Epidemiological and genetic characteristics of norovirus in Hangzhou, China, in the postepidemic era.

OBJECTIVE: Norovirus (NoV) is an important human pathogen that can cause severe gastroenteritis in vulnerable populations. This study aimed to analyze the epidemiological and genetic characteristics of 2021-2023 NoV in Hangzhou, China. METHODS: This study enrolled patients aged 0-18 years who underwent NoV RNA detection in the hospital between January 2021 and October 2023 and analyzed the epidemiological characteristics of NoV. Polymerase chain reaction (PCR) was used to detect NoV RNA. Subtype classification and whole-genome sequencing were performed. RESULTS: There was a high prevalence of NoV infection in 2023, with NoV-positive samples accounting for 63.10 % of the total number of positive samples collected during the three-year period. The prevalence was abnormally high in summer, and the number of positive samples accounted for 48.20 % of the total positive samples for the whole year, which was much greater than the level in the same period in previous years (2023, 48.20% vs 2021, 13.66% vs 2022, 15.21 %). The GⅡ.4 subtype played a leading role, followed by increased mixed infection with GⅠ.5 and GⅡ.4. Whole-genome sequencing results suggested that GII.P16-GⅡ.4 had R297H and D372N key locus mutations. The evolutionary rate was 4.29 × 10-3 for the RdRp gene and 4.84 × 10-3 for the VP1 gene. The RdRp gene and VP1 gene of NoV GII.P16-GⅡ.4 have undergone rapid population evolution during the COVID-19 epidemic. CONCLUSION: In the summer of 2023, an abnormally high incidence of NoV appeared in Hangzhou, China. The major epidemic strain GII.P16-GⅡ.4 showed a certain range of gene mutations and a fast evolutionary rate.

Mobile genetic elements mediate the cross-media transmission of antibiotic resistance genes from pig farms and their risks.

Manure composting in traditional small-scale pig farms leads to the migration and diffusion of antibiotics and antibiotics resistance genes (ARGs) along the chain of transmission to the surrounding environment, increasing the risk of environmental resistance. Understanding the transmission patterns, driving factors, and health risks of ARGs on small-scale pig farms is important for effective control of ARGs transmission. This study was conducted on a small pig farm and its surrounding environment. The cross-media transmission of ARGs and their risks in the farming habitat were investigated using Metagenomic annotation and qPCR quantitative detection. The results indicate that ARGs in farms spread with manure pile-soil-channel sediment-mudflat sediment. Pig farm manure contributed 22.49 % of the mudflat sediment ARGs. Mobile genetic elements mediate the spread of ARGs across different media. Among them, tnpA and IS26 have the highest degree. Transmission of high-risk ARGs sul1 and tetM resulted in a 50 % and 116 % increase in host risk for sediment, respectively. This study provides a basis for farm manure management and control of the ARGs spread.

Functional prediction of the potential NGLY1 mutations associated with rare disease CDG.

Genetic diseases are currently diagnosed by functional mutations. However, only some mutations are associated with disease. It is necessary to establish a quick prediction model for clinical screening. Pathogenic mutations in NGLY1 cause a rare autosomal recessive disease known as congenital disorder of deglycosylation (NGLY1-CDDG). Although NGLY1-CDDG can be diagnosed through gene sequencing, clinical relevance of a detected mutation in NGLY1 needs to be further confirmed. In this study, taken NGLY1-CDDG as an example, a comprehensive and practical predictive model for pathogenic mutations on NGLY1 through an NGLY1/Glycopeptide complex model was constructed, the binding sites of NGLY1 and glycopeptides were simulated, and an in vitro enzymatic assay system was established to facilitate quick clinical decisions for NGLY1-CDDG patients. The docking model covers 42 % of reported NGLY1-CDDG missense mutations (5/12). All reported mutations were subjected to in vitro enzymatic assay in which 18 mutations were dysfunctional (18/30). In addition, a full spectrum of functional R328 mutations was assayed and 11 mutations were dysfunctional (11/19). In this study, a model of NGLY1 and glycopeptides was built for potential functional mutations in NGLY1. In addition, the effect of potential regulatory compounds, including N-acetyl-l-cysteine and dithiothreitol, on NGLY1 was examined. The established in vitro assay may serve as a standard protocol to facilitate rapid diagnosis of all mutations in NGLY1-CDDG. This method could also be applied as a comprehensive and practical predictive model for the other rare genetic diseases.

