Correlation analysis of EV71 detection and case severity in hand, foot, and mouth disease in the Hunan Province of China.

An increase in the incidence of hand, foot and mouth disease (HFMD) cases has been observed in the Hunan province of mainland China since 2009 with a particularly higher level of severe cases in 2010-2012. Intestinal viruses of the picornaviridae family are responsible for the human syndrome associated with HFMD with enterovirus 71 (EV71) and Coxsackievirus A16 (Cox A16) being the most common causative strains. HFMD cases associated with EV71 are generally more severe with an increased association of morbidity and mortality. In this study, the etiology surveillance data of HFMD cases in Hunan province from March 2010 to October 2012 were analyzed to determine if there is a statistically relevant linear correlation exists between the detection rate of EV71 in mild cases and the proportion of severe cases among all HFMD patients. As the cases progressed from mild to severe to fatal, the likelihood of EV71 detection increased (25.78%, 52.20% and 84.18%, respectively). For all cases in the timeframe evaluated in this study, the presence of virus was detected in 63.21% of cases; among cases showing positivity for virus, EV71 infection accounted for 50.14%. These results provide evidence to support the observed higher morbidity and mortality associated with this outbreak and emphasizes the importance of early detection in order to implement necessary prevention measures to mitigate disease progression.

Evaluation of post-mortem heart blood culture in a Chinese population.

BACKGROUND: Post-mortem blood cultures have been used in a wide variety of research studies. However, their significance is still a matter of dispute among medico-legal experts. This study was aimed to determine the factors which influenced post-mortem blood culture results and to assess their value in determining the cause of death. METHODS: We retrospectively investigated 76 post-mortem cases with suspected infection and correlated pathological findings with heart blood culture results. RESULTS: We found that survival time after onset of illness was significantly associated with positive heart blood cultures (P=0.014). Blood culture results were not influenced by age, gender, prior antibiotic therapy, interval time from death to store or interval time from death to autopsy (P>0.05). In those who had heart blood cultures taken, 49 cases (64.5%) including four cases with mixed growth, showed positive results, and approximately one-third of blood cultures were sterile. The positive predictive value (PPV) was 77.6% (38/49) including two cases with mixed growth; most were genuine pathogens according to the clinical and pathological findings. However, the negative predictive value (NPV) was 59.3% (16/27). Escherichia coli, Streptococcus spp., Staphylococcus aureus, Streptococcus pneumoniae and Klebsiella spp. were isolated most often. CONCLUSION: These findings suggest that the results of post-mortem heart blood cultures, when combined with clinical and pathological findings, strengthen the understanding of the cause of death.

[Analysis on genetic characteristics of mumps virus strains circulating in Hunan province in 2011].

OBJECTIVE: To be acquainted with genetic characteristics and variation of mumps virus strains circulating in Hunan province. METHODS: Mumps virus (MV) strains were isolated using Vero/ SLAM cells. The small hydrophobic protein (SH) genes of MV isolates were sequenced, and the sequences were analysed phylogenetically between the isolated strains and other reference mumps strains. RESULTS: 4 mumps virus strains were isolated from 16 specimens collected in 2011 from different regions of Hunan province. The genotype of isolated strains were supposed to be F type. CONCLUSION: Genotype F is the main genotype of circulating strains in Hunan province in 2011 and there is no variation between genotype.

[Virological surveillance of pandemic (H1N1) 2009 virus and its genetic characteristics in Hunan Province, 2009-2011].

To understand and master the dynamic variation of the pandemic influenza A (H1N1) 2009 in Hunan province from 2009 to 2011, and to know the genetic characteristics and drug resistance of the pandemic (H1N1) 2009 viruses. Throat swab specimens of influenza-like illness patients were collected from sentinel hospitals and tested for influenza by fluorescent PCR or virus isolation methods. Partial isolates were selected for sequencing. The sequences were used for phylogenetic analysis by MEGA 5. 05 software. From the 20th week of 2009 to the 52nd week of 2011, 17 773 specimens were tested. 3 831 specimens were influenza-positive with a positive rate of 21. 6%, of which 1 794 were positive specimens of pandemic (H1N1) 2009, accounting for 46. 8%00 of the influenza-positives. There were 2 epidemic peaks of pandemic (H1N1) 2009, which were in the 41st-53rd week of 2009 and the 1st-12nd week of 2011, respectively. The HA genes of 23 strains that were selected for sequencing had close relationship; the distribution of strains in the phylogenetic tree was basically in chronological order. The complete genome sequence analysis showed that all of 8 gene segments of 7 strains were homologous to the vaccine strain, and there was no gene reassortment. The HA amino acid sites of the 23 strains were highly similar to the vaccine strain (98. 2% - 100. 0% in homology), but all 23 strains had P83S, S203T and 1321V mutations. The 222 site mutation that may lead to enhanced virulence was found in the A/Hunan/YQ30/2009 strain. The mutation was D222E. There was no oseltamivir resistance mutation found in all strains. The pandemic (H1N1) 2009 in Hunan province from 2009 to 2011 had a bimodal distribution. There was no large-scale variation of virus genes. The clinical use of oseltamivir was still effective. Key words: Pandemic (H1N1) 2009; Surveillance; Genetic characteristics

[Analysis of genetic features of influenza B virus in Hunan province from 2007 to 2010].

