Multilocus sequence typing MLST with high solution sensitivity and specificity is widely used to study the population genetic structure of pathogen by amplification and sequencing of the housekeeping genes. MLST also provides more evidence and plays an important role in parasite research. This paper reviews the principle and method of MLST and its applica-tion on population genetic structure analysis of parasites.

Object To establish an isolation method of theanine reference substance by preparative HPLC. Methods After raw material extracted by water, the water phase was extracted by chloroform saturated with water, then concentrated on a thermostat. After centrifuged, supernatant was isolated and purified by preparative HPLC. Fraction was frozen and dried by Flexi Drier. Crude product was rinsed by methanol. The purity of product was determined by analytical HPLC. Results The purity of product is higher than 98% and theanine yield from raw material by this method exceeds 60%. Conclusion The developed method is simple, rapid, and at low production cost. The product owns the quality of reference substance.

BACKGROUND:There are many complications of limb salvage surgery in patients with osteosarcoma of the middle tibia, and the limb salvage surgery is one of the current difficulties in clinical treatment. OBJECTIVE:To evaluate the clinical efficacy of reconstruction with massive al ograft bone for osteosarcoma of the middle tibia by retrospectively reviewing relevant cases. METHODS:Seven patients with osteosarcoma of the middle tibia were treated. And we analyzed their clinical data retrospectively. Al patients completed the formal preoperative adjuvant chemotherapy and we confirmed that there was no distant metastasis before surgery. Al patients received large al ogeneic bone transplantation and internal fixation, and the gastrocnemius muscle flap coveraged graft bone in surgery. The average length of al ogeneic bone was 12.5 cm. Five patients received postoperative adjuvant chemotherapy completely, and two patients received partly. RESULTS AND CONCLUSION:The fol ow-up period was 18-36 months. One patient had local tumor recurrence at 1 year after transplantation, and died of lung metastases after amputation. One patient survived after resection of lung metastases that occurred at 1.5 years after transplantation. One patient died of lung metastases at 2 years after transplantation. The rest four patients were tumor-free. The mean Musculoskeletal Tumor Society (MSTS) score was 26.5, the mean International Society of Limb Salvage (ISOLS) graft score was 31. Among four underage patients, one had leg length deformities, with limb shortening 2 cm. There were no postoperative infections and pathological fractures. Using large al ogeneic bone for the repair of bone defects after tumor surgery of the middle tibia can have a good clinical efficacy under the premise of strict indications. Using gastrocnemius muscle flap to cover the bone graft during surgery is an effective measure to reduce postoperative complications.

Objective:To study the effects and safety of metformin combined with vildagliptin on the glycemic control for patients with newly diagnosed type 2 diabetes mellitus.Methods:60 patients with type 2 diabetes mellitus who were treated from February 2015 to April 2016 were selected and divided into the control group and the observation group according to different treatment methods.The control group was treated with routine treatment.The observation group was treated with vildagliptin based on the control group.The blood glucose,glycosylated hemoglobin,two-hour postprandial blood glucose and serum as well as urinal amylase were measured before and after treatment,and the clinical curative effect of the two groups and the levels of interleukin-6,tumor necrosis factor and C-reactive protein were compared.Results:After treatment,the total effective rate of observation group was 90％,which was significantly higher than that of the control group(66.7％,P＜0.05).After treatment,the serum interleukin-6,tumor necrosis factor,C-reactive protein and fasting blood glucose,glycosylated hemoglobin and postprandial blood glucose levels were significantly lower than those of the control group[(7.63± 1.12)dvs(8.68± 1.30)d;(7.23± 0.95)d vs(7.89± 1.20)d;(11.14± 1.56)d vs(12.12± 1.89)d];[(12.12± 1.89)d vs(ll.20± 1.34)d;(6.89± 0.96)d vs(8.23± 1.10)d;(1.65± 0.23)d vs(3.65± 0.48)d] (P＜0.05).After treatment,the INS level of observation group was significantly lower than that of the control group (P＜0.05) and the GLP-1 level was significantly higher than that of the control group (P＜0.05).Conclusion:Metformin combined with vildagliptin could effectively control the blood glucose of patients with newly diagnosed type 2 diabetes and enhance the safety.

