A pan-respiratory antiviral chemotype targeting a transient host multi-protein complex.

We present a novel small molecule antiviral chemotype that was identified by an unconventional cell-free protein synthesis and assembly-based phenotypic screen for modulation of viral capsid assembly. Activity of PAV-431, a representative compound from the series, has been validated against infectious viruses in multiple cell culture models for all six families of viruses causing most respiratory diseases in humans. In animals, this chemotype has been demonstrated efficacious for porcine epidemic diarrhoea virus (a coronavirus) and respiratory syncytial virus (a paramyxovirus). PAV-431 is shown to bind to the protein 14-3-3, a known allosteric modulator. However, it only appears to target the small subset of 14-3-3 which is present in a dynamic multi-protein complex whose components include proteins implicated in viral life cycles and in innate immunity. The composition of this target multi-protein complex appears to be modified upon viral infection and largely restored by PAV-431 treatment. An advanced analog, PAV-104, is shown to be selective for the virally modified target, thereby avoiding host toxicity. Our findings suggest a new paradigm for understanding, and drugging, the host-virus interface, which leads to a new clinical therapeutic strategy for treatment of respiratory viral disease.

Validation of the Chinese version stage of recovery instrument-30 (STORI-30) for adults with severe mental illness.

BACKGROUND: Stage of Recovery Instrument-30 (STORI-30) is grounded in a five-stage model of psychological recovery, and serves as measuring recovery stage of people with mental illness. AIMS: To develop and validate the Chinese version STORI-30 on adults with severe mental illness. METHODS: STORI-30 was translated to traditional Chinese through forward-backward method. An expert panel and potential users evaluated face validity and content validity. The Chinese version STORI-30 plus other convergent and divergent scales were then administered to 113 participants for field test. RESULTS: Face and content validity were confirmed with acceptable Content Validity Index and high inter-rater agreement. Exploratory factor analysis revealed a three-factor structure. An ordinal sequence was presented among the five subscales, similar to the original version. Construct validity was supported by positive correlations with recovery and mental well-being scales, and negative correlation with self-stigma scale. Good internal consistency (Cronbach's α = 0.78-0.86) and high level of test-retest reliability (Intraclass correlation coefficient = 0.96) were obtained. CONCLUSIONS: Chinese STORI-30 presents satisfactory psychometric properties in internal consistency, construct convergent and divergent validity, and test-retest reliability. The three-factor structure revealed does not echo the original five-stage recovery model. Further studies exploring the underlying structure are warranted.

Comparison of Ganoderma boninense Isolate's Aggressiveness Using Infected Oil Palm Seedlings.

Basal stem rot incidence caused by a white-rot fungus, Ganoderma boninense, is the major disease of oil palm in Southeast Asia. The rate of disease transmission and host damage are affected by variations in pathogen aggressiveness. Several other studies have used the disease severity index (DSI) to determine G. boninense aggressiveness levels while verifying disease using a culture-based method, which might not provide accurate results or be feasible in all cases. To differentiate G. boninense aggressiveness, we employed the DSI and vegetative growth measurement of infected oil palm seedlings. Disease confirmation was performed through scanning electron microscopy and molecular identification of fungal DNA from both infected tissue and fungi isolated from Ganoderma selective medium. Two-month-old oil palm seedlings were artificially inoculated with G. boninense isolates (2, 4A, 5A, 5B, and 7A) sampled from Miri (Lambir) and Mukah (Sungai Meris and Sungai Liuk), Sarawak. The isolates were categorized into three groups: highly aggressive (4A and 5B), moderately aggressive (5A and 7A), and less aggressive (2). Isolate 5B was identified as the most aggressive, and it was the only one to result in seedling mortality. Out of the five vegetative growth parameters measured, only the bole size between treatments was not affected. The integration of both conventional and molecular approaches in disease confirmation allows for precise detection.

Sphagnum capillifolium holobiont from a subarctic palsa bog aggravates the potential of nitrous oxide emissions.

