The ribosomal transcription units of five echinostomes and their taxonomic implications for the suborder Echinostomata (Trematoda: Platyhelminthes).

Five newly obtained nuclear ribosomal transcription unit (rTU) sequences from Echinostomatidae and Echinochasmidae are presented. The inter- and intrafamilial relationships of these and other families in the suborder Echinostomata are also analyzed. The sequences obtained are the complete rTU of Artyfechinostomum malayanum (9,499 bp), the near-complete rTU of Hypoderaeum conoideum (8,076 bp), and the coding regions (from 5'-terminus of 18S to 3'-terminus of 28S rRNA gene) in Echinostoma revolutum (6,856 bp), Echinostoma miyagawai (6,854 bp), and Echinochasmus japonicus (7,150 bp). Except for the longer first internal transcribed spacer (ITS1) in Echinochasmus japonicus, all genes and spacers were almost identical in length. Comprehensive maximum-likelihood phylogenies were constructed using the PhyML software package. The datasets were either the concatenated 28S + 18S rDNA sequences (5.7-5.8 kb) from 60 complete rTUs of 19 families or complete 28S sequences only (about 3.8-3.9 kb) from 70 strains or species of 22 families. The phylogenetic trees confirmed Echinostomatoidea as monophyletic. Furthermore, a detailed phylogeny constructed from alignments of 169 28S D1-D3 rDNA sequences (1.1-1.3 kb) from 98 species of 50 genera of 10 families, including 154 echinostomatoid sequences (85 species/42 genera), clearly indicated known generic relationships within Echinostomatidae and Echinochasmidae and relationships of families within Echinostomata and several other suborders. Within Echinostomatidae, Echinostoma, Artyfechinostomum, and Hypoderaeum appeared as monophyletic, while Echinochasmus (Echinochasmidae) was polyphyletic. The Echinochasmidae are a sister group to the Psilostomidae. The datasets provided here will be useful for taxonomic reappraisal as well as studies of evolutionary and population genetics in the superfamily Echinostomatoidea, the sole superfamily in the suborder Echinostomata.

Heterologously expressed SacP23, a novel bacteriocin from Paenibacillus polymyxa #23, is active against methicillin resistant Staphylococcus aureus.

Antimicrobial peptides have the potential to be used in a range of applications, including as an alternative to conventional antibiotics for the treatment of bacterial infections of humans and animals. Therefore, there is interest in identifying novel bacteriocins which have desirable physico-chemical properties or antimicrobial activities. Paenibacillus polymyxa #23, isolated from a marine sponge, has wide spectrum antimicrobial activity against Gram-negative and Gram-positive bacteria. To explore the basis of this antimicrobial activity, the complete genome sequence of the strain was examined. Multiple genes predicted to encode antimicrobial peptides were identified. One gene was predicted to encode a novel sactipeptide bacteriocin, named SacP23. To confirm that SacP23 does have antimicrobial activity and to explore the antimicrobial spectrum of the peptide it was heterologously expressed in Bacillus subtilis. A cluster of eight genes, encoding a full complement of accessory genes as well as the structural gene expressed from the native promoter, was cloned into B. subtilis BS54A. The recombinant strain displayed antimicrobial activity against several Gram-positive bacteria, including multi-drug resistant Staphylococcus aureus. Heterologous expression of a whole gene cluster offers a powerful way to interrogate and resolve the various antimicrobial activities expressed by native strains that encode multiple compounds of interest.

Current Status and Future Trends in Removal, Control, and Mitigation of Algae Food Safety Risks for Human Consumption.

