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Objective:To propose a markerless beam's eye view (BEV) tumor tracking algorithm, which can be applied to megavolt (MV) images with poor image quality, multi-leaf collimator (MLC) occlusion and non-rigid deformation.Methods:Window template matching, image structure transformation and demons non-rigid registration method were used to solve the registration problem in MV images. The quality assurance (QA) plan was generated in the phantom and executed after manually setting the treatment offset on the accelerator, and 682 electronic portal imaging device (EPID) images in the treatment process were collected as fixed images. Meanwhile, the digitally reconstructured radiograph (DRR) images corresponding to the field angle in the planning system were collected as floating images to verify the accuracy of the algorithm. In addition, a total of 533 images were collected from 21 cases of lung tumor treatment data for tumor tracking study, providing quantitative results of tumor location changes during treatment. Image similarity was used for third-party verification of tracking results.Results:The algorithm could cope with different degrees (10%-80%) of image missing. In the phantom verification, 86.8% of the tracking errors were less than 3 mm, and 80% were less than 2 mm. Normalized mutual information (NMI) varied from 1.182±0.026 to 1.202±0.027 ( P<0.005) before and after registration and the change of Hausdorff distance (HD) was from 57.767±6.474 to 56.664±6.733 ( P<0.005). The case results were predominantly translational (-6.0 mm to 6.2 mm), but non-rigid deformation still existed. NMI varied from 1.216±0.031 to 1.225±0.031 ( P<0.005) before and after registration and the change of HD was from 46.384±7.698 to 45.691±8.089 ( P<0.005). Conclusions:The proposed algorithm can cope with different degrees of image missing and performs well in non-rigid registration with data missing images which can be applied in different radiotherapy technologies. It provides a reference idea for processing MV images with multi-modality, partial data and poor image quality.
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Objective:To evaluate the feasibility of predicting lung cancer target position by online optical surface motion monitoring.Methods:CT images obtained in different ways of stereotactic body radiotherapy (SBRT) plans from 16 lung cancer cases were selected for experimental simulation. The planned CT and the original target position were taken as the reference, and the 10 phases of CT in four dimension CT and each cone beam (CBCT) were taken as the floating objects, on which the floating target location was delineated. The binocular visual surface imaging method was used to obtain point cloud data of reference and floating image body surface, while the point cloud feature information was extracted for comparison. Based on the random forest algorithm, the feature information difference and the corresponding target area position difference were fitted, and an online prediction model of the target area position was constructed.Results:The model had a high prediction success rate for the target position. The variance explainded and root mean squared error ( RMSE) of left-right, superior-inferior, anterior-posterior directions were 99.76%, 99.25%, 99.58%, and 0.0447 mm, 0.0837 mm, 0.0616 mm, respectively. Conclusion:The online monitoring of lung SBRT target position proposed in this study is feasible, which can provide reference for online monitoring and verification of target position and dose evaluation in clinical radiotherapy.
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With the gradual increase in the diagnosis rate of early gastric cancer, clinicians must consider prevention of gastric anatomical structure and physiological function while ensuring the radical treatment of the tumor. Pylorus-preserving gastrectomy is a function- preserving operation that preserves the pylorus, inferior pyloric vessel, and the vagus nerve in patients with early middle gastric cancer. One of the major controversies at present is the thoroughness of limited lymph node dissection for pyloric-preserving gastrectomy. Various studies have reported that the lymph node metastasis rate of early middle gastric cancer was low, especially in the suprapyloric region, inferior pylorus and the upper pancreatic region. Partial lymph node dissection is required for vascular and neurological protection, which is also safe and feasible in studies reported by major centers. Many clinical studies have been carried out in Japan and Korea, and postoperative follow-up has gradually increased evidence, providing the basis for the safety of lymph node dissection. In large case studies comparing pylorus- preserving gastrectomy with traditional distal gastrectomy, the incidence of postoperative morbidity, such as dumping syndrome, bile reflux esophagitis, weight loss, and malnutrition is low. Sentinel lymph node navigation technology is gradually applied to the diagnosis and treatment of early gastric cancer, and its clinical application value still needs further research.
