Objective To construct two DNA vaccines encoding Gag-Env fusion protein and Tat-Rev-Integrase(C-half)-Vif-Nef fusion protein derived from the first HIV-1 CRF01_AE isolate(AE2f) in Chi-na and to evaluate the immunogenicity in mice. Methods Two DNA vaccines were constructed by inserting the codon optimized and synthesized gag-env fusion gene and tat-rev-integrase(c-half)-vif-nef fusion gene de-rived from AE2f into mammalian expression vector pDRVISV1. 0, the generated DNA vaccines were desig-nated as pSVAE/GE and pSVAE/TRIVN, respectively, and their in vitro expression were determined by Western blot with transfected 293T cells. Mice were i. m. immunized with either pDRVI1.0 as mock control, pSVAE/GE or pSYAE/TRIVN for 4 times at two-week interval. Two weeks following the final im-munization, cellular responses to pool of HIV-1 Env, Gag, Tat, Rev, Intergrase, Vif and Nef peptides were evaluated by ELISPOT assay. Results The construction of DNA vaccine pSVAE/GE and pSVAE/TRIVN was validated by restriction enzyme digestion and bidirectional sequencing. Western blot showed a specific band at molecular mass 220×10~3 in lane of pSVAE/GE transfeeted 293T cell and a specific band at 95×10~3 in the lane of pSVAE/TRIVN. Both DNA vaccines mounted significant specific T cell responses with (3010 ± 566) SFC/10~6 splenocytes for DNA vaccine pSV AE/GE and (948 ± 737) SFC/10~6 spleno-cytes for DNA vaccine pSVAE/TRIVN, whereas the mock control of pDRVISV1.0 only raised marginal T cell responses. Conclusion Both pSVAE/GE and pSVAE/TRIVN were capable of expressing the inserted fusion immunogen genes and able to elicit vigorous cellular immune responses, therefore, these DNA vac-cines are highly immunogenic.

Objective To investigate and compare the features of the HIV-1-specific CTL responses among three HIV-infected groups with varied infection history. Methods Three HIV-infeeted groups were enrolled in this study, including two groups infected by blood transmission (one group has been infected for more than 10 years and the other for 1-2 years) and one group of the man who have sex with man. The HIV-1-specific CTL responses were quantified by an IFN-γ based ELISPot assay with a peptide matrix system containing overlapping peptides spanning the entire HIV-1 Clade B genomic consensus sequences. Results The responding rate of CTL responses against all 17 peptide pools among the group that infected 1-2 years,the group infected more than 10 years and the group of MSM were 40% ,65% ,23%. One way ANOVO analysis showed that the responding rate of CTL responses against all 17 peptide pools were statistical significant among the three groups (F=19.96, P＜0.01);the magnitude of CTL responses of the three groups were 0-5 835 SFCs/106 PBMC, 0-7 225 SFCs/106PBMC, 0-9 740SFCs/106pBMC, Kruskal-Wallis test showed that the magnitude of CTL responses were statistical significant among the three groups( H = 101.90 , P ＜0.01);the breadth of CTL were 7 ( 2-11 ), 11(9-14) and 4 (2-6) respectively and Kruskal- Wallis test showed that the breadth of CTL had no statistical significant among the three groups( H = 34. 75 ,P ＜0. 01 ). The sequence of responding rate, magnitude and breadth of CTL from high to low was the group that had been infected for more than 10 years, the group infected 1-2 years and the sex transmission group. The common characteristics of the CTL response among the three groups were that the responding rate and the magnitude of the peptide Nef and Gag was higher than other peptide's. The magnitude of CTL responses among three different CD4count groups (CD4 ＜ 200/μl, CD4 200-500/μl, CD4 ≥500/μl,) was 0-18 475 SFCs/106pBMC, 350-34 095 SFCs/106pBMC, 490-21 550 SFCs/106 PBMC and had no statistic difference among the three different CD4 groups(H=2.93, P=0.23) while the breadth of CTL was 3(0-8), 10(2-17), 10 (1-17)respoctively and the breadth of CTL was lower in the group of CD4 count less than 200/μl than the other two groups( H = 14. 72, P ＜ 0. 01 ). The magnitude of CTL responses among three different viral load (VL)groups (VL＜ LDL, LDL ＜ VL ＜ 1 × 104 copys/ml, VL≥1 ×104 copys/ml) was 490-18 475 SFCs/106pBMC, 0-24 115 SFCs/106pBMC, 770-34 095 SFCs/106 pBMC and had no statistic difference among the three different viral load groups ( H = 0.79, P=0.67) and the breadth of the three different viral load groups CTL was 8( 1-17), 11 (0-17), 8 (1-16) and Kruskal-Wallis test showed that there was no statistic difference among the three different viral load groups (H =5.27, P =0. 07). Conclusions All groups predominantly develope T cell immune responses against Nef and Gag proteins. With the elapse of HIV infection, the CTL responses are increased in both magnitude and responding rate. This information is important for vaccine development.