With the rapid development of the economy and productivity, an increasing number of citizens are not only concerned about the nutritional value of algae as a potential new food resource but are also, in particular, paying more attention to the safety of its consumption. Many studies and reports pointed out that analyzing and solving seaweed food safety issues requires holistic and systematic consideration. The three main factors that have been found to affect the food safety of algal are physical, chemical, and microbiological hazards. At the same time, although food safety awareness among food producers and consumers has increased, foodborne diseases caused by algal food safety incidents occur frequently. It threatens the health and lives of consumers and may cause irreversible harm if treatment is not done promptly. A series of studies have also proved the idea that microbial contamination of algae is the main cause of this problem. Therefore, the rapid and efficient detection of toxic and pathogenic microbial contamination in algal products is an urgent issue that needs to be addressed. At the same time, two other factors, such as physical and chemical hazards, cannot be ignored. Nowadays, the detection techniques are mainly focused on three major hazards in traditional methods. However, especially for food microorganisms, the use of traditional microbiological control techniques is time-consuming and has limitations in terms of accuracy. In recent years, these two evaluations of microbial foodborne pathogens monitoring in the farm-to-table chain have shown more importance, especially during the COVID-19 pandemic. Meanwhile, there are also many new developments in the monitoring of heavy metals, algal toxins, and other pollutants. In the future, algal food safety risk assessment will not only focus on convenient, rapid, low-cost and high-accuracy detection but also be connected with some novel technologies, such as the Internet of Things (artificial intelligence, machine learning), biosensor, and molecular biology, to reach the purpose of simultaneous detection.

Molecular analysis reveals a distinct subgenogroup of porcine epidemic diarrhea virus in northern Vietnam in 2018-2019.

The spike protein (S) of porcine epidemic diarrhea virus (PEDV), in particular, the C-terminal domain of the S1 subunit (S1-CTD), which contains the conserved CO-26K-equivalent (COE) region (aa 499-638), which is recognized by neutralizing antibodies, exhibits a high degree of genetic and antigenic diversity. We analyzed 61 PEDV S1-CTD sequences (630 nt), including 26 from samples collected from seven provinces in northern Vietnam from 2018 to 2019 and 35 other sequences, representing the G1a and 1b, G2a and 2b, and recombinant (G1c) genotypes and vaccines. The majority (73.1%) of the strains (19/26) belonged to subgroup G2b. In a phylogenetic analysis, seven strains were clustered into an independent, distinct subgenogroup named dsG with strong nodal support (98%), separate from both G1a and G1b as well as G2a, 2b, and G1c. Sequence analysis revealed distinct changes (513T>S, 520G>D, 527V>(L/M), 591L>F, 669A>(S/P), and 691V>I) in the COE and S1D regions that were only identified in these Vietnamese strains. This cluster is a new antigenic variant subgroup, and further studies are required to investigate the antigenicity of these variants. The results of this study demonstrated the continuous evolution in the S1 region of Vietnamese PEDV strains, which emphasizes the need for frequent updates of vaccines for effective protection.

Comparison of the gut microbiome of sacbrood virus-resistant and -susceptible Apis cerana from South Korea.

Honey bees are important pollinators for the conservation of the ecosystem and agricultural products and provide a variety of products important for human use, such as honey, pollen, and royal jelly. Sacbrood disease (SD) is a devastating viral disease in Apis cerana; an effective preventive measure for SD is urgently needed. In this study, the relationship between the gut microbiome of honey bees and SD was investigated by pyrosequencing. Results revealed that sacbrood virus (SBV)-resistant A. cerana strains harbour a unique acetic acid bacterium, Bombella intestini, and the lactic acid bacteria (LAB) Lactobacillus (unclassified)_uc, Bifidobacterium longum, B. catenulatum, Lactococcus lactis, and Leuconostoc mesenteroides in larvae and Hafnia alvei, B. indicum, and the LAB L. mellifer and Lactobacillus HM215046_s in adult bees. Changes in the gut microbiome due to SBV infection resulted in loss of bacteria that could affect host nutrients and inhibit honey bee pathogens, such as Gilliamella JFON_s, Gilliamella_uc, Pseudomonas putida, and L. kunkeei in A. cerana larvae and Frischella_uc, Pantoea agglomerans, Snodgrassella_uc, and B. asteroides in adult bees. These findings provide important information for the selection of probiotics for A. cerana larvae and adults to prevent pathogenic infections and keep honey bees healthy.

Advanced materials for immobilization of purple phototrophic bacteria in bioremediation of oil-polluted wastewater.

