Target identification is highly instructive in defining the biological roles of microRNAs. However, little is known about other small noncoding RNAs; for example, tRNA-derived RNA Fragments (tRFs). Some tRFs exhibit a gene-silencing mechanism distinctly different from that of typical microRNAs. We recently demonstrated that a respiratory syncytial virus (RSV)-induced tRF, called tRF5-GluCTC, promotes RSV replication. RSV is the single most important cause of lower respiratory tract infection in children. By using biochemical screening and bioinformatics analyses, we have identified apolipoprotein E receptor 2 (APOER2) as a target of tRF5-GluCTC. The 3'-portion of tRF5-GluCTC recognizes a target site in the 3'-untranslated region of APOER2 and suppresses its expression. We have also discovered that APOER2 is an anti-RSV protein whose suppression by tRF5-GluCTC promotes RSV replication. Our report represents the first identification of a natural target of a tRF and illustrates how a virus utilizes a host tRF to control a host gene to favor its replication.

Acute Respiratory Distress Syndrome (ARDS) is a clinical syndrome characterized by diffuse alveolar damage usually secondary to an intense host inflammatory response of the lung to a pulmonary or extrapulmonary infectious or non-infectious insult often leading to the development of intra-alveolar and interstitial fibrosis. Curcumin, the principal curcumoid of the popular Indian spice turmeric, has been demonstrated as an anti-oxidant and anti-inflammatory agent in a broad spectrum of diseases. Using our well-established model of reovirus 1/L-induced acute viral pneumonia, which displays many of the characteristics of the human ALI/ARDS, we evaluated the anti-inflammatory and anti-fibrotic effects of curcumin. Female CBA/J mice were treated with curcumin (50 mg/kg) 5 days prior to intranasal inoculation with 107 pfu reovirus 1/L and daily, thereafter. Mice were evaluated for key features associated with ALI/ARDS. Administration of curcumin significantly modulated inflammation and fibrosis, as revealed by histological and biochemical analysis. The expression of IL-6, IL-10, IFNγ, and MCP-1, key chemokines/cytokines implicated in the development of ALI/ARDS, from both the inflammatory infiltrate and whole lung tissue were modulated by curcumin potentially through a reduction in the phosphorylated form of NFκB p65. While the expression of TGFß1 was not modulated by curcumin, TGFß Receptor II, which is required for TGFß signaling, was significantly reduced. In addition, curcumin also significantly inhibited the expression of α-smooth muscle actin and Tenascin-C, key markers of myofibroblast activation. This data strongly supports a role for curcumin in modulating the pathogenesis of viral-induced ALI/ARDS in a pre-clinical model potentially manifested through the alteration of inflammation and myofibroblast differentiation.

The intestinal organoid culture system is a pathbreaking working model for investigating pathogen-host interactions in the intestines. However, due to the limitations of the first generation of intestinal organoids, basal-out structure and growth in Matrigel, most pathogens can rarely attach to the apical membrane directly and hardly initiate infection. In this study, we first developed a next-generation porcine intestinal organoid culture system, characterized by an apical membrane on the surface, called apical-out. To investigate the infectivity and antiviral immune responses of this apical-out porcine intestinal organoid, a swine enteric virus, transmissible gastroenteritis virus (TGEV), was employed to inoculate the culture system. Both RT–qPCR and IFA analysis demonstrated that TGEV replicated in the apical-out porcine intestinal organoid culture system. Additionally, our results illustrated that TGEV infection significantly upregulated the expression levels of IFN-α, IFN-λ1, ISG15, ISG58, TNF-α, and IL-6 in this culture system. Hence, we successfully developed a porcine intestinal apical-out organoid culture system, which will facilitate the investigation of pathogen-host interactions in pig intestines. IMPORTANCE Intestinal organoids are a newly developed culture system for investigating pathogen-host interactions. Intestinal organoid models have been widely used since their development because the results obtained from this type of culture model better represent physiological conditions than those from well-established cell lines. The 3D porcine intestinal organoid model was reported in 2018 and 2019 for the investigation of intestinal pathogens. However, those organoid culture models were basal-out intestinal organoids, which are not suitable for porcine enteric virus research because they invade the intestines via the apical side of epithelial cells on villi. In this study, we developed a porcine apical-out intestinal organoid culture system and verified its infectivity, type I, and type III IFN antiviral responses, and inflammatory responses following infection by a swine enteric virus. Our results imply that this apical-out porcine intestinal organoid culture system is an ideal model for the investigation of interactions between swine enteric viruses and the intestines.

