosorbent assays for routine on epithelia. nonsecretor saliva, which showed diagnosis of arboviral infections. J Clin We obtained 38 saliva samples that binding to human saliva required Microbiol. 2000;38:1823–6. 9. Kuno G, Chang GJ, Tsuchiya KR, Kara- from healthy persons with ABO, A, B, or H motifs. This fi nding was batsos N, Cropp CB. Phylogeny of the Secretor, and Lewis phenotypes, and confi rmed by drastically decreased genus Flavivirus. J Virol. 1998;72:73–83. we selected confi rmed FUT2 (secretor) binding after removal of A, B, and 10. Kuno G, Chang GJ. Biological trans- and FUT3 (Lewis) genotypes (4) to H epitopes from secretor saliva by mission of arboviruses: reexamination of and new insights into components, include ABO, secretor, and Lewis treatment with specifi c glycosidases. mechanisms, and unique traits as well phenotypic diversity. Binding capacity There was no relationship with the as their evolutionary trends. Clin Micro- of 6 RHDV strains representative of Lewis status. biol Rev. 2005;18:608–37. http://dx.doi. virus diversity (2) was tested against To determine if human epithelial org/10.1128/CMR.18.4.608-637.2005 human saliva samples by using a cells were recognized by RHDV, Address for correspondence: Eric S. Halsey, method similar to that reported for binding of the G3 strain to human Naval Medical Research Unit 6, 3230 Lima Pl, human norovirus (5). tissue sections was assessed. Human Washington, DC 20521-3230, USA; email: eric. In brief, saliva samples diluted trachea, lung, and gastroduodenal [email protected] 1:1,000 or B type 2 bovine serum junction samples obtained from albumin–conjugated tetrasaccharide organ donors (before current French 1Current affi liation: University of Texas (positive control) were coated on restrictions of December 1988) were Medical Branch, Galveston, Texas, USA. ELISA plates. After blocking with used to prepare tissue microarrays. milk diluted in phosphate-buffered Tissues from 18 persons were used

