Defective interfering (DI) particles of vesicular stomatitis virus which contain covalently linked complementary [+]message and [-]anti-message RNA as a single-stranded ribonucleoprotein complex within the particle, are extremely efficient inducers of interferon. A single particle can induce a quantum yield of interferon. A single molecule of double-stranded RNA presumed to form, at least in part, on entry into the cell is thought to induce interferon synthesis. Conventional [-]RNA DI particles with the same polypeptide composition as [+/-]RNA DI particles fail to induce interferon.

A gene encoding chicken interferon (ChIFN) was cloned from a cDNA library made from primary chick embryo cells that had been "aged" in vitro so as to produce copious amounts of IFN upon induction. The coding region is predicted to produce a signal peptide of 31 amino acids and a mature protein of 162 amino acids with a molecular weight of 18,957. There are four potential N-glycosylation sites and six cysteine residues. Three disulfide bonds are possible, with two being common to most mammalian type I IFNs. A motif of 10 amino acids surrounding Cys-137 is highly conserved: It shows 80% homology with mammalian type I IFNs, but only 30% with a reported fish IFN. The T-rich 3' UTR displays the canonical element AATAAA required for polyadenylation, and contains six repeats of the octamer CTATTTAT that may be involved in down-regulating translation. Northern blots demonstrate that the accumulation of ChIFN mRNA correlates with induction of ChIFN determined by bioassay. Biologically active protein was synthesized in transfected mouse L cells using mRNA prepared in vitro from the cloned sequence. This activity was neutralized by a monoclonal antibody prepared against purified ChIFN. The ChIFN gene shows sequence identity at the amino acid/nucleotide level with consensus mammalian IFNs as follows: alpha (24/23%), beta (20/24%), omega (23/43%), tau (20/43%), gamma (3/31%), and with flatfish IFN (16/35%). The conserved features of the predicted ChIFN protein and the general similarity of predicted secondary structure suggest a molecule that fits the five alpha-helix three-dimensional topology reported for type I mammalian IFNs.(ABSTRACT TRUNCATED AT 250 WORDS)

Infectious bronchitis virus (IBV) causes an economically important respiratory disease in poultry worldwide. Previous studies have shown that CD8(+) cytotoxic T lymphocytes (CTL) are critical in controlling acute IBV infection, but the role of innate immunity is unknown. This study describes the in vitro and in vivo anti-IBV activity of natural spleen cell-derived and recombinant chicken interferon type I (rChIFN-alpha). Both natural and rChIFN-alpha inhibited replication of the Beaudette strain of IBV in chicken kidney cells (CKC) in a dose-dependent manner, with the antiviral activity of the former accounted for entirely by its content of type I IFN. IFN at 100 U/ml reduced viral replication by 50% as measured by syncytia formation. In addition, the spleen cell-derived supernatants (natural IFN) inhibited tracheal ring ciliostasis mediated by the Gray strain of IBV. Optimal protection against IBV-induced respiratory disease was obtained after intravenous or oral administration of ChIFN given 1 day before virus challenge and each of 5 days thereafter. ChIFN-I protected chicks from clinical illness by delaying the onset of the disease and decreasing the severity of illness, demonstrating its potential as an immune enhancer.

The dynamic changes in the temporal appearance and quantity of a new class of influenza virus, noninfectious cell-killing particles (niCKP), were compared to defective interfering particles (DIP). After a single high-multiplicity passage in MDCK cells of an egg-derived stock that lacked detectable niCKP or DIP, both classes of particles appeared in large numbers (>5 ؋ 10 8 /ml), and the plaque-forming particle (PFP) titer dropped ϳ60-fold. After two additional serial high-multiplicity passages the DIP remained relatively constant, the DIP/niCKP ratio reached 10:1, and the PFP had declined by about 10,000-fold. Together, the niCKP and DIP subpopulations constituted ca. 20% of the total hemagglutinating particle population in which these noninfectious biologically active particles (niBAP) were subsumed. DIP neither killed cells nor interfered with the cell-killing (apoptosis-inducing) activity of niCKP or PFP (infectious CKP), even though they blocked the replication of PFP. Relative to the UV-target of ϳ13,600 nucleotides (nt) for inactivation of PFP, the UV target for niCKP was ϳ2,400 nt, consistent with one of the polymerase subunit genes, and that for DIP was ϳ350 nt, consistent with the small DI-RNA responsible for DIP-mediated interference. Thus, niCKP and DIP are viewed as distinct particles with a propensity to form during infection at high multiplicities. These conditions are postulated to cause aberrations in the temporally regulated replication of virus and its packaging, leading to the production of niBAP. DIP have been implicated in the virulence of influenza virus, but the role of niCKP is yet unknown.

The particle-bound RNA polymerase activity of vesicular stomatitis virus (VSV) can be demonstrated in vivo. Linear synthesis of viral RNA persists for 5 to 6 hours at 34 degrees C in infected monolayers of chick embryo cells treated with cycloheximide and actinomycin D to block synthesis of protein and cell-specific RNA. At least 55 percent of the RNA made under these conditions is complementary to virion RNA. RNA synthesis mediated by VSV polymerase activity is inhibited in cells first treated with chick-derived interferon or polyriboinosinate* polyribocytidylate, but not by mouse interferon. The RNA product of VSV polymerase activity is present throughout the cytoplasm, and its synthesis is inhibited by the interferon system, as judged by autoradiographs that show the physical distribution, in cells, of RNA produced by virion polymerase in the absence of translation-a demonstration of the transcription product of the viral genome.