Design,synthesis and evaluation against Chikungunya virus of novel small-molecule antiviral agents

Chikungunya virus is a re-emerging arbovirus transmitted to humans by mosquitoes, responsible for an acute flu-like illness associated with debilitating arthralgia, which can persist for several months or become chronic. In recent years, this viral infection has spread worldwide with a previously unknown virulence. To date, no specific antivirals treatments nor vaccines are available against this important pathogen. Starting from the structures of two antiviral hits previously identified in our research group with in silico techniques, this work describes the design and preparation of 31 novel structural analogues, with which different pharmacophoric features of the two hits have been explored and correlated with the inhibition of Chikungunya virus replication in cells. Structure-activity relationships were elucidated for the original scaffolds, and different novel antiviral compounds with EC₅₀ values in the low micromolar range were identified. This work provides the foundation for further investigation of these promising novel structures as antiviral agents against Chikungunya virus.     

Sequence analysis of a soil-borne wheat mosaic virus isolate from Italy shows that it is the same virus as European wheat mosaic virus and Soil-borne rye mosaic virus

The complete sequence of the two RNAs of a furovirus isolate from durum wheat in Italy was determined. Sequence comparisons and phylogenetic analysis were done to compare the Italian virus with Soil-borne wheat mosaic virus (SBWMV) from the USA and with furovirus sequences recently published as European wheat mosaic virus (EWMV), from wheat in France, and Soil-borne rye mosaic virus (SBRMV), from rye and wheat in Germany. Over the entire genome, the Italian isolate RNA1 and RNA2 had respectively 97.5% and 98.6% nucleotide identity with EWMV, 95.5% and 85.8% with SBRMV-G and 70.6% and 64.5% with SBWMV. The Italian isolate was therefore clearly distinct from SBWMV. The European isolates all appear to belong to the same virus and the name Soil-borne cereal mosaic virus may resolve earlier ambiguities.     

Comparison between inactivated Sendai virus, polyethylene glycol and Tween 80 as permeabilizing agents for introduction of neurospora endonuclease into X-irradiated cells

Inactivated Sendai virus, polyethylene glycol and Tween 80 were employed as agents to make X-irradiated CHO cells permeable for Neurospora endonuclease, in studies designed to evaluate the influence of this enzyme on the frequencies of X-ray-induced chromosome aberrations. Polyethylene glycol and Tween 80 were found not to be very efficient in making cells permeable. Besides, polyethylene glycol was found to increase the frequencies of X-ray-induced chromosomal aberrations.     

Serological status for N. caninum, bovine viral diarrhea virus, and infectious bovine rhinotracheitis virus at pregnancy testing and reproductive performance in beef herds

Neospora caninum, infectious bovine rhinotracheitis (IBR), and bovine viral diarrhea virus (BVDV) are important differentials for the diagnosis of infectious reproductive loss in beef herds. The objective of this study was to describe the serological status of both pregnant and non-pregnant beef cows from herds with varying levels of reproductive success. The study provided an opportunity to examine whether there were any associations between serological status for BVDV, IBR, and N. caninum and pregnancy status, as well as the subsequent risk of abortion, or stillbirth. Samples were collected from 2516 cows and heifers from 66 herds; 31 herds where the proportion pregnant was <90% and 35 randomly selected herds where the proportion pregnant was ≥90%. Of these samples 5.9% were positive for antibodies to N. caninum, 20.4% had titres >1:80 to IBR, 91.8% had titres ≥1:256 to BVDV type 1, and 23.9% had titres ≥1:256 to BVDV type 2. N. caninum antibody concentration was associated with an increased individual animal risk of non-pregnancy (OR_log S/P, 1.9; 95% CI, 1.2–2.9) and abortion (OR_pos/neg, 2.8; 95% CI, 1.1–7.5). The proportion of animals at pregnancy testing with antibodies to BVDV type 2 above 1:3000 (OR_{10% change in prevalence}, 2.3; 95% CI, 1.5–3.5) was also associated with an increased risk of abortion. No other measures of antibody status were associated with reduced reproductive performance in this group of herds. Antibodies to Mycobacterium avium spp.paratuberculosis were also measured; 0.7% of samples were positive (sample to positive (S/P) >0.25) and 3.6% were suspicious (S/P, 0.10–0.25).     

