Identification of keystone taxa in rhizosphere microbial communities using different methods and their effects on compounds of the host Cinnamomum migao.

Exploring keystone taxa affecting microbial community stability and host function is crucial for understanding ecosystem functions. However, identifying keystone taxa from humongous microbial communities remains challenging. We collected 344 rhizosphere and bulk soil samples from the endangered plant C. migao for 2 years consecutively. Used high-throughput sequencing 16S rDNA and ITS to obtain the composition of bacterial and fungal communities. We explored keystone taxa and the applicability and limitations of five methods (SPEC-OCCU, Zi-Pi, Subnetwork, Betweenness, and Module), as well as the impact of microbial community domain, time series, and rhizosphere boundary on the identification of keystone taxa in the communities. Our results showed that the five methods, identified abundant keystone taxa in rhizosphere and bulk soil microbial communities. However, the keystone taxa shared by the rhizosphere and bulk soil microbial communities over time decreased rapidly decrease in the five methods. Among five methods on the identification of keystone taxa in the rhizosphere community, Module identified 113 taxa, SPEC-OCCU identified 17 taxa, Betweenness identified 3 taxa, Subnetwork identified 3 taxa, and Zi-Pi identified 4 taxa. The keystone taxa are mainly conditionally rare taxa, and their ecological functions include chemoheterotrophy, aerobic chemoheterotrophy, nitrate reduction, and anaerobic photoautotrophy. The results of the random forest model and structural equation model predict that keystone taxa Mortierella and Ellin6513 may have an effects on the accumulation of 1, 4, 7, - Cycloundecatriene, 1, 5, 9, 9-tetramethyl-, Z, Z, Z-, beta-copaene, bicyclogermacrene, 1,8-Cineole in C. migao fruits, but their effects still need further evidence. Our study evidence an unstable microbial community in the bulk soil, and the definition of microbial boundary and ecologically functional affected the identification of keystone taxa in the community. Subnetwork and Module are more in line with the definition of keystone taxa in microbial ecosystems in terms of maintaining community stability and hosting function.

Inhibitors of HIV-1 and Cathepsin L Proteases Identified from the Insect Gall of Hypericum kouytchense.

Hypericum kouytchense Lévl is a semi-evergreen plant of the Hypericaceae family. Its roots and seeds have been used in a number of traditional remedies for antipyretic, detoxification, anti-inflammatory, antimicrobial and antiviral functions. However, to date, no bioactivity compounds have been characterized from the insect gall of H. kouytchens. In this study, we evaluated the antiviral activities of different extracts from the insect gall of H. kouytchen against cathepsin L, HIV-1 and renin proteases and identified the active ingredients using UPLC-HRMS. Four different polar extracts (HW, H30, H60 and H85) of the H. kouytchense insect gall exhibited antiviral activities with IC50 values of 10.0, 4.0, 3.2 and 17.0 µg/mL against HIV-1 protease; 210.0, 34.0, 24.0 and 30.0 µg/mL against cathepsin L protease; and 180.0, 65.0, 44.0 and 39.0 µg/mL against human renin, respectively. Ten compounds were identified and quantified in the H. kouytchense insect gall extracts. Epicatechin, eriodictyol and naringenin chalcone were major ingredients in the extracts with contents ranging from 3.9 to 479.2 µg/mg. For HIV-1 protease, seven compounds showed more than 65% inhibition at a concentration of 1000.0 µg/mL, especially for hypericin and naringenin chalcone with IC50 values of 1.8 and 33.0 µg/mL, respectively. However, only hypericin was active against cathepsin L protease with an IC50 value of 17100.0 µg/mL, and its contents were from 0.99 to 11.65 µg/mg. Furthermore, we attempted to pinpoint the interactions between the active compounds and the proteases using molecular docking analysis. Our current results imply that the extracts and active ingredients could be further formulated and/or developed for potential prevention and treatment of HIV or SARS-CoV-2 infections.

Seed Dormancy Release and Germination Requirements of Cinnamomum migao, an Endangered and Rare Woody Plant in Southwest China.