Melting permafrost mounds in subarctic palsa mires are thawing under climate warming and have become a substantial source of N2O emissions. However, mechanistic insights into the permafrost thaw-induced N2O emissions in these unique habitats remain elusive. We demonstrated that N2O emission potential in palsa bogs was driven by the bacterial residents of two dominant Sphagnum mosses especially of Sphagnum capillifolium (SC) in the subarctic palsa bog, which responded to endogenous and exogenous Sphagnum factors such as secondary metabolites, nitrogen and carbon sources, temperature, and pH. SC's high N2O emission activity was linked with two classes of distinctive hyperactive N2O emitters, including Pseudomonas sp. and Enterobacteriaceae bacteria, whose hyperactive N2O emitting capability was characterized to be dominantly pH-responsive. As the nosZ gene-harboring emitter, Pseudomonas sp. SC-H2 reached a high level of N2O emissions that increased significantly with increasing pH. For emitters lacking the nosZ gene, an Enterobacteriaceae bacterium SC-L1 was more adaptive to natural acidic conditions, and N2O emissions also increased with pH. Our study revealed previously unknown hyperactive N2O emitters in Sphagnum capillifolium found in melting palsa mound environments, and provided novel insights into SC-associated N2O emissions.

Confirmation and variability of the Allee effect inDictyostelium discoideumcell populations, possible role of chemical signaling within cell clusters.

In studies of the unicellular eukaryoteDictyostelium discoideum, many have anecdotally observed that cell dilution below a certain 'threshold density' causes cells to undergo a period of slow growth (lag). However, little is documented about the slow growth phase and the reason for different growth dynamics below and above this threshold density. In this paper, we extend and correct our earlier work to report an extensive set of experiments, including the use of new cell counting technology, that set this slow-to-fast growth transition on a much firmer biological basis. We show that dilution below a certain density (around 104cells ml-1) causes cells to grow slower on average and exhibit a large degree of variability: sometimes a sample does not lag at all, while sometimes it takes many moderate density cell cycle times to recover back to fast growth. We perform conditioned media experiments to demonstrate that a chemical signal mediates this endogenous phenomenon. Finally, we argue that while simple models involving fluid transport of signal molecules or cluster-based signaling explain typical behavior, they do not capture the high degree of variability between samples but nevertheless favor an intra-cluster mechanism.

A Pan-respiratory Antiviral Chemotype Targeting a Transient Host Multiprotein Complex.

We present a small molecule chemotype, identified by an orthogonal drug screen, exhibiting nanomolar activity against members of all the six viral families causing most human respiratory viral disease, with a demonstrated barrier to resistance development. Antiviral activity is shown in mammalian cells, including human primary bronchial epithelial cells cultured to an air-liquid interface and infected with SARS-CoV-2. In animals, efficacy of early compounds in the lead series is shown by survival (for a coronavirus) and viral load (for a paramyxovirus). The drug target is shown to include a subset of the protein 14-3-3 within a transient host multi-protein complex containing components implicated in viral lifecycles and in innate immunity. This multi-protein complex is modified upon viral infection and largely restored by drug treatment. Our findings suggest a new clinical therapeutic strategy for early treatment upon upper respiratory viral infection to prevent progression to lower respiratory tract or systemic disease. One Sentence Summary: A host-targeted drug to treat all respiratory viruses without viral resistance development.

Linking prokaryotic community composition to carbon biogeochemical cycling across a tropical peat dome in Sarawak, Malaysia.

