Four spontaneously derived serologicafly distinct classes of mutants of the Paramecium bursaria chlorella virus (PBCV-1) were isolated using polydonal antiserum prepared against either intact PBCV-l or PBCV-1-derived sero-types. The oligosaccharide(s) of the viral major capsid protein and two minor glycoproteins determined virus serological specificity. Normally, viral glycoproteins arise from host-specific glycosylation of viral proteins; the glycan portion can be altered only by growing the virus on another host or by mutations in glycosylation sites of the viral protein. Neither mechanism explains the changes in the glycan(s) ofthe PBCV-1 maJor capsid protein because all of the viruses were grown in the same host alga and the predicted amino acid sequence of the major capsid protein was identical in the PBCV-1 serotypes. PBCV-1 antiserum resistance is best explained by viral mutations that block specific steps in glycosylation, possibly by inactivating glycosyltansferases.Paramecium bursaria chlorella virus (PBCV-1) is a large (==190 nm in diameter) polyhedral plaque-forming virus that replicates in certain unicellular eukaryotic exsymbiotic Chlorella-like green algae (1). The PBCV-1 genome is a linear nonpermuted 333-kb double-stranded DNA with covalently closed hairpin ends (2, 3). PBCV-1 contains at least 50 proteins and a lipid component located inside the capsid shell (4). The major capsid protein (Vp54) ofPBCV-1 is one offour proteins located on the viral surface and is one of three glycosylated viral proteins. PBCV-1 and its related viruses have recently been assigned to a virus family, named Phycodnaviridae (5). Additional features of these viruses have been reviewed (6).Chase et al. (7) showed that chlorella viruses exclude one another during dual inoculation of the host. For these studies we isolated a spontaneous mutant of PBCV-1, named EPA-1, which was resistant to PBCV-1 polyclonal antiserum (dilution of 1:4). This antiserum completely inhibited PBCV-1 infection even at a dilution of 1:1000. Polyclonal antiserum prepared against EPA-1 inhibited EPA-1 infection (dilution of 1:1000) but not PBCV-1 infection (dilution of 1:4). In this study we have addressed two questions: (i) Can additional PBCV-1 serotypes be isolated using polyclonal antisera to intact viruses? (ii) What virus component determines antiserum resistance? We have found four PBCV-1 serotypes and discovered that the antisera react primarily with oligosaccharide(s) attached to the major capsid protein plus two minor glycoproteins. More importantly, in contrast to normal virus protein glycosylation, the virus rather than the host dictates glycosylation specificity.MATERIALS AND METHODS Culture Conditions. The procedures for producing, purifying, and "plaquing" the viruses and the growth ofthe host alga (Chlorella NC64A) on MBBM medium have been described (1,8). Virus glycoproteins were specifically labeled by adding 100 ,uCi of D-[6-3H]-galactose (1 Ci = 37 GBq) to 1 ml of cells (6 x 108 cells per ml) 30 min after virus infectio...

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We studied the role of membrane-derived oligosaccharides (MDOs) in sodium dodecyl sulfate (SDS) resistance by Escherichia coli. MDOs are also known as osmoregulated periplasmic glucans. Wild-type E. coli MC4100 grew in the presence of 10% SDS whereas isogenic mdoA and mdoB mutants could not grow above 0.5% SDS. Similarly, E. coli DF214, a mutant (pgi, zwf) unable to grow on glucose, exhibited conditional sensitivity to SDS in that it grew in gluconate and glucose or galactose but not in gluconate and mannose or sorbose. DF214 requires both gluconate and glucose/galactose because the gluconate is used for energy production, while glucose/galactose is used for MDO synthesis. Finally, the fate of E. coli cells subjected to SDS shock either during growth or when used as an inoculum is dependent on the presence or absence of sufficient MDOs. In both cases, cells grown under high-osmolarity (low-MDO) conditions were rapidly lysed by 5% SDS. Based on findings from a wild-type E. coli (MC4100), two mdo mutants and strain DF214 we conclude that MDOs are required for SDS resistance.

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