Molecular epidemiology demonstrates that imported and local strains circulated during the 2014 dengue outbreak in Guangzhou,China

The dengue virus(DENV) is a vital global public health issue. The 2014 dengue epidemic in Guangzhou, China, caused approximately 40,000 cases of infection and five deaths. We carried out a comprehensive investigation aimed at identifying the transmission sources in this dengue epidemic. To analyze the phylogenetics of the 2014 dengue strains, the envelope(E) gene sequences from 17 viral strains isolated from 168 dengue patient serum samples were sequenced and a phylogenetic tree was reconstructed. All 17 strains were serotype Ⅰ strains, including 8genotype Ⅰ and 9 genotype V strains. Additionally, 6 genotype Ⅰ strains that were probably introduced to China from Thailand before 2009 were widely transmitted in the 2013 and 2014 epidemics, and they continued to circulate until 2015, with one affinis strain being found in Singapore. The other 2 genotype Ⅰ strains were introduced from the Malaya Peninsula in 2014. The transmission source of the 9 genotype Ⅴ strains was from Malaysia in 2014. DENVs of different serotypes and genotypes co-circulated in the 2014 dengue outbreak in Guangzhou. Moreover, not only had DENV been imported to Guangzhou, but it had also been gradually exported, as the viruses exhibited an enzootic transmission cycle in Guangzhou.     

The Asian Lineage of Zika Virus: Transmission and Evolution in Asia and the Americas

Since first isolation in 1947 from the Zika forest in Uganda, Zika virus(ZIKV) has been principally known as a benign agent associated with sporadic human infections in a restricted number of African countries. However, during 2015–2016,an Asian lineage of ZIKV caused an unprecedentedly large outbreak in the Americas and sizeable numbers of exported cases across the globe. In this review, we critically appraise the recent advances in molecular epidemiological studies of ZIKV performed to date, and we highlight the pivotal role played by genomic surveillance in elucidating the origins,dissemination and evolution of the Asian lineage of ZIKV in Asia and in the Americas.     

Molecular evolution of influenza A (H3N2) viruses circulated in Fujian Province, China during the 1996–2004 period

We studied the genetic and epidemic characteristics of influenza A (H3N2) viruses circulated in human in Fujian Province, south of China from 1996 to 2004. Phylogenetic analysis was carried out for genes encoding hemagglutinin1 (HA1) of influenza A virus (14 new and 11 previously reported reference se-quences). Our studies revealed that in the 8 flu seasons, the mutations of HA1 genes occurred from time to time, which were responsible for about four times of antigenic drift of influenza H3N2 viruses in Fujian, China. The data demonstrated that amino acid changes were limited to some key codons at or near antibody binding sites A through E on the HA1 molecule. The changes at the antibody binding site B or A or sialic acid receptor binding site 226 were critical for antigenic drift. But the antigenic sites might change and the key codons for antigenic drift might change as influenza viruses evolve. It seems important to monitor new H3 isolates for mutations in the positively selected codons of HA1 gene in south of Asia.     

Characterization of serogroup C meningococci isolated from 14 provinces of China during 1966—2005 using comparative genomic hybridization

Neisseria meningitidis is a major cause of bacterial meningitis and septicemia worldwide. In China, serogroup A strains were responsible for over 95% of the cases, while serogroup B strains were mainly the cause of localized outbreaks and sporadic cases. Before 2003, serogroup C strains were only re-covered from a few sporadic cases. However, a sudden increase in the number of cases due to sero-group C strains occurred during 2003—2005 in Anhui Province, China. Many cases were found in other provinces at the same time. Multilocus sequence typing (MLST) results indicated that the unique se-quence type 4821 clone meningococci, a new hyper-virulent lineage, was responsible for the serogroup C meningitis outbreaks. We have completed the project of sequencing the whole genome of the Chi-nese N. meningitidis serogroup C representative isolate 053442. We fabricated a whole-genome mi-croarray of N. meningitidis isolate 053442 and analyzed the genome composition differences among 81 serogroup C isolates which were isolated from 14 provinces of China during 1966—2005. The com-parative genomic hybridization (CGH) result shows that the genome compositions of nearly all sero-group C isolates are similar to that of 053442. The products of many absent open reading frames (ORFs) are conserved hypothetical proteins. The results will provide a valuable resource from which one can analyze the genome composition and genetic background of serogroup C meningococci in China.     

