African Swine Fever Virus MGF505-7R Interacts with Interferon Regulatory Factor 9 to Evade the Type I Interferon Signaling Pathway and Promote Viral Replication.

African swine fever (ASF) is an acute and severe infectious disease caused by the ASF virus (ASFV). The mortality rate of ASF in pigs can reach 100%, causing huge economic losses to the pig industry. Here, we found that ASFV protein MGF505-7R inhibited the beta interferon (IFN-ß)-mediated Janus-activated kinase-signal transducer and activation of transcription (JAK-STAT) signaling. Our results demonstrate that MGF505-7R inhibited interferon-stimulated gene factor 3 (ISGF3)-mediated IFN-stimulated response element (ISRE) promoter activity. Importantly, we observed that MGF505-7R inhibits ISGF3 heterotrimer formation by interacting with interferon regulatory factor 9 (IRF9) and inhibits the nuclear translocation of ISGF3. Moreover, to demonstrate the role of MGF505-7R in IFN-I signal transduction during ASFV infection, we constructed and evaluated ASFV-ΔMGF505-7R recombinant viruses. ASFV-ΔMGF505-7R restored STAT2 and STAT1 phosphorylation, alleviated the inhibition of ISGF3 nuclear translocation, and showed increased susceptibility to IFN-ß, unlike the parental GZ201801 strain. In conclusion, our study shows that ASFV protein MGF505-7R plays a key role in evading IFN-I-mediated innate immunity, revealing a new mode of evasion for ASFV. IMPORTANCE ASF, caused by ASFV, is currently prevalent in Eurasia, with mortality rates reaching 100% in pigs. At present, there are no safe or effective vaccines against ASFV. In this study, we found that the ASFV protein MGF505-7R hinders IFN-ß signaling by interacting with IRF9 and inhibiting the formation of ISGF3 heterotrimers. Of note, we demonstrated that MGF505-7R plays a role in the immune evasion of ASFV in infected hosts and that recombinant viruses alleviated the effect on type I IFN (IFN-I) signaling and exhibited increased susceptibility to IFN-ß. This study provides a theoretical basis for developing vaccines against ASFV using strains with MGF505-7R gene deletions.

A Triplex PCR Method for Distinguishing the Wild-Type African Swine Fever Virus From the Deletion Strains by Detecting the Gene Insertion.

To date, there is no effective vaccine or antiviral therapy available to prevent or treat African swine fever virus (ASFV) infections. ASFV gene deletion strains have been proposed as promising anti-ASFV vaccine candidates. In recent years, most ASFV gene deletion strains worldwide have been recombinant strains expressing EGFP or mCherry as markers. Therefore, in this study, a new triplex real-time PCR (RT-PCR) method was established for the broad and accurate differentiation of ASFV wild-type vs. gene deletion strains. We designed three pairs of primers and probes to target B646L, EGFP, and mCherry, and RT-PCR was used to detect these three genes simultaneously. The detection method prevented non-specific amplification of porcine reproductive and respiratory syndrome virus, porcine epidemic diarrhea virus, circovirus type 2, pseudorabies virus, and classical swine fever virus genes. The minimum copy number of standard plasmid DNA detected using triplex RT-PCR was 9.49, 15.60, and 9.60 copies for B646L, EGFP, and mCherry, respectively. Importantly, of the 1646 samples analyzed in this study, 67 were positive for ASFV, all corresponding to the wild-type virus. Overall, our data show that the triplex RT-PCR method established in this study can specifically identify both ASFV wild-type and gene deletion strains.

Long-Term Application of Bio-Compost Increased Soil Microbial Community Diversity and Altered Its Composition and Network.

The influence of bio-compost on the diversity, composition and structure of soil microbial communities is less understood. Here, Illumina MiSeq sequencing and a network analysis were used to comprehensively characterize the effects of 25 years of bio-compost application on the microbial diversity of soil and community composition. High dosages of bio-compost significantly increased the bacterial and fungal richness. The compositions of bacterial and fungal communities were significantly altered by bio-compost addition. Bio-compost addition enriched the relative abundance of beneficial microorganisms (such as Sphingomonas, Acidibacter, Nocardioides, etc.) and reduced the relative abundance of harmful microorganisms (such as Stachybotrys and Aspergillus). Electrical conductivity, soil organic matter and total phosphorus were the key factors in shaping soil microbial community composition. The bacterial network was more complex than fungal network, and bacteria were more sensitive to changes in environmental factors than fungi. Positive interactions dominated both the bacterial and fungal networks, with stronger positive interactions found in the bacterial network. Functional prediction suggested that bio-composts altered the soil bacterial-community metabolic function with respect to carbon, nitrogen and phosphorus cycles and fungal community trophic modes. In conclusion, suitable bio-compost addition is beneficial to the improvement of soil health and crop quality and therefore the sustainability of agriculture.

Molecular Detection, Multilocus Genotyping, and Population Genetics of Enterocytozoon bieneusi in Pigs in Southeastern China.