Tropical peat swamp forest is a global store of carbon in a water-saturated, anoxic and acidic environment. This ecosystem holds diverse prokaryotic communities that play a major role in nutrient cycling. A study was conducted in which a total of 24 peat soil samples were collected in three forest types in a tropical peat dome in Sarawak, Malaysia namely, Mixed Peat Swamp (MPS), Alan Batu (ABt), and Alan Bunga (ABg) forests to profile the soil prokaryotic communities through meta 16S amplicon analysis using Illumina Miseq. Results showed these ecosystems were dominated by anaerobes and fermenters such as Acidobacteria, Proteobacteria, Actinobacteria and Firmicutes that cover 80-90% of the total prokaryotic abundance. Overall, the microbial community composition was different amongst forest types and depths. Additionally, this study highlighted the prokaryotic communities' composition in MPS was driven by higher humification level and lower pH whereas in ABt and ABg, the less acidic condition and higher organic matter content were the main factors. It was also observed that prokaryotic diversity and abundance were higher in the more oligotrophic ABt and ABg forest despite the constantly waterlogged condition. In MPS, the methanotroph Methylovirgula ligni was found to be the major species in this forest type that utilize methane (CH4), which could potentially be the contributing factor to the low CH4 gas emissions. Aquitalea magnusonii and Paraburkholderia oxyphila, which can degrade aromatic compounds, were the major species in ABt and ABg forests respectively. This information can be advantageous for future study in understanding the underlying mechanisms of environmental-driven alterations in soil microbial communities and its potential implications on biogeochemical processes in relation to peatland management.

Genetic Diversity and Demographic History of Ganoderma boninense in Oil Palm Plantations of Sarawak, Malaysia Inferred from ITS Regions.

Ganoderma boninense causes basal stem rot (BSR) and is responsible for substantial economic losses to Southeast Asia's palm oil industry. Sarawak, a major producer in Malaysia, is also affected by this disease. Emergence of BSR in oil palm planted on peat throughout Sarawak is alarming as the soil type was previously regarded as non-conducive. Phylogenetic analysis indicated a single species, G. boninense as the cause of BSR in Sarawak. Information on evolutionary and demographic history for G. boninense in Sarawak inferred through informative genes is lacking. Hence, a haplotype study on single nucleotide polymorphisms in internal transcribed spacers (SNPs-ITS) of G. boninense was carried out. Sequence variations were analysed for population structure, phylogenetic and phylogeographic relationships. The internal transcribed spacers (ITS) region of 117 isolates from four populations in eight locations across Sarawak coastal areas revealed seven haplotypes. A major haplotype, designated GbHap1 (81.2%), was found throughout all sampling locations. Single nucleotide polymorphisms were observed mainly in the ITS1 region. The genetic structure was not detected, and genetic distance did not correlate with geographical distance. Haplotype network analysis suggested evidence of recent demographic expansion. Low genetic differences among populations also suggested that these isolates belong to a single G. boninense founder population adapting to oil palm as the host.

Microenvironmental Interplay Predominated by Beneficial Aspergillus Abates Fungal Pathogen Incidence in Paddy Environment.

Rice fungal pathogens, responsible for severe rice yield loss and biotoxin contamination, cause increasing concerns on environmental safety and public health. In the paddy environment, we observed that the asymptomatic rice phyllosphere microenvironment was dominated by an indigenous fungus, Aspergillus cvjetkovicii, which positively correlated with alleviated incidence of Magnaporthe oryzae, one of the most aggressive plant pathogens. Through the comparative metabolic profiling for the rice phyllosphere microenvironment, two metabolites were assigned as exclusively enriched metabolic markers in the asymptomatic phyllosphere and increased remarkably in a population-dependent manner with A. cvjetkovicii. These two metabolites evidenced to be produced by A. cvjetkovicii in either a phyllosphere microenvironment or artificial media were purified and identified as 2(3H)-benzofuranone and azulene, respectively, by gas chromatography coupled to triple quadrupole mass spectrometry and nuclear magnetic resonance analyses. Combining with bioassay analysis in vivo and in vitro, we found that 2(3H)-benzofuranone and azulene exerted dissimilar actions at the stage of infection-related development of M. oryzae. A. cvjetkovicii produced 2(3H)-benzofuranone at the early stage to suppress MoPer1 gene expression, leading to inhibited mycelial growth, while azulene produced lately was involved in blocking of appressorium formation by downregulation of MgRac1. More profoundly, the microenvironmental interplay dominated by A. cvjetkovicii significantly blocked M. oryzae epidemics in the paddy environment from 54.7 to 68.5% (p < 0.05). Our study first demonstrated implication of the microenvironmental interplay dominated by indigenous and beneficial fungus to ecological balance and safety of the paddy environment.

