PoRVA G9P[23] and G5P[7] infections differentially promote PEDV replication by reprogramming glutamine metabolism.

PoRVA and PEDV coinfections are extremely common in clinical practice. Although coinfections of PoRVA and PEDV are known to result in increased mortality, the underlying mechanism remains unknown. Here, we found that PoRVA infection promoted PEDV infection in vivo and in vitro and that PoRVA G9P[23] (RVA-HNNY strain) enhanced PEDV replication more significantly than did PoRVA G5P[7] (RVA-SXXA strain). Metabolomic analysis revealed that RVA-HNNY more efficiently induced an increase in the intracellular glutamine content in porcine small intestinal epithelial cells than did RVA-SXXA, which more markedly promoted ATP production to facilitate PEDV replication, whereas glutamine deprivation abrogated the effect of PoRVA infection on promoting PEDV replication. Further studies showed that PoRVA infection promoted glutamine uptake by upregulating the expression of the glutamine transporter protein SLC1A5. In SLC1A5 knockout cells, PoRVA infection neither elevated intracellular glutamine nor promoted PEDV replication. During PoRVA infection, the activity and protein expression levels of glutamine catabolism-related enzymes (GLS1 and GLUD1) were also significantly increased promoting ATP production through glutamine anaplerosis into the TCA cycle. Consistent with that, siRNAs or inhibitors of GLS1 and GLUD1 significantly inhibited the promotion of PEDV replication by PoRVA. Notably, RVA-HNNY infection more markedly promoted SLC1A5, GLS1 and GLUD1 expression to more significantly increase the uptake and catabolism of glutamine than RVA-SXXA infection. Collectively, our findings illuminate a novel mechanism by which PoRVA infection promotes PEDV infection and reveal that the modulation of glutamine uptake is key for the different efficiencies of PoRVA G9P[23] and PoRVA G5P[7] in promoting PEDV replication.

IL-10 upregulates SOCS3 to inhibit type I interferon signaling to promote PoRVA replication in intestinal epithelial cells.

Porcine group A rotavirus (PoRVA) is one of the common enteric viruses causing severe diarrhea in piglets. Although PoRVA infection has been identified to promote IL-10 production, the role of IL-10 during viral infection remains unclear. In this study, we found that elevated IL-10 levels during PoRVA infection promote viral replication by inhibiting type I interferon production and response. IL-10 treatment upregulated the expression of SOCS3 in PoRVA-infected IPEC-J2 cells, which inhibited IFN-I production by preventing the degradation of IκB and nuclear translocation of NF-κB, thereby significantly promoting PoRVA replication. Furthermore, we determined that SOCS3 also inhibited type Ⅰ interferon signaling pathway, which led to a significantly reduced ISGs after IFN-α stimulation. In PoRVA-infected cells, overexpression of SOCS3 significantly inhibits phosphorylation and heterodimerization of STAT1, thereby promoting viral replication. Finally, we demonstrated the effect of IL-10 on PoRVA replication in vivo by murine models of PoRVA infection. PoRVA replication levels were lower in the ileum of IL-10 knockout (IL-10-/-) mice than that in PoRVA-infected wild-type mice, but PoRVA replication levels were higher in the ileum of IFNAR knockout (IFNAR-/-) mice than that in PoRVA-infected wild-type mice. Taken together, our findings provide information to understand the strategies of PoRVA to evade host innate antiviral immunity.

Transgenerational Inheritance Effects of Copper Oxide Nanoparticles (CuONPs) Induced Asthenospermia and Infertility via Gamete H3K9me3 Insufficiency Pathway in Mice.

The widespread use of colloidal copper oxide nanoparticles (CuONPs) poses substantial health risks to humans. CuONPs can penetrate the blood-testis barrier and induce spermatocide, and the understanding of the adverse effects of asthenospermia on spermatogenesis, embryonic development, and transgenerational inheritance is limited. In this study, male mice were orally administered different doses of CuONPs via continuous exposure for one spermatozoon development period (35 days) and then exposed without CuONPs for another 35 days. The CuONPs that accumulated in the testes induced oxidative stress (OS), affected the progress of spermatogenesis and sperm capacitation, and compromised epigenetic modifications, resulting in asthenospermia and embryonic development anomalies in male offspring. In a mechanism, CuONP exposure impaired the self-renewal and differentiation of spermatogonial stem cells (SSCs) via the GDNF/PI3K/AKT signaling pathway under OS. Importantly, CuONP exposure was found to potentially lower H3K9me3 levels in paternal sperm, which would further transgenerational transmission and interfere with sperm mitochondrial energy metabolism and motility, leading to asthenospermia and subfertility in the offspring. Collectively, these data reveal a molecular mechanism by which CuONP exposure disturbs H3K9me3 levels via the OS pathway, which further mediates the asthenospermic effects of reproductive failure by interfering with mitochondrial arrangement and formation in the next generation.

Assessing the breeding phenology of a threatened frog species using eDNA and automatic acoustic monitoring.