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Humans , Pylorus/pathology , Stomach Neoplasms/pathology , Gastrectomy , Gastroenterostomy , Lymph Node ExcisionABSTRACT
Objective:To investigate the clinical efficacy of redo rectal resection and coloanal anastomosis.Methods:The retrospective and descriptive study was conducted. The clinicopatholo-gical data of 49 patients who underwent redo rectal resection and coloanal anastomosis for the treatment of local recurrence of tumors and failure of colorectal or coloanal anastomosis after rectal resection in the Sixth Affiliated Hospital of Sun Yat-sen University from November 2012 to December 2021 were collected. There were 32 males and 17 females, aged 57(range,31-87)years. Redo rectal resection and coloanal anastomosis was performed according to the patient′s situations. Observa-tion indicators: (1) surgical situations; (2) postoperative situations; (3) follow-up. Measurement data with normal distribution were represented as Mean± SD, and measurement data with skewed distri-bution were represented as M( Q1, Q3) or M(range). Count data were described as absolute numbers or percentages. Results:(1) Surgical situations. All 49 patients underwent redo rectal resection and coloanal anastomosis successfully, with the interval between the initial surgery and the reopera-tion as 14.2(7.1,24.3)months. The operation time and volume of intraoperative bold loss of 49 patients in the redo rectal resection and coloanal anastomosis was 313(251,398)minutes and 125(50,400)mL, respectively. Of the 49 patients, there were 38 cases receiving laparoscopic surgery including 12 cases with transanoscopic laparoscopic assisted surgery, 11 cases receiving open surgery including 2 cases as conversion to open surgery, there were 20 cases undergoing Bacon surgery, 14 cases undergoing Dixon surgery, 12 cases undergoing Parks surgery, 2 cases undergoing intersphincter resection and 1 case undergoing Kraske surgery, there were 20 cases undergoing rectum dragging out excision and secondary colonic anastomosis, 13 cases undergoing dragging out excision single anastomosis, 12 cases undergoing rectum dragging out excision double anastomosis, 4 cases undergoing first-stage manual anastomosis, there were 21 cases with enterostomy before surgery, 16 cases with prophylactic enterostomy after surgery, 12 cases without prophylactic enterostomy after surgery. The duration of postoperative hospital stay of 49 patients was (14±7)days. (2) Postoperative situations. Fifteen of 49 patients underwent postoperative complications, including 8 cases with grade Ⅱ Clevien-Dindo complications and 7 cases with ≥grade Ⅲ Clevien-Dindo complications. None of 49 patient underwent postoperative transferring to intensive care unit and no patient died during hospitalization. Results of postoperative histopathological examination in 23 patients with tumor local recurrence showed negative incision margin of the surgical specimen. (3) Follow-up. All 49 patients underwent post-operative follow-up of 90 days. There were 42 cases undergoing redo rectal resection and coloanal anastomosis successfully and 7 cases failed. Of the 37 patients with enterostomy, 20 cases failed in closing fistula, and 17 cases succeed. There were 46 patients receiving follow-up with the median time as 16.1(7.5,34.6)months. The questionnaire response rate for low anterior resection syndrome (LARS) score was 48.3%(14/29). Of the patients who underwent redo coloanal anastomosis and closure of stoma successfully, there were 9 cases with mild-to-moderate LARS.Conclusion:Redo rectal resection and coloanal anastomosis is safe and feasible for patients undergoing local recurr-ence of tumors and failure of colorectal or coloanal anastomosis after rectal resection, which can successfully restore intestinal continuity in patients and avoid permanent enterostomy.