Seed dormancy is a complex adaptive trait of plants that are influenced by several physiological and environmental factors. The endangered plant Cinnamomum migao is also known to exhibit seed dormancy and low germination, which may influence its regeneration; however, these characteristics remain unexplored. To our knowledge, this study is the first to examine the type of dormancy and improve the germination percentage of C. migao seeds. We evaluated the structure and characteristics of the embryo and endocarp of C. migao seeds as well as the effects of endogenous inhibitors. Furthermore, we assessed the effects of light, stratification, alternating temperature, and gibberellic acid 3 (GA3) on the dormancy release of these seeds. The embryo was well developed the endocarp was water-permeable, and no obvious mechanical hindrance to germination was observed. However, the endocarp and embryo contained phenols and other germination inhibitors. The seed extracts of C. migao delayed the germination of cabbage and ryegrass seeds, which indicates the presence of endogenous inhibitors. These findings suggest that C. migao seeds exhibit physiological dormancy. Light and an alternating temperature (15/20°C) did not influence germination. However, GA3 pretreatment, alternating temperatures, and warm stratification relieved dormancy. GA3 pretreatment combined with the 15°C stratification treatment was most effective in rapidly releasing the C. migao seed dormancy. Our findings may facilitate the storage and conservation of this endangered plant, which is currently underrepresented in ex situ collections.

Effect of sequential high-flow nasal cannula oxygen therapy and non-invasive positive-pressure ventilation in patients with difficult weaning from mechanical ventilation after extubation on respiratory mechanics.

BACKGROUND: Patients with difficult weaning who undergo mechanical ventilation are more likely to be at risk of reintubation and the sequential use of oxygen therapy after extubation is a concern for clinicians. Therefore, the aim of the present study was to compare the effects of transnasal high-flow nasal cannula (HFNC) oxygen therapy and non-invasive positive-pressure ventilation (NIV) on respiratory mechanics in patients with difficult weaning. METHODS: The present study was a single-center, retrospective, observational study. Twenty-nine patients with difficult weaning off invasive mechanical ventilation from the Department of Critical Care Medicine, The First Affiliated Hospital of Guangzhou Medical University, from December 2018 to April 2021, were included. Within 48 h after extubation, alternate respiratory support with HFNC and NIV was provided. Relevant indicators were recorded after each support mode had been maintained for at least 60 min. These included esophageal pressure (Pes), gastric pressure (Pga), transdiaphragmatic pressure (Pdi), pressure-time product of Pes (PTPes), pressure-time product of Pga (PTPga), pressure-time product of Pdi (PTPdi), ratio of the PTPdi to the PTPes (PTPdi/PTPes), and ratio of the Pes to the Pdi (Pes/Pdi), diaphragmatic electromyogram (EMGdi), percentage of esophageal pressure coefficient of variation (CVes%),diaphragmatic electromyogram coefficient of variation (CVEMG),inspiratory time (Ti), expiratory time (Te) and respiratory cycle time (Ttot). RESULTS: Of the 29 patients included, 22 were males and 7 were females [age: 63.97±15.34 years, Acute Physiological and Chronic Health Estimation II (APACHE II) score: 18.00±5.63]. The CVes% and the Pes/Pdi were significantly higher in patients with NIV than HFNC using 40 L/min, CVes%: 9 (-6, 20) vs. -7 (-23, 6) and Pes/Pdi: 0.17 (-0.1, 0.53), vs. -0.12 (-0.43, 0.08) (P<0.05). The remaining indicators were not statistically different. CONCLUSIONS: The sequential NIV and HFNC can be tolerated in patients with such difficult weaning off mechanical ventilation after extubation, and more patients tend to choose HFNC subjectively. Compared with HFNC, NIV reduces the work of adjunctive respiratory muscle, but the patient's Pes dispersion is high when NIV is used, and it is necessary to pay attention to patient-ventilator coordination in clinical practice. We recommend alternating HFNC and NIV during the sequential respiratory therapy after extubation.

[Study on HPLC fingerprint of Sabia parviflora].