Oil pollution which results from industrial activities, especially oil and gas industry, has become a serious issue. Cinder beats (CB), coconut fiber (CF) and polyurethane foam (PUF) are promising immobilization carriers for crude oil biodegradation because they are inexpensive, nontoxic, and non-polluting. The present investigation was aimed to evaluate this advanced technology and compare the efficiency of these immobilization carriers on supporting purple phototrophic bacterial (PPB) strains in hydrocarbon biodegradation of crude oil contaminated seawater. The surface of these biocarriers was supplemented with crude oil polluted seawater and immobilized by PPB strains, Rhodopseudomonas sp. DD4, DQ41 and FO2. Through scanning electron microscopy (SEM), the bacterial cells were shown to colonize and attach strongly to these biocarriers. The bacteria-driven carrier systems degraded over 84.2% supplemented single polycyclic aromatic hydrocarbons (PAHs). The aliphatic and aromatic components in crude oil that treated with carrier-immobilized consortia were degraded remarkably after 14 day-incubation. Among the three biocarriers, removal of the crude oil by CF-bacteria system was the highest (nearly 100%), followed by PUF-bacteria (89.5%) and CB-bacteria (86.3%) with the initial crude oil concentration was 20 g/L. Efficiency of crude oil removal by CB-bacteria and PUF-bacteria were 86.3 and 89.5%, respectively. Till now, the studies on crude oil degradation by mixture species biofilms formed by PPB on different carriers are limited. The present study showed that the biocarriers of an oil-degrading consortium could be made up of waste materials that are cheap and eco-friendly as well as augment the biodegradation of oil-contaminated seawater.

Broad spectrum antimicrobial activities from spore-forming bacteria isolated from the Vietnam Sea.

The widespread occurrence of pathogenic bacteria resistant to last-line antibiotics has resulted in significant challenges in human and veterinary medicine. There is an urgent need for new antimicrobial agents that can be used to control these life threating pathogens. We report the identification of antimicrobial activities, against a broad range of bacterial pathogens, from a collection of marine-derived spore-forming bacteria. Although marine environments have been previously investigated as sources of novel antibiotics, studies on such environments are still limited and there remain opportunities for further discoveries and this study has used resources derived from an under-exploited region, the Vietnam Sea. Antimicrobial activity was assessed against a panel of Gram-positive and Gram-negative bacteria, including several multi-drug resistant pathogens. From a total of 489 isolates, 16.4% had antimicrobial activity. Of 23 shortlisted isolates with the greatest antimicrobial activity, 22 were Bacillus spp. isolates and one was a Paenibacillus polymyxa isolate. Most of the antimicrobial compounds were sensitive to proteases, indicating that they were proteins rather than secondary metabolites. The study demonstrated that marine bacteria derived from the Vietnam Sea represent a rich resource, producing antimicrobial compounds with activity against a broad range of clinically relevant bacterial pathogens, including important antibiotic resistant pathogens. Several isolates were identified that have particularly broad range activities and produce antimicrobial compounds that may have value for future drug development.

Investigation of the gut microbiome of Apis cerana honeybees from Vietnam.

OBJECTIVES: In this study, the gut microbiome of healthy adult honeybees, Apis cerana, was investigated by sequencing the V3 - V4 region in 16S rRNA gene using Illumina Miseq platform. RESULTS: The total of 37,853 reads for 16S rRNA gene were obtained and 30,121 (79.6%) reads were valid with 25,291 (84.0%) reads that were classified into 116 species belonging to four major phyla. The relative abundances of the bacterial isolates in honeybee samples were phylum Proteobacteria (70.7%), Actinobacteria (10.7%), Firmicutes (10.3%), and Bacteroidetes (8.4%), respectively. Lactic acid bacteria comprised 18.95% with 10 groups including Bifidobacterium asteroides, B. indicum, Fructobacillus fructosus, Lactobacillus apinorum, L. apis, L. helsingborgensis, L. kimbladii, L. kullabergensis, and L. kunkeei. CONCLUSIONS: The presence of beneficial bacteria in the gut highlighted their role in the honeybee and suggested that they can be promising candidates for the development of probiotics for health improvement, infection control and disease management of honeybees.

Improved genomic resources for the black tiger prawn (Penaeus monodon).

