Direct and Rapid Identification of Vibrio Cholerae Serogroup and Toxigenicity by a Novel Multiplex Real-Time Assay.

Molecular diagnostic assays for cholera detection have superior sensitivity to conventional assays and are now being accepted as the new standard method, especially the real-time PCR/RT-PCR. However, limited throughput capacity and long detection duration prevent them from detecting more specimens and more targets in one turnaround time simultaneously. In this study, we utilized nucleic acid extraction-free, direct RT-PCR and high-speed amplification to develop a novel multiplex assay, a quadplex direct one-tube real-time RT-PCR assay, for rapid detection of the serogroup and cholera toxin toxigenicity of Vibrio cholerae targeting the epsM, ctxA, rfb-O1, and rfb-O139 genes. Performance of the multiplex assay was evaluated by comparison with the monoplex real-time PCR assay according to the China Cholera Prevention Manual. Detection data from clinical specimens showed that the new assay had good diagnostic sensitivities for epsM (100%, n = 301), ctxA (100%, n = 125), rfb-O1 (100%, n = 85), and rfb-O139 (97.87%, n = 49). Analysis of the analytical sensitivities with serial dilutions of positive standards showed that the detection limits of the new assay for Vibrio cholerae epsM,ctxA,rfb-O1, and rfb-O139 were up to 200, 590, 115, and 1052 copies per mL lower than the monoplex real-time PCR (910, 345, and 1616 copies/mL respectively, for ctxA,rfb-O1, and rfb-O139). The results indicate that the multiplex assay is a rapid, sensitive, specific, and easy-to-use detection tool for Vibrio cholerae, especially suitable for rapid identification and screening detection of mass specimens.

The Impact of Residences and Roads on Wind Erosion in a Temperate Grassland Ecosystem: A Spatially Oriented Perspective.

The existence of residences and roads is an important way in which human activity affects wind erosion in arid and semiarid environments. Studies assessing the impact of these elements on wind erosion have only focused on limited plots, and their threat of erosion to the surrounding environment has been ignored by many studies. This study was based on spatially overlayed analysis of independent wind erosion distribution simulated by the revised wind erosion equation (RWEQ) and remote-sensing-image-derived residence and road distribution data. Wind erosion at different distances from residences and roads was quantified at the landscape scale of a typical temperate grassland ecosystem, explicitly demonstrating the crucial impacts of both elements on wind erosion. The results showed that wind erosion weakened as the distance from residences and roads increased due to the priority pathways of human activities, and the wind erosion around the residence was more severe than around the road. Human activities in the buffer zones 0-200 m from the residences most frequently caused severe wind erosion, with a wind soil loss of 25 t ha-1 yr-1 and a wind soil loss of approximately 5.25 t ha-1 yr-1 for 0-60 m from the roads. The characteristics of wind erosion variation in the buffer zones were also affected by residence size and the environments in which the residences were located. The variation in wind erosion was closely related to the road levels. Human activities intensified wind erosion mainly by affecting the soil and vegetation around residences and roads. Ecological management should not be limited to residences and roads but should also protect the surrounding environments. The findings of this study are aimed towards a spatial perspective that can help implement rational and effective environmental management measures for the sustainability of wind-eroded ecosystems.

Spatially explicit quantification of total soil erosion by RTK GPS in wind and water eroded croplands.

Quantification of total soil erosion in wind and water eroded croplands is essential for assessing their contributions and the interaction between them. However, it is difficult to quantify total soil erosion amounts by the traditional monitoring and modelling approaches of wind erosion and water erosion. To address this problem, a Real Time Kinematic Global Positioning System (RTK GPS) was applied for a series of wind and water eroded croplands in the Bashang area in North China to quantify the total soil erosion amount over a period of 44 years. By comparing the elevation of the croplands with a reference surface without erosion, the total soil erosion modulus and its spatial variation were determined. Results showed that the erosion moduli of the six croplands ranged from 1.09 to 45.34 Mg ha-1 y-1 with an average modulus of 17.02 Mg ha-1 y-1. The croplands in the west suffered from more intense wind erosion compared to the middle and eastern areas; this was due to the presence of forest-grasslands, which served as wind breaks for the croplands in the middle and eastern regions. However, the croplands in the east showed the highest total erosion modulus, which was due to the influences of a gully. Within the croplands, the slope areas suffered from intense soil erosion which was mainly owing to water erosion. The reliability and uncertainty of this approach were discussed in terms of the equipment precision, results accuracy, and possible deposition on the reference surface. This study shows that when a suitable reference surface is identified and the erosion amount is considerable, RTK GPS survey can be used as a reliable and effective method to assess the spatially explicit total soil erosion in croplands influenced by both wind and water erosion.