This research was performed to establish the HPLC fingerprint of Sabia parviflora. HPLC method was carried out on a Thermo Accucore-C18(4. 6 mm×150 mm,2. 6 µm) column by 30% tetrahydrofuran in methyl alcohol-acetonitrile-0. 1% phosphate solution as mobile phase at a flow rate of 1. 0 m L·min-1,the column temperature was 30 ℃ and the detection wavelength was 360 nm. The fingerprints were further evaluated by chemometrics methods including similarity analysis,hierarchical clustering analysis,and principal component analysis. In HPLC fingerprint,15 common peaks were selected as the common peaks,and 6 contents of them were identified. The similarity degrees of 38 batches of the samples was more than 0. 710,and the samples were divided into 6 clusters by their quality difference. The method was precision,repeatable,stable,simple and reliable,which could be used for quality control and evaluation of S. parviflora.

Assessment of respiratory drive with esophageal diaphragmatic electromyography in patients with acute respiratory distress syndrome treated with prone position ventilation.

BACKGROUND: Prone position ventilation (PPV) is an important strategy for patients with severe acute respiratory distress syndrome (ARDS). This prospective study investigated the use of electromyography of the diaphragm (EMGdi) for monitoring respiratory drive in patients with moderate to severe ARDS during long-term PPV. METHODS: An integrated nostril-gastric feeding tube containing an esophageal electrode and balloon was placed in 14 patients with severe ARDS prior to PPV. EMGdi and trans-pulmonary pressure (∆PL) data were collected before PPV (baseline), every 2 h during PPV, and 2 h after the restoration of supine position ventilation (post-2 h SPV). RESULTS: In ARDS patients, the static compliance of the chest wall was significantly decreased after PPV. EMGdi levels were slightly lower in the early, middle, and late stages of PPV compared with baseline. Patients who received neuromuscular blocker experienced a greater drop in EMGdi from baseline than those who did not. CONCLUSIONS: For ARDS patients, EMGdi was slightly decreased after prolonged PPV. This is contrary to the change in diaphragm electromyography during normal body position changes. Monitoring EMGdi regularly during PPV in ARDS patients is feasible and can be used as a reference for lung protective ventilation strategies.

[Analysis of medicinal resources diversity of 33 pilots (districts) in Guizhou province].

This study is based on the data analysis of medicinal plant resources and diversity collected from the fourth Chinese traditional medicine resource survey( pilot). Through the analysis of relevant data from 33 census pioneer plots in Guizhou province( area),a total of 265 families,1 432 genera and 5 296 species of medicinal resources were reported,including algae,fungi,lichens,mosses,a total of 43 genera and 35 families,57,48 families,120 genera and 453 species of ferns,gymnosperms 11 families,22 genera and 61 species,167 families,1 243 genera and 4 721 species of angiosperms,4 genera and 4 families four medicinal animals.Compared with the data related to the third survey of traditional Chinese medicine resources,the number of ferns,gymnosperms and angiosperms in the fourth survey has increased far more than that of the third survey. From the regional distribution of medicinal resources,the composition of the genus,the type of life,and the location of the medicine,the richness of the medicinal plant resources in Guizhou province is not only reflected in many types,but also in the variety of medicinal resources. These studies provide a scientific basis for vigorously developing the Chinese herbal medicine industry and the sustainably using medicinal plant resources in Guizhou province.

[The role of human lysozyme-like protein 4 in fertilization and its enzymatic properties].

OBJECTIVE: To elucidate the possible role of human lysozyme-like protein 4 (LYZL4) in fertilization and characterize its enzymatic properties. METHODS: The localization of LYZL4 in human spermatozoa was investigated by immunofluorescence staining, the sources of LYZL4 on the sperm surface examined by RT-PCR, and the role of LYZL4 in fertilization assessed by the zona-free hamster egg penetration test. The recombinant plasmid pPIC9K-LYZL4 was constructed and its expression induced with methanol after transformed into competent Pichia pastoris GS115. The recombinant LYZL4 protein (rLYZL4) was purified from the fermentation supernatant and subsequently identified by Western blot. The hyaluronan binding ability of rLYZL4 was determined by ELISA and the muramidase activity, hyaluronidase activity, and free radical scavenging ability examined by spectrophotometric methods. RESULTS: Immunodetection with a specific antiserum localized LYZL4 on the acrosomal membrane of mature spermatozoa, which was exclusively secreted from the testis and epididymis as shown by RT-PCR. Immunoneutralization of LYZL4 significantly decreased the number of human spermatozoa bound to zona-free hamster eggs in a dose-dependent manner in vitro. The recombinant protein was expressed successfully by the P. pastoris strain GS115. Purified rLYZL4 exhibited a potent hyaluronan binding ability and a strong free radical scavenging ability but no muramidase or hyaluronidase activity. CONCLUSIONS: LYZL4 secreted from the testis and epididymis is localized on the acrosomal membrane of mature spermatozoa and plays a role in sperm-egg binding as well as in binding hyaluronan and scavenging free radicals, which suggests that it might be a multi-functional molecule contributive to sperm protection and sperm-egg binding.