Indole-3-Acetic Acid Produced by Burkholderia heleia Acts as a Phenylacetic Acid Antagonist to Disrupt Tropolone Biosynthesis in Burkholderia plantarii.

Burkholderia heleia PAK1-2 is a potent biocontrol agent isolated from rice rhizosphere, as it prevents bacterial rice seedling blight disease caused by Burkholderia plantarii. Here, we isolated a non-antibacterial metabolite from the culture fluid of B. heleia PAK1-2 that was able to suppress B. plantarii virulence and subsequently identified as indole-3-acetic acid (IAA). IAA suppressed the production of tropolone in B. plantarii in a dose-dependent manner without any antibacterial and quorum quenching activity, suggesting that IAA inhibited steps of tropolone biosynthesis. Consistent with this, supplementing cultures of B. plantarii with either L-[ring-(2)H5]phenylalanine or [ring-(2)H2~5]phenylacetic acid revealed that phenylacetic acid (PAA), which is the dominant metabolite during the early growth stage, is a direct precursor of tropolone. Exposure of B. plantarii to IAA suppressed production of both PAA and tropolone. These data particularly showed that IAA produced by B. heleia PAK1-2 disrupts tropolone production during bioconversion of PAA to tropolone via the ring-rearrangement on the phenyl group of the precursor to attenuate the virulence of B. plantarii. B. heleia PAK1-2 is thus likely a microbial community coordinating bacterium in rhizosphere ecosystems, which never eliminates phytopathogens but only represses production of phytotoxins or bacteriocidal substances.

Impact of IL28B and PNPLA3 polymorphisms on treatment outcomes in patients infected with genotype 6 hepatitis C virus.

BACKGROUND/AIMS: Interleukin-28B (IL28B) and patatin-like phospholipase domain containing 3 (PNPLA3) gene polymorphisms are associated with hepatitis C virus (HCV) clearance and fatty liver, respectively. We aimed to test if their polymorphisms are associated with virologic responses in Chinese chronic hepatitis C (CHC) patients. METHODS: This was a retrospective-prospective cohort study. Consecutive patients infected by genotype 1 and 6 HCV received antiviral therapy were included. Host IL-28B rs12979860/rs8099917 and PNPLA3 rs738409 genotype were tested. The primary outcome was sustained virologic response (sustained virologic response [SVR]: undetectable HCV RNA 24 weeks post-treatment). RESULTS: From 305 patients had positive antibody to HCV, 52 and 31 patients infected by genotype 1 and 6 HCV, respectively were recruited. Mean age was 58 ± 11 years; 70% were male. Mean baseline HCV RNA was 6.8 ± 2.7 log IU/ml. The SVR for patients infected by genotype 1 and 6 HCV was 67.3% and 90.3%, respectively. The proportions of IL28B genotypes were 78%, 21%, and 1% for TT/TG/GG at rs8099917, and 81%, 18%, and 1% for CC/TC/TT at rs12979860, respectively. The proportions of PNPLA3 rs738409 genotypes were 16%, 36%, and 48% for GG/GC/CC. IL28B genotype was significantly associated with SVR in patients infected by genotype 1 but not genotype 6 HCV, with 80% versus 38% of patients infected by genotype 1 achieved SVR carried TT versus TG/GG at rs8099917, respectively (P=0.003). PNPLA3 genotype was not associated with SVR. CONCLUSIONS: IL28B gene with rs8099917 T allele as an independent predictor of SVR in Chinese CHC patients infected by genotype 1 but not genotype 6 HCV.

Vancomycin use in hospitalized pediatric patients.