Differences in Variation of Human Immunodeficiency Virus Type 1 Sequences from Henan and Shanghai Regions of China

Illegally paid blood donation was a risk factor for HIV acquisition exclusively in Henan and Hubei Provinces of China,and not in Shanghai.Nucleotide sequences in the gag and env genes of HIV-1 were compared between isolates from Henan and Shanghai regions of China to test whether an expected higher degree of a common source of infections from this unique blood donation transmission risk would be evident as decreased variation among Henan isolates in an exploratory cross-sectional analysis.Among 38 isolates studied,23 of 23(100%)from Henan and 8 of 15(54%)from Shanghai were subtype B.In addition,fewer sequence differences were found in gp41 of subtype B isolates from Henan than from Shanghai isolates.Further studies with additional controls are therefore warranted to confirm the role of the degree of a common source of infections in differences in HIV variation across populations.     

Detection,isolation, and characterization of chikungunya viruses associated with the Pakistan outbreak of 2016–2017

The chikungunya virus(CHIKV) is a mosquito-transmitted alphavirus, which has infected millions of people in Africa, Asia, Americas, and Europe since it reemerged in India and Indian Ocean regions in 2005–2006. Starting in the middle of November 2016, CHIKV has been widely spread, and more than 4,000 cases of infections in humans were confirmed in Pakistan. Here, we report the first isolation and characterization of CHIKV from the Pakistan outbreak. Eight CHIKV strains were newly isolated from human serum samples using a cell culture procedure. A full-length genome sequence and eight complete envelope(E1) sequences of CHIKV from Pakistan were obtained in this study. Alignment of the CHIKV E1 sequences revealed that the eight new CHIKV isolates were highly homogeneous, with only two nonsynonymous substitutions found at generally conserved sites(E99 and Q235). Based on the comparison of 342 E1 sequences, the two nonsynonymous mutations were located in well-recognized domains associated with viral functions such as the cell fusion and vector specificity, suggesting their potential functional importance. Phylogenetic analysis indicated that the CHIKV strains from Pakistan originated from CHIKV circulating in the Indian region. This study helps elucidate the epidemics of CHIKV in Pakistan and also provides a foundation for studies of evolution and expansion of CHIKV in South Asia.     

Characterization of serogroup C meningococci isolated from 14 provinces of China during 1966—2005 using comparative genomic hybridization

Neisseria meningitidis is a major cause of bacterial meningitis and septicemia worldwide. In China, serogroup A strains were responsible for over 95% of the cases, while serogroup B strains were mainly the cause of localized outbreaks and sporadic cases. Before 2003, serogroup C strains were only re-covered from a few sporadic cases. However, a sudden increase in the number of cases due to sero-group C strains occurred during 2003—2005 in Anhui Province, China. Many cases were found in other provinces at the same time. Multilocus sequence typing (MLST) results indicated that the unique se-quence type 4821 clone meningococci, a new hyper-virulent lineage, was responsible for the serogroup C meningitis outbreaks. We have completed the project of sequencing the whole genome of the Chi-nese N. meningitidis serogroup C representative isolate 053442. We fabricated a whole-genome mi-croarray of N. meningitidis isolate 053442 and analyzed the genome composition differences among 81 serogroup C isolates which were isolated from 14 provinces of China during 1966—2005. The com-parative genomic hybridization (CGH) result shows that the genome compositions of nearly all sero-group C isolates are similar to that of 053442. The products of many absent open reading frames (ORFs) are conserved hypothetical proteins. The results will provide a valuable resource from which one can analyze the genome composition and genetic background of serogroup C meningococci in China.     