Microalgae biodiesel can solve these problems currently of plants materials,such as:shortage of arable land,impact of climate change for production and to lead high crop prices and so on.Constructing "engineered microalgae" through transgenic technology,the microalgae have capacity of high growth,shorter periods of growth and several times higher oil production than terrestrial plants.Furthermore,sea water can be as its natural medium for industrial production.The advantages of microalgae biodiesel,current status and progress of researches on engineered microalgae as well as product technologies of microalgal biodiesel was introduced.

OBJECTIVETo investigate the antimicrobial resistance and penicillin resistance-associated genes (TEM and pbp2B) of Streptococcus pneumoniae (S. pneumoniae) isolated from sputum specimens of Guangzhou children with respiratory tract infection.

METHODSE-test and Kirby-Bauer methods were applied to detect the antibiotic susceptibility of 44 strains of S. pneumoniae. PCR was used to detect resistance genes pbp2B and TEM, followed by DNA sequence analysis of pbp2B gene. The sequence results were compared to those of penicillin-susceptible S. pneumoniae R6.

RESULTSOf the 44 isolates of S. pneumoniae, only 5 (11.4%) were susceptible to penicillin. All strains were resistant to erythromycin but susceptible to ofloxacin and vancomycin. The resistance rate of the isolates to clindamycin and trimoxazole was more than 90%. The S. pneumoniae isolates showed a high susceptibility to amoxicillin, imipenem and ceftriaxone, with a resistance rate of 0, 2.6% and 3.9%, respectively. The sequence analysis showed that more than 99% nucleotide sequence of pbp2B gene of five penicillin-susceptible isolates was the same as penicillin-susceptible S. pneumoniae R6, without any amino acid replacement. Site mutation was found in the remaining 39 penicillin-nonsusceptible isolates with a nucleotide mutation rate ranging from 13.2% to 23.1% and amino acid replacement rate from 6.5% to 10.9%. The 39 penicillin-nonsusceptible isolates were classified into 4 types according to the mutation site between Ser391 and Thr492 of pbp2B: type I (n=30), type II (n=7), type III (n=1) and type IV (n=1). No TEM gene was detected in all the 44 S. pneumoniae isolates.

CONCLUSIONSThe S.pneumoniae isolates from Guangzhou children with respiratory tract infection are resistant to penicillin and erythromycin. Amoxicillin and the third generation cephalosporin may be recommended for treating S. pneumoniae infection. The mutation of pbp2B gene plays an important role in the development of S. pneumoniae resistance to penicillin.

Aminoacyltransferases ; genetics ; Child, Preschool ; Drug Resistance, Bacterial ; genetics ; Female ; Humans ; Infant ; Male ; Microbial Sensitivity Tests ; Penicillin Resistance ; genetics ; Penicillin-Binding Proteins ; genetics ; Respiratory Tract Infections ; microbiology ; Streptococcus pneumoniae ; drug effects ; genetics ; beta-Lactamases ; genetics

To purify recombinant human nucleoside diphosphate kinase A (rhNDPK-A) efficiently in pilot scale, cells of rhNDPK-A producing E. coli were homogenized by high pressure under 4 degrees C, 950 Pa. The insoluble debris was removed by microfiltration and the soluble portion was concentrated by ultrafiltration. The resulted crude sample was loaded on DEAE-sepharose Fast Flow. The target fraction was collected and then load on Cibacron Blue 3GA Sepharose CL-4B. Eluted with buffer containing ATP from the AC column, rhNDPK-A was polished with ultrafiltration. The results showed that after homogenized 2 rounds, 1500g cells of E. coli brought crude sample containing 47.6g NDPK-A. Treated with microfiltration and ultrafiltration, 27.3g of NDPK-A were recovered from this bacteria homogenate. After 2-step purification with column chromatography and then polished with ultrafiltration, 17.2 g rhNDPK-A were collected with purity of 96.3%. The recovery of the whole purification process was 36.2%, and the productivity of rhNDPK-A was 1.15 g per 100 g wet cells. Comparing the recovery of each purification step, it was found that the recovery of polish is higher than that of affinity chromatography, which is higher than that of ion exchange chromatography. The limit step was the process of sample pretreatment among the 4 purification steps. Combine with the fermentation results reported before, it was deduced that the productivity of rhNDPK-A was 510 mg/L. In conclusion, an easily controlled purification condition with high yield provides material for the translation researches of NDPK; In addition, it was suggested the crucial step determine the recovery of non-secretive recombinant proteins might be the process of sample pretreatment, not be the process of column chromatography.
Chromatography, Affinity ; Chromatography, Ion Exchange ; Electrophoresis, Polyacrylamide Gel ; Escherichia coli ; genetics ; Humans ; NM23 Nucleoside Diphosphate Kinases ; genetics ; metabolism ; Pilot Projects ; Recombinant Proteins ; isolation & purification ; metabolism ; Ultrafiltration

