Abstract: To investigate the distribution of avian influenza virus in environmental samples from live poultry markets (LPM) in China, samples were collected and tested by nucleic acid during 2009-2013 season. Each sample was tested by real-time RT PCR using flu A specific primers. If any real-time PCR was positive, the sample was inoculated into specific-pathogen-free (SPF) embryonated chicken eggs for viral isolation. The results indicated that the positive rate of nucleic acid in enviromental samples exhibited seasonality. The positive rate of nucleic acid was significantly higher in Winter and Spring. The positive rate of nucleic acid in LPM located in the south of China was higher than in northern China. Samples of Sewage for cleaning poultry and chopping board showed that higher positive rate of nucleic acid than other samples. The Subtype identification showed that H5 and H9 were main subtypes in the enviromental samples. Viral isolation indicated H5 subtypes was more than H9 subtypes between 2009 and 2013 while H9 subtypes increased in 2013. Our findings suggested the significance of public health based on LPM surveillance and provided the basis of prevention and early warning for avian flu infection human.
Animals ; China ; Feces ; virology ; Fresh Water ; virology ; Influenza A virus ; classification ; genetics ; isolation & purification ; Influenza in Birds ; virology ; Poultry ; Poultry Diseases ; virology ; Public Health ; Seasons ; Sewage ; virology

We wished to analyze the genetic characterization of echovirus 11 (Echo11) from samples of environmental sewage in Shandong Province (China). The VP1 coding region was typed as the strains were amplified. Phylogenetic analyses on the VP1 sequences from these isolates, strains isolated from AFP cases in the period 1994-2010 and others published in GenBank were conducted. From 2011 to 2012, 94 Echo11 strains were isolated from samples of environmental sewage in Jinan and Linyi City in Shandong Province. Numbers of Echo11 were seasonal and reached peaks in the summer and autumn in both cities; A- mong these isolates, nucleotide (nt) identities were 89.5%-100.0% whereas amino acid (aa) identities were 95.4%-100.0%. The nt and aa identities were 76.6%-79.7% and 90.4%-92.5% between those strains and the prototype (Gregory) strain of Echo11, respectively. All isolates from Shandong Province were the A genotype and the strains evolved very rapidly, which suggested that several transmission chains was co-circulating. We described the temporal fluctuation and genetic characterization of Echo11 isolates from surveillance of environmental sewage in Shandong Province, thereby providing important information for exploring the dynamic change and genetic variation of circulating human enteroviruses in this Province in China.
China ; Enterovirus B, Human ; classification ; genetics ; isolation & purification ; Phylogeny ; Sewage ; virology

To investigate the epidemic and evolutionary trends of enterovirus (EV) in the external environment of Yunnan Province, China, molecular typing was performed on 4 EV strains that were isolated from environmental sewage in Yunnan. The VP1 region of isolates was amplified by RT-PCR using universal enterovirus primers, and the amplified VP1 region was sequenced for GenBank BLAST search and genotype analysis. The 4 EV strains were identified as ECHO7. Their nucleotide and amino acid homologies with the VP1 sequences of 68 ECHO7 strains retrieved from GenBank were measured by Mega software analysis. Our findings showed that ECHO7 strains from environmental sewage and population samples were in different evolutionary branches. These strains showed typical geographical and temporal differences; In addition, there were different transmission chains at the same time and in the same area. ECHO7 strains isolated from sewage water and patients with acute flaccid paralysis during the same period in Yunnan belonged to different clusters and evolved at different speeds. Special concerns are needed for this problem. Continuous molecular biological surveillance of human EV in the external environment of Yunnan will provide strong support for early warning of EV diseases.
China ; Databases, Genetic ; Enterovirus ; genetics ; isolation & purification ; Evolution, Molecular ; Humans ; Molecular Epidemiology ; Sequence Analysis ; Sewage ; virology

OBJECTIVETo study the change of nucleic acid sequence and the germicidal effect of an E. coli bacteriophage with broad host range isolated from hospital sewage as well as to study the mechanism of phage host specificity and the effect of killed bacteria by phage-disinfectant to the samples from sewage water.

METHODSTo extract the nucleic acid from phage f(2) and phage with broad host range using anti-serum-carbamidine hydrochloride assay. Purity with agarose gel electrophoresis was then evaluated. Differences of nucleic acid sequence between phage f(2) and phage with broad host range with reverse transcription-polymerase chain reaction (RT-PCR) and random amplified polymorphic DNA (RAPD)-PCR were also comparing and analysed. Through observing the germicidal test of phage f(2) and phage with broad host range to samples from environment, different sterilization effects between the two phages were compared.

RESULTSAnalystic test for nucleic acid revealed that the two phages both belonged to 6000 bp, single-stranded RNA bacteriophage. Significant differences in their specificity of RAPD-PCR and RT-PCR were found during the changed of host range; with 26 RAPD-cDNA differential fragments found that in two phages RAPD-PCR products. The RT-PCR product of phage f(2) was 450 bp cDNA fragment, but the phage with broad host range did not show PCR product. Treating the sewage water with phage under broad host range, the germicidal test showed that the cleaning rate of E. coli bacteria and phage f(2) in water samples from environment could reach 36.75% - 56.28%, 30.84% - 47.96%, 19.19% - 35.06% and 13.05% - 27.85%, respectively.

