The impact of grandparenting on mental health among rural middle-aged and older adults in China: exploring the role of children's support.

Objectives: In the rural regions of China, characterized by a pronounced aging demographic and limited resources, a substantial proportion of middle-aged and older adults engage in grandparenting roles. Yet, the literature lacks consistent evidence regarding the effects of grandparenting on the mental health of this cohort. Accordingly, this study aimed to explore the impact of grandparenting on the mental health of rural middle-aged and older adults, as well as the underlying mechanisms. Methods: This analysis encompassed 10,881 middle-aged and older adults, utilizing data from the 2018 Harmonized China Health and Retirement Longitudinal Study (CHARLS). The mental health of participants was assessed using the Center for Epidemiological Studies Depression-10 (CESD-10) scale, while support from children was categorized into financial and emotional types. The study employed logistic and OLS regression models to identify the mediating role of child support and utilized the Karlson-Holm-Breen (KHB) method for decomposing this mediating effect. Results: The findings demonstrated that grandparenting had a significant negative impact on depression among rural middle-aged and older adults. Furthermore, children's support played a vital role in mediating this relationship, accounting for approximately one-third of the overall influence. Moreover, the decomposition analysis revealed that both emotional and economic support from adult children equally contributed to the declination of depression among rural middle-aged and older adults. Conclusion: Grandparenting significantly enhances mental well-being in rural middle-aged and older adults, with the support from adult children serving as a vital pathway for this positive impact. Both economic and emotional assistance from children hold equal importance in this dynamic. It underscores the necessity of fortifying the family support system to amplify the support provided by children, which in turn could significantly enhance the mental health of rural middle-aged and older adults.

Ntsr1 contributes to pulmonary hypertension by enhancing endoplasmic reticulum stress via JAK2-STAT3-Thbs1 signaling.

Pulmonary hypertension (PH) is a severe clinical syndrome with pulmonary vascular remodeling and poor long-term prognosis. Neurotensin receptor 1 (Ntsr1), serve as one of the G protein-coupled receptors (GPCRs), implicates in various biological processes, but the potential effects of Ntsr1 in PH development are unclear. The Sugen/Hypoxia (SuHx) or monocrotaline (MCT) induced rat PH model was used in our study and the PH rats showed aggravated pulmonary artery remodeling and increased right ventricular systolic pressure (RVSP). Our results revealed that Ntsr1 induced endoplasmic reticulum (ER) stress response via ATF6 activation contributed to the development of PH. Moreover, RNA-sequencing (RNA-seq) and phosphoproteomics were performed and the Ntsr1-JAK2-STAT3-thrombospondin 1 (Thbs1)-ATF6 signaling was distinguished as the key pathway. In vitro, pulmonary artery smooth muscle cells (PASMCs) under hypoxia condition showed enhanced proliferation and migration properties, which could be inhibited by Ntsr1 knockdown, JAK2 inhibitor (Fedratinib) treatment, STAT3 inhibitior (Stattic) treatment, Thbs1 knockdown or ATF6 knockdown. In addition, adeno-associated virus 1 (AAV1) were used to knockdown the expression of Ntsr1, Thbs1 or ATF6 in rats and reversed the phenotype of PH. In summary, our results reveal that Ntsr1-JAK2-STAT3-Thbs1 pathway can induce enhanced ER stress via ATF6 activation and increased PASMC proliferation and migration capacities, which can be mechanism of the pulmonary artery remodeling and PH. Targeting Ntsr1 might be a novel therapeutic strategy to ameliorate PH.

Zbtb16 increases susceptibility of atrial fibrillation in type 2 diabetic mice via Txnip-Trx2 signaling.