OBJECTIVE: To investigate the gene variations of influenza B virus isolated in Hunan province from 2007 to 2010. METHODS: A total of 42 strains of influenza B virus,which were isolated in the Influenza Surveillance Network Laboratories in Hunan province between year 2007 and 2010, were selected for the study. The hemagglutinin 1 (HA1) and neuraminidase (NA) genes of the selected strains were amplified by RT-PCR, and the sequence of the purified product were detected and homologically compared with the sequence of influenza vaccine strains isolated from Northern Hemisphere by WHO during the same period. In addition, the phylogenetic trees were constructed to characterize the molecular features. RESULTS: In the Victoria branch of the HA1 gene phylogenetic tree, the strains isolated from year 2007 to 2009 were included in the V1 sub-branch, as well as the vaccine strain Malaysia/2506/2004; the strains isolated in year 2010 were involved in the V2 sub-branch, similar to the vaccine strains Brisbane/60/2008. In the Yamagata branch,the strains isolated in year 2007 were in the Y1 sub-branch,different from the strains isolated between year 2008 and 2010, which were in the Y2 sub-branch, instead. All virus in NA gene phylogenetic tree were included in the Yamagata branch, indicated their Yamagata origin. The genetic sequence analysis of the 7 strains isolated in year 2010 revealed that the viruses were classified as genotype 2 and genotype 15. The results of homological comparison between HA1 molecule and the influenza vaccine strains recommended by WHO were as below: Victoria lineage, 98.6% - 99.1% in 2007, 98.6% - 99.1% in 2008, 98.1% - 99.1% in 2009, and 97.6% - 99.1% in 2010; and Yamagata lineage, 97.9% - 98.5% in 2007, 97.9% - 98.5% in 2009 and 97.9% - 98.2% in 2010. The major mutations of the strains isolated in year 2007 were found in sites R48K, K88R, P108A, D197N and S230G. While the major mutations of the strains isolated between year 2009 and 2010 were sited in K88R, S150I, N166Y, D197N and S230G. CONCLUSION: The prevalent influenza B virus isolated in Hunan province from 2007 to 2010 has mutated and evolved continuously.

[Molecular characterization of highly pathogenic avian influenza A (H5N1) viruses isolated from humans in Hunan province, 2006 - 2009].

OBJECTIVE: To understand the possible origins, genetic re-assortment and molecular characterization of 4 highly pathogenic avian influenza A (H5N1) viruses isolated from humans in Hunan province, between 2006 and 2009. METHODS: H5N1 PCR test-positive specimens were inoculated in embryonated eggs while H5N1 virus was isolated and genomes sequenced. Genome homology and genetic molecular characterization were analyzed by BLAST and MEGA 4.0. RESULTS: All gene segments of the 4 viruses were avian in origin. No re-assortment was found between avian influenza A (H5N1) viruses and human seasonal influenza viruses. Viruses that isolated from domestic poultry shared high similarity with the 4 human viruses in gene homology. Data from the whole genome phylogenetic analysis showed that the 4 viruses were in clade 2.3.4, while 2 viruses belonged to genotype V, and another 2 were new genotypes. Results from molecular characterization showed that amino acid sequences of HA cleavage site of the 4 viruses were PLRERRKR/G. All 4 viruses had A160T mutation in HA, a 20 amino acid deletion in the neuraminidase (NA) stalk at position 49 - 68, and a 5 amino acid deletion in the non-structural protein 1 (NS1). Most sites in the HA molecules showed that the viruses preferentially bound to avian influenza virus receptor. However, T192I mutation that might enhance the α 2, 6-linked sialic acid human influenza receptor binding had emerged in HN/1/09 and HN/2/09. D701N mutation of PB2 that increased the virulence in mice was found in HN/1/08. Analysis on drug resistance gene amino acid showed that all 4 viruses were sensitive to amantadine and oseltamivir. CONCLUSION: Highly pathogenic avian influenza A (H5N1) viruses isolated from humans in Hunan province from 2006 to 2009 were avian in origin, and the 4 viruses belonged to different genotypes. Some mutations that related to virulence and receptor binding positions had emerged in some of the strains.