Cantaloupe line CZW-30 containing coat protein genes of cucumber mosaic virus,zucchini yellow mosaic virus,and watermelon mosaic virus-2 is resistant to these three viruses in the field

Cantaloupe line CZW-30 containing coat protein gene constructs of cucumber mosaic cucumovirus (CMV), zucchini yellow mosaic potyvirus (ZYMV), and watermelon mosaic virus 2 potyvirus (WMV-2) was investigated in the field over two consecutive years for resistance to infections by CMV, ZYMV, and/or WMV-2. Resistance was evaluated under high disease pressure achieved by mechanical inoculations and/or natural challenge inoculations by indigenous aphid vectors. Across five different trials, homozygous plants were highly resistant in that they never developed systemic symptoms as did the nontransformed plants but showed few symptomatic leaves confined close to the vine tips. Hemizygous plants exhibited a significant delay (2–3 weeks) in the onset of disease compared to control plants but had systemic symptoms 9–10 weeks after transplanting to the field. Importantly, ELISA data revealed that transgenic plants reduced the incidence of mixed infections. Only 8% of the homozygous and 33% of the hemizygous plants were infected by two or three viruses while 99% of the nontransformed plants were mixed infected. This performance is of epidemiological significance. In addition, control plants were severely stunted (44% reduction in shoot length) and had poor fruit yield (62% loss) compared to transgenic plants, and most of their fruits (60%) were unmarketable. Remarkably, hemizygous plants yielded 7.4 times more marketable fruits than control plants, thus suggesting a potential commercial performance. This is the first report on extensive field trials designed to assess the resistance to mixed infection by CMV, ZYMV, and WMV-2, and to evaluate the yield of commercial quality cantaloupes that are genetically engineered.     

Semliki Forest virus membrane proteins, preparation and characterization of spike complexes soluble in detergent-free medium

After Triton X-100 delipidation and subsequent Triton X-100 removal in a sucrose gradient the membrane protein spikes of Semliki Forest virus remained soluble in aqueous buffers. It was shown they were present as octameric complexes with a molecular weight of 95 · 10⁴ and that they contain less than 4% lipid and detergent by weight. In electron microscopy after negative staining they appeared as “rosette”-shaped particles. Part of the protein could also be found associated in ordered paracrystalline arrays.     

Reactivation of herpes simplex virus in a cell line inducible for simian virus 40 synthesis

The reactivation of UV-irradiated herpes simplex virus (HSV) was investigated in irradiated and unirradiated transformed hamster cells in which infectious simian virus 40 (SV40) can be induced. Reactivation was enhanced when the cells were treated with UV light or mitomycin C prior to infection with HSV. The IV dose-response curve of this enhanced reactivation was strikingly similar to that found for induction of SV40 virus synthesis in cells treated under identical condictions. This is the first time that two SOS functions described in bacteria have been demonstrated in a single mammalian cell line.     

Crystallization of belladonna mottle virus

Belladonna mottle virus belongs to the turnip yellows mosaic virus group of the small spherical plant viruses. It contains 180 protein subunits, which are arranged in a T = 3 icosahedral surface lattice. The top and bottom viral components crystallize isomorphously in hexagonal space group R3 ($\text{a = b = 296}\text{A}\text{?}\text{, c = 729}\text{A}\text{?}$). The unit cell contains three virus particles, while the crystallographic asymmetric unit consists of only one-third of a particle. X-ray diffraction data from the crystals extend to at least 3.8 Å resolution.     

Selective inactivation of the infectivity of freeze-dried Sendai virus by heat

The infectivity of freeze-dried Sendai virus was destroyed after heating at 100 ° C for 20 min while the hemagglutinin (HA) titer and the hemolytic (HL) activity were not affected. The HA titer was unaltered after heating at up to 140 ° C for 30 min. The HL activity was increased after freeze-drying, further increased after heating of freeze-dried virus at 115 ° C for 20 min, but was destroyed after heating for 30 min at 140 ° C.The selective heat inactivation of freeze-dried Sendai virus could be of use in the production of myxovirus vaccines and inactivated virus for cell-fusion studies.     

