Activation of CaMKII via ER-stress mediates coxsackievirus B3-induced cardiomyocyte apoptosis.

Cardiomyocyte apoptosis contributes to the development of coxsackievirus B3 (CVB3)-induced myocarditis, but the mechanism for the apoptosis by CVB3 infection remains unclear. Here, we showed that CVB3-induced endoplasmic reticulum (ER) stress response and apoptosis in cultured H9c2 cardiomyocytes. We found that Ca2+ -calmodulin-dependent kinase II (CaMKII) was activated by ER stress-dependent intracellular Ca2+ overload in the CVB3-infected H9c2 cardiomyocytes. Treatment with an inhibitor of ER stress, 4-phenylbutyric acid (4-PBA), attenuated intracellular Ca2+ accumulation indirectly and reduced CaMKII activity. Inhibition of CaMKII with pharmacological inhibitor (KN-93) or short hairpin RNA reduced CVB3-induced H9c2 apoptosis and repressed cytochrome c release from mitochondria to cytoplasm; whereas overexpression of the activated mutant of CaMKII (CaMKII-T287D) enhanced CVB3-induced H9c2 apoptosis and mitochondrial cytochrome c release, which could be alleviated by blocking of mitochondrial Ca2+ uniporter or mitochondrial permeability transition pore. Further in vivo investigation revealed that blocking of CaMKII with KN-93 prevented cardiomyocytes apoptosis and improved cardiac contractile function in CVB3-infected mouse heart. Collectively, these findings provide a novel evidence that CaMKII plays a vital role in the promotion of CVB3-induced cardiomyocyte apoptosis, which links ER stress and mitochondrial Ca2+ uptake.

Effects of PGC1α on myocardial ischemia reperfusion injury and the underlying mechanisms. / PGC1αå¯¹å¿ƒè‚Œç¼ºè¡€å†çŒæ³¨æŸä¼¤çš„ä½œç”¨åŠå ¶æœºåˆ¶.

OBJECTIVES: Peroxisome proliferator-activated receptor gamma coactivator 1α (PGC1α) controls mitochondrial biogenesis, but its role in cardiovascular diseases is unclear. The purpose of this study is to explore the effect of PGC1α on myocardial ischemia-reperfusion injury and the underlying mechanisms. METHODS: The transverse coronary artery of SD rat was ligated for 30 minutes followed by 2 hours of reperfusion. Triphenyltetrazolium chloride (TTC) staining was performed to measure the area of myocardial infarction. Immunohistochemistry and Western blotting were used to detect the PGC1α expression in myocardium. The rat cardiomyocyte H9C2 was subjected to hypoxia/reoxygenation (H/R) with the knockdown of PGC1α or hypoxia- inducible factor 1α (HIF-1α), or with treatment of metformin. Western blotting was used to detect the expression of PGC1α, HIF-1α, p21, BAX, and caspase-3. CCK-8 was performed to detect cell viability, and flow cytometry was used to detect apoptosis and mitochondrial superoxide (mitoSOX) release. RT-qPCR was used to detect the mRNA expression of PGC1α and HIF-1α. Besides, chromatin immunoprecipitation (ChIP)-qPCR and luciferase reporter gene assay were applied to detect the transcriptional regulation effect of HIF-1α on PGC1α. RESULTS: After I/R, the PGC1α expression was increased in infarcted myocardium. H/R induced H9C2 cell apoptosis (P<0.001). The release of mitoSOX (P<0.001) and protein expression of PGC1α, and apoptosis-related p21, BAX, and caspase-3 were increased. However, knockdown of PGC1α reduced apoptosis (P<0.01) and mitoSOX release (P<0.001), and decreased protein expression of apoptosis-related gene. HIF-1α is bound to the promoter region of PGC1α. Knockdown of HIF-1α inhibited the transcription and protein expression of PGC1α and further to reduce the apoptosis (all P<0.001) and mitoSOX release (P<0.01). While overexpression of PGC1α reversed the effects caused by HIF-1α knockdown (all P<0.001). Metformin effectively reduced H/R-induced apoptosis (P=0.013), mitoSOX release (P<0.001), HIF-1α, PGC1α and apoptosis-related protein expression, recovered the cell viability (P<0.001), and reduced myocardial infarction (P=0.003). CONCLUSIONS: After I/R, HIF-1α up-regulates the expression of PGC1α, leading to an increase in ROS production and aggravation of injury. Metformin can inhibit the accumulation of HIF-1α during hypoxia and effectively protect myocardium from ischemia/reperfusion injury.