ObjectiveTo evaluate the effect and safety of different thrombolytic therapies for acute cerebral infarction due to occlusion of middle cerebral artery(MCA).MethodsOne hundred and thirty-two cases of acute cerebral infarction in territory of MCA were randomly divided into 3 groups,all of which were treated with alteplase.Group A (48 cases) was treated by intra-venous therapy with alteplase,group B (43 cases) was treated by infusing alteplase at the site of the internal carotid artery,and group C(41 cases) was treated by infusing alteplase into the thrombus.The improvement of neurological function,complications and mortality rate were recorded and statistically compared,with analysis of variance for counting data of normal distribution,x2 test for quantitative data,and the mean difference was significant at the 0.05level.ResultsThe effective rates of group A,B and C at 2 h,24 h,2 w were 18.8％ (9/48),39.6％ ( 19/48),45.8％ (22/48) ;39.5％ (17/43),53.5％ (23/43),58.1％ (25/43) ;78.0％ (32/41),85.4％ (35/41 ),87.8％ (36/41)respectively.The effective rate of group C was obviously better than group A( x2 =12.809,9.979,9.289,P ＜ 0.01 ) and B (x2 =31.295,19.425,17.161,P ＜ 0.01 ) with statistical significance.The effective rate of group B was better than group A at 2 h after thrombolytic therapy with statistical significance (x2 =4.801,P ＜ 0.05 ).The effective rate of group A and B did not have significant difference at 24 h,2 w after therapy ( x2 =1.765,1.375,P ＞ 0.05 ).The hemorrhage rates of group A,B and C were 14.6％ (7/48),14.0％ (6/43),7.3％ (3/41 ),the mortality rates of group A,B and C were 6.2％ (3/48),4.6％ (2/43),2.4％ (1/41),and there was no significant difference among the 3 groups ( x2 =1.328,0.786,P ＞ 0.05 ).ConclusionIt is suggested that the thrombus-imbeded thrombolytic therapy is a better way in treating acute cerebral infraction due to occlusion of MCA for its rapid and better therapeutic effect.

Osteosarcoma is a highly malignant bone tumor that predominantly affects children and adolescents. Standard chemotherapy currently used is usually poor, with a lower survival rate of patients with recurrence or metastasis. And it is necessary to find precise targeted therapy drugs from a genic perspective. This paper reviews the progress of genome research on germline mutation and somatic mutation of osteosarcoma in recent years, summarizes mutations and pathways that are closely related to osteosarcoma and can reflect the characteristics of osteosarcoma in order to provide a hope for the next generation of molecular targeted therapy.