The effect of nonpharmaceutical interventions on influenza virus transmission.

Background: The use of nonpharmaceutical interventions (NPIs) during severe acute respiratory syndrome 2019 (COVID-19) outbreaks may influence the spread of influenza viruses. This study aimed to evaluate the impact of NPIs against SARS-CoV-2 on the epidemiological features of the influenza season in China. Methods: We conducted a retrospective observational study analyzing influenza monitoring data obtained from the China National Influenza Center between 2011 and 2023. We compared the changes in influenza-positive patients in the pre-COVID-19 epidemic, during the COVID-19 epidemic, and post-COVID-19 epidemic phases to evaluate the effect of NPIs on influenza virus transmission. Results: NPIs targeting COVID-19 significantly suppressed influenza activity in China from 2019 to 2022. In the seventh week after the implementation of the NPIs, the number of influenza-positive patients decreased by 97.46% in southern regions of China and 90.31% in northern regions of China. However, the lifting of these policies in December 2022 led to an unprecedented surge in influenza-positive cases in autumn and winter from 2022 to 2023. The percentage of positive influenza cases increased by 206.41% (p < 0.001), with high positivity rates reported in both the northern and southern regions of China. Conclusion: Our findings suggest that NPIs against SARS-CoV-2 are effective at controlling influenza epidemics but may compromise individuals' immunity to the virus.

Talaromyces marneffei endocarditis initially detected by Next Generation Sequencing: A case report.

BACKGROUND: Talaromyces marneffei (T. marneffei) is a thermal dimorphic fungus, which can cause lung or blood stream infection in patients, often life-threatening. However, endocarditis caused by T. marneffei has not been reported. For elderly patients with implanted cardiac devices or artificial valves, the prevention and treatment of infective endocarditis should not be ignored. METHODS: This is a descriptive study of a T. marneffei endocarditis by joint detection of cardiac ultrasound examination, peripheral blood DNA metagenomics Next Generation Sequencing (mNGS), and in vitro culture. RESULTS: We describe an 80-year-old female patient with an unusual infection of T. marneffei endocarditis. After intravenous drip of 0.2 g voriconazole twice a day for antifungal treatment, the patient showed no signs of improvement and their family refused further treatment. CONCLUSION: Infective endocarditis is becoming more and more common in the elderly due to the widely use of invasive surgical procedures and implantation of intracardiac devices. The diagnosis and treatment of T. marneffei endocarditis is challenging because of its rarity. Here, we discussed a case of T. marneffei endocarditis, and emphasized the role of mNGS in early diagnosis, which is of great significance for treatment and survival rate of patients.

Nocardia pseudobrasiliensis infection in a patient with arthritis.

Nocardia pseudobrasiliensis is a new taxon constituting an emerging species of human pathogenic Nocardia, which shares morphological features with N. brasiliensis. However, N. pseudobrasiliensis is more invasive and more easily disseminated, and it exhibits distinctive antibiotic susceptibility. Few clinical cases related to N. pseudobrasiliensis infection have been reported, and N. pseudobrasiliensis hydrarthrosis has not been described. Here, we analyzed the case information, diagnostic process, treatment, and prognosis of a patient with N. pseudobrasiliensis hydrarthrosis who received treatment in Zhejiang Provincial People's Hospital. Magnetic resonance imaging showed joint cavity effusion and soft tissue swelling with high signal on proton density-fat saturated images and low signal on T1-weighted images. Oil microscopy revealed abundant acid-fast-positive filaments in hydrarthrosis puncture fluid. The pathogen was identified as N. pseudobrasiliensis by matrix-assisted laser desorption ionization-time of flight mass spectrometry. In contrast to the 100% ciprofloxacin resistance displayed by N. brasiliensis, this clinical isolate of N. pseudobrasiliensis was completely susceptible. In summary, this is the first report of N. pseudobrasiliensis in joint effusion from a patient with arthritis.