Complete genome sequence and comparative genome analysis of a new special Yersinia enterocolitica.

Yersinia enterocolitica is the most diverse species among the Yersinia genera and shows more polymorphism, especially for the non-pathogenic strains. Individual non-pathogenic Y. enterocolitica strains are wrongly identified because of atypical phenotypes. In this study, we isolated an unusual Y. enterocolitica strain LC20 from Rattus norvegicus. The strain did not utilize urea and could not be classified as the biotype. API 20E identified Escherichia coli; however, it grew well at 25 °C, but E. coli grew well at 37 °C. We analyzed the genome of LC20 and found the whole chromosome of LC20 was collinear with Y. enterocolitica 8081, and the urease gene did not exist on the genome which is consistent with the result of API 20E. Also, the 16 S and 23 SrRNA gene of LC20 lay on a branch of Y. enterocolitica. Furthermore, the core-based and pan-based phylogenetic trees showed that LC20 was classified into the Y. enterocolitica cluster. Two plasmids (80 and 50 k) from LC20 shared low genetic homology with pYV from the Yersinia genus, one was an ancestral Yersinia plasmid and the other was novel encoding a number of transposases. Some pathogenic and non-pathogenic Y. enterocolitica-specific genes coexisted in LC20. Thus, although it could not be classified into any Y. enterocolitica biotype due to its special biochemical metabolism, we concluded the LC20 was a Y. enterocolitica strain because its genome was similar to other Y. enterocolitica and it might be a strain with many mutations and combinations emerging in the processes of its evolution.

[Risk assessments and control strategies of plague in five key surveillance counties, Zhejiang province].

OBJECTIVE: To analyze the epidemiology data on plague in five counties in Zhejiang province and to evaluate the risk of plague in theses areas. METHODS: We selected five monitoring stations as a risk assessment (Qingyuan county, Longquan city, Yiwu city, Wencheng county, and Ruian city) in Zhejiang province where the plague epidemic more serious in the history. At least one constant site and 1-4 variable sites where plague occurred in history were selected for monitoring. We collected the five counties (cities) surveillance data of indoor rat density, indoor Rattus flavipectus density, the Xenopsylla cheopis index of rat, the Xenopsylla cheopis index of Rattus flavipectus in 1995-2014. Isolation of Yersinia pestis was conducted among 171,201 liver samples and F1 antibody were detected among 228,775 serum samples. Risk matrix, Borda count method, and Delphi approach were conducted to assess risk of the plague of five counties (cities) in Zhejiang province. RESULTS: Indoor rat density in Qingyuan county, Longquan city, Yiwu city, Wencheng county, Ruian city was 1.58%-5.50%, 1.13%-9.76%, 0.56%-3.67%, 2.83%-16.08%, 7.16%-15.96%, respectively; Indoor Rattus flavipectus density of five counties (cities) was 0.08%-2.23%, 0-2.02%, 0-0.54%, 0.71%-5.58%, 0.55%-4.92%, respectively. The Xenopsylla cheopis index of rat in Qingyuan county and Wencheng county was 0.011-0.500 and 0.015-0.227, respectively; The Xenopsylla cheopis index of Rattus flavipectus of Qingyuan county and Wencheng county was 0.119-3.412 and 0.100-1.430, respectively; Ruian City and Yiwu city cannot collected Xenopsylla cheopis, Long quan city only collected the Xenopsylla cheopis index of rat in the five years. Yersinia pestis were not isolated in five counties (cities).There were 3 Apodemus agrarius samples positive of plague F1 antibody test, in Longquan city and Yiwu city in 2005. Borda count method to assess the Longquan city, Yiwu (Borda point were both 321) plague risk was higher than three other regions; Delphi approach to evaluation five counties (cities) belong to the plague had a lower risk areas, according to the level of risk score (Pf) Longquan city and Yiwu (Pf was 0.314, 0.292, respectively) plague risk were higher than three other regions (Pf were all 0.292). CONCLUSION: The main host and media were lower in five key plague surveillance counties (cities) of Zhejiang province; The result of Borda count method and Delphi approach for risk assessment indicated that endogenous plague recrudescence was at lower level, but Longquan city and Yiwu city risk were higher than other counties (cities).