A large impact of tropical biomass burning on CO and CO2 in the upper troposphere

A large interannual variation of biomass burning emissions from Southeast Asia is asso-ciated with the ENSO events. During 1997/98 and 1994 El Nino years, uncontrolled wildfires of tropical rainforests and peat lands in Indonesia were enlarged due to a long drought. EnhancedCO injection into the upper troposphere from the intense Indonesian fires was clearly observed in the 8-year measurements from a regular flask sampling over the western Pacific using a JAL air-liner between Australia and Japan. This airliner observation also revealed that upper tropospheric CO_2 cycle largely changed during the 1997 El Nino year due partly to the biomass burning emis-sions. Widespread pollution from the biomass burnings in Southeast Asia was simulated using aCO tracer driven by a 3D global chemical transport model. This simulation indicates that tropical deep convections connected to rapid advection by the subtropical jet play a significant role in dis-persing biomass-burning emissions from Southeast Asia on a global scale.     

Clinical and biochemical indexes from 2019-nCoV infected patients linked to viral loads and lung injury

The outbreak of the 2019-nCoV infection began in December 2019 in Wuhan,Hubei province,and rapidly spread to many provinces in China as well as other countries.Here we report the epidemiological,clinical,laboratory,and radiological characteristics,as well as potential biomarkers for predicting disease severity in 2019-nCoV-infected patients in Shenzhen,China.All 12 cases of the 2019-nCoV-infected patients developed pneumonia and half of them developed acute respiratory distress syndrome (ARDS).The most common laboratory abnormalities were hypoalbuminemia,lymphopenia,decreased percentage of lymphocytes (LYM) and neutrophils (NEU),elevated C-reactive protein (CRP) and lactate dehydrogenase (LDH),and decreased CD8 count.The viral load of 2019-nCoV detected from patient respiratory tracts was positively linked to lung disease severity.ALB,LYM,LYM (%),LDH,NEU (%),and CRP were highly correlated to the acute lung injury.Age,viral load,lung injury score,and blood biochemistry indexes,albumin (ALB),CRP,LDH,LYM (%),LYM,and NEU (%),may be predictors of disease severity.Moreover,the AngiotensinⅡlevel in the plasma sample from 2019-nCoV infected patients was markedly elevated and linearly associated to viral load and lung injury.Our results suggest a number of potential diagnosis biomarkers and angiotensin receptor blocker (ARB) drugs for potential repurposing treatment of 2019-nCoV infection.     

Taxonomy is the cornerstone of biodiversity conservation-SEABRI reports on biological surveys in Southeast Asia

The Southeast Asia Biodiversity Research Institute (SEABRI) is an international scientific research and education organization affiliated directly to the Chinese Academy of Sciences (CAS) and managed by the Xishuangbanna Tropical Botanical Garden (XTBG).By harnessing its connections with all CAS institutes,local institutes and international agencies and leveraging on their resources,it seeks to make a significant contribution to biodiversity conservation in Southeast Asia.SEABRI intends to project itself as a crucial networking platform for biodiversity researchers from both China and Southeast Asia,an internationally respected research and conservation institution on Southeast Asian biodiversity,and a model on cooperation on research and education between China and Southeast Asian countries.     

SARS-CoV Genome Polymorphism： A Bioinformatics Study

A dataset of 103 SARS-CoV isolates （101 human patients and 2 palm civets） was investigated on different aspects of genome polymorphism and isolate classification. The number and the distribution of single nucleotide variations （SNVs） and insertions and deletions, with respect to a “profile”, were determined and discussed （“profile” being a sequence containing the most represented letter per position）. Distribution of substitution categories per codon positions, as well as synonymous and non-synonymous substitutions in coding regions of annotated isolates, was determined, along with amino acid （a.a.） property changes. Similar analysis was performed for the spike （S） protein in all the isolates （55 of them being predicted for the first time）. The ratio Ka/Ks confirmed that the S gene was subjected to the Darwinian selection during virus transmission from animals to humans. Isolates from the dataset were classified according to genome polymorphism and genotypes. Genome polymorphism yields to two groups, one with a small number of SNVs and another with a large number of SNVs, with up to four subgroups with respect to insertions and deletions. We identified three basic nine-locus genotypes： TTTT/TTCGG, CGCC/TTCAT, and TGCC/TTCGT, with four subgenotypes. Both classifications proposed are in accordance with the new insights into possible epidemiological spread, both in space and time.     