Background: Climate change has driven shifts in breeding phenology of many amphibians, causing phenological mismatches (e.g., predator-prey interactions), and potentially population declines. Collecting data with high spatiotemporal sensitivity on hibernation emergence and breeding times can inform conservation best practices. However, monitoring the phenology of amphibians can be challenging because of their cryptic nature over much of their life cycle. Moreover, most salamanders and caecilians do not produce conspicuous breeding calls like frogs and toads do, presenting additional monitoring challenges. Methods: In this study, we designed and evaluated the performance of an environmental DNA (eDNA) droplet digital PCR (ddPCR) assay as a non-invasive tool to assess the breeding phenology of a Western Chorus Frog population (Pseudacris maculata mitotype) in Eastern Ontario and compared eDNA detection patterns to hourly automatic acoustic monitoring. For two eDNA samples with strong PCR inhibition, we tested three methods to diminish the effect of inhibitors: diluting eDNA samples, adding bovine serum albumin to PCR reactions, and purifying eDNA using a commercial clean-up kit. Results: We recorded the first male calling when the focal marsh was still largely frozen. Chorus frog eDNA was detected on April 6th, 6 days after acoustic monitoring revealed this first calling male, but only 2 days after males attained higher chorus activity. eDNA signals were detected at more sampling locales within the marsh and eDNA concentrations increased as more males participated in the chorus, suggesting that eDNA may be a reasonable proxy for calling assemblage size. Internal positive control revealed strong inhibition in some samples, limiting detection probability and quantification accuracy in ddPCR. We found diluting samples was the most effective in reducing inhibition and improving eDNA quantification. Conclusions: Altogether, our results showed that eDNA ddPCR signals lagged behind male chorusing by a few days; thus, acoustic monitoring is preferable if the desire is to document the onset of male chorusing. However, eDNA may be an effective, non-invasive monitoring tool for amphibians that do not call and may provide a useful complement to automated acoustic recording. We found inhibition patterns were heterogeneous across time and space and we demonstrate that an internal positive control should always be included to assess inhibition for eDNA ddPCR signal interpretations.

The S protein of a novel recombinant PEDV strain promotes the infectivity and pathogenicity of PEDV in mid-west China.

Porcine epidemic diarrhoea virus (PEDV) is an emerging and re-emerging swine enterovirus that causes highly contagious diarrhoea and mortality in piglets. To better understand the current prevalence of PEDV in mid-west China, and to find out the reason for the re-emergence of PEDV from the viral genomic characteristics. Herein, we firstly investigated epidemiology of PEDV in mid-west China from 2019 to 2020. A total of 62.23% (257/413) of diarrhoea samples were positive for PEDV, and the PEDV-positive cases were mainly detected in winter. Then, we selected the SXSL strain as a representative strain to study the genetic and pathogenic characterization of PEDV pandemic strains in mid-west China. The recombination analysis showed that SXSL strain was a recombinant strain, and the major and minor parent strains of the recombination are CH/SCZJ/2018 strain and GDS48 strain, respectively. Complete genome sequencing and homology analysis showed that the S protein of SXSL strain contained multiple amino acid indels and mutations compared to the PEDV representative strains. Furthermore, we evaluated the effect of S protein on the infectivity and pathogenicity of PEDV by the PEDV reverse genetics system, and results showed that SXSL S protein increased the infectivity and pathogenicity of chimeric virus. Overall, our findings provided important information for understanding the roles of S protein in the prevalence of PEDV in mid-west China and developing vaccines based on PEDV pandemic strains.

Depth-Dependent Spatiotemporal Dynamics of Overwintering Pelagic Microcystis in a Temperate Water Body.

Cyanobacteria in the genus Microcystis are dominant components of many harmful algal blooms worldwide. Their pelagic-benthic life cycle helps them survive periods of adverse conditions and contributes greatly to their ecological success. Many studies on Microcystis overwintering have focused on benthic colonies and suggest that sediment serves as the major inoculum for subsequent summer blooms. However, the contemporaneous overwintering pelagic population may be important as well but is understudied. In this study, we investigated near-surface and near-bottom pelagic population dynamics of both microcystin-producing Microcystis and total Microcystis over six weeks in winter at Dog Lake (South Frontenac, ON, Canada). We quantified relative Microcystis concentrations using real-time PCR. Our results showed that the spatiotemporal distribution of overwintering pelagic Microcystis was depth dependent. The abundance of near-bottom pelagic Microcystis declined with increased depth with no influence of depth on near-surface Microcystis abundance. In the shallow region of the lake (<10 m), most pelagic Microcystis was found near the lake bottom (>90%). However, the proportion of near-surface Microcystis rose sharply to over 60% as the depth increased to approximately 18 m. The depth-dependent distribution pattern was found to be similar in both microcystin-producing Microcystis and total Microcystis. Our results suggest the top of the water column may be a more significant contributor of Microcystis recruitment inoculum than previously thought and merits more attention in early CHAB characterization and remediation.