Atrial fibrillation (AF) is the most prevalent sustained cardiac arrhythmia, and recent epidemiological studies suggested type 2 diabetes mellitus (T2DM) is an independent risk factor for the development of AF. Zinc finger and BTB (broad-complex, tram-track and bric-a-brac) domain containing 16 (Zbtb16) serve as transcriptional factors to regulate many biological processes. However, the potential effects of Zbtb16 in AF under T2DM condition remain unclear. Here, we reported that db/db mice displayed higher AF vulnerability and Zbtb16 was identified as the most significantly enriched gene by RNA sequencing (RNA-seq) analysis in atrium. In addition, thioredoxin interacting protein (Txnip) was distinguished as the key downstream gene of Zbtb16 by Cleavage Under Targets and Tagmentation (CUT&Tag) assay. Mechanistically, increased Txnip combined with thioredoxin 2 (Trx2) in mitochondrion induced excess reactive oxygen species (ROS) release, calcium/calmodulin-dependent protein kinase II (CaMKII) overactivation, and spontaneous Ca2+ waves (SCWs) occurrence, which could be inhibited through atrial-specific knockdown (KD) of Zbtb16 or Txnip by adeno-associated virus 9 (AAV9) or Mito-TEMPO treatment. High glucose (HG)-treated HL-1 cells were used to mimic the setting of diabetic in vitro. Zbtb16-Txnip-Trx2 signaling-induced excess ROS release and CaMKII activation were also verified in HL-1 cells under HG condition. Furthermore, atrial-specific Zbtb16 or Txnip-KD reduced incidence and duration of AF in db/db mice. Altogether, we demonstrated that interrupting Zbtb16-Txnip-Trx2 signaling in atrium could decrease AF susceptibility via reducing ROS release and CaMKII activation in the setting of T2DM.

Isolation and molecular characteristics of a recombinant feline calicivirus from Qingdao, China.

Feline calicivirus (FCV) is a highly contagious pathogen seriously affecting the upper respiratory tract and producing oral diseases in the feline. Despite widespread vaccination, the prevalence of FCV remains high. In this study, the FCV qingdao (qd)/2019/china was isolated from a domestic feline oropharyngeal swab collected from Qingdao, China. The virus was purified using the plaque assay and identified using the Polymerase chain reaction and indirect immunofluorescence assay methods, the capsid amino acid, VP1 of qd/2019/china, showed sequence identity with the other isolates ranging between 83.90% (ym3/2001/jp) and 91.10% (CH-JL4). The sequence of the capsid amino acid revealed qd/2019/china to be closely related to CH-JL4 and clustered with CH-JL4 in the phylogenetic tree. The phylo-genetic analysis indicated that the complete genomes (GenBank® accession No. MZ322896) of qd/2019/china and CH-JL4 were also classified into the same cluster. The recombination analysis with Simplot indicated that the qd/2019/china originated from the recombination of CH-JL4 and HRB-SS, and the region 3,821 - 5,301 nt originated from HRB-SS. Further, the region 3,821 - 5,301 nt were found to belong to the protease-polymerase (PP) of HRB-SS. Here, we isolated a new recombinant virus, FCV qd/2019/china. Therefore, these results would be beneficial for better understanding of the evolution and epidemiology of FCV.

Fecal and oral microbiome analysis of snakes from China reveals a novel natural emerging disease reservoir.

Introduction: The gastrointestinal tract and oral cavity of animal species harbor complex microbial communities, the composition of which is indicative of the behavior, co-evolution, diet, and immune system of the host. Methods: This study investigated the microbial composition in snakes from varying altitudinal ranges by assessing the fecal and oral bacterial communities in Protobothrops mucrosquamatus, Elaphe dione, and Gloydius angusticeps from Sichuan Province, China, using metagenomic sequencing. Results and discussion: It was revealed that Bacteroidetes, Proteobacteria, Firmicutes, and Fusobacteria were the core microbial phyla in fecal samples across all three species, while Proteobacteria, Bacteroidetes, Actinobacteria, and Firmicutes were the core microbial phyla in oral samples across all three species. Notably, the dominance of Armatimonadetes was documented for the first time in the feces of all three species. Comparative analysis of the microbiomes of the three species indicated distinct microbiological profiles between snakes living at low- and high-altitude regions. Furthermore, 12 to 17 and 22 to 31 bacterial pathogens were detected in the oral and fecal samples, respectively, suggesting that snakes may serve as a novel reservoir for emerging diseases. Overall, this study provides a comparative analysis of the fecal and oral microbiomes in three snake species. Future investigations are anticipated to further elucidate the influence of age, genetics, behavior, diet, environment, ecology, and evolution on the gut and oral microbial communities of snakes.

Pseudorabies Virus Associations in Wild Animals: Review of Potential Reservoirs for Cross-Host Transmission.

