Comparative pathogenesis of turkey reoviruses.

Turkey reoviruses have been implicated in multiple disease syndromes resulting in significant economic losses to the turkey industry. It has been known for decades that turkey enteric reovirus (TERV) is involved in poult enteritis complex, but turkey arthritis reovirus (TARV), the causative agent of tenosynovitis in turkeys, emerged in 2011. In 2019, we isolated reovirus from several cases of hepatitis in turkeys and tentatively named it turkey hepatitis reovirus (THRV). The comparative pathogenesis of these viruses, and correlation with their genetic make-up (if any), is not known. In this study, we inoculated nine groups of 1-week-old turkey poults with two THRV, five TARV and two TERV via oral route. A tenth group served as a negative control. A subset of birds from each group was euthanised at 3, 5, 7, 14, 21, and 28 days post-inoculation (dpi). Tissues were collected for histology and real-time RT-PCR. All nine viruses were found to be enterotropic; the virus gene copy number in the intestine reached a peak at 5 dpi followed by a sharp decline at 7 dpi. All viruses caused a significant decline in body weight gain of birds as compared to the negative control group. Both TARV and THRV strains replicated in tendons and produced histologic lesions consistent with tenosynovitis. Hepatic lesions were produced by THRV only and the virus was re-isolated from liver and spleen of inoculated birds fulfilling Koch's postulates. The results of this study should be helpful in facilitating diagnosis and designing future mitigation plans.

Detection and molecular characterization of astroviruses in turkeys.

This study was conducted to determine the prevalence and molecular characteristics of turkey astrovirus 1 (TAstV-1) and avian nephritis virus (ANV) in turkeys with light turkey syndrome (LTS), which is characterized by lower body weight in market-age turkeys than their standard breed character. We collected pools of fecal samples from four LTS and two non-LTS turkey flocks in Minnesota at 2, 3, 5 and 8 weeks of age. Of the 80 LTS pools tested, 16 (20.0 %) and 11 (13.8 %) were positive for TAstV-1 and ANV, respectively. For non-LTS flocks, these numbers were 8 (20.0 %) and 5 (12.5 %), respectively. The maximum number of birds was positive at five weeks of age. We also tested 130 fecal samples of poult enteritis syndrome (PES) cases submitted to the Minnesota Veterinary Diagnostic Laboratory and found 19 and 11 positive for TAstV-1 and ANV, respectively. RdRp gene sequences were determined for a total of 29 TAstV-1 and 22 ANV samples. Phylogenetic analysis of the RdRp gene revealed 92-100 % and 88-100 % nucleotide sequence identity among TAstV-1 and ANV sequences, respectively. A large number of nucleotide and amino acid substitutions were observed in LTS and PES flocks than in non-LTS flocks. One of the PES sequences grouped with ANV-like sequences detected in chickens, indicating that regular screening of birds should be continued. Further, complete genome analysis should be conducted to determine whether this virus is a novel divergent strain or a recombinant of chicken and turkey ANV-like viruses. The detection of TAstV-1 and ANV in a considerable number of non-LTS cases emphasizes the need for further studies on the transmission pattern and pathogenesis of these viruses to determine their role as pathogens of turkeys.

Survival of Porcine Reproductive and Respiratory Syndrome Virus (PRRSV) in the Environment.

Porcine reproductive and respiratory syndrome (PRRS) is one of the most economically important diseases of swine, with losses due to poor reproductive performance and high piglet and growing pig mortality. Transmission of porcine reproductive and respiratory syndrome virus (PRRSV) may occur by both direct and indirect routes; the latter includes exposure to PRRSV-contaminated fomites, aerosols, and arthropod vectors. This review has collected available data on the ex-vivo environmental stability and persistence of PRRSV in an effort to highlight important sources of the virus and to determine the role of environmental conditions on the stability of the virus, especially temperature. The ex-vivo settings include fomites (solid, porous, and liquid fomites), insects, people, and pork meat, as well as the role of environmental conditions on the stability of the virus, especially temperature.

Inactivation of Three Subtypes of Influenza A Virus by a Commercial Device Using Ultraviolet Light and Ozone.