Objective To investigate the clinical effect of using pedicled flap to reconstruct skin and soft tis-sue defect after resection of sarcoma around extremities joint. Methods Retrospectively analysis 103 cases of sar-coma around extremities joint who was treated from June, 2012 to June, 2017 and used the pedicled fasciocutaneous flaps or pedicled musculocutaneous flaps to reconstruct the defect after sarcoma resection. There were 61 males and 42 females, with a mean age of 52.3 (14-82 ) years old. Among these patients, there were 32 initial diagnosis, 30 had been resected in other units, and 41 recurrent tumor. There were 13 patients that tumor around wrist:reconstructed by radial wrist upper branch fasciocutaneous flap, dorso-ulnar flap based on supracarpal perforator, and posterior in-terosseous artery flap. Eight patients with tumor around elbow: reconstructed by lateral arm flap and media arm flap. Eight patients with tumor around shoulder: reconstructed by latissimus dorsi musculocutaneous flap and lateral tho-racic flap. Fourteen patients with tumor around hip: rectus femoris musculocutaneous flap, tensor fascia lata musculo-cutaneous flap, and posterior thigh fasciocutaneous flap. Sixteen patients that tumor around knee:reconstructed by an-terolateral low leg flap, and sural calf fasciocutaneous flap. Forty-four patients that tumor around ankle: reconstructed by saphenous neuro-veno-fasciocutaneous flap, sural neuro-veno-fasciocutaneous flap, lateral supramalleolar flap, and medial plantar flap. The defect area after tumor resection were:3 cm ×4 cm-18 cm×25 cm. After surgery, observe the situation and fitness of flap, complication, joint mobility, and tumor recurrence and metastasis. Results Allflaps survived except 6 died or partial died, with a survival rate of 94.2％, of which dead flap were replaced by a new flap or covered by free skin graft. Infection occurred in 4 cases (2 in donor site and 2 in recipient site), were cured by dressing and antibiotic. Except for 12 cases dropped, all rest 91 cases were followed-up by 33 months on average (1-5 years), flaps' shape and texture were good, without swollen and fester, and fit well. Superficial sensibility of direct fasciocuta-neous flaps and musculocutaneous flaps were normal, protective sensation of reverse fascial flaps were recovered. The mobility of affected joint show no difference compared with the other side. No ulcer occurred on flaps of plantar and but-tocks, that helps normal sit position and gait. There were 17 recurrences and 32 metastasis occurred, 21 patients died so far. There was a statistical difference between recurrent cases with first shot(10.3％) second widely resection(11.5％) and recurrent resection(30.6％)(P<0.05). Conclusion Using pedicled flap to reconstruct skin defect after the resection of sarcoma around extremities joint, can get good clinical result, with simple and reliable operation. It is worth to popularize.

Background: /Aim: It is known that an imbalance in the intestinal f lora plays a crucial role in colorectal cancer (CRC), but the effect of food consumption patterns on the types of intestinal flora remains to be clarified. We aimed to analyze the associations between food intake and intestinal flora in healthy and CRC individuals. Methods: Food intake data were recorded using the Food Frequency Questionnaire (FFQ). The composition and diversity of the intestinal flora detected by 16S rRNA gene sequencing, and the data were analyzed by R version 3.1.1 software. Results: Higher intake of red meat or pickled foods, and lower intake of white meat, fruits, vegetables, beans, nuts were found in the CRC group compared with the healthy group. Higher levels of Fusobacteria and Proteobacteria, and lower levels of Firmicutes were observed in the CRC group. Partial correlation analysis revealed that the intake of fruits, beans, and nuts was negatively correlated with Proteobacteria and Fusobacteria, but pickled food was positively correlated with Fusobacteria (p < 0.05). Fish, beans, and nuts intake was negatively correlated with Escherichia (p = 0.01). Multiple regression analysis revealed that vegetable oil (odds ratio [OR], 0.26; 95% confidence interval [CI], 0.13 to 0.82), vegetables (OR, 0.26; 95% CI, 0.10 to 0.64), eggs (OR, 0.26; 95% CI, 0.10 to 0.69), pickled foods (OR, 21.02; 95% CI, 6.02 to 73.45), and red meat (OR, 4.23; 95% CI, 1.68 to 10.60) had an impact on CRC risk. Conclusions The species and abundance of intestinal flora varies between CRC and healthy individuals and may be affected by their food preference.