The role of microbial interactions in dental caries: Dental plaque microbiota analysis.

OBJECTIVES: Dental caries is a result of the ecological dysfunction of the polymicrobial community on the tooth surface, which evolves through microbial interactions. In this study, we conducted a thorough analysis of the dental plaque microbiome to comprehend its multi-microbial aetiology. MATERIALS AND METHOD: In this study, plaque was collected from healthy tooth surfaces, shallow carious teeth and deep carious teeth, and bacterial composition and abundance were assessed using 16S rRNA high-throughput sequencing. Random forest and LEfSe were used to profile various microorganisms at each stage. Additionally, we developed a molecular ecological network (MEN) based on random matrix theory (RMT) to examine microbial interactions for the first time. RESULTS: Our results reveal that Scardovia wiggsiae, Streptococcus mutans, and Propionibacterium acidifaciens may be associated with initial caries, and Propionibacterium acidifaciens differentiates between shallow and deep caries. As caries progressed, the alpha diversity index declined, indicating a decrease in microbial variety. The network topological indices such as centralization betweenness revealed that the caries network had become more complex, involving more microbial interactions. The shallow network revealed a high negative correlation ratio across nodes, indicating that microbes competed heavily. In contrast, the positive correlation ratio of deep network nodes was high, and microorganisms transitioned from a competitive to a synergistic state. CONCLUSIONS: This study suggests that microbial diversity and interactions are critical to caries progression and that future caries research should give greater consideration to the role of microbial interaction factors in caries progression.

Quantitative Risk Assessment of Five Foodborne Viruses in Shellfish Based on Multiplex qPCR.

Foodborne diseases are currently the most critical food safety issue in the world. There are not many hazard identification and exposure assessments for foodborne viruses (Norovirus GI, GII, Hepatitis A Virus, Rotavirus, Adenovirus) in shellfish. Multiplex qPCR for the simultaneous detection of five foodborne viruses was established and used to assess infection risk based on a 1-year pathogenesis study. The sensitivity, specificity and reproducibility of the multiplex qPCR method are consistent with that of conventional qPCR, which saves more time and effort. Overall, 37.86% of shellfish samples had one or more foodborne viruses. Risk assessment formulae and matrices were used to develop risk assessments for different age groups, different seasons and different shellfish. The annual probability of contracting a foodborne virus infection from shellfish is greater than 1.6 × 10-1 for all populations, and even for infants aged 0-4 years, it is greater than 1.5 × 10-2, which is much higher than the risk thresholds recommended by WHO (10-6) and the US EPA (10-4). High risk (level IV) is associated with springtime, and medium risk (level III) is associated with Mussel consumption. This study provides a basis for the risk of foodborne viral infections in people of different ages, in different seasons, and by consuming different shellfish.

Vertical migration and dissipation of oxytetracycline induces the recoverable shift in microbial community and antibiotic resistance.

Antibiotic resistance gene (ARG) spread in anthropogenic polluted soils is believed to be accelerated by the incidental inputs of antibiotics via fertilizing and irrigation, and endangering food and human health. However, due to the complex nature of substrates and uncertain microbial responses, the primary drivers of ARG dissemination remain unclear. To address this concern, the effects of antibiotic inputs on soil microbes and antibiotic resistance under simulated natural conditions was investigated in this study. Specifically, four flow-through reactors with gravity flow were established, and the oxytetracycline (OTC) a typical antibiotic in agricultural soils was studied at environmental concentrations (i.e. 0.1, 1 and 10 mg/kg) for 31 days. The vertical distribution and dissipation of OTC were profiled by measuring the residuals in layers over time. Correspondingly, the effects of antibiotic exposure on microbial communities and ARG abundances were studied. The results showed that the average exposure intensity of OTC in different soil layers ranged in 0.03-6.45 mg/kg, and resulted in different dissipation kinetics. In addition, top layer was found to be the main site of OTC reduction, where OTC dissipated at magnitude of 74.0-96.6 %, depending on the initial OTC concentration. OTC migration and dissipation resulted in the shift of community composition to the extent of 0.25-0.33 in terms of Bray-Curtis distance, which partially recovered over time. And the achievement of alternative community compositions was supposed to be largely affected by the microbial interaction. Along with the community changes, a short-term accumulation of resistance genes was detected, while the relative abundance of indicator ARGs, i.e. tetG and mexB, rising up to 10-fold higher than the initial, although eventually decayed. Collective findings of this study indicated that antibiotics at environmental concentrations might trigger extra microbial interactions and thereby reducing the demand for ARGs accumulation. It provided valuable understandings in the risk of antibiotic spillage, especially for the incident exposure at the environmentally relevant concentrations.

