High Throughput Sequencing-Aided Survey Reveals Widespread Mixed Infections of Whitefly-Transmitted Viruses in Cucurbits in Georgia,USA

Viruses transmitted by the sweet potato whitefly (Bemisia tabaci) have been detrimental to the sustainable production of cucurbits in the southeastern USA. Surveys were conducted in the fall of 2019 and 2020 in Georgia, a major cucurbit-producing state of the USA, to identify the viruses infecting cucurbits and their distribution. Symptomatic samples were collected and small RNA libraries were prepared and sequenced from three cantaloupes, four cucumbers, and two yellow squash samples. An analysis of the sequences revealed the presence of the criniviruses cucurbit chlorotic yellows virus (CCYV), cucurbit yellow stunting disorder virus (CYSDV), and the begomovirus cucurbit leaf crumple virus (CuLCrV). CuLCrV was detected in 76%, CCYV in 60%, and CYSDV in 43% of the total samples (n = 820) tested. The level of mixed infections was high in all the cucurbits, with most plants tested being infected with at least two of these viruses. Near-complete genome sequences of two criniviruses, CCYV and CYSDV, were assembled from the small RNA sequences. An analysis of the coding regions showed low genetic variability among isolates from different hosts. In phylogenetic analysis, the CCYV isolates from Georgia clustered with Asian isolates, while CYSDV isolates clustered with European and USA isolates. This work enhances our understanding of the distribution of viruses on cucurbits in South Georgia and will be useful to develop strategies for managing the complex of whitefly-transmitted viruses in the region.     

High throughput sequencing reveals distinct microbial populations within the mucosal and luminal niches in healthy individuals

Background: The intestinal microbiota is associated with human health and diseases. The luminal microbiota (LM) and the mucosal-associated microbiota (MAM) are 2 distinct ecosystems with different metabolic and immunological functions. Aim: To characterize the intestinal LM and MAM in humans using high throughput sequencing of the 16S rRNA gene. Methods: Fresh fecal samples and distal colonic mucosal biopsies collected from 24 healthy subjects before (fecal) and during (mucosa) a flexible sigmoidoscopy of an un-prepared bowel. High throughput sequencing of the 16S rRNA gene was used to characterize bacterial communities. Sequences were processed using the QIIME pipeline. Results: LM and MAM populations were significantly different (ANOSIM: R = 0.49, P = 0.001). The LM displayed tighter clustering compared to the MAM (average weighted UniFrac distances 0.27 ± 0.05 vs. 0.43 ± 0.09, P < 0.001, respectively), and showed higher diversity (Shannon diversity index: 4.96 ± 0.37 vs 4.14 ± 0.56, respectively, P < 0.001). The dominant phyla in the LM and MAM were significantly different: Firmicutes (41.4% vs. 29.1%, FDR < 0.0001, respectively), Bacteroidetes (20.2% vs. 26.3%, FDR < 0.05, respectively), Actinobacteria (22% vs. 12.6%, FDR < 0.0001, respectively) and Proteobacteria (9.3% vs. 19.3%, FDR < 0.0001, respectively). The abundance of 56 genera differed significantly (FDR < 0.1) between the 2 niches. All of the genera in the fecal microbiota were present in the MAM while 10 genera were found to be unique to the MAM. Conclusion: The LM and MAM are distinct microbial ecosystems that differ significantly from each other in microbial diversity and composition. These two microbial niches should be investigated independently to better understand the role of the intestinal microbiota in health and disease.     

High throughput sequencing reveals distinct microbial populations within the mucosal and luminal niches in healthy individuals

Background: The intestinal microbiota is associated with human health and diseases. The luminal microbiota (LM) and the mucosal-associated microbiota (MAM) are 2 distinct ecosystems with different metabolic and immunological functions. Aim: To characterize the intestinal LM and MAM in humans using high throughput sequencing of the 16S rRNA gene. Methods: Fresh fecal samples and distal colonic mucosal biopsies collected from 24 healthy subjects before (fecal) and during (mucosa) a flexible sigmoidoscopy of an un-prepared bowel. High throughput sequencing of the 16S rRNA gene was used to characterize bacterial communities. Sequences were processed using the QIIME pipeline. Results: LM and MAM populations were significantly different (ANOSIM: R = 0.49, P = 0.001). The LM displayed tighter clustering compared to the MAM (average weighted UniFrac distances 0.27 ± 0.05 vs. 0.43 ± 0.09, P < 0.001, respectively), and showed higher diversity (Shannon diversity index: 4.96 ± 0.37 vs 4.14 ± 0.56, respectively, P < 0.001). The dominant phyla in the LM and MAM were significantly different: Firmicutes (41.4% vs. 29.1%, FDR < 0.0001, respectively), Bacteroidetes (20.2% vs. 26.3%, FDR < 0.05, respectively), Actinobacteria (22% vs. 12.6%, FDR < 0.0001, respectively) and Proteobacteria (9.3% vs. 19.3%, FDR < 0.0001, respectively). The abundance of 56 genera differed significantly (FDR < 0.1) between the 2 niches. All of the genera in the fecal microbiota were present in the MAM while 10 genera were found to be unique to the MAM. Conclusion: The LM and MAM are distinct microbial ecosystems that differ significantly from each other in microbial diversity and composition. These two microbial niches should be investigated independently to better understand the role of the intestinal microbiota in health and disease.     

Identification of a New Badnavirus in the Chinaberry (Melia azedarach) Tree and Establishment of a LAMP-LFD Assay for Its Rapid and Visual Detection

The Chinaberry tree, a member of the Meliaceae family, is cultivated in China for use in traditional medicines. In 2020, Chinaberry trees with leaf deformation symptoms were found in Hangzhou, Zhejiang province, China. In order to identify possible pathogenic viruses, a symptomatic sample was subjected to deep sequencing of small interfering RNAs. Assembly of the resulting sequences led to the identification of a novel badnavirus, provisionally designated Chinaberry tree badnavirus 1 (ChTBV1). With the recent development of China’s seedling industry and increasing online shopping platforms, the risk of tree virus transmission has increased substantially. Therefore, it is important to detect the occurrence of ChTBV1 to ensure the safety of the Chinaberry tree seedling industry. Here, we describe the development and validation of a sensitive and robust method relying on a loop-mediated isothermal amplification (LAMP) assay, targeting a 197 nt region, to detect ChTBV1 from Chinaberry tree leaves. The LAMP assay was also adapted for rapid visualization of results by a lateral flow dipstick chromatographic detection method.     

Rapid spread of influenza A(H1N1)pdm09 viruses with a new set of specific mutations in the internal genes in the beginning of 2015/2016 epidemic season in Moscow and Saint Petersburg (Russian Federation)

A dramatic increase of influenza activity in Russia since week 3 of 2016 significantly differs from previous seasons in terms of the incidence of influenza and acute respiratory infection (ARI) and in number of lethal cases. We performed antigenic analysis of 108 and whole‐genome sequencing of 77 influenza A(H1N1)pdm09 viruses from Moscow and Saint Petersburg. Most of the viruses were antigenically related to the vaccine strain. Whole‐genome analysis revealed a composition of specific mutations in the internal genes (D2E and M83I in NEP, E125D in NS1, M105T in NP, Q208K in M1, and N204S in PA‐X) that probably emerged before the beginning of 2015/2016 epidemic season.