Pseudorabies virus (PRV) has received widespread attention for its potential health effects on humans, wildlife, domestic animals, and livestock. In this review, we focus on PRV dynamics in wildlife, given the importance of wild-origin PRV transmission to domestic and farm animals. Wild boars, pigs, and raccoons can serve as reservoirs of PRV, with viral transmission to domestic livestock occurring via several routes, such as wild herd exposure, contaminated meat consumption, and insect vector transmission. Many endangered feline and canine species can be infected with PRV, with acute disease and death within 48 h. The first confirmed human case of PRV infection in mainland China was reported in 2017. Thus, PRV exhibits potentially dangerous cross-host transmission, which is likely associated with inappropriate vaccination, poor awareness, and insufficient biosecurity. Currently, no vaccine provides full protection against PRV in all animals. Here, we summarize the epidemiology and pathogenesis of PRV infection in wild, domestic, and farmed animals, which may facilitate the design of novel therapeutics and strategies for controlling PRV infection and improving wildlife protection in China.

Molecular Characterization and Phylogenetic Analysis of a Variant Recombinant Porcine Epidemic Diarrhea Virus Strain in China.

Since 2010, a variant of porcine epidemic diarrhea virus (PEDV) has re-emerged in several provinces of China, resulting in severe economic losses for the pork industry. Here, we isolated and identified a variant PEDV strain, SC-YB73, in Guangdong Province, China. The pathological observations of jejunum showed atrophy of villi and edema in the lamina propria. The sequence analysis of the viral genome identified a six-nucleotide insertion in the E gene, which has not previously been detected in PEDV strains. Furthermore, 50 nucleotide sites were unique in SC-YB73 compared with 27 other PEDV strains. The phylogenetic analysis based on the complete genome showed that SC-YB73 was clustered in variant subgroup GII-a, which is widely prevalent in the Chinese pig population. The recombination analysis suggested that SC-YB73 originated from the recombination of GDS47, US PEDV prototype-like strains TW/Yunlin550/2018, and COL/Cundinamarca/2014. In the present study, we isolated and genetically characterized a variant PEDV strain, thus providing essential information for the control of PED outbreaks in China.

Development and Characterization of Microsatellite Markers Based on the Chloroplast Genome of Tree Peony.

Tree peony (Paeonia suffruticosa Andr.) is a famous ornamental and medicinal flowering species. However, few high-efficiency chloroplast microsatellite markers have been developed for it to be employed in taxonomic identifications and evaluation of germplasm resources to date. In the present study, a total of 139 cpSSR loci were identified across eleven tree peony plastomes. Dinucleotide repeat SSRs (97.12%) were most abundantly repeated for the AT motif (58.27%), followed by the TA motif (30.94%) and the TC motif (7.91%). Twenty-one primer pairs were developed, and amplification tests were conducted for nine tree peony individuals. Furthermore, 19 cpSSR markers were amplified on 60 tree peony accessions by a capillary electrophoresis test. Of 19 cpSSR markers, 12 showed polymorphism with different alleles ranging from 1.333 to 3.000. The Shannon's information index and polymorphism information content values ranged from 0.038 to 0.887 (mean 0.432) and 0.032 to 0.589 (mean 0.268), respectively. The diversity levels for twelve loci ranged from 0.016 (at loci cpSSR-8 and cpSSR-26) to 0.543 (at locus cpSSR-15), averaging 0.268 for all loci. A total of 14 haplotypes (23.33%) were detected in the three populations. The haplotypic richness ranged from 0.949 to 1.751, with a mean of 1.233 per population. The genetic relationship suggested by the neighbor-joining-based dendrogram divided the genotypes into two clusters. The Jiangnan population was allotted to Cluster II, and the other two populations were distributed into both branches. These newly developed cpSSRs can be utilized for future breeding programs, population genetics investigations, unraveling the genetic relationships between related species, and germplasm management.

ECG Utilization Patterns of Patients With Arrhythmias During COVID-19 Epidemic and Post-SARS-CoV-2 Eras in Shanghai, China.