Successful treatment with amoxicillin-clavulanic acid: cutaneous nocardiosis caused by Nocardia brasiliensis.

AIM: To emphasize the role of non-sulfonamides in the treatment of Nocardia infection and reduce the adverse reactions caused by sulfonamides. METHODS: We retrospectively analyzed a case of cutaneous nocardiosis in an immunocompetent individual. The colonies obtained by staining the pus in the lesion with antacid and culturing the agar plates were identified by flight mass spectrometry. The pathogenic identification showed Nocardia brasiliensis infection and the patient was treated with amoxicillin-clavulanic acid. RESULTS: After treatment with amoxicillin and clavulanic acid, the ulcer gradually peeled and crusted, leaving dark pigmentation. The patient has finally recovered. CONCLUSION: Sulfonamides are the first-line antibacterial agents for years in treatment of nocardiosis but are of great toxicity and side effects. This patient was successfully treated with amoxicillin-clavulanic acid and it provided a reference protocol for patients with sulfonamide-resistant Nocardia or sulfonamides intolerance.

Dabie bandavirus infection induces macrophagic pyroptosis and this process is attenuated by platelets.

Severe fever with thrombocytopenia syndrome (SFTS) is an emerging tick-borne infection with a high mortality rate in humans, which is caused by Dabie bandavirus (DBV), formerly known as SFTS virus. Clinical manifestations of SFTS are characterized by high fever, thrombocytopenia, leukopenia, hemorrhage, gastrointestinal symptoms, myalgia and local lymph node enlargement with up to 30% case fatality rates in human. Macrophage depletion in secondary lymphoid organs have important roles in the pathogenic process of fatal SFTS, but its exact cell death mechanism remains largely unknown. Here, we showed for the first time that DBV infection induced macrophagic pyroptosis, as evidenced by swollen cells, pore-forming structures, accumulation of gasdermin D N-terminal (GSDMD-NT) as well as the release of lactate dehydrogenase (LDH) and IL-1ß in human macrophages. In addition to the upregulation of pyronecrosis genes, the expressions of pyroptosis-related proteins (GSDMD, caspase-1 and IL-1ß) were also elevated. To be noted, platelets were found to play a protective role in DBV-derived pyroptosis. Transcriptome analysis and in vitro studies demonstrated that platelets significantly reduced the gene expressions and protein production of pro-pyroptotic markers and inflammatory cytokines in macrophages, whereas platelets conferred a propagation advantage for DBV. Collectively, this study demonstrates a novel mechanism by which DBV invasion triggers pyroptosis as a host defense to remove replication niches in human macrophages and platelets provide an additional layer to reduce cellular death. These findings may have important implications to the pathogenesis of lethal DBV, and provide new ideas for developing novel therapeutics to combat its infection.

Immunogenicity and Blocking Efficacy of Norovirus GII.4 Recombinant P Protein Vaccine.

Noroviruses (NoVs) are the main cause of acute gastroenteritis in all ages worldwide. The aim of this study was to produce the recombinant P protein of norovirus and to demonstrate its blocking effect. In this study, the engineered strains were induced to express the P protein of NoVs GII.4, which was identified using SDS-PAGE and ELISA as having the capacity to bind to histo-blood group antigens (HBGAs). Rabbits were immunized to obtain neutralizing antibodies. ELISA and ISC-RT-qPCR were used to determine the blocking efficacy of the neutralizing antibody to human norovirus (HuNoV) and murine norovirus (MNV). The recombinant P protein (35 KD) was obtained, and the neutralizing antibody was successfully prepared. The neutralizing antibody could block the binding of the P protein and HuNoV to HBGAs. Neutralizing antibodies can also block MNV invasion into host cells RAW264.7. The recombinant P protein expressed in E. coli can induce antibodies to block HuNoV and MNV. The recombinant P protein of NoVs GII.4 has the value of vaccine development.

