Journal of Fish Diseases 1995,18, 519-527 Low-pH increases the binding of haemorrhagic septicaemia rhabdovirus to membrane phospholipids J. M. COLL INIA, SanidadAnimal, aSA-Valdeolmos. Madrid, Spain Abstract. The rhabdovirus causing viral haemorrhagic septicaemia (VHS), a disease of coldwater fish, especially salmonids, binds to sonicated radioactively labelled membrane phospholipids [phosphatidylserine (PS), phosphatidylethanolamine (PE) and phosphatidylcholine (PC)J. The extent of binding of VHS V to membrane phospholipids is dependent on pH, occurring at physiological pH, but being maximal at about pH 5 5 and suggesting that a pH-induced conformational change is needed for the VHSV to maximally bind phosphohpids. VHSV bound PS more effectively than either PE or PC, as demonstrated by ultracentrifugation and competition experiments. Introduction The lack of vaccines against fish rhabdoviruses (viral haemorrhagic septicaemia, VHS; infectious haematopoietic necrosis, IHN; spring viaremia of carp, SVC) make these among the most damaging diseases of international aquaculture. We are trying to understand the determinants of these viral infec- tions to help in the design of recombinant vaccines (Leong & Fryer 1993; Lorenzen, Olesen, Vcstergaard- Jorgensen, Etzerodt, Holtet & Thorgersen 1993; Estepa, Thiry & Coll 1994) and/or viral inhibition agents. Since available evidence suggests that the mammalian rhabdovirus-cellular membrane interactions which lead to some of the initial steps in viral infections seem to involve viral-membrane phospholipid(s) inter- actions, the study of rhabdovirus-flsh cellular membrane phospholipid interactions (Bernard, Keroualt & de Kinkelin 1984) could help design new methods for control of these viruses. Phospholipid-detergent micelles extracted from cellular membranes are capable of inhibiting the at- tachment and the infection of some mammalian rhabdoviruses, including rabies (Superti, Seganti, Tsiang & Orsi 1984) and vesicular stomatitis virus (VSV) (Schlegel, Willigan & Pastan 1982). Of all the lipids tested, phosphatidylserine (PS) is the strongest inhibitor of VSV attachment to the cells (Schlegel, Sue, Willingham & Pastan 1983) at physiological pH, but 10 times more rabies or VSV attach to the cellular membranes at pH <6-5 than at higher pH (Wunner, Reagan & Koprowski 1984). After viral attachment, the internationalization of the virus by a temperature-dependent process occurs immediately. This is followed by the fusion of viral and cellular membranes at pH <5-5, as suggested by inhibition of fusion by lysosomotropic amines (Rigaut, Birk & Lenard 1991; Schlegel et al. 1982; Superti et al. 1984). Mammalian rhabdoviruses possess a unique glycoprotein G present in the virion as a homotrimeric integral membrane protein forming about 400 protruding 83 A spikes (Gaudin, Ruigrok, Tuffereau, Knossow & Ramand 1992). The trimers of glycoprotein G are responsible for the virus attachment to the cellular membrane and they also contain the fusion properties of the virus in both mammalian (Gaudin, Ruigrok, Knossow & Flamand 1993) and fish (Lecocq-Xhonneux, Thiry, Dheur, Rossius, Vanderheijden, Martial & DeKinkelin 1994) rhabdoviruses. Exposure of isolated trimers of the rabies glycoprotein G to low pH induces conformational and size changes (Gaudin et al. 1993) that expose some of its hydrophobic re- gions to the surface of the trimer, possibly allowing further interactions with the target cellular membrane (Bourhy, Kissi & Tordo 1993). In influenza, the membrane fusion step also depends on the exposure of a Correspondence: Dr J. M. Coll, INIA, Sanidad Animal, CIS A - Valdeolmos, 28130- Madrid, Spain. 519