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Objective:To propose a machine learning-based markerless beam′s eye view (BEV) tumor tracking algorithm that can be applied to low-quality megavolt (MV) images with multileaf collimator (MLC)-induced occlusion and non-rigid deformation.Methods:This study processed the registration of MV images using the window template matching method and end-to-end unsupervised network Voxelmorph and verified the accuracy of the tumor tracking algorithm using dynamic chest models. Phantom QA plans were executed after the treatment offset was manually set on the accelerator, and 682 electronic portal imaging device (EPID) images obtained during the treatment were collected as fixed images. Moreover, the digitally reconstructed radiography (DRR) images corresponding to the portal angles in the planning system were collected as floating images for the study of target volume tracking. In addition, 533 pairs of EPID and DRR images of 21 lung tumor patients treated with radiotherapy were collected to conduct the study of tumor tracking and provide quantitative result of changes in tumor locations during the treatment. Image similarity was used for third-party validation of the algorithm.Results:The algorithm could process images with different degrees (10%-80%) of data missing and performed well in non-rigid registration of images with data missing. As shown by the phantom verification, 86.8% and 80% of the tracking errors were less than 3 mm and less than 2 mm, respectively, and the normalized mutual information (NMI) varied from 1.18 ± 0.02 to 1.20 ± 0.02 after registration ( t = -6.78, P = 0.001). The tumor motion of the clinical cases was dominated by translation, with an average displacement of 3.78 mm and a maximum displacement of 7.46 mm. The registration result of the cases showed the presence of non-rigid deformations, and the corresponding NMI varied from 1.21 ± 0.03 before registration to 1.22 ± 0.03 after registration ( t = -2.91, P = 0.001). Conclusions:The tumor tracking algorithm proposed in this study has reliable tracking accuracy and high robustness and can be used for non-invasive and real-time tumor tracking requiring no additional equipment and radiation dose.
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In recent years， with the change in life style， social environment， and national childbearing policy， the proportion of high-risk pregnant women has increased significantly， triggering the spectrum of obstetric diseases to constantly change， which has brought new challenges to the diagnosis and treatment of obstetrics. Traditional Chinese medicine （TCM） has been proved effective in dealing with a variety of obstetric diseases， and various treatment methods are available， which can serve as alternative means for solving refractory obstetric diseases. However， most obstetric clinicians are currently less aware of the therapeutic effects of TCM， which has significantly hindered its participation in clinical treatment. Therefore， the China Association of Chinese Medicine （CACM） organized the outstanding young obstetricians of TCM and western medicine to discuss 15 obstetric diseases responding specifically to TCM or integrated TCM and western medicine， including hyperemesis gravidarum， threatened abortion， ectopic gestation， cough during pregnancy， pregnancy-induced hypertension syndrome， maternal-fetal ABO incompatibility， postpartum hypogalactia， residual pregnancy tissue in uterine cavity， puerperal infection， pantalgia after childbirth， hematoma/undesirable healing after caesarean section， postpartum urinary retention， ileus after cesarean section， pelvic floor dysfunction， and postnatal depression. The suggestions for their treatment with TCM or integrated TCM and western medicine were also proposed， aiming to provide patients with effective and personalized treatments in clinical practice and improve the diagnosis and treatment effects of obstetric diseases， thus benefiting the public. At the same time， more obstetrical clinicians are expected to understand the therapeutic effects and advantages of TCM and draw on the strengths of both TCM and western， thereby promoting the establishment of an obstetric diagnosis and treatment system with Chinese characteristics.
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Objective Tofacilitatetheradiologiststoinquireinformationrelatedtobreastdiseasesandimprovetheefficiencyand accuracyofsearchingdata.Methods Inthispaper,amultilevelstrategyhybridquestion-and-answermodelwasusedtodesignanautomatic question-and-answersysteminthefieldofbreastdiseasesimaging.Thefirstlayerofthismodelwasthequick matchingofquestion sentencesandFAQknowledgebase.Thesecondlayerwastoobtaintheanswersinthebasicknowledgebaseaccordingtothethresholdsetby processingthequestionsandcalculatingthesimilarityofthequestions.Thethirdlayerwastoacquiretheanswerparagraphfromthe webdocumentreturnedbythefull-textsearchengine.Results Thetestresultsshowedthattheaccuracyofthesystemansweracquisitionreached 85%.Thesystemcouldgivesatisfactoryanswerstotheproblemsexistingintheknowledgebase.Conclusion Theintelligentquestion answeringsystemforbreastdiseaseisconvenientandfast,anditisaneffectivetoolforradiologiststoinquiretheknowledgeofbreast diseases.