Soluble extracellular polymeric substance (SEPS) of histo-blood group antigen (HBGA) expressing bacterium Sphingobacterium sp. SC015 influences the survival and persistence of norovirus on lettuce.

Foodborne norovirus (NoV) outbreaks linked to leafy greens are common due to a lack of efficient strategies to prevent NoV spread from contaminated surfaces. We previously found that Sphingobacterium sp. SC015 in lettuce phyllosphere expresses histo-blood group antigen (HBGA)-like substances in soluble extracellular polymeric substances (SEPS) that contribute to NoV adherence on lettuce. Here, we extracted SEPS from bacterium SC015 (SEPS-SC015), analyzed their chemical composition, and examined their roles in the survival and protection of NoV and surrogates [murine norovirus (MNV-1) and Tulane virus (TuV)] on lettuce. Presence of SEPS-SC015 significantly increased survival and persistence of human NoV (HuNoV), MNV-1, and TuV at days 7 and 14, compared with virus alone. HuNoV, TuV, and MNV-1 seeded with SEPS-SC015 were more resistant to heat (70 °C, 2 min) than these viruses alone. SEPS-SC015 also increased viral resistance to sodium hypochlorite inactivation by treatment with 30 and 300 ppm bleach at 26 °C for 10 min. However, SEPS-SC015 was not effective at protecting these viruses under UV inactivation. Binding of TuV to SC015 bacteria and SEPS-SC015, visualized using transmission electron microscopy, suggests that protection might be related to direct interaction between SEPS-SC015 and viral particles. This study provides important insights that will help inform strategies to improve food safety.

Effect of Direct Viral-Bacterial Interactions on the Removal of Norovirus From Lettuce.

Norovirus (NoV) is the main non-bacterial pathogen causing outbreaks of gastroenteritis and is considered to be the leading cause of foodborne illness. This study aims to determine whether lettuce-encapsulated bacteria can express histo-blood group antigen (HBGA)-like substances to bind to NoV and, if so, to explore its role in protecting NoV from disinfection practices. Fifteen bacterial strains (HBGA-SEBs) were isolated from the lettuce microbiome and studied as they were proved to have the ability to express HBGA-like substances through indirect ELISA detection. By using attachment assay, HBGA-SEBs showed great abilities in carrying NoVs regarding the evaluation of binding capacity, especially for the top four strains from genera Wautersiella, Sphingobacterium, and Brachybacterium, which could absorb more than 60% of free-flowing NoVs. Meanwhile, the direct viral-bacterial binding between HBGA-like substance-expressing bacteria (HBGA-SEB) and NoVs was observed by TEM. Subsequently, results of simulated environmental experiments showed that the binding of NoVs with HBGA-SEBs did have detrimental effects on NoV reduction, which were evident in short-time high-temperature treatment (90°C) and UV exposure. Finally, by considering the relative abundance of homologous microorganisms of HBGA-SEBs in the lettuce microbiome (ca. 36.49%) and the reduction of NoVs in the simulated environments, we suggested putting extra attention on the daily disinfection of foodborne-pathogen carriers to overcome the detrimental effects of direct viral-bacterial interactions on the reduction of NoVs.

Survey of pathogenic bacteria of biofilms in a metropolitan drinking water distribution system.