518 Emerging Infectious Diseases • www.cdc.gov/eid • Vol. 18, No. 3, March 2012 LETTERS and represented the following Acknowledgments 6. Capucci L, Fallacara F, Grazioli S, La- phenotypes: O secretor Lewis+ (n We thank the cell and tissue imaging vazza A, Pacciarini ML, Brocchi E. A further step in the evolution of rabbit = 8), A secretor Lewis+ (n = 3), B core facility of Institut Fédératif de hemorrhagic disease virus: the appearance secretor Lewis+ (n = 2), O secretor Recherche Thérapeutique de Nantes for of the fi rst consistent antigenic variant. Lewis– (n = 1), and O nonsecretor preparing tissue microarrays; Ghislaine Virus Res. 1998;58:115–26. http://dx.doi. Lewis+ (n = 4). Deparafi nated and Le Gall-Reculé, Anses, Ploufragan, Pedro org/10.1016/S0168-1702(98)00106-3 7. Tan M, Jiang X. Norovirus-host in- endogen peroxidase–blocked sections Esteves, and Joana Abrantes for providing teraction: multi-selections by human were incubated overnight at 4°C with whole liver extracts of infected animals; histo-blood group antigens. Trends Mi- the G3 strain from an infected liver and Lorenzo Capucci for providing crobiol. 2011;19:382–8. http://dx.doi. extract at a concentration of 2 × 109 monoclonal antibody 2G3. org/10.1016/j.tim.2011.05.007 8. Gould AR, Kattenbelt JA, Lenghaus C, genome copies/mL. This study was supported by a grant Morrissy C, Chamberlain T, Collins BJ, et Binding was detected by using from the Région des Pays de la Loire al. The complete nucleotide sequence of monoclonal antibody 2G3 against rabbit haemorrhagic disease virus (Czech Calilago and by the Fédération Nationale RHDV, biotinylated anti-mouse IgG, strain V351): use of the polymerase chain des Chasseurs. reaction to detect replication in Aus- horseradish peroxidase–conjugated tralian vertebrates and analysis of viral avidin, and 3-amino-9-ethylcarbazole population sequence variation. Virus Res. substrate with hemalum counter- Kristina Nyström, 1997;47:7–17. http://dx.doi.org/10.1016/ staining, as described (2). Staining Béatrice Le Moullac-Vaidye, S0168-1702(96)01399-8 Nathalie Ruvoën-Clouet, 9. Merchán T, Rocha G, Alda F, Silva E, of epithelial cells of stomach or Thompson G, de Trucios SH, et al. De- trachea of secretors, but not those and Jacques Le Pendu tection of rabbit haemorrhagic disease of nonsecretors, was observed. This Author affi liations: Institut National de virus (RHDV) in nonspecifi c vertebrate fi nding indicated that attachment la Santé et de la Recherche Médicale, hosts sympatric to the European wild rab- Nantes, France; and University of Nantes, bit (Oryctolagus cuniculus). Infect Genet factors for RHDV are present on Evol. 2011;11:1469–74. http://dx.doi. human cells that constitute potential Nantes org/10.1016/j.meegid.2011.05.001 points of entry for RHDV (online DOI: http://dx.doi.org/10.3201/eid1803.111402 10. Kitchen A, Shackelton LA, Holmes EC. Technical Appendix Figure, panels Family level phylogenies reveal modes of macroevolution in RNA viruses. Proc Natl B–E). References Acad Sci U S A. 2011;108:238–43. http:// Attachment to HBGAs of human dx.doi.org/10.1073/pnas.1011090108 calicivirus strains represents a fi rst 1. Abrantes J, van der Loo W, Le Pendu J, Esteves PJ. Rabbit haemorrhagic disease Address for correspondence: Jacques Le Pendu, step of the infection process (7). (RHD) and rabbit haemorrhagic disease In this study, we have shown that virus (RHDV): a review. Vet Res. 2012. In Unité 892, Institut National de la Santé et de cross-species recognition of HBGAs press. la Recherche Médicale, 8 quai Moncousu, BP in cells that may be likely points of 2. Nyström K, Le Gall-Recule G, Grassi 70721, 44007 Nantes Cedex 1, France; email: P, Abrantes J, Ruvoën-Clouet N, Le [email protected] entry of RHDV into human cells. Moullac-Vaidye B, et al. Histo-blood RHDV infection has been shown to group antigens act as attachment factors of be rabbit specifi c (8), which indicates rabbit hemorrhagic disease virus infection Letters that other molecular elements not in a virus strain-dependent manner. PLoS Letters commenting on recent articles shared by rabbits and other mammals Pathog. 2011;7:e1002188. http://dx.doi. org/10.1371/journal.ppat.1002188 as well as letters reporting cases, restrict its host range. Nevertheless, 3. Woolhouse MEJ, Haydon DT, Antia R. outbreaks, or original research are RHDV RNA was recently isolated Emerging pathogens: the epidemiology welcome. Letters commenting on ar- from sympatric wild small mammals, and evolution of species jump. Trends ticles should contain no more than which suggested that the species range Ecol Evol. 2005;20:238–44. http://dx.doi. org/10.1016/j.tree.2005.02.009 300 words and 5 references; they are of RHDV may not be as limited as 4. Azevedo M, Eriksson S, Mendes N, Serpa more likely to be published if submitted previously believed (9). In addition, J, Figueiredo C, Resende LP, et al. Infec- within 4 weeks of the original article’s recent phylogenetic analysis showed tion by Helicobacter pylori expressing the publication. Letters reporting cases, that caliciviruses exhibit high levels BabA adhesin is infl uenced by the secretor phenotype. J Pathol. 2008;215:308–16. outbreaks, or original research should of host switching (10). Therefore, http://dx.doi.org/10.1002/path.2363 contain no more than 800 words and surveillance of RHDV and studies 5. de Rougemont A, Ruvoën-Clouet N, Si- 10 references. They may have 1 to decipher molecular mechanisms mon B, Estienney M, Elie-Caille C, Aho Figure or Table and should not be di- involved in its extreme pathogenicity S, et al. Qualitative and quantitative analysis of the binding of GII.4 norovirus vari- vided into sections. All letters should are warranted. ants onto human blood group antigens. contain material not previously pub- J Virol. 2011;85:4057–70. http://dx.doi. lished and include a word count. org/10.1128/JVI.02077-10

Emerging Infectious Diseases • www.cdc.gov/eid • Vol. 18, No. 3, March 2012 519 Article DOI: http://dx.doi.org/10.3201/eid1803.111402 Shared Human/Rabbit Ligands for Rabbit Hemorrhagic Disease Virus

Technical Appendix A Technical Appendix Figure. Binding of rabbit hemorrhagic disease virus (RHDV) to human saliva samples. A) Histo-blood group antigens of saliva samples are shown at the bottom of each panel, corresponding to binding of strains G1–G6. O, A, and B, secretors (separated by shading); se, nonsecretors. B type 2 represents synthetic B type 2 tetrasaccharide conjugated to bovine serum albumin and used as a positive control. Bars show optical density at 450 nm

(OD450). Signal above background values was not observed when whole liver extract from an uninfected animal was used. B–E) staining of human tissues for the G3 strain of RHDV. B) Trachea of an O secretor, showing staining of the entire epithelial layer and vascular endothelium in underlying connective B C tissue. C) Trachea of an O secretor, showing staining of only basal epithelial cells and vascular endothelium. D) Pyloric area of gastroduodenum junction of an O secretor, showing staining of surface epithelium. E) Pyloric area of an O nonsecretor, showing absence of staining of D E surface epithelium (monoclonal antibody 2G3 against RHDV, biotinylated anti-mouse IgG, horseradish peroxidase–conjugated avidin, and 3-amino-9-ethylcarbazole–stained with hemalum [hematoxylin and alum] counterstain.) Original magnifications ×400 in B and C and ×100 in D and E.

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