World production of farmed crustaceans was 7.8 million tons in 2016. While only making up approximately 10% of world aquaculture production, crustaceans are generally high-value species and can earn significant export income for producing countries. Viet Nam is a major seafood producing country earning USD 7.3 billion in 2016 in export income with shrimp as a major commodity. However, there is a general lack of genomic resources available for shrimp species, which is challenging to obtain due to the need to deal with large repetitive genomes, which characterize many decapod crustaceans. The first tiger prawn (P. monodon) genome assembly was assembled in 2016 using the standard Illumina PCR-based pair-end reads and a computationally-efficient but relatively suboptimal assembler, SOAPdenovo v2. As a result, the current P. monodon draft genome is highly fragmented (> 2 million scaffolds with N50 length of <1000 bp), exhibiting only moderate genome completeness (< 35% BUSCO complete single-copy genes). We sought to improve upon the recently published P. monodon genome assembly and completeness by generating Illumina PCR-free pair-end sequencing reads to eliminate genomic gaps associated with PCR-bias and performing de novo assembly using the updated MaSurCA de novo assembler. Furthermore, we scaffolded the assembly with low coverage Nanopore long reads and several recently published deep Illumina transcriptome paired-end sequencing data, producing a final genome assembly of 1.6 Gbp (1,211,364 scaffolds; N50 length of 1982 bp) with an Arthropod BUSCO completeness of 96.8%. Compared to the previously published P. monodon genome assembly from China (NCBI Accession Code: NIUS01), this represents an almost 20% increase in the overall BUSCO genome completeness that now consists of more than 90% of Arthropod BUSCO single-copy genes. The revised P. monodon genome assembly (NCBI Accession Code: VIGR01) will be a valuable resource to support ongoing functional genomics and molecular-based breeding studies in Vietnam.

Enhanced Degradation of Diesel Oil by Using Biofilms Formed by Indigenous Purple Photosynthetic Bacteria from Oil-Contaminated Coasts of Vietnam on Different Carriers.

Oil pollution in marine environment caused by oil spillage has been a main threat to the ecosystem including the ocean life and to the human being. In this research, three indigenous purple photosynthetic strains Rhodopseudomonas sp. DD4, DQ41, and FO2 were isolated from oil-contaminated coastal zones in Vietnam. The cells of these strains were immobilized on different carriers including cinder beads (CB), coconut fiber (CF), and polyurethane foam (PUF) for diesel oil removal from artificial seawater. The mixed biofilm formed by using CB, CF, and PUF as immobilization supports degraded 90, 91, and 95% of diesel oil (DO) with the initial concentration of 17.2 g/L, respectively, after 14 days of incubation. The adsorption of DO on different systems was accountable for the removal of 12-16% hydrocarbons for different carriers. To the best of our knowledge, this is the first report on diesel oil degradation by purple photosynthetic bacterial biofilms on different carriers. Moreover, using carriers attaching purple photosynthetic bacteria to remove diesel oil in large scale is considered as an essential method for the improvement of a cost-effective and efficient bioremediation manner. This study can be a promising approach to eliminate DO from oil-contaminated seawater.

Isolation of endophytic fungi and screening of Huperzine A-producing fungus from Huperzia serrata in Vietnam.

Huperzine A (HupA), a natural Lycopodium alkaloid derived from Huperzia serrata (Thunb. ex Murray) Trev. plants, is a highly active acetylcholinesterase inhibitor and a key compound used for treating Alzheimer's disease (AD). Recently, HupA has been reported in various endophytic fungi isolated from H. serrata. In the present study, 153 endophytic fungi were isolated from healthy tissues of H. serrata collected from natural populations in Lam Dong province of Central Vietnam. The endophytic fungi were identified based on morphological characteristics and Internal Transcribed Spacer sequences. Among them, 34 strains were classified into seven genera belonging to Ascomycota, including Alternaria, Fusarium, Trichoderma, Penicillium, Paecilomyces, and Phoma, and eight strains belonging to the genus Mucor (Zygomycota). The other strains remained unidentified. According to the results of thin-layer chromatography and high-performance liquid chromatography, only one of the 153 strains, Penicillium sp. LDL4.4, could produce HupA, with a yield 1.38 mg l-1 (168.9 µg g-1 dried mycelium) when cultured in potato dextrose broth, which was considerably higher than that of other reported endophytic fungi. Such a fungus is a promising candidate and alternative to presently available HupA production techniques for treating AD and preventing further memory decline.

Whole-genome sequencing and characterization of an antibiotic resistant Neisseria meningitidis B isolate from a military unit in Vietnam.