Bacteria, especially pathogenic bacteria, were detected in order to estimate the safety of drinking water distribution systems (DWDSs). Sixteen biofilms and 12 water samples (six retained and six flowing) were collected from a city DWDS in eastern China. Biofilms were observed using scanning electron microscopy. Cultivable bacteria of biofilms were counted by heterotrophic plate counts, ranging from 3.61 × 101 to 1.67 × 106 CFU·cm-2. Coliforms, Salmonella, Shigella, Vibrio and Legionella were separated by Eosin-Methylene Blue (EMB) agar, Salmonella chromogenic medium, Shigella chromogenic medium, Thiosulfate Citrate Bile Salts Sucrose (TCBS) agar and Buffered Charcoal Yeast Extract (BCYE) agar and 13/16, 8/16, 7/16, 6/16, 0/16 biofilm samples were found to be positive, respectively. Retained and flowing water samples were collected to estimate the influence of hydrodynamic conditions on biofilm detachment. All six retained water samples were positive for bacteria, the count ranged from 1.2 × 103 to 2.8 × 104 CFU·mL-1 and 2/6, 3/6, 2/6, 0/6, 0/6 samples were positive for coliforms, Salmonella, Shigella, Legionella and Vibrio, respectively. While only three of six flowing water samples were bacteria positive, the counts ranged from 102 to 103 CFU·mL-1, 2/6 were coliform positive and no pathogens were detected under testing. The results show that there are pathogens in DWDS biofilms, which can cause health-related problems if detached from their surfaces.

Antiviral Effects of Houttuynia cordata Polysaccharide Extract on Murine Norovirus-1 (MNV-1)-A Human Norovirus Surrogate.

Houttuynia cordata is an herbal plant rich in polysaccharides and with several pharmacological activities. Human noroviruses (HuNoVs) are the most common cause of foodborne viral gastroenteritis throughout the world. In this study, H. cordata polysaccharide (HP), with a molecular weight of ~43 kDa, was purified from H. cordata water extract (HWE). The polysaccharide HP was composed predominantly of galacturonic acid, galactose, glucose, and xylose in a molar ratio of 1.56:1.49:1.26:1.11. Methylation and NMR analyses revealed that HP was a pectin-like acidic polysaccharide mainly consisting of α-1,4-linked GalpA, ß-1,4-linked Galp, ß-1,4-linked Glcp, and ß-1,4-linked Xylp residues. To evaluate the antiviral activity of H. cordata extracts, we compared the anti-norovirus potential of HP with HWE and ethanol extract (HEE) from H. cordata by plaque assay (plaque forming units (PFU)/mL) for murine norovirus-1 (MNV-1), a surrogate of HuNoVs. Viruses at high (8.09 log10 PFU/mL) or low (4.38 log10 PFU/mL) counts were mixed with 100, 250, and 500 µg/mL of HP, HWE or HEE and incubated for 30 min at room temperature. H. cordata polysaccharide (HP) was more effective than HEE in reducing MNV-1 plaque formation, but less effective than HWE. When MNV-1 was treated with 500 µg/mL HP, the infectivity of MNV-1 decreased to an undetectable level. The selectivity indexes of each sample were 1.95 for HEE, 5.74 for HP, and 16.14 for HWE. The results of decimal reduction time and transmission electron microscopic revealed that HP has anti-viral effects by deforming and inflating virus particles, thereby inhibiting the penetration of viruses in target cells. These findings suggest that HP might have potential as an antiviral agent in the treatment of viral diseases.

Evaluation of an Extraction Method for the Detection of GI and GII Noroviruses in Fruit and Vegetable Salads.

Human norovirus (HuNoV) is a major foodborne virus causing gastroenteritis outbreaks in humans. Salad products can be vectors of transmission for foodborne viruses such as HuNoV when these products are contaminated naturally or through unsanitary food handling. Therefore, development of simple, reliable and sensitive techniques for the detection of HuNoV in salad products is needed to ensure food safety. The purpose of our study was to optimize a method for the detection of HuNoV in artificially contaminated salad products. To this end, 2 different kinds of salads (fruit salads and vegetable salads) were experimentally inoculated with HuNoV GI, HuNoV GII, and MS2 suspensions. The selected method was based on treatment with pectinase followed by Trizol-chloroform purification, and the recovery efficiencies were 6.07% to 26.52% for HuNoV GI and 5.54% to 37.36% for HuNoV GII. MS2 was used as the process control, and the recovery efficiencies for fruit salad and vegetable salad samples were 38.57% and 41.13%, respectively. The optimized method could be applied in diagnostic laboratories to identify NoV contamination in composite foods, such as salad products, should an event of foodborne outbreak occur.

Antiviral activity of phenanthrenes from the medicinal plant Bletilla striata against influenza A virus.