Background: The COVID-19 pandemic has led to concerns around its subsequent impact on global health. Objective: To investigate the health-seeking behavior, reflected by ECG utilization patterns, of patients with non-COVID-19 diseases during and after COVID-19 epidemic. Methods: Taking advantage of the remote ECG system covering 278 medical institutions throughout Shanghai, the numbers of medical visits with ECG examinations during the lockdown (between January 23 and April 7, 2020), post-lockdown (between April 8 and December 31, 2020) and post-SARS-CoV-2 (between January 23 and April 7, 2021) periods were analyzed and compared against those during the same periods of the preceding years (2018 and 2019). Results: Compared with the same period during pre-COVID years, the number of medical visits decreased during the lockdown (a 38% reduction), followed by a rebound post-lockdown (a 17% increase) and a fall to the baseline level in post-SARS-CoV-2 period. The number of new COVID-19 cases announced on a given day significantly correlated negatively with the numbers of medical visits during the following 7 days. Medical visit dynamics differed for various arrhythmias. Whereas medical visits for sinus bradycardia exhibited a typical decrease-rebound-fallback pattern, medical visits for atrial fibrillation did not fall during the lockdown but did exhibit a subsequent increase during the post-lockdown period. By comparison, the volume for ventricular tachycardia remained constant throughout this entire period. Conclusion: The ECG utilization patterns of patients with arrhythmias exhibited a decrease-rebound-fallback pattern following the COVID-19 lockdowns. Medical visits for diseases with more severe symptoms were less influenced by the lockdowns, showing a resilient demand for healthcare.

Dimethyl Fumarate Ameliorates Doxorubicin-Induced Cardiotoxicity By Activating the Nrf2 Pathway.

Doxorubicin (DOX) is limited in clinical application because of its cardiotoxicity. Oxidative stress and apoptosis are crucial in DOX-induced cardiac injury. Dimethyl fumarate (DMF) is an FDA-approved oral drug with powerful effects to reduce oxidative stress and apoptosis through the Nrf2 pathway. This study was aimed to determine whether DMF can protect against DOX-induced cardiac injury. We used both neonatal rat cardiomyocytes (NRCMs) in vitro and DOX-induced cardiac toxicity in vivo to explore the effects of DMF. The results showed that DMF significantly improved cell viability and morphology in NRCMs. In addition, DMF alleviated DOX-induced cardiac injury in rats, as evidenced by decreased CK-MB, LDH levels, improved survival rates, cardiac function, and pathological changes. Moreover, DMF significantly inhibited cardiac oxidative stress by reducing MDA levels and increasing GSH, SOD, and GSH-px levels. And DMF also inhibited DOX-induced cardiac apoptosis by modulating Bax, Bcl-2 and cleaved caspase-3 expression. Moreover, DMF exerted its protective effects against DOX by promoting Nrf2 nuclear translocation, which activated its downstream antioxidant gene Hmox1. Silencing of Nrf2 attenuated the protective effects of DMF in NRCMs as manifested by increased intracellular oxidative stress, elevated apoptosis levels, and decreased cell viability. In addition, DMF showed no protective effects on the viability of DOX-treated tumor cells, which suggested that DMF does not interfere with the antitumor effect of DOX in vitro. In conclusion, our data confirmed that DMF alleviated DOX-induced cardiotoxicity by regulating oxidative stress and apoptosis through the Nrf2 pathway. DMF may serve as a new candidate to alleviate DOX-related cardiotoxicity in the future.

Characterization of the bacterial microbiota in different gut and oral compartments of splendid japalure (Japalura sensu lato).

BACKGROUND: Gut and oral microbes form complex communities and play key roles in co-evolution with their hosts. However, little is understood about the bacterial community in lizards. RESULTS: In this study, we investigated the gut and oral bacterial communities in Japalura sensu lato from Sichuan Province, China, using 16S rRNA gene sequencing. Results showed that Bacteroidota (36.5%) and Firmicutes (32.8%) were the main phyla in the gut, while Proteobacteria, Bacteroidota, Firmicutes, and Actinobacteriota were the dominant phyla in the oral cavity. 16 S rRNA sequencing analysis of fecal samples showed that: (1) Bacteroidota was the most abundant in Japalura sensu lato, which was different from the bacterial community of insectivorous animals; (2) Bacteroidota, Firmicutes, Actinobacteriota, Fusobacteriota, and Cyanobacteria were the most abundant phylum in Japalura sensu lato. (3) Proteobacteria was the dominant phylum in Japalura sensu lato and other domestic insectivorous lizards (Shinisaurus crocodilurus, Phrynocephalus vlangalii, and Takydromus septentrionalis); (4) Comparing with the bacterial community of Shinisaurus crocodilurus, Phrynocephalus vlangalii, Takydromus septentrionalis, Liolaemus parvus, L. ruibali, and Phymaturus williamsi, Desulfobacterota was uniquely present in the gut of Japalura sensu lato. 16 S rRNA sequencing of oral samples showed that Chloroflexi and Deinococcota phyla were enriched in the oral cavity, which may have a significant influence on living in extreme environments. CONCLUSIONS: Thus, based on 16 S rRNA sequencing analysis of the community composition of the gut and oral microbiomes, this study firstly represents a foundation for understanding the gut and oral microbial ecology of Japalura sensu lato, and constitutes a detail account of the diversity of the microbiota inhabiting the gut and oral cavity of Japalura sensu lato. Further researches will continue to reveal how gut and oral microbial communities may be impacting the ecology and evolution of lizards.