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Objective To detect the serum levels of calpainin (S100A11) using nanomagicbeabs sorting-time resolved fluoroimmuno assay (NMBS-TRFIA) and evaluate its diagnostic value in pancreatic carcinoma.Methods 88 patients with pancreatic carcinoma,50 patients with acute pancreatitis,10 patients with pancreatic cyst and 20 healthy controls were selected as the study subjects.The human peripheral serum blood was sorted with S100A11 antibody coupled nanomagicbeabs,and the concentration of S100A11 was detected by TRFIA method.The area under the receiver operating characteristic (ROC) curve (AUC) was used to determine the cut-off level for diagnosis of pancreatic carcinoma,in order to evaluate the sensitivity and specificity for diagnosis of pancreatic carcinoma.Results S100A11 showed a linear relationship within the range of 6.08～ 500 ng/ml using NMBS-TRFIA method,intraassay CV≤6.35％,inter-assay CV≤7.12％,and the average recovery rate was 104.7％.The serum levels of S100A11 in patients with pancreatic carcinoma,patients with acute pancreatitis and patients with pancreatic cyst were 185.53 ± 161.19,106.06±113.83 and 68.99± 47.83 ng/ml respectively.Compared with the normal control group (37.98±25.14 ng/ml),the differences were statistically significant (t=-8.065,-3.375,-2.266,all P ＜0.01).The serum levels of S100A11 in patients with pancreatic carcinoma was significantly higher than those in patients with acute pancreatitis and patients with pancreatic cyst (all P＜0.05).According to the ROC curve,ROCAUC=0.985 (95％ CI:0.972 ～ 0.997),the best cut-off level for the diagnosis of pancreatic carcinoma was 89.5 ng/ml (sensitivity 81.8 ％,specificity 67.5 ％).Conclusion NMBS-TRFIA can enrich S100A11 in serum and improve the detection sensitivity of serum S100A11,and the method is simple and easy to be popularized.Serum S100A11 has a high sensitivity and specificity in the diagnosis of pancreatic carcinoma,and is a new serum marker for the diagnosis of early pancreatic carcinoma.
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ST-segment elevation myocardial infarction (STEMI) in patients with multivessel disease is a common situation, and suggests poor prognosis. After the reperfusion of infarct-related blood vessels in the primary PCI (PPCI), how to deal with the non-infarct related vessel has been a hot issue. In recent years, more and more evidence suggests that compared with treatment of infarct-related vessels only, revascularization of non-infarct related vessels will result in a better prognosis. But the timing and the choice of non-infarct-related vessel revascularization are still controversial. This article reviews the latest clinical evidence to improve the diagnosis and treatment of STEMI patients with multivessel disease.
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A method of identification of C=C location and relative quantitation of unsaturated phosphatidylcholine ( PC ) isomers in breast cells by online photochemical reaction-pulsed directed current electrospray-tandem mass spectrometry ( PB-pulsed-dc-ESI-MS/MS) was established with benzophenone ( BP) as a photochemical reactant. The three-phase extraction method was used to extract the lipids in the cells, and then the C=C in the unsaturated PC and the carbonyl in BP were specifically cycled under the irradiation of 254 nm ultraviolet light (Paternò-Büchi, PB reaction). The PB products were ionized and mass-isolated for low-energy collision dissociation through the non-contact pulsed-dc-ESI ionization method. The double bond position and the relative content of the location isomers were obtained from the resulting ions in the MS/MS spectrum. The C=C location of 8 kinds of unsaturated PCs in MCF-7 and MCF-10A was detected, and the relative contents of 4 kinds of C=C location isoforms were analyzed. It was found that the relative abundance of △9 isomer in PC 16:018:1 was not significantly different between the two cells. The relative abundance of△9 isomers in PC 18:018:1 and PC 18:118:1 was slightly different. However, there is a big difference of △9 in LPC 18:1 between the cancer cell and normal cell (56. 0% ± 1. 3% vs. 71. 7% ± 6. 8%). The establishment of such a rapid and easy mass spectrometry method can analyze the C=C location and the relative content of location isomers, and it is expected to be a powerful tool to identify different cell states and different disease states.
