To reconstruct the heat-labile enterotoxin subunit B (LTB) gene of Escherichia coil in order to increase the outputs of the prokaryotic expression on recombinant LTB (rLTB), and to determine its immune adjuvant activity on mucosa,the nucleotide sequence of the whole length of LTB gene was synthesized according to the preferred codons of E. coli, and the prokaryotic expression plasmid pET32a-rLTB and its expression system in E. coli BL21DE3 were reconstructed. The recombinant plasmid was extracted and the inserted sequence of rLTB gene was determined. Meanwhile, the expression quantity of the reconstructed rLTB was identified by SDS-PAGE and BioRad agarose image analyzing system, and compared with that of the un-reconstructed rLTB. The abilities of the reconstructed rLTB and the un-reconstructed rLTB to bind with bovine GM1 were determined by means of GM1-ELISA assay. By using the recombinant urease subunit B as antigen, the effects of the reconstructed and the un-reconstructed rLTB on the improvement of immune protection of BALB/c mice infected with Helicobacter pylori strain SS1 and the induction of S-IgAs in infected mice were assayed. The experimental results showed that the expression quantity of the reconstructed rLTB approached upto 35.4% of the total bacterial proteins after induction with 1 mmol/L IPTG for pET32a-rLTB-E. coliBL21DE3 and to be 12.6 times higher than that of the un-reconstructed rLTB (2.8 %). In addition, both the abilities of the recombinant reconstructed LTB and the un-reconstructed rLTB to bind with bovine GM1 could be demonstrated by GM1-ELISA. The immune protection rate of the recombinant urease subunit B in the infected mice was 66.7%; and it could reach up to 91.7% with a significantincrease of the specific S-IgA level, when it was immunized with the reconstructed or the un-reconstructed rLTB. It is concluded that the reconstructed LTB gene in the present study shows a remarkable increased outputs of expression of this gene with a strong immune adjuvant activity on mucosa.

Objective To observe the influences of Shendan Sanjie capsule on chemotherapy effect and immune index to patients with advanced non-small cell lung cancer.MethodsDividing 76 patients with advanced non-small cell lung cancer from department of oncology from January 2014 to May 2016,each with 38 cases.The control group recei-ved western chemotherapy,observation group received western chemotherapy with Shendan Sanjie capsule.The differences of related indicators were compared.Results①After chem-otherapy,in the control group, CD4+decreased,CD8+ increased and CD4+/CD8+ decreased,and before chemotherapy, the difference was significant(P<0.05).In observation group,there was no significant difference in CD4+, CD8+, CD4+ / CD8+ levels.The above indicators of observation group were better than that of control group.②After chemotherapy,RR(CR+PR)rate of observation group were 57.89%, higher than the control group with 36.84%(P<0.05).③During chemotherapy,toxic side effect ratio of observation group was lower than that of the control group(P<0.05).ConclusionComparing Shen Dan Sanjie capsule can adjust patient immunity and reduce side effects of chemotherapy during chemotherapy.

Objective To understand the predominant β-lactamase genotypes and their carrying modes ofEscherichia coli isolates in Zhejiang province,and the effects of β-1actam antibiotics on inducing or histidine kinase inhibitor closantel (CLO) on inhibiting the expression of β-1actamase genes.Methods Micro-dilution method and E-test were applied to measure the resistant rate and minimal inhibitory concentration (MIC) in E.coli isolates against β-1actam antibiotics.PCR and sequence analysis of PCR products were conducted to detect the β-lactamase genotypes and their carrying modes.Real-time fluorescent quantitative RT-PCR and β-lactamase confirmation test were performed to determine the influence of 1/4 MIC penicillin and cefotaxime,and CLO on the transcription and expression of β-lactamase genes in the resistant E.coli isolates.Results Among the 462 E.coli strains isolated in Zhejiang,285 (61.7％) were resistant to penicillin,ampicillin,cefoxitin,cefotaxim and ceftazidime.In the 285 resistant isolates,the detection rate of TEM or CTX-M β-1actamase gene (83.2％ or 75.1％) was significantly higher than that of KPC,SHV or OXA β-lactamase gene (1.4％-10.2％) (P＜0.01) and the carrying rate of two or more β-1actamase genes (68.8％) was also significantly higher than that of single β-1actamase gene (31.2％) (P＜0.01),and 61.4％ of the resistant isolates carried TEM + CTX-M genes (P＜0.01).Except KPC gene,1/4 MIC of cefotaxim and penicillin induced a rapid increase of TEM-mRNA,CTX-M-mRNA,SHV-mRNA or OXA-mRNA levels (P＜0.01),but 50-500 μg/ml CLO inhibited these levels (P＜0.01).After pre-treatment with 100 μg/ml CLO,82.8％-85.6％ of the resistant isolates became sensitive to β-lactam antibiotics (P＜0.01),while the detection rate of β-lactamases was also decreased from 95.1％ to 16.1％ (P＜0.01).Conclusion TEM and CTX-M are the predominant β-lactamase genotypes in E.coli isolates in Zhejiang and TEM+CTX-M is the predominant carrying mode of β-lactamase genes.Low concentrations of β-lactam antibiotics can up-regulate the expression levels of β-lactamase genes in E.coli through bacterial two-component signaling systems,but this effect can be inhibited by CLO,a histidine kinase inhibitor.