Homology analysis of pathogenic Yersinia species Yersinia enterocolitica, Yersinia pseudotuberculosis, and Yersinia pestis based on multilocus sequence typing.

We developed a multilocus sequence typing (MLST) scheme and used it to study the population structure and evolutionary relationships of three pathogenic Yersinia species. MLST of these three Yersinia species showed a complex of two clusters, one composed of Yersinia pseudotuberculosis and Yersinia pestis and the other composed of Yersinia enterocolitica. Within the first cluster, the predominant Y. pestis sequence type 90 (ST90) was linked to Y. pseudotuberculosis ST43 by one locus difference, and 81.25% of the ST43 strains were from serotype O:1b, supporting the hypothesis that Y. pestis descended from the O:1b serotype of Y. pseudotuberculosis. We also found that the worldwide-prevalent serotypes O:1a, O:1b, and O:3 were predominated by specific STs. The second cluster consisted of pathogenic and nonpathogenic Y. enterocolitica strains, two of which may not have identical STs. The pathogenic Y. enterocolitica strains formed a relatively conserved group; most strains clustered within ST186 and ST187. Serotypes O:3, O:8, and O:9 were separated into three distinct blocks. Nonpathogenic Y. enterocolitica STs were more heterogeneous, reflecting genetic diversity through evolution. By providing a better and effective MLST procedure for use with the Yersinia community, valuable information and insights into the genetic evolutionary differences of these pathogens were obtained.

Evaluation of the effect of concurrent chronic total occlusion and successful staged revascularization on long-term mortality in patients with ST-elevation myocardial infarction.

AIMS: To investigate the impact of chronic total occlusion (CTO) in non-infarct-related artery (IRA) on the long-term prognosis and evaluate the clinical significance of staged revascularization in patients with ST-segment elevation myocardial infarction (STEMI). METHODS: 1266 STEMI patients with primary percutaneous coronary intervention (PCI) were categorized as single-vessel disease (SVD), multivessel disease (MVD) without and with CTO. We study the clinical outcomes of patients after primary PCI in the following 3 years. Additionally, patients with CTO received staged revascularization, and major adverse cardiac events (MACE) during 3-year follow-up were recorded. RESULTS: Presence of CTO was a predictor of both early mortality [hazard ratio (HR) 3.4, 95% confidence interval (CI) 2.4-4.5, P < 0.01] and late mortality (HR 1.9, 95% CI 1.4-3.6, P < 0.01), whereas MVD without CTO was only a predictor of early mortality (HR 1.7, 95% CI 1.3-2.3, P < 0.05). In CTO group, 100 patients had successful CTO recanalization, and 48 patients failed. During 3-year follow-up, patients with failed procedure had higher cardiac mortality (22.9% versus 9.0%, P = 0.020) and lower MACE-free survival (50.0% versus 72.0%, P = 0.009) compared to patients with successful procedure. CONCLUSION: The presence of CTO and not MVD alone is associated with long-term mortality. Successful revascularization of CTO in the non-IRA is associated with improved clinical outcomes in patients with STEMI undergoing primary PCI.

Case Report: A case of Chlamydia psittaci infection in an HIV patient.

Chlamydia psittaci is the pathogen of psittacosis and infects a wide range of birds and even humans. Human infection occurs most commonly in those with a history of contact with birds or poultry. We describe a case of psittacosis in a human immunodeficiency virus infected patient in Zhejiang Province for the first time. C. psittaci infection was confirmed by nested polymerase chain reaction (PCR) and Real-Time PCR. Phylogenetic analysis revealed that the sequences from the patient's samples clustered with genotype A in the same branch. Our study highlights the possibility of diagnosing psittacosis in patients with a chronic disease such as HIV-infected patients, and should increase awareness and surveillance of psittacosis in China.

Anion-induced self-assembly of luminescent and magnetic homoleptic cyclic tetranuclear Ln4(salen)4 and Ln4(salen)2 complexes (Ln = Nd, Yb, Er, or Gd).