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Objective:To compare the correlation of four drug-resistance interpretation system including HIVDB, ANRS, REGA and HIV GRADE for testing HIV-1 genotype drug resistance. Methods:Trendy subtypes and restructuring model of HIV-1 including B’,CRF01_AE and CRF07_BC were selected,each genotype was 200 series,600 samples were analyzed by four drug?resistance interpretation system, the results were divided into three levels including resistance (R), possible drug?resistance (I) and susceptible (S). Results:Four drug?resistance interpretation system had a high correlation in genotypic drug?resistance consequence (rs>0.57, P<0.01), CRF07_BC had the highest correlation, followed by subtype B, CRF01_AE was slightly lower.Conclusion:Four drug?resistance interpretation system has a high correlation in analyzing antiviral drug resistance for major epidemic of HIV in China.
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As highly active anti-retroviral therapy continues, large numbers of drug-resistant strains appeared in human immunodeficiency virus ( HIV) infectors, which always leads to a decline of treatment results or even a treatment failure.The genotypic resistance analysis technique of the bioinformatics is a powerful tool to forecast the HIV resistance and also determines the success or failure of antiviral therapy.This review will be focus on the advantages and disadvantages, influence factor of the genotype resistance prediction and improve measure of the primary HIV drug resistance genotype interpretation system, depending on the principle and characteristics of the genotypic drug resistance analysis as the breakthrough point, in order to provide guidance for reasonable application of different genotypes drug resistance analysis system in China.
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Objective To evaluate the efficacy and safety of adjuvant IFN therapy for viral hepatitis-related hepatocellular carcinoma(HCC)after treatment with surgical resection or transarterial chemoembolization(TACE).Methods Controlled trials of adjuvant treatment with IFN for patients with HCC published between 2000 and 2012 were searched electronically in MEDLINE,PubMed,Cochrane Library,and EMBASE databases.According to the heterogeneity of the studies,two different models-the fixed-effect model and the random-effect model-were applied to analyze the results. Results Ten trials were screened according to inclusion and exclusion standards.Eight randomized,controlled trials and two non-randomized,controlled trials were included.These ten trials with a total of 1 ,029 subjects were eventually involved in the meta-analysis;528 HCC patients were treated with adjuvant treatment with IFN and 501 patients with placebo.Compared to the control group,the recurrence rates of HCC in IFN group were significantly lower CR=0.66;95% CI=0.50 to 0.86;P=0.02),especially after TACE treatment according to subgroup analysis(OR=0.73;95%CI=0.52 to 1.01;P=0.06 for surgical resection;and OR=0.54;95%CI=0.33 to 0.86,P=0.01 for TACE).The death rates in the IFN group also significantly decreased according to not only total events analysis(OR=0.42;95%CI=0.32 to 0.56;P<0.01)but also subgroup analysis(OR=0.51;95%CI=0.36 to 0.72;P=0.0002 for surgical resection;and OR=0.33;95%CI=0.21 to 0.50;P<0.01 for TACE).Conclusion Adjuvant IFN therapy may significantly reduce the recurrence rates of patients with viral hepatitis-related HCC and improve the survival of patients after surgical resection or TACE.The ideal dose mostly selected is 3 MIU/mL,three times per week,which can make patients tolerate the adverse reactions of IFN better and maintain effective concentrations for a long time.
