The W195 Residue of the Newcastle Disease Virus V Protein Is Critical for Multiple Aspects of Viral Self-Regulation through Interactions between V and Nucleoproteins.

The transcription and replication of the Newcastle disease virus (NDV) strictly rely on the viral ribonucleoprotein (RNP) complex, which is composed of viral NP, P, L and RNA. However, it is not known whether other viral non-RNP proteins participate in this process for viral self-regulation. In this study, we used a minigenome (MG) system to identify the regulatory role of the viral non-RNP proteins V, M, W, F and HN. Among them, V significantly reduced MG-encoded reporter activity compared with the other proteins and inhibited the synthesis of viral mRNA and cRNA. Further, V interacted with NP. A mutation in residue W195 of V diminished V-NP interaction and inhibited inclusion body (IB) formation in NP-P-L-cotransfected cells. Furthermore, a reverse-genetics system for the highly virulent strain F48E9 was established. The mutant rF48E9-VW195R increased viral replication and apparently enhanced IB formation. In vivo experiments demonstrated that rF48E9-VW195R decreased virulence and retarded time of death. Overall, the results indicate that the V-NP interaction of the W195 mutant V decreased, which regulated viral RNA synthesis, IB formation, viral replication and pathogenicity. This study provides insight into the self-regulation of non-RNP proteins in paramyxoviruses.

Chicken ISG12(2) attenuates Newcastle disease virus and enhances the efficiency of Newcastle disease vaccine via activating immune pathways.

Low virulence and strong immunogenicity are quite important for Newcastle disease virus (NDV) producing Newcastle disease (ND) living-attenuated vaccine. However, immunogenicity of NDV positively correlates to its virulence. Usually, the velogenic NDV induces stronger immune responses of poultry than the lentogenic strain, but virulent NDV poses a risk for chicken. In this study, we identified the chicken interferon (IFN)-stimulated gene 12-2 (ISG12(2)) not only attenuated NDV, but also increased immunogenicity of ND vaccine strain. First, we found that NDV infection or IFNs stimulation induced expression of chicken ISG12(2) that reinforced expression of IFNs. Over-expression or knock-down proved that chicken ISG12(2) inhibited NDV replication. Then, recombinant NDV LaSota strains (rLaSota/Fmut/ISG12(2) and rLaSota/ISG12(2)), expressing ISG12(2), were rescued. Pathogenicity tests showed that ISG12(2) expression attenuated NDV. RNA-seq or RT-qPCR demonstrated that, comparing to rLaSota/Fmut and rLaSota, rLaSota/Fmut/ISG12(2) and rLaSota/ISG12(2) induced hosts to produce cytokines enriching in innate and adaptive immune pathways in vitro and in vivo. Finally, we showed that rLaSota/ISG12(2) vaccination improved immune condition of chicken to quickly respond NDV infection and then enhance protection. These results suggest that chicken ISG12(2) is a potential novel molecular adjuvant to regulate immune responses, which decrease virulence and increase immunogenicity of NDV. The chicken ISG12(2) may contribute to the development of high efficient poultry vaccine.

[Soil bacterial community characteristics and ecological function prediction of alfalfa and crop rotation systems in the Loess Plateau, Northwest China]. / 黄土高原苜蓿-粮食作物轮作下土壤细菌群落特征和生态功能预测.

In order to understand the effects of lucerne cropping rotation on the bacterial community of loess soil, a long-term field experiment was conducted in rain-fed agricultural area of Loess Plateau. The cropping systems included continuous lucerne (Medicago sativa, LC), lucerne removed and rotated with spring wheat (Triticum aestivum, LFW), lucerne removed and rotated with corn (Zea mays, LFC), lucerne removed and rotated with potato (Solanum tuberosum, LP), and lucerne removed and rotated with continuous millet (Panicum miliaceum, LM). Based on 16S rRNA high-throughput sequencing technology, we investigated soil bacterial community structure and diversity in different cropping systems, and predicted ecological function using PICRUSt method. The results showed that the dominant phyla of loess soil bacteria were Actinomycetes (20.3%-32.0%), Proteobacteria (19.2%-23.0%), Acidobacteria (12.4%-14.2%) and Chloroflexus (11.0%-12.7%). The dominant genus was Bacillus (1.9%) in lucerne-corn system and Pseudarthrobacter (2.5%) in other treatments. Rotation with annual crops decreased the relative abundance of Actinobacteria and increased that of Chloroflexi and Firmicutes. Redundancy analysis showed that the main soil factors driving soil bacterial community structure were nitrate, ammonium, and total nitrogen. PICRUSt function prediction results showed that metabolism (78.6%-79.1%) was the main function of soil bacterial communities in loess soil. Rotation with continued annual crops significantly decreased the abundance of soil bacterial carbohydrate metabolism functional genes, and significantly increased the abundance of functional genes for soil bacterial cofactors and vitamin metabolism, neurodegenerative diseases, and immune system. In conclusion, lucerne removed and rotated with continuous annual crops changed soil bacterial community structure and ecological functions. This study provided theoretical reference to explore succession characteristics of soil bacteria and to select succeeding crops for alfalfa in loess soil.

[Soil nematode community characteristics of alfalfa field with different growing ages in the semi-arid Loess Plateau of Central Gansu, Northwest China]. / 陇中黄土高原半干旱区不同种植年限紫花苜蓿土壤线虫群落分布特征.

To clarify the impacts of long-term alfalfa plantation on the soil nematode community, soil samples were collected from different alfalfa growing ages (2 a, 9 a, 18 a) in the semi-arid area of Loess Plateau in Central Gansu by Illumina Miseq sequencing technology. The main controlling factors affecting its community change were also explored. The results showed that soil nematode belongs to 2 classes, 7 orders, 16 families and 21 genera. Among them, Chromadorea was the dominant group (44.6%-81.4%), the relative abundance of which decreased with alfalfa growing ages. Paratylenchus, Helicotylenchus, Xiphinema, Pristionchus, Ditylenchus, Panagrolaimus, Longidorus, Aprutides, Isolaimium and Aglenchus were the special nematode species of alfalfa, among which Paratylenchus (54.1%), Helicotylenchus (23.9%) and Xiphinema (21.9%) were the dominant nematodes in 2 a, 9 a and 18 a alfalfa soil respectively. Plant-parasitic nematode was the dominant group in alfalfa soil (31.8%-67.1%), and its relative abundance decreased at first and then increased with alfalfa growing ages. Results of redundancy analysis showed that soil available phosphorus and total nitrogen were the dominant environmental factors affecting community structure of soil nematodes in the region.

Comparative biology of two genetically closely related Newcastle disease virus strains with strongly contrasting pathogenicity.

A lentogenic strain of Newcastle disease virus (NDV) with an intracerebral pathogenicity index (ICPI) of 0.36 was derived by the passage of a mesogenic NDV isolate with an original ICPI of 1.04. Animal experiments showed that the original strain caused much severer clinical signs and mortality than the derived strain in chickens. To elucidate the molecular reason for this virulence change, the complete viral genomes of the original and derived strains were sequenced. Molecular analysis showed that both viruses contained the same fusion (F) protein with a cleavage site (Fcs) motif that is usually associated with velogenic viruses. Molecular comparison revealed five amino acid (aa) differences in nucleoprotein (NP) (aa 426), hemagglutinin-neuraminidase (HN) (aas 215 and 430), and large protein (L) (aas 1694 and 1767), accompanied by the changes of relevant biological activities in membrane fusion and replication. Thus, we believe that the virulence changes may induced by these mutations. Our findings make a foundation for more in-depth investigations of the molecular mechanism underlying virulence.