Unique homoleptic cyclic tetranuclear Ln(4)(Salen)(4) complexes [Ln(4)(L)(2)(HL)(2)(µ(3)-OH)(2)Cl(2)]·2Cl (Ln = Nd, 1; Ln = Yb, 2; Ln = Er, 3; Ln = Gd, 4) or Ln(4)(Salen)(2) complexes [Ln(4)(L)(2)(µ(3)-OH)(2)(OAc)(6)] (Ln = Nd, 5; Ln = Yb, 6; Ln = Er, 7; Ln = Gd, 8) have been self-assembled from the reaction of the hexadentate Salen-type Schiff-base ligand H(2)L with LnCl(3)·6H(2)O or Ln(OAc)(6)·6H(2)O (Ln = Nd, Yb, Er, or Gd), respectively (H(2)L: N,N'-bis(salicylidene)cyclohexane-1,2-diamine). The result of their photophysical properties shows that the strong and characteristic NIR luminescence for complexes 1-2 and 5-6 with emissive lifetimes in microsecond ranges are observed, and the sensitization arises from the excited state (both (1)LC and (3)LC) of the hexadentate Salen-type Schiff-base ligand with the flexible linker. Temperature dependence (1.8-300 K) magnetic susceptibility studies of the eight complexes suggest the presence of an antiferromagnetic interaction between the Ln(3+) ions.

A cluster of Psittacosis cases in Lishui, Zhejiang Province, China, in 2021.

Introduction: Psittacosis, caused by Chlamydia psittaci, is widespread throughout the world. In humans, C. psittaci infection may lead to severe conditions and complications, including sepsis and multiple organ failure. We report a cluster of cases caused by C. psittaci in Zhejiang Province, 2021, which led to one death and three cases of hospitalization. Methods: The cases were confirmed by nest-PCR, RT-PCR, and mNGS. Results: The four cases were related and the sequences obtained from the samples were closely correlated with those from Taiwan. Discussion: This study is the first to report on the case of death from psittacosis in Zhejiang Province, and our results help to assess the disease and recommend effective measures to prevent further spread of C. psittaci.

Bacteroides fragilis Supplementation Deteriorated Metabolic Dysfunction, Inflammation, and Aorta Atherosclerosis by Inducing Gut Microbiota Dysbiosis in Animal Model.

BACKGROUND: The gut microbial ecosystem is an important factor that regulates host health and the onset of chronic diseases, such as inflammatory bowel diseases, obesity, hyperlipidemia, and diabetes mellitus, which are important risk factors for atherosclerosis. However, the links among diet, microbiota composition, and atherosclerotic progression are unclear. METHODS AND RESULTS: Four-week-old mice (-/- mice, C57Bl/6) were randomly divided into two groups, namely, supplementation with culture medium (control, CTR) and Bacteroides fragilis (BFS), and were fed a high-fat diet. The gut microbiota abundance in feces was evaluated using the 16S rDNA cloning library construction, sequencing, and bioinformatics analysis. The atherosclerotic lesion was estimated using Oil Red O staining. Levels of CD36, a scavenger receptor implicated in atherosclerosis, and F4/80, a macrophage marker in small intestine, were quantified by quantitative real-time PCR. Compared with the CTR group, the BFS group showed increased food intake, fasting blood glucose level, body weight, low-density lipoprotein level, and aortic atherosclerotic lesions. BFS dramatically reduced Lactobacillaceae (LAC) abundance and increased Desulfovibrionaceae (DSV) abundance. The mRNA expression levels of CD36 and F4/80 in small intestine and aorta tissue in the BFS group were significantly higher than those in the CTR group. CONCLUSIONS: gut microbiota dysbiosis was induced by BFS. It was characterized by reduced LAC and increased DSV abundance and led to the deterioration of glucose/lipid metabolic dysfunction and inflammatory response, which likely promoted aorta plaque formation and the progression of atherosclerosis.

De Winter syndrome as an emergency electrocardiogram sign of ST-elevation myocardial infarction: a case report.