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Objective To observe the effects of Haizao Yuhu Decoction on the CXCR 4 expression of TT cell tumor-bearing mice. Methods Culture the human thyroid TT cells in vitro, and inoculate them into 30 healthy mice BALB/C. Subsequently, Haizao Yuhu Decoction was given to the tumor-bearing mice. They are grouped into three groups, including tumor-bearing group (n=10), Haizao Yuhu Decoction high dosage group (n=10) and Haizao Yuhu Decoction low dosage group (n=10) to observe the tumor growth to calculate the anti-tumor rate. After the tumor has been reaped, immunohistochemical is adopted to test the expressions of chemotactic factor receptor-CXCR 4 on protein level. RT-PCR is used to test the expressions of CXCR 4 on genetic level. Results Comparing with the tumor-bearing group, HZYHD high-dosage group has decreased the expressions of CXCR 4 on both the protein and genetic levels, which is to certain degree dependent on concentration.Conclusion Haizao Yuhu Decoction could decrease the expression of CXCR 4.
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Execution is one of the key factors which influence the management level of hospital libraries. The paper introduces the concept, performance and synthesis patterns of execution, and points out that execution is the main means to enhance performance of li-brary, then puts forward concrete strategies to enhance execution of library from two levels: individuals and libraries.
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<p><b>OBJECTIVE</b>To investigate the clinical curative effect of the combination between medicine and acupoint flow on SOD, NO in patients with ulcerative colitis.</p><p><b>METHOD</b>Two hundred sixty two patients with ulcerative colitis were randomly divided into four groups: the patients in traditional Chinese medicine group were fed with changpikang, the patients in ear acupoint group were pasted and pressed spleen, large intestine, sympathesis, subcortex; the patients in medicine and acupoint group were taken with Changpikang and ear acupoint; the patients in treatment group were treated by taking sulfasalazine, with a treatment course of four weeks. The changes of SOD, NO before and after treatments were recored.</p><p><b>RESULT</b>There was a significant difference between four groups before and after treatment (P < 0.05), medicine and acupoint group was superior to that others (P < 0.05).</p><p><b>CONCLUSION</b>The method of combination between medicine and acupoint could increase the contents of SOD, decrease the level of NO, elevate the potential of organism to resist oxygen free radical and promote intestinal tract ulcer concrescence.</p>
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Adult , Aged , Female , Humans , Male , Middle Aged , Young Adult , Acupuncture, Ear , Methods , Colitis, Ulcerative , Drug Therapy , Metabolism , Therapeutics , Drugs, Chinese Herbal , Therapeutic Uses , Medicine, Chinese Traditional , Methods , Nitric Oxide , Metabolism , Sulfasalazine , Therapeutic Uses , Superoxide Dismutase , Metabolism , Treatment OutcomeABSTRACT
<p><b>OBJECTIVE</b>To characterize CRF01_AE strains of recombinant human immunodeficiency virus type-1 (HIV-1) found in the Second National Molecular Epidemiology Study on HIV in China and to analyze its sequence variation in the env V3-C3 region during the First National Molecular Epidemiology Study (NMES1, 1996 - 1998) to the Second National Molecular Epidemiology Study (NMES2, 2001 - 2002).</p><p><b>METHODS</b>DNA was extracted from peripheal blood mononuclear cells of the subjects with HIV infection. The env C2-V4 region of HIV-1 was amplified with nested polymerase chain reaction (n-PCR). PCR products were directly sequenced using ABI 377 DNA sequencer, then the gene-based phylogenetic tree was constructed and its variation of amino acids was analyzed with GCG software.