Analysis of characteristics of HIV cases aged 15-24 in Jiaxing City / 中国学校卫生

Objective@#To understand the epidemiological characteristics and differences of HIV-positive cases among 15-24 years old in Jiaxing city and provide evidence for the development of targeted prevention and control measures.@*Methods@#A descriptive epidemiological method was used to analyze the data of HIV cases aged 15-24 reported in Jiaxing from 1999 to 2018.@*Results@#A total of 375 cases of young HIV were reported in 1999-2018, with an average age of 21.29±1.90 years, of which 42 were students. The ratio of male to female was 2.47∶1. The proportion of foreign household registration was higher (76%, 285 cases). The proportion of off-campus youth cases in total cases showed a downward trend(χ2=8.26, P=0.00), but the proportion of student cases showed an upward trend(χ2=15.73, P<0.01). Off-campus youth cases were mainly heterosexual transmission(59.16%, 197 cases), and the students’ cases were mainly homosexual transmission(88.10%, 37 cases). There were significant differences in gender, age, household registration, education level, route of transmission, late detection, CD4 level and source of detection among students and off-campus adolescents(P<0.05).@*Conclusion@#The prevalence of AIDS in adolescents and students is worthy of attention. The characteristics of adolescents inside and outside the school are different. Targeted prevention measures should be taken to reduce the harm of AIDS to young people.

A Real-Time PCR with Melting Curve Analysis for Molecular Typing of Vibrio parahaemolyticus.

Foodborne disease caused by Vibrio parahaemolyticus is a serious public health problem in many countries. Molecular typing has a great scientific significance and application value for epidemiological research of V. parahaemolyticus. In this study, a real-time PCR with melting curve analysis was established for molecular typing of V. parahaemolyticus. Eighteen large variably presented gene clusters (LVPCs) of V. parahaemolyticus which have different distributions in the genome of different strains were selected as targets. Primer pairs of 18 LVPCs were distributed into three tubes. To validate this newly developed assay, we tested 53 Vibrio parahaemolyticus strains, which were classified in 13 different types. Furthermore, cluster analysis using NTSYS PC 2.02 software could divide 53 V. parahaemolyticus strains into six clusters at a relative similarity coefficient of 0.85. This method is fast, simple, and conveniently for molecular typing of V. parahaemolyticus.

A multiplex liquid-chip assay based on Luminex xMAP technology for simultaneous detection of six common respiratory viruses.

We utilized one-step multiplex reverse transcription-PCR (RT-PCR) and Luminex xMAP technology to develop a respiratory multiplex liquid-chip assay (rMLA) for simultaneous detection of 6 common respiratory viruses, including influenza virus type A (FluA) and type B (FluB), para-influenza virus type 3 (PIV-3), respiratory syncytial virus (RSV), human metapneumovirus (MPV) and a threatening virus to China, Middle East Respiratory Syndrome coronavirus (MERS-CoV). Performance of rMLA was evaluated by comparing with real-time RT-PCR. Detection data from clinical specimens showed that the rMLA had diagnostic sensitivities of 97.10% for FluA, 94.59% for FluB, 98.68% for PIV-3, 94.87% for RSV and 95.92% for MPV (No Data for MERS-CoV due to the lack of positive specimens). Data of analytical sensitivities showed that the detection limits of the rMLA assay were 5-25 viral RNA copies per µl for FluA, FluB, PIV-3 and MERS-CoV, approximate to the real-time RT-PCR assay; while the values were 8 and 22copies/µl for MPV and RSV, lower than the real-time RT-PCR(78 and 114 copies/µl respectively). The results indicated that the rMLA is a sensitive, specific detection tool and comparable to real-time RT-PCR, especially suitable for high-throughput detection of respiratory specimens.

Development and Application of A Duplex Real-Time RT-PCR Assay for Simultaneous Detection of Coxsackievirus A6 and Coxsackievirus A10.