The case report aims to reveal de Winter's electrocardiogram (ECG) pattern as an equivalent to anterior ST-segment elevation myocardial infarction (STEMI). We report a case of a 49-year-old man with a history of smoking who presented to the emergency department with a 1 day history of chest pain that was exacerbated 5 h prior to presentation. Detailed clinical investigations and coronary angiographic characteristics were recorded. The first ECG of the patient was consistent with de Winter syndrome. Acute coronary artery angiography showed that the proximal left anterior descending coronary artery was completely occluded after the first diagonal branch artery was given off. A percutaneous coronary intervention was immediately performed. Our case indicates that early identification and diagnosis of such ECGs and timely reperfusion therapy of de Winter syndrome as a STEMI equivalent are required to improve the prognosis of such patients.

[Establishment and evaluation of identification method for Yersinia pestis and Yersinia pseudotuberculosis].

OBJECTIVE: To establish a gene identification method of Yersinia pestis and Yersinia pseudotuberculosis for plague surveillance. METHODS: According to the specific genomic sequences of Y. pestis and Y. pseudotuberculosis, i.e. "pestis Island (PeI)" and "pseudotuberculosis Island (PsI)" and the published genomic sequences of 12 strains of Y. pestis and 4 strains of Y. pseudotuberculosis, the specific identification primers of these sequences were designed. RESULTS: A total of 52 strains of Y. pestis and 57 strains of Y. pseudotuberculosis and other intestinal bacteria strains were tested with PCR. Of the 5 pairs of Y. pestis identification primers, PeI2 and PeI11 were specific for Y. pestis. Besides Y. pestis, the primers PeI1, PeI3 and PeI12 could detect part of 57 Y. pseudotuberculosis strains. Of the 5 pairs of Y. pseudotuberculosis identification primers, PsI1 could detect all the 52 strains of Y. pestis and 57 strains of Y. pseudotuberculosis. PsI7, PsI16, PsI18 and PsI19 were specific for Y. pseudotuberculosis. CONCLUSION: The primers PsI1, PeI 2 and PeI11, PsI7, PsI16, PsI18 and PsI19 can be used in the rapid identification of Y. pestis and Y. pseudotuberculosis, which can be also used to explore the circulation of atypical Y. pestis in quiescent plague foci.

Ecology and geographic distribution of Yersinia enterocolitica among livestock and wildlife in China.

The results in this study show the prevalence of Yersinia enterocolitica varies in different animal species and regions of China. The highest prevalence is among pigs (12.91%), followed by dogs (9.80%), Ochotona curzoniae (plateau pica) (6.76%), chickens (4.50%), rodents (3.40%), cattle (2.78%) and sheep (0.89%). Pathogenic isolates comprised the majority of the Y. enterocolitica recovered from pigs (73.50%) and dogs (59.44%); whereas the nonpathogenic Y. enterocolitica made up most of poultry and wildlife recovered strains. A correlation analysis comparing the prevalence and geographic factors showed the isolation rate of Y. enterocolitica in pigs and dogs was negatively correlated with elevation (r=-0.50, P<0.05) and annual average air temperature (r=-0.43, P<0.05), but there was positive correlation with annual precipitation (r=0.43, P<0.05); conversely, the isolation rate from wildlife is positively correlated with elevation (r=0.3, P<0.05) contrary to the result seen in livestock. Twelve novel biotype 2 pathogenic Y. enterocolitica carried ail and ystB virulence genes, and one biotype 1A nonpathogenic strain positive with ail, ystB and ystA genes were isolated from Microtus fuscus (Qinghai vole) on plague foci of the Qinghai-Xizang plateau. The PFGE pattern K6GN11C30021 was predominant in pigs (44.25%) and patients (41.18%); K6GN11C30068 was predominant in dogs (40.16%). Animal isolates from the same region shared the same pattern (K6GN11C30021 and K6GN11C30012), indicating they may be from the same clone and arose through cross infection. Moreover, the identical PFGE pattern among local animals and diarrhea patients suggested that the animals may be the source of infections in these areas.

Achalasia treatment: improved outcome of laparoscopic myotomy with operative manometry.