Molecular characterization of the FCoV-like canine coronavirus HLJ-071 in China.

BACKGROUND: According to the differences of antigen and genetic composition, canine coronavirus (CCoV) consists of two genotypes, CCoV-I and CCoV-II. Since 2004, CCoVs with point mutations or deletions of NSPs are contributing to the changes in tropism and virulence in dogs. RESULTS: In this study, we isolated a CCoV, designated HLJ-071, from a dead 5-week-old female Welsh Corgi with severe diarrhea and vomit. Sequence analysis suggested that HLJ-071 bearing a complete ORF3abc compared with classic CCoV isolates (1-71, K378 and S378). In addition, a variable region was located between S gene and ORF 3a gene, in which a deletion with 104 nts for HLJ-071 when compared with classic CCoV strains 1-71, S378 and K378. Phylogenetic analysis based on the S gene and complete sequences showed that HLJ-071 was closely related to FCoV II. Recombination analysis suggested that HLJ-071 originated from the recombination of FCoV 79-1683, FCoV DF2 and CCoV A76. Finally, according to cell tropism experiments, it suggested that HLJ-071 could replicate in canine macrophages/monocytes cells. CONCLUSION: The present study involved the isolation and genetic characterization of a variant CCoV strain and spike protein and ORF3abc of CCoV might play a key role in viral tropism, which could affect the replication in monocyte/macrophage cells. It will provide essential information for further understanding the evolution in China.

Evaluating angiotensin-converting enzyme 2-mediated SARS-CoV-2 entry across species.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic represents a global threat, and the interaction between the virus and angiotensin-converting enzyme 2 (ACE2), the primary entry receptor for SARS-CoV-2, is a key determinant of the range of hosts that can be infected by the virus. However, the mechanisms underpinning ACE2-mediated viral entry across species remains unclear. Using infection assay, we evaluated SARS-CoV-2 entry mediated by ACE2 of 11 different animal species. We discovered that ACE2 of Rhinolophus sinicus (Chinese rufous horseshoe bat), Felis catus (domestic cat), Canis lupus familiaris (dog), Sus scrofa (wild pig), Capra hircus (goat), and Manis javanica (Malayan pangolin) facilitated SARS-CoV-2 entry into nonsusceptible cells. Moreover, ACE2 of the pangolin also mediated SARS-CoV-2 entry, adding credence to the hypothesis that SARS-CoV-2 may have originated from pangolins. However, the ACE2 proteins of Rhinolophus ferrumequinum (greater horseshoe bat), Gallus gallus (red junglefowl), Notechis scutatus (mainland tiger snake), or Mus musculus (house mouse) did not facilitate SARS-CoV-2 entry. In addition, a natural isoform of the ACE2 protein of Macaca mulatta (rhesus monkey) with the Y217N mutation was resistant to SARS-CoV-2 infection, highlighting the possible impact of this ACE2 mutation on SARS-CoV-2 studies in rhesus monkeys. We further demonstrated that the Y217 residue of ACE2 is a critical determinant for the ability of ACE2 to mediate SARS-CoV-2 entry. Overall, these results clarify that SARS-CoV-2 can use the ACE2 receptors of multiple animal species and show that tracking the natural reservoirs and intermediate hosts of SARS-CoV-2 is complex.

Molecular Characterization of a Novel Budgerigar Fledgling Disease Virus Strain From Budgerigars in China.

