Characterization of a new calicivirus isolated from feces of a dog.

Canine calicivirus (CaCV), isolated from feces of a dog with diarrhea, was readily propagated in cultures of canine cells and in a dolphin cell line. Serologic evidence indicated many dogs in at least one geographic area had been infected with CaCV, but its role as an etiologic agent of disease was not established. In cell culture most CaCV virions were strongly cell-associated making purification difficult. CaCV was established as a member of the Caliciviridae by morphology and physicochemical properties of virions (density, sedimentation rate, single major polypeptide, RNA genome size), although some of the properties differed slightly from those of previously described caliciviruses; evidence was also obtained for caliciviral RNA species in infected cells. Based on tests with antisera to numerous caliciviruses and presumed caliciviruses, CaCV appeared to be not closely related to any previously described virus except the stunting syndrome agent of chickens.

Solid-phase radioimmunoassay for detection of plague antigen in animal tissue.

A rapid and sensitive radioimmunoassay for the detection of soluble Yersinia pestis fraction I antigen using antibody-coated beads and radiolabeled immunoglobulin was applied to spleen-liver extracts, rendered noninfectious by either treatment or filtration, from rodents dead of plague. The procedure was capable of detecting nanogram amounts of soluble plague antigen.

Assay of antibodies to caliciviruses by radioimmune precipitation using staphylococcal protein A as IgG adsorbent.

A radioimmune assay method designated St-RIP using a staphylococcal IgG adsorbent, which potentially has broad applications to viral (and nonviral) antigen-antibody systems, was applied to detection of calicivirus antibodies. Purified 125I-labeled virions of San Miguel sea lion virus serotypes 4 (SMSV-4) and 5 (SMSV 5) were incubated with sera; the immune complexes were reacted with an immunoadsorbent, formaldehyde-fixed staphylococci (Staphylococcus aureus protein A producer, strain Cowan I), and collected by centrifugation. Broad cross-reactivity was observed among serotypes of SMSV and vesicular exanthema of swine virus (VESV), but there was no reaction with antisera to six noncaliciviruses. Antibody production in a rabbit inoculated with SMSV-5 polypeptide was monitored by St-RIP assay; reactivity with intact SMSV-4 virion antigen was slightly less than, but closely paralleled, reactivity with SMSV-5 virion antigen. Applicability of the St-RIP test to serologic survey was demonstrated with pinniped, swine, and human (laboratory personnel) sera; numerous positive St-RIP reactions suggested the occurrence of widespread contacts with caliciviruses.

Single major polypeptide of a calicivirus: characterization by polyacrylamide gel electrophoresis and stabilization of virions by cross-linking with dimethyl suberimidate.

A calicivirus, San Miguel sea lion virus serotype 4, isolate 15FT, externally labelled with 125I, was shown by gel electrophoresis to possess a single major polypeptide. The polypeptide migrated anomalously upon electrophoresis in two sodium dodecyl sulfate (SDS) systems: more slowly than bovine serum albumin in a continuous phosphate-buffered system and more rapidly than bovine serum albumin in a discontinuous system. Estimated molecular weights in the two systems were approximately 71,000 and 64,000, respectively. There was no clear evidence for a minor virion polypeptide. Treatment of purified San Miguel sea lion virions with dimethyl suberimidate, a cross-linking reagent, preserved virion integrity during long-term storage at 4 degrees C. Oligomeric species of the polypeptide were observed upon electrophoresis of products from cross-linked virions. Based upon a preferred polypeptide molecular weight estimate of 71,000 and distribution of oligomeric species, a calicivirion model with 120 monomeric protein units is proposed as an alternative to a 180-unit model.

Survival of airborne influenza virus: effects of propagating host, relative humidity, and composition of spray fluids.

Influenza A virus, strain WSNH, propagated in bovine, human and chick embryo cell cultures and aerosolized from the cell culture medium, was maximally stable at low relative humidity (RH), minimally stable at mid-range RH, and moderately stable at high RH. Most lots of WSNH virus propagated in embryonated eggs and aerosolized from the allantoic fluid were also least stable at mid-range RH, but two preparations after multiple serial passage in eggs showed equal stability at mid-range and higher RH's. Airborne stability varied from preparation to preparations of virus propagated both in cell culture and embryonal eggs. There was no apparent correlation between airborne stability and protein content of spray fluid above 0.1 mg/ml, but one preparation of lesser protein concentration was extremely unstable at 50 to 80 per cent RH. Polyhydroxy compounds exerted a protective effect on airborne stability.

Biophysical comparisons of calicivirus serotypes isolated from pinnipeds.

Biophysical properties of three new San Miquel sea lion virus isolates from pinnipeds were examined and compared with those of previously characterized serotypes. The caliciviruses showed an identical sedimentation rate of 183S in 5-20% sucrose. Buoyant densities in CsCl were in the range of 1.35-1.39 g/ml, with differences among the serotypes.

Liquid scintillation radioassay in disposable microcentrifuge tubes: radioimmune precipitates and other applications.

A simple, economical radioassay system employing disposable polypropylene microcentrifuge tubes was developed. Plastic adapters permitted automatic operation in liquid scintillation spectrometers. Counting efficiencies of (3)H, (14)C, (32)P, and (125)I in liquid scintillation cocktails and of (32)P by Cerenkov radiation (at lower efficiency in absence of added scintillator) were comparable to those in standard vials. Multipurpose use of the microtubes made the system versatile and expedient, e.g., collection of precipitates and radioassay in the same container. Collection of radioimmune precipitates was aided by a carrier inorganic precipitate, Mg(2)P(2)O(7).

Electrophoretic analysis of ribosomal and viral ribonucleic acids with a simple technique for slicing low-concentration polyacrylamide gels.

The electrophoretic mobilities of ribosomal ribonucleic acids (RNA) from cultured mammalian (HeLa, Vero, MDBK), avian (chick embryo), and bacterial (Escherichia coli) cells, and RNA species extracted from selected viruses (Sindbis, polio, tobacco mosaic, Sendai) were compared, employing a simple, inexpensive technique for slicing low-concentration polyacrylamide gels. The procedure provides for rapid fractionation of gels used for characterization of RNA, incorporating extrusion and serial sectioning of frozen gels. Among 28S ribosomal RNA species, Vero and MDBK were indistinguishable, whereas HeLA RNA had a slightly lower mobility (higher apparent molecular weight) and chick RNA had a higher mobility (lower apparent molecular weight). The 18S ribosomal RNA species of the three mammalian sources were indistinguishable, but chick 18S RNA had a slightly lower apparent molecular weight. The inverse relation between mobility and log-molecular weight among the ribosomal and viral RNA species, though not highly precise, demonstrates the applicability of the technique to the study of molecular weights of viral RNA species.