Avian influenza (AI) is a highly contagious disease that can be transmitted to naïve birds through fomites. The survival of AI viruses (AIV) on nonporous and porous fomites also dictates how long the fomite can serve as a vehicle for virus transmission. AIVs are known to be inactivated by ozone and ultraviolet (UV) light. However, the combined effect of UV light and ozone in combating AIV on different fomites has not been investigated. This study was undertaken to determine AIV inactivation by a commercial device called the BioSec shoe sanitizing station. This device generates both ozone and UV light for 8 sec when activated. We evaluated this device against three different subtypes of AIVs applied on seven different fomites. In general, the device inactivated all three AIV subtypes loaded on all fomites but to varying degrees of inactivation. The percentage of virus reduction on nonporous fomites (98.6%-99.9%) was higher than on porous fomites (90.0%-99.5%). In conclusion, this new device has the potential to help reduce the risk of transmission of AIV.

Inactivación de cuatro subtipos del virus de la influenza A mediante un dispositivo comercial usando luz ultravioleta y ozono. La influenza aviar (IA) es una enfermedad altamente contagiosa que puede transmitirse a aves susceptibles a través de fómites. La supervivencia de los virus de la influenza aviar en fómites porosos y no porosos también determina cuánto tiempo el fómite puede servir como vehículo para la transmisión del virus. Se sabe que los virus de influenza aviar son inactivados por el ozono y la luz ultravioleta (UV). Sin embargo, no se ha investigado el efecto combinado de la luz ultravioleta y el ozono para inactivar el virus de la influenza aviar en diferentes fómites. Este estudio se llevó a cabo para determinar la inactivación del virus de la influenza aviar mediante un dispositivo comercial llamado estación de desinfección de calzado BioSec. Este dispositivo genera ozono y luz ultravioleta durante 8 segundos cuando se activa. Se evaluó este dispositivo frente a cuatro subtipos diferentes del virus influenza aviar aplicados en siete fómites diferentes. En general, el dispositivo inactivó los cuatro subtipos de influenza aviar inoculados en todos los fómites, pero con distintos grados de inactivación. El porcentaje de reducción de virus en fómites no porosos (98.6%­99.9%) fue mayor que en fómites porosos (90.0%­99.5%). En conclusión, este nuevo dispositivo tiene el potencial de ayudar a reducir el riesgo de transmisión del virus de la influenza aviar.

Development, Optimization, and In Vitro/In Vivo Evaluation of Azelaic Acid Transethosomal Gel for Antidermatophyte Activity.

Treatment of dermatophytosis is quite challenging. This work aims to investigate the antidermatophyte action of Azelaic acid (AzA) and evaluate its efficacy upon entrapment into transethosomes (TEs) and incorporation into a gel to enhance its application. Optimization of formulation variables of TEs was carried out after preparation using the thin film hydration technique. The antidermatophyte activity of AzA-TEs was first evaluated in vitro. In addition, two guinea pig infection models with Trichophyton (T.) mentagrophytes and Microsporum (M.) canis were established for the in vivo assessment. The optimized formula showed a mean particle size of 219.8 ± 4.7 nm and a zeta potential of -36.5 ± 0.73 mV, while the entrapment efficiency value was 81.9 ± 1.4%. Moreover, the ex vivo permeation study showed enhanced skin penetration for the AzA-TEs (3056 µg/cm2) compared to the free AzA (590 µg/cm2) after 48 h. AzA-TEs induced a greater inhibition in vitro on the tested dermatophyte species than free AzA (MIC90 was 0.01% vs. 0.32% for T. rubrum and 0.032% for T. mentagrophytes and M. canis vs. 0.56%). The mycological cure rate was improved in all treated groups, specially for our optimized AzA-TEs formula in the T. mentagrophytes model, in which it reached 83% in this treated group, while it was 66.76% in the itraconazole and free AzA treated groups. Significant (p < 0.05) lower scores of erythema, scales, and alopecia were observed in the treated groups in comparison with the untreated control and plain groups. In essence, the TEs could be a promising carrier for AzA delivery into deeper skin layers with enhanced antidermatophyte activity.

In vitro virucidal activity of a commercial disinfectant against viruses of domestic animals and poultry.

Outbreaks of viral diseases in animals are a cause of concern for animal welfare and economics of animal production. One way to disrupt the cycle of infection is by combating viruses in the environment and prohibiting them from being transmitted to a new host. Viral contamination of the environment can be reduced using well-tested and efficacious disinfectants. Duplalim is a commercially available disinfectant consisting of 12% glutaraldehyde and 10% quaternary ammonium compounds. We evaluated this disinfectant for its efficacy against several viruses in poultry (n = 3), pigs (n = 5), dogs (n = 2), and cattle (n = 4). In suspension tests, 1:100 dilution of Duplalim was found to inactivate more than 99% of these 14 viruses in 15 min or less. The titers of a majority of these viruses decreased by ≥99.99% in <60 min of contact time. In conclusion, the ingredient combination in Duplalim is very effective in inactivating common viruses of domestic animals and poultry.