Budgerigar fledgling disease virus (BFDV) is the causative polyomavirus of budgerigar fledgling disease, an important avian immunosuppressive disease in budgerigars (Melopsittacus undulatus). In the current study, we explored the etiological role and molecular characteristics of BFDV. We identified a novel BFDV strain, designated as SC-YB19, belonging to a unique cluster with three other domestic strains (WF-GM01, SD18, and APV-P) and closely related to Polish isolates based on complete sequences. Sequence analysis showed that SC-YB19 had an 18-nucleotide (nt) deletion in the enhancer region, corresponding to the sequence position 164-181 nt, which differed significantly from all other BFDV strains. Based on sequence alignment, three unique nucleotide substitutions were found in VP4 (position 821), VP1 (position 2,383), and T-antigen (position 3,517) of SC-YB19, compared with SD18, WF-GM01, QDJM01, HBYM02, APV7, and BFDV1. Phylogenetic analyses based on complete sequences suggested that SC-YB19, along with the domestic WF-GM01, SD18, and APV-P strains, formed a single branch and were closely related to Polish, Japanese, and American isolates. These results demonstrate that BFDV genotype variations are co-circulating in China, thus providing important insight into BFDV evolution.

Development of a recombinase polymerase amplification fluorescence assay to detect feline coronavirus.

Feline coronavirus (FCoV) is classified into two pathotypes: the avirulent feline enteric coronavirus (FECV), and the virulent feline infectious peritonitis virus (FIPV). Rapid pathogen detection, which is efficient and convenient, is the best approach for early confirmatory diagnosis. In this study, we first developed and evaluated a rapid recombinase polymerase amplification (RPA) detection method for FCoV that can detect FCoV within 15 min at 39 °C. The detection limit of that assay was 233 copies/µL DNA molecules per reaction. The specificity was high: it did not cross-react with canine distemper virus (CDV), canine coronavirus (CCoV), canine adenovirus (CAV), feline calicivirus (FCV), feline herpesvirus (FHV), or feline parvovirus (FPV). This assay was evaluated using 42 clinical samples (30 diarrhea samples and 12 ascites samples). The coincidence rate between FCoV-RPA and RT-qPCR for detection in clinical samples was 95.2%. In summary, FCoV-RPA analysis provides an efficient, rapid, and sensitive detection method for FCoV.

Exosomal LINC01005 derived from oxidized low-density lipoprotein-treated endothelial cells regulates vascular smooth muscle cell phenotypic switch.

Phenotype switch of vascular smooth muscle cells (VSMCs) plays an important role in the development of atherosclerosis (AS). Endothelial cells can regulate VSMC phenotypic switch by secreting exosomes, crucial mediators of intracellular communication. This study aimed to determine whether exosomal LINC01005 from oxidized low-density lipoprotein (ox-LDL)-treated human umbilical vein endothelial cells (HUVECs) plays a role in regulating VSMC phenotypic switch and to validate the underlying molecular mechanism. Exosomes were extracted from ox-LDL-treated HUVECs (ox-LDL-Exo) and then administered into VSMCs. VSMC phenotypic switch was assessed by determining VSMC phenotypic markers using western blot. VSMC cell proliferation and migration were evaluated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and wound healing assay, respectively. The interaction between miR-128-3p and LINC01005 or Krüppel-like factor 4 (KLF4) was analyzed by luciferase reporter assay. ox-LDL-Exo contained high expression of LINC01005. Inhibition of LINC01005 expression in ox-LDL-Exo abrogated the ox-LDL-Exo-induced VSMC phenotypic switch, proliferation, and migration. Furthermore, LINC01005 acted as a sponge of miR-128-3p to upregulate KLF4 expression. Moreover, miR-128-3p overexpression and KLF4 silencing in VSMCs attenuated the ox-LDL-Exo-induced VSMC phenotypic switch, proliferation, and migration. Collectively, exosomal LINC01005 from ox-LDL-treated HUVECs promotes VSMC phenotype switch, proliferation, and migration by regulating the miR-128-3p/KLF4 axis.

Molecular characterization of HLJ-073, a recombinant canine coronavirus strain from China with an ORF3abc deletion.