[Serological survey of pandemic influenza A (H1N1) virus infection among population in Hunan Province].

OBJECTIVE: To understand the infection condition and analytical methods of Influenza A (H1N1) virus in the population of Hunan Province during different periods. METHODS: Quick surveys on the positive rate of Influenza A (H1N1) virus hemagglutination inhibition (HI) test have been conducted for 5 times successively from November 2009 to March 2010 in 14 medical and health institutions of Changsha city, whose results were then compared with those from the sampling surveys of whole Hunan province. RESULTS: 2131 subjects were involved in this study; the total population standardized rates of antibody positive investigated for 5 times were 9.32% , 14.62%, 31.08%, 28.43% and 22.80% respectively; the population of 6-17-years-old has the highest rate of antibody positive; only 9.84% of the antibody positive subjects attributed to vaccine inoculation; there was no significant difference in the standardized positive rates between the quick serological surveys and the corresponding sampling survey of Hunan province (P > 0.05). CONCLUSION: The positive rate of A (H1N1) virus antibody reached the peak in late January 2010; quick investigations in small region could be used to evaluate the infection prevalence during pandemic of infectious diseases.

[Surveillance on the etiology and genetic characteristics of rabies in Hunan province, from 2008 to 2009].

OBJECTIVE: To analyze the etiology of rabies in Hunan province and the genetic characteristics of rabies N gene isolated from 2008 to 2009. METHODS: Direct immunofluorescence assay (DFA) and nested PCR were employed to detect the monitoring samples including brain tissues of dogs and saliva, serum or urine which were collected in 2008 to 2009, from the rabies patients. Positive samples were sequenced by ABI3730 gene analyzer for the full length of the N gene target. The homology and hpylogeography of the rabies virus were analyzed after the phylogenetic tree was constructed by Blast, Clustal W and Mega 4.0 software. RESULTS: Of the 1451 tissue samples from the dogs' brain, 31 were positive under DFA and the positive rate was 2.14%. The DFA positive samples were redetected by RT-PCR and the positive rate was 1.17%. 56 samples of saliva, serum and urine samples were detected by RT-PCR from the rabies patients, with 3 positives and the positive rate was 5.36%. The length of nest PCR products were 255 bp. The rates of homology to the nucleotide and the amino acid of rabies N gene were 87.2% - 87.9% after compared to the pasture strain. The phylogenetic tree was successfully built and 20 strains isolated lately belonged to the rabies gene type I. CONCLUSION: The epidemic situation of human and dogs rabies in Human were relatively stable, with all the isolated rabies virus belonging to genotype I, without any variation.

[Pathogenic diagnosis of an unexplained epidemic event of infectious diarrhea].

OBJECTIVE: To determine the pathogen of an unexplained epidemic event of infectious diarrhea by laboratory diagnosis of suspected cases samples. METHODS: 28 samples from 28 suspected cases (22 fecal samples, 3 vomitus samples, 3 anus swab samples) were tested for Norovirus by RT-PCR. Sequencing and phylogenetic analysis were acomplished of 5 positive samples. RESULTS: 160 of 5694 population were ill with an attack rate of 2.81%. The peak period was 7-9, March. 14 of 28 samples were tested Norovirus positive.Sequencing and phylogenetic analysis showed Norovirus type GII/4 was the causative agent and it had highest identity (97. 9%) with epidemic strain 2006b. CONCLUSION: The epidemic event ofinfectious diarrhea were caused by GII/4 Norovirus strains.

[Viral surveillance on rabies in Hunan province, in 2006].

OBJECTIVE: To understand the source and distribution of rabies virus (RV) in Hunan province with viral surveillance in order to provide scientific measures for prevention and control on rabies. METHODS: Brain samples from healthy-looking domestic dogs were collected in the agricultural markets at the districts of high, middle, and low incidence rates and detected by direct Immunofluorescence assay (DFA). Positive samples would be further detected by RT-PCR and the surveillance samples were detected by RT-PCR. The positive samples detected by RT-PCR were sequenced with N gene. RESULTS: The infection rate of those healthy-looking domestic dogs with rabies virus was 2.78% in Hunan province in 2005. 23 positive samples' N gene were sequenced and their similarities were 88.8% - 100.0%. The results indicated that Hunan rabies virus N gene aberrance was mainly synonymous aberrance and did not carry obvious regional characteristics. The rabies virus were circulating among different districts in Hunan province, and the neighboring provinces such as Guizhou, Hubei, Guangxi, Jiangsu and Henan. There were no positive samples detected in salivary, blood and urine samples. There was one positive sample detected in two skin samples. CONCLUSION: There are dogs infected with rabies virus found in Hunan province and this study showed that rabies virus detected in Hunan had a close genetic relationship with those rabies identified in other provinces, suggesting that study on the immunity and management of dog related rabies should be strengthened.