BACKGROUND: Invasive meningococcal disease (IMD) persists in military units in Vietnam despite the availability of antibiotics and vaccines. A hindrance to reducing the incidence of IMD in Vietnam is a lack of molecular data from isolates of the causative agent, Neisseria meningitidis from this country. Here, we characterized key genetic and epidemiological features of an invasive N. meningitidis isolate from a military unit in Vietnam using whole-genome sequencing. METHODS: Neisseria meningitidis was isolated from a conscript admitted for meningitis and tested against seven antibiotics. DNA from the isolate was extracted and sequenced using the Illumina HiSeq platform. Denovo assembly and scaffolding were performed to construct a draft genome assembly, from which genes were predicted and functionally annotated. Genome analysis included epidemiological characterization, genomic composition and identification of antibiotic resistance genes. RESULTS: Susceptibility testing of the isolate showed high levels of resistance to chloramphenicol and diminished susceptibility to ampicillin and rifampicin. A draft genome of ~ 2.1 Mb was assembled, containing 2451 protein coding sequences, 49 tRNAs and 3 rRNAs. Fifteen coding sequences sharing ≥ 84% identity with known antibiotic resistance genes were identified. Genome analysis revealed abundant repetitive DNAs and two prophages. Epidemiological typing revealed newly described sequence type, antigenic finetype and Bexsero® Antigen Sequence Typing (BAST). The BAST profile showed no coverage by either Bexsero® or Trumenba®. CONCLUSIONS: Our results present the first genome assembly of an invasive N. meningitidis isolate from a military unit in Vietnam. This study illustrates the usefulness of whole genome sequencing (WGS) analysis for epidemiological and antibiotic resistance studies and surveillance of IMD in Vietnam.

Integration Process for Protein Extraction from Microalgae Using Liquid Biphasic Electric Flotation (LBEF) System.

Microalgae are the most promising sources of protein, which have high potential due to their high-value protein content. Conventional methods of protein harnessing required multiple steps, and they are generally complex, time consuming, and expensive. Currently, the study of integration methods for microalgae cell disruption and protein recovery process as a single-step process is attracting considerable interest. This study aims to investigate the novel approach of integration method of electrolysis and liquid biphasic flotation for protein extraction from wet biomass of Chlorella sorokiniana CY-1 and obtaining the optimal operating conditions for the protein extraction. The optimized conditions were found at 60% (v/v) of 1-propanol as top phase, 250 g/L of dipotassium hydrogen phosphate as bottom phase, crude microalgae loading of 0.1 g, air flowrate of 150 cc/min, flotation time of 10 min, voltage of 20 V and electrode's tip touching the top phase of LBEF. The protein recovery and separation efficiency after optimization were 23.4106 ± 1.2514% and 173.0870 ± 4.4752%, respectively. Comparison for LBEF with and without the aid of electric supply was also conducted, and it was found that with the aid of electrolysis, the protein recovery and separation efficiency increased compared to the LBEF without electrolysis. This novel approach minimizes the steps for overall protein recovery from microalgae, time consumption, and cost of operation, which is beneficial in bioprocessing industry.

Escherichia coli modular coculture system for resveratrol glucosides production.

In bio-based fermentation, the overall bioprocess efficiency is significantly affected by the metabolic burden associated with the expression of complete biosynthetic pathway as well as precursor and cofactor generating enzymes into a single microbial cell. To attenuate such burden by compartmentalizing the enzyme expression, recently synthetic biologists have used coculture or poly-culture techniques for biomolecules synthesis. In this paper, coculture system of two metabolically engineered Escherichia coli populations were employed which comprises upstream module expressing two enzymes converting para-coumaric acid into resveratrol and the downstream module expressing glucosyltransferase to convert the resveratrol into its glucosidated forms; polydatin and resveratroloside. Upon optimization of the initial inoculum ratio of two E. coli populations, 92 mg resveratrol glucosides/L (236 µM) was produced i.e. achieving 84% bioconversion from 280 µM of p-coumaric acid in 60 h by 3 L fed batch fermentor. This is the report of applying coculture system to produce resveratrol glucosides by expressing the aglycone formation pathway and sugar dependent pathway into two different cells.

Peptide Fraction pOh2 Exerts Antiadipogenic Activity through Inhibition of C/EBP-α and PPAR-γ Expression in 3T3-L1 Adipocytes.