Molecular characterization of a novel Camelus dromedarius papillomavirus.

Papillomaviruses affect both human and non-human hosts. In camels, papillomatosis is caused by Camelus dromedarius papillomavirus type 1 and 2 (CdPV1 and CdPV2, respectively). In late 2018, an outbreak of camelpox occurred in a herd of fattening camels in Egypt. Several animals were found to be co-infected with camelpox and camel papillomaviruses. The morbidity with papillomatosis was 35 %. The infection was confirmed by PCR then Illumina sequencing revealed the presence of a complete genome of two CdPVs. One of these was CdPV1 (MT130101) and the other was a putative novel virus, tentatively named as CdPV3 (MT130100). Seven ORFs and a long upstream regulatory region were identified in the genomes of both viruses. Pairwise comparisons of L1 gene revealed 98.92 % nt identity between MT130101/CdPV1/Egypt/2018 and HQ912790/CdPV1/Sudan/2009 with 100 % coverage. However, MT130100/CdPV3/ Egypt/2018 showed only 68.99 % nt identity with the closest genome HQ912791/CdPV2/Sudan/2009. Phylogenetic analyses indicated that CdPV1 and CdPV3 belonged to the genus Deltapapillomavirus. These results should be useful for future CdPVs molecular surveillance and construction of evolutionary characteristics of this virus.

Enteric Viruses Associated with Mid-growth Turkey Enteritis.

Since August 2014, the University of Minnesota Veterinary Diagnostic Laboratory has received cases of turkey enteritis that are clinically different from previously described cases of poult enteritis syndrome and light turkey syndrome. The birds develop dark green and extremely foul-smelling diarrhea starting at 8-10 wk of age, which may last up to 15-16 wk of age. The affected turkey flocks show poor uniformity, and feed conversion and market weights are reduced. Multiple-age farms are affected more often than the single-age farms. Morbidity varies from flock to flock and in some cases reaches 100%. At necropsy, undigested feed with increased mucus is observed in the intestines along with prominent mucosal congestion and/or hemorrhage. Microscopically, lymphocytic infiltrates expand the villi in duodenum and jejunum to form lymphoid follicles, which are often accompanied by heterophils. Next generation sequencing (Illumina Miseq) on a pool of feces from affected birds identified genetic sequences of viruses belonging to Astroviridae, Reoviridae, Picornaviridae, Picobirnaviridae, and Adenoviridae. On testing pools of fecal samples from apparently healthy (16 pools) and affected birds (30 pools), there was a higher viral load in the feces of affected birds. Picobirnavirus was detected only in the affected birds; 20 of 30 pools (66.7%) were positive. These results indicate that a high viral load of turkey picobirnavirus alone, or in association with novel picornaviruses, may be a cause of this new type of turkey enteritis.

Detection and molecular characterization of kobuvirus from diarrheic goats in Minnesota.

Kobuvirus infections are common among humans, rodents, carnivores, pigs, and ruminants. We report herein the complete genome sequence of a novel caprine kobuvirus (MN604700) from diarrheic kids in Minnesota. Whole-genome sequencing revealed a kobuvirus genome of 8,139 nt with a single ORF region encoding a polyprotein of 2,480 amino acids. Further analysis revealed nt substitutions along the genome compared with that of the caprine kobuvirus reference strain, with 93% identity. Phylogenetic analysis indicated that the clade of the caprine kobuvirus was most closely related to porcine kobuviruses rather than bovine or ovine kobuviruses. Using primers designed from this genome, caprine kobuvirus was identified in the stools of other goats. Sanger sequencing of PCR products indicated 3D and VP1 gene nucleotides of this latter strain were 95% and 91% identical with those of MN604700, respectively. There were 35 and 101 nt substitutions in 3D and VP1 genes, respectively. Findings of kobuvirus over a 2-y period may indicate an endemic state, which needs further research. In addition, screening for kobuviruses over large geographic areas is needed to identify the evolutionary connections among different strains.