[Laboratory diagnosis and molecular characterization of highly pathogenic avian influenza virus (H5N1) in human in Hunan Province in 2005-2006].

To determine the etiologic agents of two atypical pneumonia human cases in Hunan Province in 2005-2006 and to study their pathogenic potential, the patients' respiratory tract samples and sera were collected. The respiratory tract samples were tested by real-time RT-PCR and RT-PCR methods, and the sera by hemagglutination-inhibition assay. Virus was isolated from case 2 and its genome was sequenced and analyzed. Results showed the H5 nucleic acid tests of two cases were positive. The H5-specific antibody titer of the convalescence serum of case 1 showed a 4-fold greater rise than that of the acute phase. And case 2's antibody titer of acute phase was negative. The two atypical pneumonia cases were confirmed as the avian influenza A (H5N1) infection cases. Viral strain A/Hunan/1/2006 was isolated from case 2. Phylogenetic and molecular analysis suggested that 8 gene segments of A/Hunan/1/2006 originated from avian viruses. And A/Hunan/1/2006 was similar with viruses isolated from avian in Hunan Province. The isolated virus did not recombine with human influenza viruses and no obvious variation was observed.

[Virological surveillance on hemorrhagic fever with renal syndrome in Hunan province in 2006].

OBJECTIVE: To understand the rate of viral carrying status among rodents as well as genotypes and distribution of Hantaviruses (HV) isolated in Hunan province. METHODS: With DFA, the HV antigen in lung tissues of rodents was detected. The total viral RNA was extracted from the lung tissues of the HV infected rats and amplified with reverse transcrition-polymerase chain reaction (RT-PCR), using the HV genotype specific primers. The amplified genes were then sequenced and subjected to genotyping and homologic analysis. RESULTS: The average density of rodents was 3.15% and the virus carrying rate among rodents was 1.31%. Data from genotype analysis showed that the HV isolated from seven lung specimens taken from Rattus norvgicus, Apodemus agraius, Mus musculus, Rattus flavipectus among indoor rodents in Shaodong and Liuyang belonged to HV type II (SEOV), and one isolated from Apodemus agraius in Shaungfen belonged to HV type I (HTNV) among outdoor rodents. Six strains were sequenced successfully and the homology between six srains was 88.3%-100%. The homology of HN1, HN2, HN4, HN6 came from Liuyang and the HN7 and HN8 from Shaodong were both 100% while the homology between L99 and the strains from Liuyang and Shaodong were 94.4% and 88.3% respectively. CONCLUSION: HV type II (SEOV) and the HV type I (HTNV) were all existed in Hunan province while SEOV was the main genotype.

[Laboratory diagnoses of the first confirmed human case of avian influenza A (H5N1) in the mainland of China].

BACKGROUND: To determine the etiologic agent of an atypical pneumonia human case admitted to Xiangtan City hospital, Hunan Province in Oct. 2005. METHODS: The patient's respiratory tract samples and serum were collected. Throat swabs were tested by microneutralization and hemagglutination-inhibition assays. RESULTS: The results of nucleic acid detection of all respiratory samples were negative and virus isolation was also negative. The H5-specific antibodies of convalescence showed a 4-fold greater rise than acute phase. CONCLUSION: The atypical pneumonias case was confirmed as the first human case of avian influenza A (H5N1) infection in the mainland of China.

[Treating high strength antibiotic wastewater by micro-aerobic hydrolysis and acidification process].

The effect of micro-aerobic hydrolysis and acidification to high strength antibiotic wastewater treatment is studied. The results demonstrate that micro oxygen enhanced the physiological metabolizability of facultative hydrolytic and acidogenic bacteria, and aerating stirring improved the hydraulic condition. Degree of acidification (AD) and volatile fatty acid (VFA) in the effluent reached 58.64% and 4825 mg/L with the shortest HRT 10 h and the maximal OLR 20 kg/(m3 x d), respectively. Wastewater biodegradability was improved greatly and rising BOD5/COD was about 17%, which offered good substrate for post aerobic treatment. The effluent quality was relative stable with the fluctuant influent, and COD and SS were 7000-8000 mg/L and 150-300 mg/L, respectively, while the removal efficiencies of COD and SS were 15%-30% and 90%-95%. The changement of VFA lagged behind that of AD in the effluent, and AD would represent the effect of hydrolysis and acidification process more properly. The sludge bed in the bottom of the reactor was the main reaction field for VFA production, and the height of the reactor for stable VFA production increased with the increase of OLR. The filling field mainly aimed to entrap SS in the effluent and benefited little to VFA production. Sludge in the reactor was mainly little sludge particles with the size of 0.5-1.0 mm and flocculent sludge.