Hand, foot and mouth disease (HFMD) is a serious public health problem. Generally, it is considered that HFMD is mainly caused by enterovirus 71 (EV71) and coxsackievirus A16 (CVA16). Nevertheless, the incidence of HFMD caused by coxsackievirus A6 (CVA6) and coxsackievirus A10 (CVA10) has increased significantly and CVA6 and CVA10 have become major causes of HFMD epidemic. This study develops a duplex real-time reverse transcript polymerase chain reaction (RT-PCR) assay for simultaneous detection of CVA 6 and CVA 10. The specificity, sensitivity, and reproducibility of this assay were analyzed. No cross-reactions with other viruses or false positives were observed. The detection limit of this assay was as low as 11.935 copies for CVA6 and 17.591 copies for CVA10 per reaction (concentration giving a positive duplex real-time RT-PCR result in 95% of samples). The coefficients of variation of the intra- and inter-assay reproducibility for CVA 6 and CVA 10 were both lower than 2%. Our results showed that this duplex real-time RT-PCR assay was a simple, rapid, and cost-effective method for simultaneous identification of CVA6 and CVA10.

Genetic characterization and antiretroviral resistance mutations among treatment-naive HIV-infected individuals in Jiaxing, China.

The aim of this study was to characterize HIV-1 genotypes and antiretroviral resistance mutations among treatment-naive HIV-infected individuals in Jiaxing, China. The HIV-1 partial polymerase (pol) genes in 93 of the 99 plasma samples were successfully amplified and analyzed. Phylogenetic analysis revealed the existence of five HIV-1 genotypes, of which the most prevalent genotype was CRF01_AE (38.7%), followed by CRF07_BC (34.4%), CRF08_BC (16.1%), subtype B/B' (5.4%), and CRF55_01B (2.1%). Besides, three types of unique recombination forms (URFs) were also observed, including C/F2/A1, CRF01_AE/B, and CRF08_BC/CRF07_BC. Among 93 amplicons, 46.2% had drug resistance-associated mutations, including 23.7% for protease inhibitors (PIs) mutations, 1.1% for nucleoside reverse transcriptase inhibitors (NRTIs) mutations, and 20.4% for non-nucleoside reverse transcriptase inhibitors (NNRTIs) mutations. Six (6.5%) out of 93 treatment-naive subjects were identified to be resistant to one or more NNRTIs, while resistance to NRTIs or PIs was not observed. Our study showed the genetic diversity of HIV-1 strains circulating in Jiaxing and a relative high proportion of antiretroviral resistance mutations among treatment-naive patients, indicating a serious challenge for HIV prevention and treatment program.

Investigation of rare chronic lipoid pneumonia associated with occupational exposure to paraffin aerosol.

OBJECTIVES: Occupational exposure to paraffin is an infrequent cause of lipoid pneumonia (LP) and related data are scare. We investigated the possible relationship between three rare cases of chronic LP and occupational exposure to paraffin aerosol in an iron foundry. METHODS: The three cases of LP and their workplaces were investigated using data from field investigations, air monitoring, pulmonary radiological examinations, cell staining, and lung biopsies. RESULTS: The patients had long-term occupational exposure to paraffin. X-ray diffraction testing revealed that the raw material from the workshop was paraffin crystal. The air concentrations of paraffin aerosol in workplaces were significantly higher than outdoor background levels. Small diffuse and miliary shadows with unclear edges were observed throughout the whole lungs via radiography. Computed tomography revealed diffuse punctate nodules and a high density of stripe-like shadows in both lungs (ground-glass opacity in a lower lobe, and a mass-like lesion and high translucent area near the bottom of the lung). Lipid-laden macrophages were found in the sputum and bronchial lavage. A broadened alveolar septum and local focal fibrosis were also discovered via lung biopsy. The inflammatory reaction in the lung tissues appeared to resolve over time. CONCLUSIONS: These three rare cases of chronic LP in workers during molding and repair processes were associated with occupational paraffin aerosol exposure. Therefore, primary prevention is essential for molding or repairing workers in the iron foundry, and a differential diagnosis of occupational chronic LP (vs. pneumoconiosis) should be considered when treating these workers.

Dynamic quantification of avian influenza H7N9(A) virus in a human infection during clinical treatment using droplet digital PCR.