Virulence factors and antibiograms of Escherichia coli isolated from diarrheic calves of Egyptian cattle and water buffaloes.

Diarrhea caused by Escherichia coli in calves is an important problem in terms of survivability, productivity and treatment costs. In this study, 88 of 150 diarrheic animals tested positive for E. coli. Of these, 54 samples had mixed infection with other bacterial and/or parasitic agents. There are several diarrheagenic E. coli pathotypes including enteropathogenic E. coli (EPEC), Shiga-toxin producing E. coli (STEC), enterotoxigenic E. coli (ETEC) and necrotoxigenic E. coli (NTEC). Molecular detection of virulence factors Stx2, Cdt3, Eae, CNF2, F5, Hly, Stx1, and ST revealed their presence at 39.7, 27.2, 19.3, 15.9, 13.6, 9.0, 3.4, and 3.4 percent, respectively. As many as 13.6% of the isolates lacked virulence genes and none of the isolate had LT or CNF1 toxin gene. The odds of isolating ETEC from male calves was 3.6 times (95% CI: 1.1, 12.4; P value = 0.042) that of female calves, whereas the odds of isolating NTEC from male calves was 72.9% lower (95% CI: 91.3% lower, 15.7% lower; P value = 0.024) than that in females. The odds of isolating STEC in winter was 3.3 times (95% CI: 1.1, 10.3; P value = 0.037) that of spring. Antibiograms showed 48 (54.5%) of the isolates to be multi-drug resistant. The percent resistance to tetracycline, streptomycin, ampicillin, and trimethoprim-sulfamethoxazole was 79.5, 67.0, 54.5, and 43.0, respectively. Ceftazidime (14.8%), amoxicillin-clavulanic acid (13.6%) and aztreonam (11.3%) showed the lowest resistance, and none of the isolates was resistant to imipenem. The results of this study can help improve our understanding of the epidemiological aspects of E. coli infection and to devise strategies for protection against it. The prevalence of E. coli pathotypes can help potential buyers of calves to avoid infected premises. The antibiograms in this study emphasizes the risks associated with the random use of antibiotics.

Epidemiological and ultrasonographic investigation of bovine fascioliasis in smallholder production system in Eastern Nile Delta of Egypt.

Regular updating of our knowledge on the epidemiological determinants of bovine fascioliasis is necessary to increase the awareness of the disease's significance and subsequently, improve the control measures. The objectives of this study were (1) to estimate the prevalence of bovine fascioliasis, and identify the association of epidemiological characteristics under traditional householders' production systems, (2) to describe the association between the clinical picture, Fasciola spp. egg count and hepatobiliary ultrasonography findings. In total, 270 faecal samples were examined microscopically for the presence or absence of Fasciola spp. egg, using the sedimentation-flotation method. Copro-positive animals were subjected to ultrasonographic examination. Overall prevalence of copro-positive animals was 27.4% (22.4-33.0%, 95% CI). The final multivariate analysis showed that there was a significant association between fascioliasis and animal species (P < 0.03), and administration of anthelmintic (P < 0.0001). Cattle have a less chance of being positive to Fasciola spp. by 0.55 (95% CI: 0.30 - 0.99) compared to water buffaloes. Administration of anthelmintic to animals on a regular basis decreased the risk of copro-positivity to Fasciola spp by 0.17 (95% CI: 0.07 - 0.36) compared to animals received anthelmintic on an irregular basis. Infected animals having different Fasciola spp. egg burden revealed different clinical symptoms associated with hepatobiliary changes on ultrasonographic examination ranged from normal hepatic parenchyma and bile system in low faecal egg load to hyperechogenic hepatic parenchyma, hyperechogenic with distal shadowing bile duct, and distended gallbladder in high faecal egg load of Fasciola spp. In conclusion, the prevalence of bovine fascioliasis is high under the traditional household's production system. Regular administration of anthelmintic significantly reduces the animal's chance of being copro-positive to Fasciola spp. Ultrasound poses a valuable prognostic technique for assessment of bovine fascioliasis.

Outbreaks of foot and mouth disease in Egypt: Molecular epidemiology, evolution and cardiac biomarkers prognostic significance.