HYPOTHESIS: Operative manometry detects residual esophagogastric junction (EGJ) high pressure, ensuring complete myotomy. DESIGN: Consecutive patients undergoing laparoscopic myotomy. SETTING: Tertiary care academic medical center. PATIENTS: From 1997 to 2003, 139 patients with achalasia underwent laparoscopic myotomy. INTERVENTIONS: We assessed myotomy completeness by operative endoscopy and performed operative manometry to measure pressure across the EGJ myotomy. Residual high pressure was isolated and intact muscle divided. MAIN OUTCOME MEASURES: Esophageal manometry, quality of life, and dysphagia severity score. RESULTS: Median lower esophageal sphincter pressure was 27 mm Hg preoperatively; 10 patients had sigmoid esophagus and 57 had previous dilation and/or toxin injection. There were 136 laparoscopic myotomies and 3 conversions to open procedures (2%). Operative endoscopy was performed in all patients. Operative manometry, completed in 132 patients (95%), identified residual EGJ high pressure leading to myotomy revision in 45 patients (31 in the first 70 treated). Small perforations occurred in 19 patients, associated with previous dilation and/or toxin injection in 12 patients. One-month follow-up was available in 136 patients (98%); 126 patients had minimal symptoms (93%), whereas 1 had recurrent EGJ high pressure, 5 esophagitis, 3 sigmoid esophagus, and 1 paraesophageal hernia. In 60 patients with complete 1-year follow-up, quality of life and dysphagia improved (P <.05); mean lower esophageal sphincter pressure decreased to 7.6 mm Hg (P <.05). CONCLUSIONS: Operative manometry identifies residual EGJ high pressure and reduces the incidence of incomplete myotomy. Laparoscopic myotomy improves quality of life and dysphagia symptoms and may be the treatment of choice in most patients with achalasia.

Does laryngectomy improve swallowing after chemoradiotherapy? A case study.

Organ preservation protocols of high-dose chemoradiotherapy have become fairly common to treat head and neck cancers. However, significant swallowing problems can occur. This study examines swallowing, oral tongue pressures, and tongue base-to-pharyngeal wall pressures in a patient who underwent total laryngectomy for improvement of swallowing after chemoradiotherapy for treatment of a hypopharyngeal tumor. The patient underwent concurrent videofluorographic and manometric examination of swallowing and examination of oral tongue pressures after the laryngectomy. One healthy subject was used as a control. After the laryngectomy, the patient no longer aspirated; however, he could swallow only liquids and pureed foods. He demonstrated difficulty with bolus clearance through the oral cavity and pharyngocervical esophagus. Pharyngeal pressures were reduced compared with those of the control subject. While total laryngectomy will stop unremitting aspiration, swallowing after chemoradiation may be severely compromised. This may not be overcome by total laryngectomy.

Two novel ail-positive biotype 1A strains of Yersinia enterocolitica isolated in China with unequal adhesion and invasion properties.

Yersinia enterocolitica is an enteric pathogen having six biotypes: 1A, 1B, 2, 3, 4, and 5. Different bioserotypes have been associated with varying pathogenicity, and the strains of biotype 1A lack the virulence-associated pYV-bearing genes and were once considered to be avirulent. However, there is growing epidemiological, clinical, and experimental evidence to suggest some biotype 1A isolates are virulent and can cause gastrointestinal disease. Here, we describe two biotype 1A strains discovered from 3807 isolates that carry the ail (attachment and invasion locus) gene. The two strains showed unique PFGE patterns compared to all other isolates in the Chinese Y. enterocolitica isolate PFGE database. Strain SDWL-003 isolated from a sheep shared ail sequence identical to A1 pattern, and the foxA (ferrioxamine receptor) sequence was identical to the pathogenic F5 pattern, besides, the PFGE patterns of SDWL-003 was also cluster to pathogenic branch; however it does not attach to or invade Hep-2 cells. The ail sequence of strain 2006RAT isolated from a Microtus fortis showed several mutations compared to other published genomes, and therefore formed an entirely new pathogenic pattern. Though it clustered to non-pathogenic block with foxA sequence polymorphism analysis or PFGE assay, the strain 2006RAT showed adhesion properties. The data here bring new insights into the molecular genetics of Y. enterocolitica biotype 1A, show some isolates of 1A biotype gaining potential pathogenicity using the function of the virulence gene - ail, and indicate the lateral gene transfer of ail virulence genes proceeded between pathogenic and nonpathogenic Y. enterocolitica.

