The human respiratory syncytial virus matrix protein is required for maturation of viral filaments.

An experimental system was developed to generate infectious human respiratory syncytial virus (HRSV) lacking matrix (M) protein expression (M-null virus) from cDNA. The role of the M protein in virus assembly was then examined by infecting HEp-2 and Vero cells with the M-null virus and assessing the impact on infectious virus production and viral protein trafficking. In the absence of M, the production of infectious progeny was strongly impaired. Immunofluorescence (IF) microscopy analysis using antibodies against the nucleoprotein (N), attachment protein (G), and fusion protein (F) failed to detect the characteristic virus-induced cell surface filaments, which are believed to represent infectious virions. In addition, a large proportion of the N protein was detected in viral replication factories termed inclusion bodies (IBs). High-resolution analysis of the surface of M-null virus-infected cells by field emission scanning electron microscopy (SEM) revealed the presence of large areas with densely packed, uniformly short filaments. Although unusually short, these filaments were otherwise similar to those induced by an M-containing control virus, including the presence of the viral G and F proteins. The abundance of the short, stunted filaments in the absence of M indicates that M is not required for the initial stages of filament formation but plays an important role in the maturation or elongation of these structures. In addition, the absence of mature viral filaments and the simultaneous increase in the level of the N protein within IBs suggest that the M protein is involved in the transport of viral ribonucleoprotein (RNP) complexes from cytoplasmic IBs to sites of budding.

Fecal survey of parasites in free-roaming cats in northcentral Oklahoma, United States.

Cats can be infected by various intestinal parasites, some that are zoonotic. Although surveys of parasite prevalence in owned and shelter cats have been published, none addressed free-roaming, wild-trapped, domestic cat (Felis catus) populations. An opportunity to determine the prevalence of intestinal parasites in wild-trapped, free-roaming cats in northcentral Oklahoma, United States occurred through a trap-neuter-return (TNR) program conducted at Oklahoma State University, Boren Veterinary Medical Hospital, between February 2015 and April 2016. Approximately 1 g to 5 g of feces was collected from 846 free-roaming cats either from cage traps, when available, or rectally using disposable fecal loops and examined by centrifugal fecal flotation tests with 33% zinc sulfate solution. Parasite infections were confirmed by microscopic detection of eggs, cysts, or oocysts and visual detection of proglottids. Approximately 63.9% (541/846) of free-roaming cats were infected by at least one parasite, and 24.9% (211/846) of cats were infected by multiple parasites. The most common intestinal parasite infections detected were: Toxocara cati (44.6%; 377/846), followed by Alaria (13.4%; 113/846), Ancylostoma (11.2%; 95/846), Cystoisospora (9.7%; 82/846), taeniids (7.7%; 65/846), Dipylidium caninum (4.5%; 38/846), Physaloptera (2.2%; 19/846), Eucoleus aerophilus (1.4%; 12/846), Giardia (1.2%; 10/846), and a small (10-12 µm in diameter) Toxoplasma-like oocyst (0.1%; 1/846). A few ectoparasites, Demodex gatoi (0.5%; 4/846) and Cheyletiella (0.1%; 1/846), were also detected by fecal flotation. Our findings indicate that a higher prevalence of parasite infections occurs in free-roaming cats in Oklahoma than in owned cats, and these free-roaming cats contribute to contamination of the environment with several zoonotic parasites. Regional data on the prevalence of parasites in free-roaming cats can aid in the justification for parasite control programs in owned cats since both can share the same environment and supports the current practice of routine broad-spectrum anthelmintic and ectoparasite treatments for owned cats.

Prevalence of Alaria infection in companion animals in north central Oklahoma from 2006 through 2015 and detection in wildlife.

OBJECTIVE To determine the prevalence of Alaria infection in cats and dogs in north central Oklahoma over various periods and investigate whether wild animal species in this region were also infected. DESIGN Combined cross-sectional study and case series. SAMPLE Results of parasitological testing of fecal samples from 5,417 client-owned dogs and 1,246 client-owned cats (2006 through 2014); fecal samples from 837 shelter or rescue dogs and 331 shelter or rescue cats (2013 and 2014) and 268 feral cats (2015); tongue or jowl samples from cadavers of 43 wild pigs, 3 opossums, and 1 raccoon; and intestinal tract segments from cadavers of 48 cats and 5 coyotes. PROCEDURES Various parasite recovery techniques were performed to detect various Alaria stages in samples. Recovered adult trematodes and mesocercariae were used for PCR assay and sequencing of the 28S rRNA gene. RESULTS Prevalence of Alaria infection was significantly higher in feral cats (9.0%) than in shelter or rescue cats (0.6%) and client-owned cats (1.4%) and in shelter or rescue dogs (1.8%) than in client-owned dogs (0.2%). Mesocercariae were recovered from tissue samples from 11 (26%) wild pigs and 1 opossum. Amplicon sequences from adult trematodes and mesocercariae were 100% identical to each other and 99% homologous to GenBank sequences of Alaria alata and Alaria mustelae. CONCLUSIONS AND CLINICAL RELEVANCE Prevalence of Alaria infection in the study area has increased in dogs and cats since 1990, when infections were rare. Prevalence in wild pigs was similar to that in Eurasia, where A alata is considered an emerging zoonotic parasite.

High-throughput screening of a diversity collection using biodefense category A and B priority pathogens.

One of the objectives of the National Institutes of Allergy and Infectious Diseases (NIAID) Biodefense Program is to identify or develop broad-spectrum antimicrobials for use against bioterrorism pathogens and emerging infectious agents. As a part of that program, our institution has screened the 10 000-compound MyriaScreen Diversity Collection of high-purity druglike compounds against three NIAID category A and one category B priority pathogens in an effort to identify potential compound classes for further drug development. The effective use of a Clinical and Laboratory Standards Institute-based high-throughput screening (HTS) 96-well-based format allowed for the identification of 49 compounds that had in vitro activity against all four pathogens with minimum inhibitory concentration values of ≤16 µg/mL. Adaptation of the HTS process was necessary to conduct the work in higher-level containment, in this case, biosafety level 3. Examination of chemical scaffolds shared by some of the 49 compounds and assessment of available chemical databases indicates that several may represent broad-spectrum antimicrobials whose activity is based on novel mechanisms of action.