CONCLUSIONThe cleaning rates to E. coli and bacteria by phage with broad host range were obviously higher than phage f(2) (P = 0.000). Analytic test for nucleic acid indicated that host-specific lytic effect of phage with broad host range had been changed at genetic level.

Bacteriophages ; genetics ; isolation & purification ; physiology ; Colony Count, Microbial ; Escherichia coli ; virology ; F Factor ; RNA Phages ; genetics ; Sewage ; microbiology ; virology ; Water Microbiology

To investigate the genetic characteristics of poliovirus isolates from environmental sewage surveillance in Shandong province, we collected sewage samples in Jinan and Linyi City. Serotyping and VP1/ 3D sequencing were performed on polioviruses isolated from the concentrated sewage samples, and VP1 mutation and recombination were analyzed. Thirty-two of sewage samples were collected, and polioviruses were detected in 10 of the samples with a positive rate of 31.3%. Eighteen Sabin strains were isolated including three type 1, nine type 2, and six type 3 polioviruses, and the number of nucleotide substitutions in VP1 coding region varied from 0 to 4. Recombination was found in three Sabin 2 and four Sabin 3 polioviruses. Analysis of neurovirulence sites of VP1 revealed that one Sabin 1 vaccine strain had a nucleotide change of A to G at nt 2749, one Sabin 2 strain had a nucleotide change of A to G at nt 2908, three Sabin 2 strains had a nucleotide change of U to C at nt 2909, and all six Sabin 3 strains had a nucleotide change of C to U at nt 2493. Poliovirus vaccine strains could be isolated from environmental sewage with a high rate of gene recombination and back mutation of neuvirulence-associated sites. None of wild-type poliovirus or vaccine-derived poliovirus was detected.
Amino Acid Sequence ; Base Sequence ; China ; Humans ; Molecular Sequence Data ; Mutation ; Poliomyelitis ; prevention & control ; virology ; Poliovirus ; genetics ; isolation & purification ; Population Surveillance ; Sewage ; virology

OBJECTIVEIn order to explore the existence of SARS coronavirus (Co-V) and/or its RNA in sewage of hospitals administered SARS patients.

METHODSA novel electropositive filter was used to concentrate the SARS-CoV from the sewage of two hospitals administered SARS patients in Beijing, including twelve 2,500 ml sewage samples from the hospitals before disinfection, and ten 25,000 ml samples after disinfection; as well as cell culture, RT-PCR and sequencing of gene to detect and identify the viruses from sewage.

RESULTSThere was no live SARS-CoV detected in the sewage in this study. The nucleic acid of SARS-CoV had been found in the 12 sewage samples before disinfection from both hospitals by semi-nested PCR. After disinfection, SARS-CoV RNA could only be detected from the samples from the 309th Hospital, and the others were negative.

CONCLUSIONIt provides evidence that there is no live SARS-Cov in the sewage from hospitals with SARS patients though SARS-CoV RNA can be detected.

Hospitals ; Humans ; Nucleocapsid ; analysis ; RNA, Viral ; analysis ; Reverse Transcriptase Polymerase Chain Reaction ; SARS Virus ; genetics ; isolation & purification ; Severe Acute Respiratory Syndrome ; virology ; Sewage ; virology

COVID-19/virology* ; Cities ; Environmental Monitoring ; Genome, Viral ; Humans ; Infection Control/methods* ; Pakistan ; RNA, Viral/genetics* ; SARS-CoV-2/genetics* ; Sewage/virology*

In order to investigate the transmission risk of H5N1 avian influenza viruses (AIV) from sewage in Changsha poultry markets, the evolution relationship and molecular characteristics of non-structural (NS) genes of H5N1 AIV from sewage were analyzed. Nine H5N1 AIV environmental sewage specimens were collected from Changsha poultry markets. The NS genes were amplifyed by PCR and then sequenced with TA cloning. Amino acid(aa) sequence alignment and phylogenetic tree analysis were conducted by Lasergene and Mega5 software. Eight NS genes TA cloning were constructed successfully. Phylogenetic tree indicated that they were belonged to the allele A subgroup. Aa homology analysis showed 90.1% 92.5% identity in NS1 proteins and 91.0% - 92.6% identity in NS2 proteins compared with reference viruses of the allele A (A/chicken/ Hubei/ w h/ 1999). The homologies of the amino sequences of NS1 and NS2 in this study were 93.8%-100.0% and 98.4%-100.0%, respectively. The C terminal of all eight H5N1 NS1 proteins from sewage in poultry markets carried a ESEV of PL motif and the 92 amino acids were E, furthermore, the 80 to 84aa were missed which were the characteristics of highly pathogenic AIV. The NS genes of H5N1 AIV from sewage in poultry markets have molecular characteristics of highly pathogenic and have the potential risk of H5N1 virus spreading.
Amino Acid Sequence ; Animals ; Influenza A Virus, H5N1 Subtype ; chemistry ; classification ; genetics ; isolation & purification ; Influenza in Birds ; transmission ; virology ; Molecular Sequence Data ; Phylogeny ; Poultry ; Sequence Homology, Amino Acid ; Sewage ; virology ; Viral Nonstructural Proteins ; chemistry ; genetics