</p><p><b>RESULTS</b>Totally, 169 strains of recombinant HIV-1 CRF01_AE were identified from blood samples collected from different high risk groups in 17 of 31 provinces, municipalities and autonomous regions all over China by the end of 2002. Although sexual transmission still dominated during NMES1 (62.2%, 23/37) and NMES2 (55.3%, 73/132), prevalence of HIV-1 CRF01_AE in intravenous drug users (IDUs) increased to 41.6% (57/137) during NMES2 from 27% (10/37) during NMES1. Phylogenetic tree analysis showed that HIV-1 CRF01_AE strains prevalent in IDUs during NMES2 did not cluster with those prevalent in the subjects infected by sexual transmission during NMES2 and those in IDUs during NMES1. The amino acid residues of V3 region of HIV-1 CRF01_AE in IDUs were relatively conservative, but the sixth, eighth, ninth, tenth, twelfth, fifteenth, sixteenth amino acid residues of C3 region displayed regular changes.</p><p><b>CONCLUSIONS</b>HIV-1 CRF01_AE strain has been introduced into inland provinces from southeastern coast areas and southwestern border areas, with an increasing prevalence in IDUs. The sequence of env V3-C3 region of recombinant HIV-1 CRF01_AE strains prevalent in IDUs during NMES2 was obviously different from that during NMES1, suggesting that HIV-1 CRF01_AE strains prevalent in IDUs during NMES2 might come from a new source and have a potential to spread.</p>
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Humans , Amino Acid Sequence , China , Epidemiology , Genes, env , Genetics , Genetic Variation , Genome, Viral , HIV Infections , Epidemiology , Genetics , Virology , HIV-1 , Classification , Genetics , Molecular Epidemiology , Molecular Sequence Data , Phylogeny , RNA, Viral , Genetics , Recombination, Genetic , Sequence Homology, Amino Acid , Substance Abuse, Intravenous , VirologyABSTRACT
<p><b>OBJECTIVE</b>The current available assays for HIV subtyping, such as sequence-based phylogenetic analysis or heteroduplex mobility assay (HMA), are labor-intensive and time-consuming. The authors have just developed a simple and rapid subtype-screening assay for subtypes B, C, and CRF01-AE using a single nested multiplex PCR.</p><p><b>METHODS</b>Proviral DNA from HIV-positive samples was extracted and subjected to first round PCR with universal primers for gag region that can detect HIV-1 M group isolates. In the second round PCR, three pairs of subtype-specific primers, respectively detecting subtype B, C and CRF01-AE, were added into one tube. The PCR products of different subtypes could be distinguished in agarose-gel electrophoresis. Another pair of primers exclusively detecting the prevalent recombinant B/C strains CRF07-BC and CRF08-BC were designed and used. Additionally, all of these samples were sequenced and analyzed phylogenetically.</p><p><b>RESULTS</b>Phylogenetic analysis showed that out of 119 samples, there were 43 subtype B samples (Euro-American B 11, Thailand B 32), 54 subtype C, 17 CRF01-AE, 3 subtype A and 2 subtype D samples. The subtype B, C, and CRF01-AE specific primer sets detected 35 (81.4%), 46 (85.2%), and 13(76.5%) samples with accuracy and specificity. Non-specific bands occasionally appeared but did not interfere with interpretation of the results. The primer pairs for CRF07-BC and CRF08-BC amplified target sequences were confirmed by sequencing and phylogenetic analysis. The specificity of all these subtype-specific primers was found to be 100%.</p><p><b>CONCLUSION</b>A simple and rapid assay was developed for screening subtypes B, C, CRF01-AE, CRF07-BC and CRF08-BC in China. This assay may have potential application in HIV laboratories in China and worldwide.</p>
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Humans , Acquired Immunodeficiency Syndrome , Virology , Base Sequence , China , DNA Primers , HIV-1 , Classification , Genetics , Phylogeny , Polymerase Chain Reaction , Methods , Sensitivity and SpecificityABSTRACT