[Microstructure characteristics and identification significance of leaf epidermis of medicinal plants from Sabia genus].

OBJECTIVE: To study the feasibility and significance of the method to identify medicinal plants through the observation and statistics of 9 species of Sabia medical plants from Guizhou province. METHODS: Leaf epidermis characteristics were observed, measured by optical microscope and analyzed by SPSS 17.0 software. RESULTS: All of these plants had some differences in indumentum, cell morphology and size, and had significant difference in the length and circumference of lower epidermis cells. CONCLUSION: The method combining microscopic observation with statistics can be used as the classification and identification basis of medicinal plants and materials of Sabia genus.

Apigenin enhances the cytotoxic effects of tumor necrosis factor-related apoptosis-inducing ligand in human rheumatoid arthritis fibroblast-like synoviocytes.

Activated rheumatoid arthritis (RA) fibroblast-like synoviocytes (RAFLSs) play a central role in both initiating and driving RA. Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) has been documented to induce apoptosis only in a small proportion of RAFLSs, which is followed by an induction of proliferation in surviving cells. Apigenin, a chemopreventive bioflavonoid, exhibits proapoptotic activity in many types of cells. In the present study, we sought to determine whether apigenin could enhance the cytotoxic effect of TRAIL on activated RAFLSs. Human RAFLSs isolated from patients with RA were treated with TRAIL (1 nM), apigenin (20 µM), or their combination, and subjected to apoptosis analysis after a 24-h incubation and proliferation analysis after a 72-h incubation. Apoptosis assay revealed that TRAIL or apigenin alone induced a marked apoptosis in RAFLS and their combination yielded a synergistic increase in RAFLS apoptosis. Immunoblotting analysis of apoptosis regulators demonstrated that combined treatment with apigenin increased caspase-3 expression and activity and decreased the Bcl-2/Bax ratio relative to treatment with TRAIL alone. The presence of apigenin significantly restrained TRAIL-induced RAFLS proliferation, coupled with restoration of the expression of two cell-cycle inhibitors p21 and p27. Moreover, the combination with apigenin blunted TRAIL-induced activation of the phosphatidylinositol 3-kinase (PI3-K)/Akt pathway. Our data collectively demonstrate that apigenin sensitizes RAFLS to TRAIL-induced apoptosis and counteracts TRAIL-dependent RAFLS proliferation, which is likely mediated through inactivation of PI3-K/Akt signaling pathway.

Evaluation of a novel fusion protein antigen for rapid serodiagnosis of tuberculosis.

The detection of Mycobacterium tuberculosis-specific antibodies in human sera has been a rapid and important diagnostic aid for tuberculosis (TB) control and prevention. However, any single antigen is not enough to be used to cover the antibody profiles of all TB patients. In this study, a novel fusion protein was constructed using gene splicing by overlap extension (SOEing), and then the antibody level against it in 171 TB patients and 86 controls was evaluated by enzyme-linked immunosorbent assay. Compared with the three individual antigen (16 kDa: sensitivity 19.9%, specificity 96.5%; MPT64: sensitivity 75.4%, specificity 34.9%; 38 kDa: sensitivity 33.3%, specificity 83.7%), the fusion protein antigen (sensitivity 42.1%, specificity 89.5%) gave the best diagnostic performance with the largest receiver operating characteristic curve area 0.656 (95% confidence interval [CI], 0.590-0.721; P<0.01). These results suggested that the novel fusion protein antigen successfully constructed by gene SOEing provided the improved diagnostic performance for TB, and other mycobacterial multiepitope fusion proteins may also be worthy of investigation for further enhancing the detection sensitivity.

Effects of berberine on human rheumatoid arthritis fibroblast-like synoviocytes.