This study involved a human infection with avian influenza H7N9(A) virus in Zhejiang province, the first one after implementing the closure measures of living poultry markets in China. The clinical symptoms, epidemiological and virological characteristics of the case were described briefly, and as the emphasis, H7N9 virus was detected quantitatively and continuously from the collected samples in 10 different periods of the patient's treatment in order to reveal changes of viral load in patient's body during the treatment. This study first used reverse-transcription droplet digital PCR (RT-ddPCR) assays to monitor viral load dynamically for human H7N9 infection, synchronously performing real-time RT-PCR as a reference technology to obtain more comprehensive data for comparison. Our results indicated that RT-ddPCR compared to real-time RT-PCR is more sensitive and accurate for quantifying H7N9 viral load without the use of standard curves. Furthermore it can provide reference data for clinical policies including infectivity judgement, ward transferring and therapy adjustment for the patient during treatment.

Socioeconomic Status and Physical Activity in Chinese Adults: A Report from a Community-Based Survey in Jiaxing, China.

OBJECTIVES: This study examines the associations of socioeconomic status (SES) with intensity of different types of physical activity (PA) in Chinese adults, aimed at outlining and projecting socioeconomic disparities in PA among the population undergoing a rapid nutrition transition. METHODS: A community-based survey was conducted among 3,567 residents aged 30-65 years old in Jiaxing, China, in 2010. SES and PA were assessed by a structured questionnaire. SES was assessed as socioeconomic index (SEI) score based on self-reported educational attainment, household income and occupation. Metabolic equivalents (METs) were calculated for each subject to quantify the total amount of PA from occupation, exercise, transportation and housework. RESULTS: Intensity of overall PA in this population was 165 MET-hours/week, in which energy expenditure in occupational PA accounted for 82%. Both types and intensity of PA were significantly different by SES: middle SES groups had higher intensity of occupational activities; lower SES subjects engaged in more household work; whereas higher SES subjects were more likely to exercise, more active during commuting and had longer sedentary time. All the three components of SES, education attainment, income and occupation, contributed to socioeconomic disparities in PA in this population. CONCLUSIONS: Our results suggest an overall insufficiency and socioeconomic inequalities in PA among Chinese adults in Jiaxing, a typical city experiencing a rapid urbanization in China. There is an urgent need to promote leisure-time activities in this population.

Multiplex real-time PCR assay for detection of pathogenic Vibrio parahaemolyticus strains.

Foodborne disease caused by pathogenic Vibrio parahaemolyticus has become a serious public health problem in many countries. Rapid diagnosis and the identification of pathogenic V. parahaemolyticus are very important in the context of public health. In this study, an EvaGreen-based multiplex real-time PCR assay was established for the detection of pathogenic V. parahaemolyticus. This assay targeted three genetic markers of V. parahaemolyticus (species-specific gene toxR and virulence genes tdh and trh). The assay could unambiguously identify pathogenic V. parahaemolyticus with a minimum detection limit of 1.4 pg genomic DNA per reaction (concentration giving a positive multiplex real-time PCR result in 95% of samples). The specificity of the assay was evaluated using 72 strains of V. parahaemolyticus and other bacteria. A validation of the assay with clinical samples confirmed its sensitivity and specificity. Our data suggest the newly established multiplex real-time PCR assay is practical, cost-effective, specific, sensitive and capable of high-throughput detection of pathogenic V. parahaemolyticus.

[Study on the prevalence of prehypertension among residents living in the communities in Zhejiang].

OBJECTIVE: To investigate the prevalence and related risk factors of prehypertension in Hangzhou, Shaoxing,Jiaxing city, Zhejiang province. METHODS: 3200 people were selected by stratified cluster random sampling method, and statistical methods including chi-square test, and logistic regression through SAS 9.0 were used. RESULTS: The prevalence of pre- hypertension was 45.9%, higher for males and urban population, with significant differences seen between males and female(49.0% vs. 48.0%, P < 0.05), urban and rural areas(59.31% vs. 44.15%, P < 0.05). Data from the multiple factor logistic analysis showed that risk factors of prehypertension would include: older age, types of profession, under low education level, being urban residents, overweight and obesity, hyper triglyceride, and family history of hypertension, with ORs and 95% CI as 0.99 (0.98-0.99), 1.28(1.07-1.28), 1.31(1.10-1.56), 1.50(1.11-2.02), 1.33(0.98-1.81), 1.60 (1.19-2.16)and 1.18(1.00-1.39), respectively. CONCLUSION: Prehypertension prevalence was found high in the studied district, especially in urban residents with low education level. Strategies including reduction on risk factors as obesity and hyper triglyceride through health education as well as lifestyle modification should be taken to hold back the increasing trend on prehypertension in Zhejiang.