Objective To explore the application of post setting and performance management in the central sterile supply.Methods The study was introduced on January 1,2015,for a period of one year,and ended on December 31,2015.56 staffs in sterile supply center were divided into four levels and three levels to develop their duties,theory and skills training,and comparative analysis of the results of training was done;on the other hand,quantitative performance evaluation standards were developed.Comparison of work quality and self evaluation,theoretical and practical assessment,and satisfaction rate in clinical departments was done.Results Before and after the intervention for 12 months statistics,it is found that,in post setting and performance management in the implementation of post-intervention compared with the pre-intervention in the center,the quality and efficiency and the overall quality of staff,the department satisfaction has improved significantly,and the difference is statistically significant (P＜0.05).After the intervention,high quality and efficiency staff in Disinfection Supply Center increases from 14.29％ up to 80.36％,and the effect is significant.After intervention,excellent rate of staff assessment theory and skills assessment at different levels increases by 20％ ～ 50％.Satisfaction Degree of clinical departments increases from about 65％ before intervention to 80％ and above after intervention.Conclusion Post setting and performance management helps to improve disinfection center staff's enthusiasm,and improve job performance continuously to improve and enhance the overall quality of the staff.

OBJECTIVETo investigate the outcome of orthotopic heart transplantation for patient with end-stage hypertrophic cardiomyopathy.

METHODSThis retrospective review analyzed the clinical data of nine patients (7 males) undergoing orthotopic heart transplantation for end-stage hypertrophic cardiomyopathy in our center. All patients received induced therapy protocols peri-operative and standard triple maintenance immunosuppressive therapy postoperative.

RESULTSOne recipients developed acute renal failure due to renal artery embolism and allograft rejection in the early posttransplantive course, symptoms and signs were improved under continuous renal replacement therapy and steroid-pulse therapy, this patient died of sudden cardiac arrest at 32 months post transplantation. Another recipient developed demyelinating disease in frontal and parietal lobe and finally recovered with medical therapy. Eight patients survived the operation with good quality of life and there was no episode of rejection or infection or chronic graft arteriosclerosis during follow-up time. Three recipients developed left ventricular hypertrophy and there were no signs of grapg-vessel diseases in the survivals.

CONCLUSIONHeart transplantation is the best therapeutic option for selected patients with end-stage hypertrophic cardiomyopathy.

Adolescent ; Adult ; Cardiomyopathy, Hypertrophic ; surgery ; Female ; Heart Transplantation ; Humans ; Male ; Middle Aged ; Postoperative Period ; Retrospective Studies ; Treatment Outcome ; Young Adult

BACKGROUNDTo isolate and identify pathogen of atypical pneumonia in Guangdong.

METHODSPathogens were isolated from variety of samples collected from atypical pneumonia patient by using MDCK cells, and identified with serological and molecular methods.

RESULTSA novel coronavirus was isolated from patients with atypical pneumonia, from which an RNA fragment of 279 nt was amplified by nested RT-PCR. And sequence assay showed that only 39-65 percent of sequence of the virus was homogenous to known coronavirus, but almost 100% homogenous (with one base exception, 12a to t) to SARS-associated coronavirus isolated from patients outside Guangdong, such as in Beijing, Hong Kong, Taiwan, Germany, Italy and so on. Indirect immunofluorescence test showed a specific antigen-antibody reactivity between the coronavirus and convalescent-phase sera of SARS patients.

CONCLUSIONThe pathogen of the atypical pneumonia in Guangdong province was a novel type of coronavirus, which could be isolated by using MDCK cells.

Animals ; Base Sequence ; Cell Line ; China ; Dogs ; Humans ; Molecular Sequence Data ; Phylogeny ; Pneumonia, Viral ; virology ; SARS Virus ; classification ; genetics ; isolation & purification ; Severe Acute Respiratory Syndrome ; virology

OBJECTIVETo evaluate the feasibility and efficacy of a new method of endoscopic esophageal variceal ligation combined with partial splenic embolization (EVL-PSE) for the patients with portal hypertension.

METHODSFrom May 1999 to February 2003, sixty-eight patients with portal hypertension underwent EVL-PSE, and hemodynamics of the portal trunk (PT), the left gastric vein and azygos vein, including maximum velocity, flow volume, vein diameter, were assessed using color ultrasound Doppler.

RESULTSThe esophageal varices and hypersplenism were greatly ameliorated after operation in patients who had undergone EVL-PSE. Postoperative portal trunk flow volume and velocity were significantly reduced (P < 0.05), and flow volume of the left gastric vein as well as the azygos vein were also reduced after operation. During 2 - 24-month follow-up, no recurrent bleeding was found.

CONCLUSIONSEVL-PSE is less traumatic with less complications, and results in marked eradication of esophageal varices, it can be carried out safely in the clinical treatment for patients with portal hypertension.

Adult ; Aged ; Combined Modality Therapy ; Embolization, Therapeutic ; Esophageal and Gastric Varices ; therapy ; Esophagoscopy ; Female ; Humans ; Hypertension, Portal ; therapy ; Ligation ; Male ; Middle Aged

Acute Kidney Injury ; physiopathology ; Adolescent ; Cardiomyopathies ; pathology ; Crush Syndrome ; physiopathology ; Earthquakes ; Humans ; Male