<p><b>OBJECTIVE</b>To collect background information on drug-resistant HIV-1 strains in various regions before the start of nation-wide antiretroviral therapy in China.</p><p><b>METHODS</b>Twenty percent of the 2,000 blood samples from antiretroviral therapy naive patients collected for the 2nd national HIV molecular epidemiology survey (NHMES) in 2002 were randomly sampled for this study. The entire protease gene and 20-230 amino acids of the reverse transcriptase gene were amplified by PCR from provirus DNA and sequenced. The results were analyzed with HIV db-Drug Resistance Algorithm and genotypic resistance mutations were determined to particular anti-HIV drugs.</p><p><b>RESULTS</b>Totally 164 protease gene sequences and 138 reverse transcriptase gene sequences were obtained from patients; 0.61% of 164 sequences displayed primary resistance mutations in the protease gene, whereas 99.39% carried 1 or more secondary mutations. Genotypic resistance to at least one nucleoside reverse transcriptase inhibitors (NRTI) was present in 5.80%,and resistance to at least one non-nucleo side reverse transcriptase inhibitors (NNRTI) was present in 1.45% of samples.</p><p><b>CONCLUSION</b>The prevalence of genotypic drug resistance is very low in drug-naive HIV infected patients from 21 provinces of China tested in this study. Laboratories participated in the NHMES have organized a network to provide drug resistance monitoring service in the current nation-wide antiviral treatment program in China.</p>
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Humans , Anti-HIV Agents , Therapeutic Uses , China , Epidemiology , Drug Resistance, Viral , Genotype , HIV Infections , Drug Therapy , Epidemiology , Virology , HIV Protease , Genetics , HIV Protease Inhibitors , Therapeutic Uses , HIV Reverse Transcriptase , Genetics , HIV-1 , Genetics , Mutation , Reverse Transcriptase Inhibitors , Therapeutic Uses , Sentinel SurveillanceABSTRACT
<p><b>OBJECTIVE</b>To identify variations in the env gene of human immunodeficiency virus type 1 (HIV-1) subtype CRF01-AE strains circulating in China and to elucidate the potential relationship between genetic variation and evolutionary pressure.</p><p><b>METHODS</b>Fragments of the HIV-1 env gene were amplified by nested-polymerase chain reaction (n-PCR) from the whole blood of HIV-1 infected individuals from four provinces in Southeast China (Guangdong, Hunan, Jiangsu and Jiangxi). The PCR products were then directly sequenced by ABI 377 DNA sequencers. The sequences covering the env V3-V4 region of 34 HIV-1 subtype CRF01-AE strains were selected to analyse phylogenetic trees and amino acid mutations. The accumulation of synonymous (Ks) and antonymous (Ka) substitutions as well as Ks/Ka ratios were calculated using DIVERGE.</p><p><b>RESULTS</b>Phylogenetic trees showed that the 34 HIV-1 subtype CRF01-AE strains from China clustered with the Chinese AE reference strain (AE.97CNGX2F), as well as with the reference strains from Thailand (AE.CM240 and AE.93TH253). The amino acid sequences of the env V4 and C3 regions in the samples were highly variable, compared with those of V3 and V3-downstream regions. The V3 loop central motif in the majority (87.5%) of the strains was GPGQ. The majority of strains did not contain positively charged amino acids at positions 306 and 320 in V3 loop. The N-linked glycosylation sites in the V3-V4 region and flanking regions in these strains were relatively conserved. Analysis of the entire region showed that the mean Ks values were significantly higher than that of the Ka values (P < 0.001), with the Ks/Ka significantly higher than 1.0 (P < 0.001). In contrast, the Ks/Ka ratio in the V4 region was significantly lower than 1.0 (P < 0.01).</p><p><b>CONCLUSIONS</b>Our study indicated that the majority of HIV-1 subtype CRF01-AE strains circulating in China were highly homogeneous. The amino acid sequences of the V4 and C3 regions were significantly more variable than those of the V3 loop. Our analysis also suggested that the phenotype of nearly all strains was likely to be non-syncytium inducing (NSI). Finally, the variation found in the V3-V4 sequence was significantly influenced by functional constraints as opposed to positive selective pressure, while the variability of the lone V4 region was strongly related to positive selective pressure.</p>