Many studies have comprehensively examined the venom of Ophiophagus hannah snake. Its venom comprises different compounds exhibiting a wide range of pharmacological activities. In this investigation, four peptide fractions (PFs), ranging from 3 kDa to 10 kDa, isolated from the Vietnamese snake venom of O. hannah were separated by HPLC and investigated for their inhibitory activity on adipogenesis in 3T3-L1 adipocytes. The most effective PF was then further purified, generating two peptides, pOh1 and pOh2. Upon investigation of these two peptides on 3T3-L1 adipocytes, it was revealed that, at 10 µg/mL, pOh2 was able to inhibit the lipid accumulation in 3T3-L1 adipocytes by up to 56%, without affecting cell viability. Furthermore, the pOh2 downregulated the gene expression of important transcription factors C/EBP-α and PPAR-γ. In addition, aP2 and GPDH adipocyte-specific markers were also significantly reduced compared to untreated differentiated cells. Taken together, pOh2 inhibited the expression of key transcription factors C/EBP-α and PPAR-γ and their target genes, aP2 and GPDH, thereby blocking the adipocyte differentiation. In conclusion, this novel class of peptide might have potential for in vivo antiobesity effects.

Molecular characterization and phylogenetic analysis of deformed wing viruses isolated from South Korea.

Deformed wing virus (DWV) is one of the most common viral infection in honeybees. Phylogenetic trees were constructed for 16 partial nucleotide sequences of the structural polyprotein region and the RNA helicase region of South Korean DWVs. The sequences were compared with 10 previously reported DWV sequences from different countries and the sequences of two closely related viruses, Kakugo virus (KGV) and Varroa destructor virus-1 (VDV-1). The phylogeny based on these two regions, the Korean DWV genomes were highly conserved with 95-100% identity, while they also shared 93-97% similarity with genotypes from other countries, although they formed a separate cluster. To investigate this phenomenon in more detail, the complete DWV genome sequences of Korea-1 and Korea-2 were determined and aligned with six previously reported complete DWV genome sequences from different countries, as well as KGV and VDV-1, and a phylogenetic tree was constructed. The two Korean DWVs shared 96.4% similarity. Interestingly, the Korea-2 genome was more similar to the USA (96.5%) genome than the Korea-1. The Korean genotypes highly conserved with USA (96%) but low similarity with the United Kingdom3 (UK3) genome (89%). The end of the 5' untranslated region (UTR), the start of the open reading frame (ORF) region, and the 3' UTR were variable and contained several substitutions/transitions. This phenomenon may be explained by intramolecular recombination between the Korean and other DWV genotypes.

Seroprevalence of Toxoplasma gondii and Trichinella spiralis infections in wild boars (Sus scrofa) in Korea.

Toxoplasma gondii and Trichinella spiralis are important zoonotic pathogens with worldwide distributions. In Korea, several outbreaks of human toxoplasmosis and trichinellosis due to the consumption of infected wild animals have been reported. The purpose of this study was to determine the seroprevalence of T. gondii and T. spiralis infections in wild boars killed in Korea from December 2009 to October 2011. A total of 521 wild boars hunted in eight provinces were examined for antibodies to T. gondii and T. spiralis by using commercial ELISA kits. Overall, 25.1% of serum samples from individual boars were seropositive for T. gondii and 1.7% were seropositive for T. spiralis. Seropositive for T. gondii was found in the boars in all the eight provinces investigated and for T. spiralis in four provinces. This is the first report on the seroprevalence of T. gondii and T. spiralis infections in wild boars in Korea. The consumption of undercooked wild boar meat may expose humans to a high risk of infection.

Molecular characterization of a H5N1 highly pathogenic avian influenza virus clade 2.3.2.1b circulating in Vietnam in 2011.