[Clinical application of DNA sequencing for detecting point mutations in hepatitis B virus associated with drug resistance].

OBJECTIVE: [corrected] To assess the specificity and applicability of direct PCR sequencing in the detection of point mutations in hepatitis B virus (HBV) associated with drug resistance. METHODS: Serum samples were obtained from 120 patients with hepatitis B treated with nucleoside analogus for at least 2 years to detect the point mutations in HBV genome in association with drug resistance using nested PCR and direct DNA sequencing. RESULTS: Forty out of the 120 patients were found to have one or two point mutations associated with drug resistance, including 17 with L180M and M204V/I mutations (42.5%), 10 with M204V/I mutation (25%), 8 with N236T mutation (20%), 3 with L180M mutation (7.5%), and 1 with both A181V/T and N236T mutations (2.5%), and 1 with A181V/T mutation(2.5%). CONCLUSION: DNA sequencing is a good method to detect all known point mutations associated with HBV drug resistance.

Hetero-binuclear near-infrared (NIR) luminescent Zn-Nd complexes self-assembled from the benzimidazole-based ligands.

With the compound [Zn(HL(1))(2)(Py)] (H(2)L(1)=2-(1H-benzo[d]imidazol-2-yl)-6-methoxyophenol), Py=pyridine) or [Zn(HL(2))(2)(Py)] (H(2)L(2)=2-(1H-benzo[d]imidazol-2-yl)-4-bromo-6-methoxyphenol) as the precursor, complexes [ZnLn(HL(1))(2)(Py)(NO(3))(3)] (Ln=Nd, 1; Ln=Gd, 2) or [ZnLn(HL(2))(2)(Py)(NO(3))(3)] (Ln=Nd, 3; Ln=Gd, 4) were obtained by the further reaction with Ln(NO(3))(3)·6H(2)O (Ln=Nd or Gd). The result of their photophysical properties shows that the strong and characteristic near-infrared (NIR) luminescence of Nd(3+) ions for complexes 1 and 3 with emissive lifetimes in microsecond range, has been sensitized from the excited state ((1)LC and (3)LC) of the benzimidazole-based ligands, and the involvement of heavy atoms (Br) at the ligand endows the enhanced NIR luminescent property.

[Clone expression of the caf1 gene of Yersinia pestis and immunological evaluation on recombinant F1 antigen].

OBJECTIVE: To get recombinant F1 antigen (rF1) and to construct the detection dipstick of plague antibody. METHODS: The caf1 gene removing the signal peptide coding sequence was cloned into plasmid pET32a(+) by double-digested sites of BamHI and Not I. Recombinant plasmid caf1-pET32a(+) was transformed into BL21 (DE3) and the rF1 was expressed. Expression products were purified by affinity chromatography. Dual detection dipstick of plague antibody was constructed with purified rF1 and natural F1, and evaluated with 528 human serum samples of Zhejiang province. RESULTS: The fusion protein rF1 of 35.5 KD was expressed by BL21 strains containing caf1-pET32a(+). The sensitivity of rF1 showed equivalent to or higher than the natural F1 antigen in detecting plague antibody. It seemed that there was a better consistency of 97.9% (kappa = 0.466) when 528 human sera was detected by rF1 and natural F1. CONCLUSION: We successfully extracted the rF1 with good immunological activity that might be used to detecting Yersinia pestis.

[Study on the application and evaluation of methods for gene and antigen detection in plague surveillance program].

OBJECTIVE: To apply and evaluate new methods regarding specific gene and antigen detection in plague surveillance program. METHODS: 1798 samples from natural foci of plague were tested, using internal quality control multiple-polymerase chain reaction, F1 antigen marked by immuno chromatographic assay and enzyme linked immunosorbent assay. Culture of Yersinia pestis and reverse indirect hemagglutination assay were used as reference diagnostic methods. RESULTS: The overall positive rate of culture on Yersinia pestis together with gene and antigen detection was 7.34%, showing an 16.81% increase when comparing to 6.28% using Yersinia pestis culture method alone. The rate of coincidence was 97.13%. CONCLUSION: The new standard being used for specific gene and antigen detection could increase the positive rate of diagnosis on plague.