Effect of periodate oxidation on hemagglutinating and antibody-producing capacities of certain enteroviruses and reoviruses. (1966)

Experiments in this laboratory have been concerned with the nature of the functional units of the capsids of reoviruses and enteroviruses.Hemagglutinating capacity of reoviruses types 1, 2, and 3 and of coxsackie virus group B, type 5, was destroyed after incubations with dilute potassium periodate.1Moreover, the in- fectivity of reovirus type 2 was also tested after similar periodate treatment, and was found to be markedly reduced.Hemagglutination (HA) by reoviruses and a number of enteroviruses was inhibited by digestions with f-glucosidase and incuba- tions with certain simple sugars.2' 3 The patterns and quantities of sugars necessary for these inhibition reactions were distinct for each virus.Smaller aldoses and alde- hydes also inhibited these HA reactions but their polyhydroxyl counterparts did not.Radioactive studies with carbon-14labeled sugars, aldoses, and aldehydes indicated that compounds which avidly inhibited these HA reactions attached to erythrocytes, but similar nonactive compounds did not.These active sugars, aldoses, and alde- hydes did not attach to purified preparations of the viruses.4These data have suggested to us that the capsids of reoviruses and enteroviruses may contain pre- viously unsuspected glycoproteins, and that virus-erythrocyte union may involve a free carbonyl group on oligosaccharide side chains of the viruses.Here we report that periodate oxidation simultaneously affects HA by a number of enteroviruses and reoviruses, reduces their infectivity, and alters these viruses in such a manner as to inhibit markedly their abilities to produce homotypic anti- bodies.Materials and Methods.-Viruses:Suspensions of rhesus kidney cells were obtained from Parke-Davis and Co. through the courtesy of Dr. W. Rightsel, and grown in 32-oz bottles with Eagle's medium and 10% calf serum.These cultures were inoculated with stocks of echoviruses types 3 (NIH-1), 7 (NIH-1), 11 (1867), 12 (NIH-1), and 19 (Burke); coxsackie viruses group B, types 1 (201468) and 5 (201482); and poliovirus type 1 (Brunhilde).When cytopathic effects were complete, cells which remained attached to glass were removed with a rubber policeman, and the virus-cell suspension was transferred to a 250-ml centrifuge bottle.It was then rapidly frozen and thawed three times in acetone and dry ice to lyse cell membranes and release intracellular virus.Reoviruses types 1 (Lang), 2 (988), and 3 (Abney) were produced in L-60 cells by methods previously described.,L-cell debris containing these viruses was treated with 100 Lg/ml of sodium deoxycholate for 2 hr (4°C) to release reoviruses which are attached to lipids of the sediment.Concentration and purification of viruses: Supernates from coxsackie, echo-, and polioviruses were treated 2 hr with deoxyribonuclease (50 Ag/ml), ribonuclease 50 Mg/ml), and trypsin (2%) at 40C, and the sediment was separated by centrifugation for 10 min at 1200 g (4°C).These final supernatant fluids were centrifuged in the Spinco model L ultracentrifuge at 27,000 rpm (19°F) for 5 hr.Pellets containing these viruses were taken up in 2-3 ml of Eagle's medium.These concentrated suspensions of coxsackie and echoviruses were further purified by ultracentrifugation in 15-65% sucrose gradients for 14 hr in the SW-25 swinging bucket rotor at 23,000 rpm (-8°C).Fractions were collected in 50-drop aliquots by puncturing the bottom of the cellulose