Human noroviruses are responsible for more than 95% of nonbacterial epidemic gastroenteritis worldwide. Both onset and resolution of disease symptoms are rapid, suggesting that components of the innate immune response are critical in norovirus control. While the study of the human noroviruses has been hampered by the lack of small animal and tissue culture systems, our recent discovery of a murine norovirus (MNV) and its in vitro propagation have allowed us to begin addressing norovirus replication strategies and immune responses to norovirus infection. We have previously demonstrated that interferon responses are critical to control MNV-1 infection in vivo and to directly inhibit viral replication in vitro. We now extend these studies to define the molecular basis for interferon-mediated inhibition. Viral replication intermediates were not detected in permissive cells pretreated with type I interferon after either infection or transfection of virion-associated RNA, demonstrating a very early block to virion production that is after virus entry and uncoating. A similar absence of viral replication intermediates was observed in infected primary macrophages and dendritic cells pretreated with type I IFN. This was not due to degradation of incoming genomes in interferon-pretreated cells since similar levels of genomes were present in untreated and pretreated cells through 6 h of infection, and these genomes retained their integrity. Surprisingly, this block to the translation of viral proteins was not dependent on the well-characterized interferon-induced antiviral molecule PKR. Similar results were observed in cells pretreated with type II interferon, except that the inhibition of viral translation was dependent on PKR. Thus, both type I and type II interferon signaling inhibit norovirus translation in permissive myeloid cells, but they display distinct dependence on PKR for this inhibition.

No abstract

Methylmercury (MeHg) is recognized as one of the major water quality concerns in the Florida Everglades. Degradation of MeHg in the water is thought to be one of the most important processes to the cycling of MeHg, but there is a lack of quantitative estimations of its effect on the distribution and cycling of MeHg in this ecosystem. Stable isotope (Me201Hg) addition method was implemented to investigate the degradation of MeHg in the Everglades. By combining these results with the field monitoring data, effects of photodegradation on MeHg distribution and its contribution to MeHg cycling were estimated. The results indicate that degradation of MeHg in Everglades water is mediated by sunlight and that UV-A and UV-B radiations are the principal driver. The spatial pattern of MeHg photodegradation potential (PPD) generally illustrated an increasing trend from north to south in the Everglades, which was opposite to the distribution of MeHg in water column. Correlation analysis shows that MeHg concentration in the water had a significant negative relation to PPD, suggesting that photodegradation could play an important role in controlling the distribution of MeHg in Everglades water. Furthermore, about 31.4% of MeHg input into the water body was removed by photodegradation, indicating its importance in the biogeochemical cycling of MeHg in the Everglades. This percent reduction is much lower than that reported for other ecosystems, which could be caused by the higher concentration of DOC in the Everglades. The relatively slower degradation of MeHg could be one of the main reasons for the high ratio of MeHg to total mercury (THg) in this ecosystem.

In the presence of iron (Fe), dissolved organic matter (DOM) may bind considerable amounts of arsenic (As), through formation of Fe-bridged As-Fe-DOM complexes and surface complexation of As on DOM-stabilized Fe-colloids (collectively referred to as As-Fe-DOM complexation). However, direct (e.g., chromatographic and spectroscopic) evidence and fundamental kinetic and stability constants have been rarely reported for this As-Fe-DOM complexation. Using a size exclusion chromatography (SEC)-UV-inductively coupled plasma mass spectrometry (ICP-MS) technique, arsenite (AsIII)-Fe-DOM complexation was investigated after adding AsIII into the priorly prepared Fe-DOM. A series of evidence, including coelution of As, Fe, and DOM from the SEC column and coretention of As, Fe, and DOM by 3 kDa MWCO centrifugal filtration membrane, demonstrated the occurrence of AsIII-Fe-DOM complexation. The kinetic data of AsIII-Fe-DOM complexation were well described by a pseudo-first order rate equation (R2 = 0.95), with the rate constant (k′) being 0.17±0.04 1/h. Stability of AsIII-Fe-DOM complexation was characterized by apparent stability constant (Ks) derived from two-site ligand binding model, with log Ks ranging from 4.4±0.2 to 5.6±0.4. Considering the kinetics (within hours) and stability (similar to typical metal-humates) of AsIII-Fe-DOM complexation, this complexation needs to be included when evaluating As mobility in Fe and DOM rich environments.

The major enteric RNA viruses in pigs include porcine epidemic diarrhoea virus (PEDV), transmissible gastroenteritis virus (TGEV), porcine rotavirus A (PRV‐A), porcine kobuvirus (PKV), porcine sapovirus (PSaV) and porcine deltacoronavirus (PDCoV). For differential diagnosis, a multiplex RT‐PCR method was established on the basis of the N genes of TGEV, PEDV and PDCoV, the VP7 gene of PRV‐A, and the polyprotein genes of PKV and PSaV. This multiplex RT‐PCR could specifically detect TGEV, PEDV, PDCoV, PRV‐A, PKV and PSaV without cross‐reaction to any other major viruses circulating in Chinese pig farms. The limit of detection of this method was as low as 100–101 ng cDNA of each virus. A total of 398 swine faecal samples collected from nine provinces of China between October 2015 and April 2017 were analysed by this established multiplex RT‐PCR. The results demonstrated that PDCoV (144/398), PSaV (114/398), PEDV (78/398) and PRV‐A (70/398) were the main pathogens, but TGEV was not found in the pig herds in China. In addition, dual infections, for example, PDCoV + PSaV, PDCoV + PRV‐A, PRA‐V + PSaV and PEDV + PDCoV, and triple infections, for example, PDCoV + PRV‐A + PSaV and PEDV + PDCoV + PKV, were found among the collected samples. The multiplex RT‐PCR provided a valuable tool for the differential diagnosis of swine enteric viruses circulating in Chinese pig farms and will facilitate the prevention and control of swine diarrhoea in China.