OBJECTIVES: To assess vancomycin utilization at children's hospitals, to determine risk factors for vancomycin use and length of therapy, and to facilitate adapting recommendations to optimize vancomycin prescribing practices in pediatric patients. METHODS: Two surveys were conducted at Pediatric Prevention Network hospitals. The first (Survey I) evaluated vancomycin control programs. The second (Survey II) prospectively reviewed individual patient records. Each hospital was asked to complete questionnaires on 25 consecutive patients or all patients for whom vancomycin was prescribed during a 1-month period. RESULTS: In Survey I, 55 of 65 (85%) hospitals reported their vancomycin control policies. Three quarters had specific policies in place to restrict vancomycin use. One half had at least 3 vancomycin restriction measures. In Survey II, personnel at 22 hospitals reviewed 416 vancomycin courses, with 2 to 25 (median = 12) patients tracked per hospital. Eighty-two percent of the vancomycin prescribed was for treatment of neonatal sepsis, fever/neutropenia, fever of unknown origin, positive blood culture, pneumonia, or meningitis. In an additional 6% (26/416), vancomycin was prescribed for patients with beta-lactam allergies and in 13% (56/416) for prophylaxis. Median duration of prophylaxis was 2 days (range: 1-15 days). Almost half (196, 47%) of the patients who received vancomycin were in intensive care units; 27% of the vancomycin courses were initiated by neonatologists and 19% by hematologists/oncologists. The predominant risk factor at the time of vancomycin initiation was the presence of vascular catheters (322, 77%); other host factors included cancer chemotherapy (55, 13%), transplant (30, 7%), shock (24, 6%), other immunosuppressant therapy (17, 4%), or hyposplenic state (2, <1%). Other clinical considerations were severity of illness (96, 23%), uncertainty about diagnosis (51, 12%), patient not responding to current antibiotic therapy (40, 10%), or implant infection (13, 3%). When vancomycin was initiated, blood cultures were positive in 85 patients (20%); cultures from other sites were positive in 45 (11%), and Gram stains of body fluids were positive in 37 (9%). In 29 (7%) patients, organisms sensitive only to vancomycin were isolated before vancomycin initiation. Reasons for discontinuing vancomycin included: therapeutic course completed (125, 30%), negative cultures (106, 25%), alternative antibiotics initiated (75, 18%), illness resolved (14, 3%), or patient expired (13, 3%). Final results of blood culture isolates resistant to beta-lactam antibiotics included 48 coagulase-negative staphylococcus, 5 Staphylococcus aureus, and 10 other species. CONCLUSIONS: At children's hospitals, vancomycin is initiated for therapy in patients who have vascular catheters and compromised host factors. Only 7% had laboratory-confirmed beta-lactam-resistant organisms isolated at the time vancomycin was prescribed. Efforts to modify empiric vancomycin use in children's hospitals should be targeted at intensivists, neonatologists, and hematologists. Initiatives to decrease length of therapy by decreasing the number of surgical prophylaxis doses and days of therapy before laboratory results may decrease vancomycin exposure.

Human intoxication by microcystins during renal dialysis treatment in Caruaru-Brazil.

In February 1996, an outbreak of illness occurred at a hemodialysis clinic in Caruaru, Pernambuco State-Brazil. At this clinic 116 (89%) of 131 patients experienced visual disturbances, nausea, vomiting, and muscle weakness, following routine haemodialysis treatment. Subsequently, 100 patients developed acute liver failure. As of December 1996, 52 of the deaths could be attributed to a common syndrome now called 'Caruaru Syndrome'. Examination of previous years' phytoplankton counts showed that cyanobacteria were dominant in the water supply reservoir since 1990. Analyses of carbon and other resins from the clinic's water treatment system plus serum and liver tissue of patients led to the identification of two groups of hepatotoxic cyanotoxins: microcystins (cyclic heptapeptides) in all of these samples and cylindrospermopsin (alkaloid hepatotoxic) in the carbon and resins. Comparison of victims symptoms and pathology with animal studies on these two cyanotoxins, leads us to conclude that the major contributing factor to death of the dialysis patients was intravenous exposure to microcystins, specifically microcystin-YR, -LR and -AR. In 2000, a review of the Brazilian regulation for drinking water quality, promoted by Brazilian Health Ministry with collaboration of PAHO, incorporated cyanobacteria and cyanotoxins into this new regulation as parameters that must to be monitored for water quality control.