Enterocytozoon bieneusi is an important opportunistic pathogen widely distributed in humans and animals that causes diarrhea or fatal diarrhea in immunocompromised hosts. To examine the infection status and molecular characteristics of E. bieneusi in pigs, 725 fecal samples were collected from pigs in six areas of Fujian Province. The E. bieneusi genotypes were identified based on the internal transcribed spacer (ITS) regions of the ribosomal RNA (rRNA) gene by nested PCR, and its population genetics were analyzed by multilocus sequence typing (MLST). The results showed that the infection rate of E. bieneusi was 24.4% (177/725), and 11 known genotypes (EbpC, EbpA, CHN-RR2, KIN-1, CHG7, CHS5, CM11, CHG23, G, PigEBITS, and D) and 2 novel genotypes (FJF and FJS) were identified. All the genotypes were found to be clustered into zoonotic Group 1. Moreover, 52 positive samples were successfully amplified at minisatellite and microsatellite loci and formed 48 distinct multilocus genotypes (MLGs). Further population structure analyses showed strong genetic linkage disequilibrium (LD) and several recombination events (Rm), indicating that E. bieneusi has a clonal population structure. This study is the first to investigate the prevalence and molecular characteristics of E. bieneusi in Fujian Province and could provide baseline data to control E. bieneusi infection in pigs and humans and deepen our understanding of the zoonotic risk of E. bieneusi and its distribution in China.

Prevalence and Genetic Identification of Three Entamoeba Species in Pigs in Southeastern China.

Parasitic Entamoeba spp. can infect many classes of vertebrates including humans and pigs. Entamoeba suis and zoonotic Entamoeba polecki have been identified in pigs, and swine are implicated as potential reservoirs for Entamoeba histolytica. However, the prevalence of Entamoeba spp. in pigs in southeastern China has not been reported. In this study, 668 fecal samples collected from 6 different regions in Fujian Province, southeastern China, were analyzed to identify three Entamoeba species by nested PCR and sequencing analysis. The overall prevalence of Entamoeba spp. was 55.4% (370/668; 95% CI 51.6% to 59.2%), and the infection rate of E. polecki ST1 was the highest (302/668; 45.2%, 95% CI 41.4% to 49.0%), followed by E. polecki ST3 (228/668; 34.1%, 95% CI 30.5% to 37.7%) and E. suis (87/668; 13.0%, 95% CI 10.5% to 15.6%). E. histolytica was not detected in any samples. Moreover, the coinfection rate of E. polecki ST1 and ST3 was 25.1% (168/668; 95% CI 21.9% to 28.4%), the coinfection rate of E. polecki ST1 and E. suis was 3.7% (25/668; 95% CI 2.3% to 5.2%), the coinfection rate of E. polecki ST3 and E. suis was 0.3% (2/668), and the coinfection rate of E. polecki ST1, E. polecki ST3, and E. suis was 4.0% (27/668; 95% CI 2.5% to 5.5%). A representative sequence (MK347346) was identical to the sequence of E. suis (DQ286372). Two subtype-specific sequences (MK357717 and MK347347) were almost identical to the sequences of E. polecki ST1 (FR686383) and ST3 (AJ566411), respectively. This is the first study to survey the occurrence and to conduct molecular identification of three Entamoeba species in southeastern China. This is the first report regarding mixed infections with E. suis, E. polecki ST1, and E. polecki ST3 in China. More research studies are needed to better understand the transmission and zoonotic potential of Entamoeba spp.

[Effects of Rutin on Myocardial Enzyme and Cardiac Morphology in Diabetic Mice].

OBJECTIVE: To study the effects of rutin on myocardial enzyme in serum and myocardial morphology in diabetic mice induced by streptozotocin (STZ). METHODS: A model of type 1 diabetic mice was established in 48 male kunming mice with daily intraperitoneal injection of streptozotocin (STZ) 62.5 mg/kg for 5 consecutive days. 12 male Kunming mice were selected as normal group randomly. The established successfully diabetic mice were randomly divided into the model group, irbesartan group [45 mg/ (kg?d)], low-, high-dose rutin groups [50, 100 mg/ (kg?d)]. The mice in the normal and the model groups were given sodium carboxymethyl cellulose solution (CMC-Na, 1 g/L) by intragastric administration respectively. After administration for 8 weeks, the levels of creatine kinase (CK), creatine kinase isoenzymes (CK-MB), hydroxybutyrate dehydrogenase (HBDH), and lactate dehydrogenase (LDH) in serum were detected by blood biochemical analyzer. The cardiac myocardial morphology was observed by HE staining, Masson trichrome staining and electron microscope. RESULTS: Compare to the normal group, the levels of the myocardial enzyme in serum evidently increased in the model group (P<0.01); Compare to the model group, the levels of the myocardial enzyme in serum decreased in low-, high-dose rutin groups; The levels of HBDH and LDH declined remarkably in the high-dose rutin group relative to the low-dose group (P<0.05); Compared to the high-dose group, the level of LDH increased in the irbesartan group (P<0.05).Moreover, relative to the model group, the morphology of myocardial tissue, the degree of fibrosis and the ultrastructure of myocardial tissue in mice were significantly improved in low-, high-dose rutin groups, and the effects were more significant in the high-dose rutin group. CONCLUSION: Rutin could decrease the levels of myocardial enzyme in serum in diabetic mice, improve the cardiac cell morphology and alleviate myocardial injury.