Molecular and in vitro Characterization of Field Isolates of Bovine Herpesvirus-1

Bovine Herpesvirus-1 (BoHV-1) is distributed worldwide and is a major pathogen in cattle, being the causal agent of a variety of clinical syndromes. The aim of this study was to isolate and to characterize (molecular and biological characterization) BoHV-1 from 29 immunosuppressed animals. It was possible to obtain 18 isolates, each from a different animal, such as from the respiratory and reproductive tracts. In some cases the cytopathic effect was visible 12 hours post-inoculation, and became characteristic after 36-48 hours. Biological characteristics were evaluated and compared with Iowa and Colorado-1 reference strains, and differences were found in plaque size, virus titer measured by TCID50 and PFU/mL, and one step virus curves. These results showed that some isolates had a highly virulent-like behavior in vitro, compared to the reference strains, with shorter eclipse periods, faster release of virus into the supernatants, and higher burst size and viral titer. There were no differences in glycoprotein expression of BoHV-1 isolates, measured by Western blot on monolayers. Moreover, using restriction endonucleases analysis, most of the viruses were confirmed as BoHV-1.1 and just one of them was confirmed as BoHV-1.2a subtype. These findings suggest that some wild-type BoHV-1 isolates could be useful as seeds to develop new monovalent vaccines.     

Invasion history of Solenopsis invicta （Hymenoptera： Formicidae） in Fujian,China based on mitochondrial DNA and its implications in development of a control strategy

One of the most invasive species worldwide, Solenopsis invicta Buren, has been described in China since 2003. Recent studies have suggested that China populations are the result of introductions from the USA; however, detailed molecular studies need to be performed in order to understand the expansion and potential multiple introductions from other countries into China. As there were populations of red imported fire ant, S. invicta in different areas and with different methods of introduction, mitochondrial cytochrome oxidase Ⅰ gene was used as a marker from 12 populations in four cities in Fujian Province, China, to determine the relationship of invasion among these populations. The three most common haplotypes previously describe in invasive populations of S. invicta： H5, H22 and H36, were found in Fujian. However, frequencies in each city were different. For instance, three populations from Longyan city which invaded with waste plastics, shared haplotype H5. Populations from Xiamen city and Jinjiang city which dispersed with nursery stock, sward and scrap leather, shared haplotype H22. The population from Nanyan village of Xinluo district, Longyan city, bore haplotype H36. Mitochondrial data reveals that the invasion history of S. invicta in Fujian Province is complex, including multiple invasions probably from other provinces within China. Security measures to prevent S. invicta spreading within China are as important as from overseas.     

Sequence Variation in the Gpl20 region of SHIV-CN97001 during in vivo Passage

SHIV-CN97001 played an important role in assessing the immune effect and strategy of the AIDS vaccine which included genes of the predominant prevalent HIV-1 strain in China. In this study, SHIV-CN97001 was in vivo passaged serially to construct pathogenic SHIV-CN97001/rhesus macaques model. To identify variation in the gp120 region of SHIV-CN97001 during passage, the fragments of gp120 gene were amplified by RT-PCR from the plasma of SHIV-CN97001 infected animals at the peak viral load time point and the gene distances （divergence, diversity） were calculated using DISTANCE. The analysis revealed that the genetic distances of SHIV-CN97001 in the third passage animals were the highest during in vivo passage. It had a relationship between viral divergence from the founder strain and viral replication ability. The nucleic acid sequence of the V3 region was highly conservative. All of the SHIV-CN97001 strains had V3 loop central motif （GPGQ） and were predicted to be using CCR5 co-receptor on the basis of the critical amino acids within V3 loop. These results show that there was no significant increase in the genetic distance during serial passage, and SHIV-CN97001 gp120 gene evolved toward ancestral states upon transmission to a new host. This could partly explain why there was no pathogenic viral strain obtained during in vivo passage.     