The emergence of highly pathogenic avian influenza (HPAI) H5N1 viruses and the risk of a human pandemic have highlighted the need for advance stockpiling of vaccine. Current vaccines may be sub-optimally matched to the actual pandemic virus due to the rapid dissemination and ongoing evolution of avian H5N1 viruses. We report here the evaluation of efficacy of NIBRG-14 vaccine (clade 1 A/Vietnam/1194/2004) against the H5N1 HPAI virus strains circulating in Vietnam. The birds were either vaccinated with a single or booster dose of vaccine by subcutaneous injection; then challenged with three H5N1 HPAI viruses (clade 1, clade 2.3.2.1a and clade 2.3.2.1b) at day 21 post-vaccination (p. v.). The results showed that NIBRG-14 vaccine protected birds from clade 1 and clade 2.3.2.1a infections. Notably, we observed that NIGRG-14 vaccine did not confer protection against clade 2.3.2.1b challenge virus. To get new insights of how H5N1 clade 2.3.2.1b (A/Duck/QuangNgai/1037/11) virus can escape from the host immune response induced by the vaccine, we further analyzed the HA gene - a key virulence determinant of the virus. Amino acid sequence analysis indicated that this virus contained the sequence SPQRERRRK-R/G at the cleavage site in the HA molecule, indicating its high virulence. Additionally, we identified numerous mutations with amino acid substitutions in the hemagglutinin: M226I, I239S located at N-link glycosylation site and 2H, 45N, 53K 120D, 133A and 14N mutations at antigenic site, which can affect receptor specificity as well as viral pathogenicity. Notably, I239S and S133A mutations are unique to A/Duck/QuangNgai/1037/2011, suggesting that it may involve in the virus' ability to evade the host immune system. Taken together, phylogenetic analysis showed that continual mutations in the HA gene may have generated a novel antigenic strain and that probably changed the virulence of the virus and made the H5N1 clade 2.3.2.1b resistant to NIBRG-14 vaccine.

Phylogenetic analysis of black queen cell virus genotypes in South Korea.

The black queen cell virus (BQCV), a picorna-like honeybee virus, was first isolated from queen larvae and pupae of honeybees found dead in their cells. BQCV is the most common cause of death in queen larvae. Phylogenetic analysis of two Apis cerana and three Apis mellifera BQCV genotypes collected from honeybee colonies in different regions of South Korea, central European BQCV genotypes, and a South African BQCV reference genotype was performed on a partial helicase enzyme coding region (ORF1) and a partial structural polypeptide coding region (ORF2). The phylogeny based on the ORF2 region showed clustering of all the Korean genotypes corresponding to their geographic origin, with the exception of Korean Am str3 which showed more similarity to the central European and the South African reference genotype. However, the ORF1-based tree exhibited a different distribution of the Korean strains, in which A. cerana isolates formed one cluster and all A. mellifera isolates formed a separate cluster. The RT-PCR assay described in this study is a sensitive and reliable method for the detection and classification of BQCV strains from various regions of Korea. BQCV infection is present in both A. cerana and A. mellifera colonies. With this in mind, the present study examined the transmission of honeybee BQCV infections between A. cerana and A. mellifera.

Novel interactions between NFATc1 (Nuclear Factor of Activated T cells c1) and STAT-3 (Signal Transducer and Activator of Transcription-3) mediate G protein-coupled receptor agonist, thrombin-induced biphasic expression of cyclin D1, with first phase influencing cell migration and second phase directing cell proliferation.

Thrombin, a G protein-coupled receptor agonist, induced a biphasic expression of cyclin D1 in primary vascular smooth muscle cells. Although both phases of cyclin D1 expression require binding of the newly identified cooperative complex, NFATc1·STAT-3, to its promoter, the second phase, which is more robust, depends on NFATc1-mediated recruitment of p300 onto the complex and the subsequent acetylation of STAT-3. In addition, STAT-3 is tyrosine-phosphorylated in a biphasic manner, and the late phase requires NFATc1-mediated p300-dependent acetylation. Furthermore, interference with acetylation of STAT-3 by overexpression of acetylation null STAT-3 mutant led to the loss of the late phase of cyclin D1 expression. EMSA analysis and reporter gene assays revealed that NFATc1·STAT-3 complex binding to the cyclin D1 promoter led to an enhanceosome formation and facilitated cyclin D1 expression. In the early phase of its expression, cyclin D1 is localized mostly in the cytoplasm and influenced cell migration. However, during the late and robust phase of its expression, cyclin D1 is translocated to the nucleus and directed cell proliferation. Together, these results demonstrate for the first time that the dual function of cyclin D1 in cell migration and proliferation is temperospatially separated by its biphasic expression, which is mediated by cooperative interactions between NFATc1 and STAT-3.