Background: /Aim: It is known that an imbalance in the intestinal f lora plays a crucial role in colorectal cancer (CRC), but the effect of food consumption patterns on the types of intestinal flora remains to be clarified. We aimed to analyze the associations between food intake and intestinal flora in healthy and CRC individuals. Methods: Food intake data were recorded using the Food Frequency Questionnaire (FFQ). The composition and diversity of the intestinal flora detected by 16S rRNA gene sequencing, and the data were analyzed by R version 3.1.1 software. Results: Higher intake of red meat or pickled foods, and lower intake of white meat, fruits, vegetables, beans, nuts were found in the CRC group compared with the healthy group. Higher levels of Fusobacteria and Proteobacteria, and lower levels of Firmicutes were observed in the CRC group. Partial correlation analysis revealed that the intake of fruits, beans, and nuts was negatively correlated with Proteobacteria and Fusobacteria, but pickled food was positively correlated with Fusobacteria (p < 0.05). Fish, beans, and nuts intake was negatively correlated with Escherichia (p = 0.01). Multiple regression analysis revealed that vegetable oil (odds ratio [OR], 0.26; 95% confidence interval [CI], 0.13 to 0.82), vegetables (OR, 0.26; 95% CI, 0.10 to 0.64), eggs (OR, 0.26; 95% CI, 0.10 to 0.69), pickled foods (OR, 21.02; 95% CI, 6.02 to 73.45), and red meat (OR, 4.23; 95% CI, 1.68 to 10.60) had an impact on CRC risk. Conclusions The species and abundance of intestinal flora varies between CRC and healthy individuals and may be affected by their food preference.

Objective:To assess the effectiveness of percutaneous acetabuloplasty (PA) in combination with radiotherapy for the treatment of acetabular metastases.Methods:A retrospective analysis of medical records from 43 patients with acetabular metastases admitted between May 2017 and May 2022 were performed, with 24 cases meeting inclusion criteria. The study cohort consisted of 9 males and 15 females, with an average age of 56.0 years (range: 40-85 years). There were 12 cases on the left side and 12 cases on the right side. The primary cancer types were breast cancer (8 cases, 33%), lung cancer (7 cases, 29%), prostate cancer (4 cases, 17%), bowel cancer (2 cases, 8%), cervical cancer (1 case, 4%), kidney cancer (1 case, 4%), and liver cancer (1 case, 4%). All patients had multiple bone metastases, with 16 cases (67%) also presenting with metastases in other organs. Among the bone metastases, 19 cases (79%) were osteolytic lesions, and 5 cases (21%) were mixed lesions. Lesion distribution included 11 cases in the acetabulum, 2 cases in the acetabulum and anterior column, 8 cases in the acetabulum and posterior column, and 3 cases in the acetabulum, anterior column, and posterior column. Lesion sizes ranged from 2.0 cm×1.5 cm×2.0 cm to 4.5 cm×4.0 cm×11.0 cm. Cortical defects were observed in 11 cases (46%), and soft tissue masses were present around the acetabular metastasis in 8 cases (33%). PA was performed under local or general anesthesia, followed by local radiotherapy within 1 week after surgery (external radiotherapy, 30 Gy, 10 d). Various clinical parameters, including primary lesion location, time of tumor diagnosis, time of bone metastasis diagnosis, number and nature of bone metastases, distribution area, lesion size, presence of cortical defects and soft tissue masses, presence of other organ metastases, surgical site, operation duration, filling effect, complications, visual analog scale (VAS) scores, walking scores, Eastern Cooperative Oncology Group (ECOG) scores, and long-term complications, were recorded and compared before surgery, after surgery, after radiotherapy, at 1, 3, and 6 months post-surgery, and during the last follow-up. The median follow-up period was 18 months.Results:Among the 24 patients, the procedure was successfully completed in 23 cases and failed in 1 case due to puncture needle-related complications. At the last follow-up, 92% (22/24) of patients showed no local symptom progression. VAS score, walking score, and ECOG score improved from 7.2±1.1, 1.4±1.4, 2.5±0.7 before surgery to 2.6±1.9, 2.5±1.4, 2.0±0.8 at 48 hours post-operation. There were no significant differences in scores between 48 hours post-surgery and 48 hours post-radiotherapy.Conclusion:PA can rapidly restore acetabular stability, alleviate pain, and enhance the quality of life for patients. In cases of poor response to PA, radiotherapy may not be effective, but it can effectively control local symptom progression. The combination of these two interventions can yield satisfactory clinical outcomes for patients with acetabular metastases.