Activated rheumatoid arthritis fibroblast-like synoviocytes (RAFLSs) play an important role in the initiation and progression of rheumatoid arthritis (RA). Rapid proliferation and defective apoptosis of RAFLSs are two main mechanisms contributing to synovial hyperplasia. Berberine, the major constituent of Coptidis Rhizoma, has been widely used as an antitumor and anti-inflammation agent. Here we show that berberine significantly inhibited cell proliferation of serum-starved human RAFLSs in a dose-dependent manner. Cell cycle analysis of berberine-treated RAFLSs indicated a cell cycle arrest at the G0/G1 phase. The inhibitory effects of berberine correlated with an induction of cyclin-dependent kinase (CDK) inhibitors Cip1/p21 and Kip1/p27 and a reduction of CDK2, CDK4 and CDK6, and cyclins D1, D2 and E. Furthermore, an apoptosis assay showed that berberine treatment increased apoptotic death of RAFLSs, which was associated with an increased expression of proapoptotic protein Bax and decreased expression of antiapoptotic proteins Bcl-2 and Bcl-xl, disruption of mitochondrial membrane potential, and activation of caspase-3, caspase-9 and poly (ADP-ribose) polymerase. Taken together, these results demonstrate that berberine exerts antiproliferative effects against RAFLSs, likely through deregulation of numerous cell cycle and apoptosis regulators, thus having potential therapeutic implications in the treatment of RA.

Complete genome sequence of a novel clinical isolate, the nontuberculous Mycobacterium strain JDM601.

Mycobacteriosis is on the increase. Nontuberculous mycobacteria (NTM) are resistant to most antituberculosis drugs naturally. We determined the complete genome sequence of a novel NTM strain, JDM601, of the Mycobacterium terrae complex, which was isolated from a patient with tuberculosis-like disease and with various antibiotic resistances.

[Long-term and stable expression of transgene mediated by piggyBac transposon in gynecological malignant tumor cells].

OBJECTIVE: To investigate the expression of exogenous gene transferred by piggyBac (PB) transposon in various gynecological malignant cell lines and reveal its potential application of gene therapy in gynecological cancer. METHODS: Amplified herpes simplex virus thymidine kinase (HSV-tk) gene coding region by PCR and integrated it into PB expression vector, PB[Act-RFP]DS, for reconstructing PB[Act-RFP, HSV-tk]DS (pPB/TK). By using different transfection reagents: FuGENE HD, jetPEI, lipofectamine 2000, pPB/TK together with helper plasmid Act-PBase were cotransfected into four mostly common gynecological malignant tumor cell lines HeLa, JEG-3, SKOV3 and HEC-1B. The mRFP1 report gene expressions was observed and detected by fluorescence microscope and flow cytometry to analyze transfection efficiency. The expressions of HSV-tk and mRFP1 gene were detected by reverse transcription PCR (RT-PCR). The cytotoxic effect of various concentration of pro-drug ganciclovir (GCV) for transfected cells was detected by methyl thiazole tetrazolium assay. The transfected cells were positive sorted by flow cytometry and limiting diluted to obtain the stable transfected cell line. The insertion sites of foreign gene transferred by PB transposon in genome were analyzed by inverse PCR. RESULTS: (1) Double digests analysis and sequences test demonstrated that pPB/TK vector was reconstructed successfully. (2) Using three different transfective reagents, PB transposon transferred HSV-tk gene and mRFP1 gene into HeLa, HEC-1B, SKOV3 and JEG-3 cell efficiently, and the transfection efficiency of pPB/TK for the same cell was different by using different transfective reagents; in Hela cell, the transfection efficiency of FuGENE HD [(78.7+/-9.2)%] was higher than that of lipofectamine 2000 [(54.1+/-11.4)%] and jetPEI [(46.5+/-7.4)%, all P<0.05]; using the same transfective reagent, the transfection efficiency of pPB/TK was also different on various cell lines, using FuGENE HD, the transfection efficiency of pPB/TK on HeLa, JEG-3 and SKOV3 cell was (78.7+/-9.2)%, (74.4+/-8.9)% and (83.2+/-9.7)% respectively, which all were higher than that on HEC-1B [(39.5+/-8.7)%, P<0.05]. (3) RT-PCR showed that there were the mRNA expression of HSV-tk and mRFP1 in all cell lines. (4) 50% inhibitory concentration of GCV for transfected cells, HeLa, JEG-3, SKOV3 and HEC-1B, was 1.29, 3.35, 0.09 and 13.28 microg/ml respectively. Inhibitory effect of GCV (10 microg/ml) on SKOV3 transfected with pPB/TK was (86+/-9)%, which was superior to that transfected with pORF-HSVtk alone [(52+/-12)%, P<0.05]. (5) The insertion sites of PB transposon in the target cells genome were located at TTAA sites. mRFP1 expression still could be detected in three months after transfected. CONCLUSIONS: PB transposon could transfer exogenous gene into various gynecological malignant cells, which could integrated into genome and obtain a long-term and stable expression. It is expected that PB transposon may supply a more efficient and safer transgene technology platform for gene therapy in gynecological cancer.