BACKGROUND: Influenza represents a serious public health concern. The emergence of resistance to anti-influenza drugs underlines the need to develop new drugs. This study aimed to evaluate the anti-influenza viral activity and possible mechanisms of 12 phenanthrenes from the medicinal plant Bletilla striata (Orchidaceae family). METHODS: Twelve phenanthrenes were isolated and identified from B. striata. Influenza virus A/Sydney/5/97 (H3N2) propagated in embryonated chicken eggs was used. Phenanthrenes mixed with the virus were incubated at 37 °C for 1 h and then inoculated into 9-day-old embryonated chicken eggs via the allantoic route to survey the antiviral activity in vivo. A (3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium) (MTS)-based assay was performed to evaluate the reduction of cytopathic effect induced by H3N2 on Madin-Darby canine kidney (MDCK) cells. The hemagglutination inhibition assay was used to study the blockage of virus receptors by the phenanthrenes, and the neuraminidase (NA) inhibition assay to evaluate the effects of the release of virus. The synthesis of influenza viral matrix protein mRNA in response to compound treatment was measured by real-time polymerase chain reaction. RESULTS: This study showed that phenanthrenes 1, 2, 3, 4, 6, 9, 10, 11, and 12 significantly inhibited the viruses in vivo, with inhibition rates of 20.7, 79.3, 17.2, 34.5, 34.5, 34.5, 44.8, 75.9, and 34.5%, respectively. In MDCK models, the phenanthrenes did not show significant antiviral activity when administered as pretreatment, while phenanthrenes 2, 3, 4, 6, 7 10, and 11 exhibited inhibitory activities as simultaneous treatment with 50% inhibition concentration (IC50) ranging from 14.6 ± 2.4 to 43.3 ± 5.3 µM. The IC50 ranged from 18.4 ± 3.1 to 42.3 ± 3.9 µM in the post-treatment assays. Compounds 1, 3, 4, 6, 10, and 11 exhibited an inhibitory effect on NA; and compounds 2, 3, 4 6, 7, 10, and 11 resulted in the reduced transcription of virus matrix protein mRNA. However, no compound could inhibit hemagglutination by the influenza virus. CONCLUSION: Phenanthrenes from B. striata had strong anti-influenza viral activity in both embryonated eggs and MDCK models, and diphenanthrenes seemed to have stronger inhibition activity compared with monophenanthrenes.

Bacterial community radial-spatial distribution in biofilms along pipe wall in chlorinated drinking water distribution system of East China.

Biofilms in the pipe wall may lead to water quality deterioration and biological instability in drinking water distribution systems (DWDSs). In this study, bacterial community radial-spatial distribution in biofilms along the pipe wall in a chlorinated DWDS of East China was investigated. Three pipes of large diameter (300, 600, and 600 mm) were sampled in this DWDS, including a ductile cast iron pipe (DCIP) with pipe age of 11 years and two gray cast iron pipes (GCIP) with pipe ages of 17 and 19 years, and biofilms in the upper, middle, and lower parts of each pipe wall were collected. Real-time quantitative polymerase chain reaction (qPCR) and culture-based method were used to quantify bacteria. 454 pyrosequencing was used for bacterial community analysis. The results showed that the biofilm density and total solid (TS) and volatile solid (VS) contents increased gradually from the top to the bottom along the pipe wall. Microorganisms were concentrated in the upper and lower parts of the pipe wall, together accounting for more than 80 % of the total biomass in the biofilms. The bacterial communities in biofilms were significantly different in different areas of the pipe wall and had no strong interaction. Compared with the upper and lower parts of the pipe wall, the bacterial community in the middle of the pipe wall was distributed evenly and had the highest diversity. The 16S rRNA genes of various possible pathogens, including Escherichia coli, Staphylococcus epidermidis, Pseudomonas aeruginosa, and Salmonella enterica, were detected in the biofilms, and the abundances of these possible pathogens were highest in the middle of the pipe wall among three areas. The detachment of the biofilms is the main reason for the deterioration of the water quality in DWDSs. The results of this study suggest that the biofilms in the middle of the pipe wall have highly potential risk for drinking water safety, which provides new ideas for the study of the microbial ecology in DWDS.