The variable codons of H5N1 avian influenza A virus haemagglutinin genes

We investigated the selection pressures on the haemagglutinin genes of H5N1 avian influenza viruses using fixed effects likelihood models. We found evidence of positive selection in the sequences from isolates from 1997 to 2007, except viruses from 2000. The haemagglutinin sequences of viruses from southeast Asia, Hong Kong and mainland China were the most polymorphic and had similar nonsyn-onymous profiles. Some sites were positively selected in viruses from most regions and a few of these sites displayed different amino acid patterns. Selection appeared to produce different outcomes in vi-ruses from Europe, Africa and Russia and from different host types. One position was found to be positively selected for human isolates only. Although the functions of some positively selected posi-tions are unknown, our analysis provided evidence of different temporal, spatial and host adaptations for H5N1 avian influenza viruses.     

Isolation and partial purification of fungal ligninolytic enzymes from the forest soil fungi isolated from Bhadra Wildlife Sanctuary

Screening was done for the isolation of effective lignin degraders from the forest soil samples, by providing lignin as a carbon source through the enrichment method, which leads to the isolation of 8 effective fungal isolates among 14 isolates. Submerged fermentation was done for the production of ligninolytic enzymes with the effective microorganisms by providing Guiaicol as a carbon source. The assay of laccase, lignin peroxidise activity and specific activity was done after the incubation intervals of 2, 4, 6, 7, 8, 10 and 12 days at 27±2℃ under shake culture condition. Partially purified protein content was estimated by using Lowry＇s method. Pleurotus sp. and Phanerochaetae chrysosporium are more effective at the 2nd and 7th days of incubation for the production of laccase and lignin peroxidases among the effective isolates.     

Genetic and Molecular Characterization of H9N2 Avian Influenza Viruses Isolated from Live Poultry Markets in Hubei Province,Central China, 2013–2017

H9N2 subtype avian influenza virus(AIV) is an influenza A virus that is widely spread throughout Asia, where it jeopardizes the poultry industry and provides genetic material for emerging human pathogens. To better understand the epidemicity and genetics of H9 subtype AIVs, we conducted active surveillance in live poultry markets(LPMs) in Hubei Province from 2013 to 2017. A total of 4798 samples were collected from apparent healthy poultry and environment. Realtime RT-PCR revealed that the positivity rate of influenza A was 26.6%(1275/4798), of which the H9 subtype accounted for 50.3%(641/1275) of the positive samples. Of the 132 H9N2 viral strains isolated, 48 representative strains were subjected to evolutionary analysis and genotyping. Phylogenetic analysis revealed that all H9N2 viral genes had 91.1%–100% nucleotide homology, clustered with genotype 57, and had high homology with human H9N2 viruses isolated from2013 to 2017 in China. Using a nucleotide divergence cutoff of 95%, we identified ten distinct H9N2 genotypes that continued to change over time. Molecular analysis demonstrated that six H9N2 isolates had additional potential glycosylation sites at position 218 in the hemagglutinin protein, and all isolates had I155 T and Q226 L mutations. Moreover, 44 strains had A558 V mutations in the PB2 protein and four had E627 V mutations, along with H9N2 human infection strains A/Beijing/1/2016 and A/Beijing/1/2017. These results emphasize that the H9N2 influenza virus in Hubei continues to mutate and undergo mammalian adaptation changes, indicating the necessity of strengthening the surveillance of the AIV H9N2 subtype in LPMs.