Development and evaluation of a novel multiple-antigen ELISA for serodiagnosis of tuberculosis.

In this study, we describe the development and evaluation of a novel multiple-antigen ELISA for rapid diagnosis and screening of active tuberculosis (TB). The humoral immune responses of 136 active TB patients and 57 healthy subjects against antigens Rv3425, 38kDa and lipoarabinomannan (LAM) from Mycobacterium tuberculosis H37Rv were examined by ELISA. Three essential results were obtained. (i) Rv3425 antigen is a potential candidate for serodiagnosis of active TB. Of 136 active TB patients, Rv3425 antigen provided a sensitivity of 31.6%, lower than that of LAM antigen, but higher than that of 38kDa antigen, with an overall specificity of 100%. (ii) For 62 smear-negative pulmonary TB patients and 15 extra-pulmonary TB patients, the multiple-antigen test provided a sensitivity of 43.5% and 26.7%, respectively, representing an improvement over acid-fast bacilli (AFB) smear-based diagnosis. (iii) Compared with the single-antigen ELISA and the two available commercial kits, the multiple-antigen test offered the highest accuracy (71.0%). In conclusion, the multiple-antigen ELSIA test based on Rv3425, 38kDa, and LAM antigens is a potentially useful tool for the serodiagnosis and screening of active TB. Combinations of Rv3425 with other mycobacterial antigens may also be worthy of further investigation.

[Research progress on geographic distribution new records of medicinal plants in Guizhou].

To introduce the advance on the species, ecological environment, distribution areas, the number of the species and efficacy of geographic distribution new records of medicinal plants in Guizhou. This article provides a basis for the collection and conservation as well as reasonable development of the genetic resources of medicinal plants.

Influence of meteorological factors and air pollution on the outbreak of severe acute respiratory syndrome.

OBJECTIVES: To understand the association between the outbreak of severe acute respiratory syndrome (SARS) and meteorological factors and air pollution. STUDY DESIGN: An ecological study was conducted. METHODS: Three hundred and fifty primary probable SARS cases diagnosed in mainland China between 1 January and 31 May 2003, and their 6727 close contacts during the period of their clinical symptoms before admission, were included in this study. Of the 6727 close contacts, 135 (2.0%) later developed clinical symptoms and were diagnosed as probable SARS cases. The daily meteorological data and daily air pollution data during the same SARS outbreak period in mainland China were used in the data analysis. Logistic regression analyses were conducted to explore the association between the secondary attack rate of SARS and meteorological factors and air pollution. RESULTS: In univariate analyses, daily average temperature (DAT), daily average air pressure (DAAP), and daily average relative humidity (DARH) were inversely associated with secondary attack rate (P<0.001); a significant positive association was found for daily hours of sunshine (DHS) (P<0.001). In multivariate analyses, factors associated with secondary attack rate were DAAP (odds ratio (OR)=0.53, 95% confidence interval (CI): 0.42, 0.66), DARH (OR=0.73, 95% CI: 0.53, 1.00), and daily average wind velocity (DAWV; OR=0.81, 95% CI: 0.68, 0.96). Adjustment for the onset time of a primary case led to little change in the results. In addition, in Hebei Province, a major affected area in China, only DAWV (OR=0.38, 95% CI: 0.20, 0.72) was a significant predictor of secondary attack rate with adjustment for the onset time of primary case. In Inner Mongolia, another major affected area in China, DAWV (OR=0.50, 95% CI: 0.26, 0.94) and DHS (OR=0.27, 95% CI: 0.09, 0.81) were significant predictors of secondary attack rate with adjustment for the onset time of primary case. CONCLUSIONS: Our results suggest that the SARS outbreak was significantly associated with DAWV, and that DAAP, DARH and DHS may also have influenced the SARS outbreak to some extent. However, because of ecological fallacy and uncontrolled confounding effects that may have biased the results, the association between the SARS outbreak and these meteorological factors and air pollution deserve further investigation.