Canine enteric coronaviruses (CCoVs) are important enteric pathogens of dogs. CCoVs with different variations are typically pantropic and pathogenic in dogs. In this study, we isolated a CCoV, designated HLJ-073, from a dead 6-week-old male Pekingese with gross lesions and diarrhea. Interestingly, sequence analysis suggested that HLJ-073 contained a 350-nt deletion in ORF3abc compared with reference CCoV isolates, resulting in the loss of portions of ORF3a and ORF3c and the complete loss of ORF3b. Phylogenetic analysis based on the S gene showed that HLJ-073 was more closely related to members of the FCoV II cluster than to members of the CCoV I or CCoV II cluster. Furthermore, recombination analysis suggested that HLJ-073 originated from the recombination of FCoV 79-1683 and CCoV A76, which were both isolated in the United States. Cell tropism experiments suggested that HLJ-073 could effectively replicate in canine macrophages/monocytes and human THP-1 cells. This is the first report of the isolation of strain HLJ-073 in China, and this virus has biological characteristics that are different from those of other reported CCoVs.

One-step triplex PCR/RT-PCR to detect canine distemper virus, canine parvovirus and canine kobuvirus.

To rapidly distinguish Canine distemper virus (CDV), canine parvovirus (CPV), and canine kobuvirus (CaKoV) in practice, a one-step multiplex PCR/RT-PCR assay was developed, with detection limits of 102.1 TCID50 for CDV, 101.9 TCID50 for CPV and 103 copies for CaKoV. This method did not amplify nonspecific DNA or RNA from other canine viruses. Therefore, the assay provides a sensitive tool for the rapid clinical detection and epidemiological surveillance of CDV, CPV and CaKoV in dogs.

Feline Infectious Peritonitis Virus Nsp5 Inhibits Type I Interferon Production by Cleaving NEMO at Multiple Sites.

Feline infectious peritonitis (FIP), caused by virulent feline coronavirus, is the leading infectious cause of death in cats. The type I interferon (type I IFN)-mediated immune responses provide host protection from infectious diseases. Several coronaviruses have been reported to evolve diverse strategies to evade host IFN response. However, whether feline infectious peritonitis virus (FIPV) antagonizes the type I IFN signaling remains unclear. In this study, we demonstrated that FIPV strain DF2 infection not only failed to induce interferon-ß (IFN-ß) and interferon-stimulated gene (ISG) production, but also inhibited Sendai virus (SEV) or polyinosinic-polycytidylic acid (poly(I:C))-induced IFN-ß production. Subsequently, we found that one of the non-structural proteins encoded by the FIPV genome, nsp5, interrupted type I IFN signaling in a protease-dependent manner by cleaving the nuclear factor κB (NF-κB) essential modulator (NEMO) at three sites-glutamine132 (Q132), Q205, and Q231. Further investigation revealed that the cleavage products of NEMO lost the ability to activate the IFN-ß promoter. Mechanistically, the nsp5-mediated NEMO cleavage disrupted the recruitment of the TRAF family member-associated NF-κB activator (TANK) to NEMO, which reduced the phosphorylation of interferon regulatory factor 3 (IRF3), leading to the inhibition of type I IFN production. Our research provides new insights into the mechanism for FIPV to counteract host innate immune response.

Identification of Feline Interferon Regulatory Factor 1 as an Efficient Antiviral Factor against the Replication of Feline Calicivirus and Other Feline Viruses.

Interferons (IFNs) can inhibit most, if not all, viral infections by eliciting the transcription of hundreds of interferon-stimulated genes (ISGs). Feline calicivirus (FCV) is a highly contagious pathogen of cats and a surrogate for Norwalk virus. Interferon efficiently inhibits the replication of FCV, but the mechanism of the antiviral activity is poorly understood. Here, we evaluated the anti-FCV activity of ten ISGs, whose antiviral activities were previously reported. The results showed that interferon regulatory factor 1 (IRF1) can significantly inhibit the replication of FCV, whereas the other ISGs tested in this study failed. Further, we found that IRF1 was localized in the nucleus and efficiently activated IFN-ß and the ISRE promoter. IRF1 can trigger the production of endogenous interferon and the expression of ISGs, suggesting that IRF1 can positively regulate IFN signalling. Importantly, the mRNA and protein levels of IRF1 were reduced upon FCV infection, which may be a new strategy for FCV to evade the innate immune system. Finally, the antiviral activity of IRF1 against feline panleukopenia virus, feline herpesvirus, and feline infectious peritonitis virus was demonstrated. These data indicate that feline IRF1 plays an important role in regulating the host type I IFN response and inhibiting feline viral infections.