Characterization and description of a virus causing salivary gland hyperplasia in the housefly,Musca domestica

Abstract. A double-stranded DNA virus was isolated from hyperplasic salivary glands of male and female houseflies, Musca domestica L. (Diptera: Muscidae), collected from a dairy in Alachua County, Florida, U.S.A. Sodium dodecyl sulphate (SDS)–polyacrylamide gel electrophoresis (PAGE) of this housefly salivary gland hyperplasia (SGH) virus revealed the presence of two major and eight minor structural polypeptides. Restriction endonuclease analysis indicated that the c. 137 kilobase pair DNA was double-stranded. Weekly sweep-net sampling of the fly population throughout the season (May-October, 1991) showed that 1.5-18.5% of the dissected flies possessed hyperplasic salivary glands. The virus replicated within the nuclei of the salivary gland cells and was transmitted per os to newly-emerged healthy adult flies.     

Crystallization of tobacco ringspot virus

Tobacco ringspot virus belongs to the nepovirus group of the small spherical plant viruses. It has been suggested that the protein capsid, which displays icosahedral symmetry, consists of 240 protein subunits which are arranged on a T = 4 surface lattice. The virus crystallizes in orthorhombic space group P2₁2₁2₁ ($\text{a = 388}\text{A}\text{?}\text{, b = 396}\text{A}\text{?}\text{,}\text{and}\text{c = 405}\text{A}\text{?}$). The unit cell contains four virus particles while the crystallographic asymmetric unit consists of one complete virion. X-ray diffraction data from the crystals extend to at least 3.3 Å resolution.     

Resistance to potato virus Y and potato virus X in Solanurn brevidens

Although Solanum brevidens could be infected with potato virus X (PVX), potato virus Y⁰ (PVY⁰) and PVY^N, no symptoms of infection were apparent and tests by double antibody sandwich ELISA, electron microscopy and sap transmission to local lesion test plants indicated that the titres of PVX were less than a tenth of those of PVY⁰ and PVY^N were less than a hundredth of those in infected plants of PDH40, a susceptible dihaploid clone of S. tuberosum cv. Pentland Crown. Furthermore, PVY⁰- and PVY^N- infected leaves of S. brevidens were a poor source of inoculum in aphid transmission tests. The possibility of a common mechanism and genetic basis of resistance to PVY, PVX and potato leaf roll virus in S. brevidens is discussed.     

Structural investigations of influenza B virus

Purified preparations of influenza B/Hong Kong/5/72 have been characterized by hydrodynamical measurements, electron microscopy and small-angle neutron scattering. As judged by these techniques the preparations are highly monodisperse, the virus particles being spherical and of molecular weight about 200 × 10⁶. The lipid bilayer is located at a radius of 425 Å and its molecular weight is estimated to be 60 × 10⁶, constituting about 30% of the total virus mass. The external radius is about 580 Å.     

Crystallization of cowpea chlorotic mottle virus

Cowpea chlorotic mottle virus belongs to the brome mosaic virus group of the small spherical plant viruses. It contains 180 protein subunits, which are arranged on a T = 3 icosahedral surface lattice. The virus crystallizes in orthorhombic space group P2₁2₁2 ($\text{a = 394}\text{A}\text{?}\text{, b = 382}\text{A}\text{?}\text{,}\text{and}\text{c = 397}\text{A}\text{?}$). The unit cell contains four virus particles, while the crystallographic asymmetric unit consists of one complete virion. X-ray diffraction data from the crystals extend to nearly 3.0 Å resolution.     

Identification of new protein kinase-related genes in three herpesviruses, herpes simplex virus, varicella-zoster virus, and Epstein-Barr virus.

By using amino acid sequence patterns (motifs) diagnostic of conserved regions within the catalytic domains of protein kinases, homologous open reading frames of three herpesviruses were identified as protein kinase-related genes. The three sequences, herpes simplex virus gene UL13, varicella-zoster virus gene 47, and Epstein-Barr virus gene BGLF4, resemble serine/threonine kinases rather than tyrosine kinases.     

Sialylatin of glycoproteins of murine mammary tumor virus, murine leukemia virus, and Mason-Pfizer monkey virus.

Neuraminidase treatment of mouse mammary tumor virus, Rauscher murine leukemia virus, and Mason-Pfizer monkey virus resulted in loss of their capacity to inhibit hemagglutination of influenza virus. Hemagglutination-inhibition activity of these RNA tumor viruses could be restored by in vitro resialylation catalyzed by sialyl transferase. The major glycoprotein in the intact envelope of desialylated and, to some extent, native virions could be specificallly labeled in vitro with CMP-(14C) sialic acid. These studies further characterize the individual glycoproteins of mouse mammary tumor virus, Rauscher murine leukemia virus, and Mason-Pfizer monkey virus.