[Effects of mild hypothermia on expression of N-methyl-D-aspartate receptor-1 in hippocampus neurons after cardiopulmonary resuscitation in rats].

OBJECTIVE: To explore the effects of mild hypothermia on expression of N-methyl-D-aspartate receptor-1 (NMDAR1) in hippocampus neurons after cardiopulmonary resuscitation (CPR) in rats. METHODS: Twenty-four male SD rats were randomly divided into normal control group, normal temperature group, and mild hypothermia group, with 8 rats in each group. The cerebral edema model after CPR was replicated by the sealed bottle method in rats in both normal temperature group and mild hypothermia group. The rats in the mild hypothermia group were treated with mild hypothermia after the model was established. The change in expression of NMDAR1 in hippocampus neurons in rat was determined with semi-quantitative reverse transcription polymerase chain reaction (RT-PCR), and pathologic changes in brain tissue were observed in both groups. RESULTS: The cerebral edema was ameliorated, NMDAR1 mRNA and protein in cerebral hippocampus neurons were significantly lower in hypothermia group than control group with significant difference (NMDAR1 mRNA: 80.48+/-0.03 vs. 80.64+/-0.18, P<0.05 ). CONCLUSION: Mild hypothermia can down regulate the expression of NMDAR1 mRNA and protein level, lower positive ion concentration and thus decrease cerebral edema, so mild hypothermia can serve as a treatment measure for cerebral edema after CPR.

Refined estimate of the incubation period of severe acute respiratory syndrome and related influencing factors.

Many epidemiologists have agreed that a refined estimate of the incubation period of severe acute respiratory syndrome (SARS) would need a sample size of about 200 cases and appropriate statistical methods enabling the inclusion of cases with defined periods of exposure. However, no such studies have been reported so far. Besides, determinants of the SARS incubation period remain unclear. In this study, 209 probable SARS cases with documented episodes of exposure between March 1 and May 31, 2003, in mainland China were included. A nonparametric method was used to analyze these data with defined periods of exposure to obtain the refined estimate of the SARS incubation period. Furthermore, the authors also explored the influence of various factors on the SARS incubation period by analysis of variance, linear regression analysis, and analysis of covariance. The estimates of mean and variance of the SARS incubation period were 5.29 days and 12.33 days(2), respectively; 90% of patients would have an incubation period of less than 11.58 days with a probability of 0.8, and 99% of patients would have an incubation of less than 22.22 days with a probability of 0.9. The affected area showed a highly significant effect on the incubation period (p < 0.001), but the contact pattern, occupation, gender, and age did not.

Prenatal fetal sex diagnosis by detecting amelogenin gene in maternal plasma.

OBJECTIVES: To provide a new, reliable noninvasive method for fetal sex determination. METHODS: Fetal sex was detected in 32 early pregnant women by identifying the amelogenin gene in maternal plasma using nested PCR analysis. First, the 122/128 bp of X-Y homologous region containing 6 bp deletions in the intron 3 of amelogenin gene in X chromosome was amplified, and then the nested PCR was carried out, whose 3' end of the upstream primer is just located in the deletion region. The fetus was male or female, depending on whether it had the 89-bp nested PCR product or not. RESULTS: The 89 bp of nested PCR product was detected in 19 plasma samples obtained from pregnant women, deducing they bear the male fetus and the remaining pregnant women bear female. When compared with the birth outcome, two samples were pseudo-positive. The coincidence was 93.8%. This method had high sensitivity that even trace amount of target fetal DNA (10 pg) could be detected. CONCLUSIONS: This conventional nested PCR analysis of amelogenin gene promises to be a reliable method for noninvasive fetal sex determination at early pregnancy using maternal plasma DNA.