Foot and mouth disease virus (FMDV) was isolated from sloughed tongue epithelium of Egyptian cattle presenting with mouth lesions and ropy salivation in two Egyptian governorates (El-Fayoum and Dakahlia). The virus was isolated in Madin-Darby bovine kidney (MDBK) cells and identified by reverse transcription-polymerase chain reaction (RT-PCR). The complete genome was obtained by next generation sequencing. The strains isolated from El-Fayoum and Dakahlia were serotype A and O, respectively and both isolates had identity with the previously reported Egyptian strains. This study reports successive outbreaks of FMDV that occurred in Egypt during 2015-2016 and describes the dynamics of two outbreaks in addition to the use of cardiac biomarkers in the diagnosis of FMD-related myocarditis in calves and its clinical relevance. Serum cardiac troponin1 (cTn I) and creatinine kinase myocardial band (CK-MB) were measured. Mean serum cardiac troponin1 (cTn I) showed significant increase (P < 0.001) in FMDV-infected calves. The increase in fatal and recovered cases was (2.794 ±â€¯0.502 ng/mL) and (1.196 ±â€¯0.443 ng/mL), respectively, compared to the healthy control cases (0.014 ±â€¯0.002 ng/mL). Thus, the serum cTn-I successfully diagnosed FMD-associated myocarditis in calves but not prognostic for the fatal cases. The FMDV sequences described in this study should further help in studying FMDV endemicity in Egypt, tracking the source of infection, selection of control strategies and vaccine updates. The study also determines the clinical relevance of cardiac biomarkers in diagnosis of FMDV-related myocarditis in infected calves.

Surveillance, isolation and complete genome sequence of bovine parainfluenza virus type 3 in Egyptian cattle.

Parainfluenza virus type 3 (PIV-3) can infect a wide variety of mammals including humans, domestic animals, and wild animals. In the present study, bovine parainfluenza virus type 3 (BPIV-3) was isolated from nasal swabs of Egyptian cattle presenting with clinical signs of mild pneumonia. The virus was isolated in Madin-Darby bovine kidney (MDBK) cells and confirmed by reverse transcription-polymerase chain reaction (RT-PCR). The complete genome of Egyptian BPIV-3 strain was sequenced by using next generation (Illumina) sequencing. The new isolate classified with genotype A of BPIV-3 and was closely related to the Chinese NM09 strain (JQ063064). Subsequently in 2015-16, a molecular surveillance study was undertaken by collecting and testing samples from cattle and buffaloes with respiratory tract infections. The survey revealed a higher rate of BPIV-3 infection in cattle than in buffaloes. The infection was inversely proportional to the age of the animals and to warm weather. This report should form a basis for further molecular studies on animal viruses in Egypt.

A Newly Emergent Turkey Arthritis Reovirus Shows Dominant Enteric Tropism and Induces Significantly Elevated Innate Antiviral and T Helper-1 Cytokine Responses.

Newly emergent turkey arthritis reoviruses (TARV) were isolated from tendons of lame 15-week-old tom turkeys that occasionally had ruptured leg tendons. Experimentally, these TARVs induced remarkable tenosynovitis in gastrocnemius tendons of turkey poults. The current study aimed to characterize the location and the extent of virus replication as well as the cytokine response induced by TARV during the first two weeks of infection. One-week-old male turkeys were inoculated orally with TARV (O'Neil strain). Copy numbers of viral genes were estimated in intestines, internal organs and tendons at ½, 1, 2, 3, 4, 7, 14 days Post inoculation (dpi). Cytokine profile was measured in intestines, spleen and leg tendons at 0, 4, 7 and 14 dpi. Viral copy number peaked in jejunum, cecum and bursa of Fabricius at 4 dpi. Copy numbers increased dramatically in leg tendons at 7 and 14 dpi while minimal copies were detected in internal organs and blood during the same period. Virus was detected in cloacal swabs at 1-2 dpi, and peaked at 14 dpi indicating enterotropism of the virus and its early shedding in feces. Elevation of IFN-α and IFN-ß was observed in intestines at 7 dpi as well as a prominent T helper-1 response (IFN-γ) at 7 and 14 dpi. IFN-γ and IL-6 were elevated in gastrocnemius tendons at 14 dpi. Elevation of antiviral cytokines in intestines occurred at 7dpi when a significant decline of viral replication in intestines was observed. T helper-1 response in intestines and leg tendons was the dominant T-helper response. These results suggest the possible correlation between viral replication and cytokine response in early infection of TARV in turkeys. Our findings provide novel insights which help elucidate viral pathogenesis in turkey tendons infected with TARV.