Objective To validate the performance of a PMA-based assay for detection of INH-resistant mutations in MTB and investigate the mutation characteristic of INH-resistance.Methods The MTB standard strain H37Rv was from National Tuberculosis Reference Laboratory,1 wild-type strain and 1 katG S315T ACC mutant strain were from Xiamen CDC,7 MTB INH-resistant strains with known INH resistance mutations were from Shenzhen Center for Chronic Disease Control,Henan CDC,No.309 Hospital of PIA and Xiamen CDC.707 MTB clinical isolates were from Xiamen CDC,Xiamen No.1 Hospital and Zhangzhou CDC,126 sputum samples were from Xiamen Tongan CDC.The genomic DNA of the MTB standard strain H37Rv,7 MTB INH-resistant strains and 833 clinical samples,were extracted with Xiamen Zeesan Mycobacterium tuberculosis Isoniazid-resistance Mutation Test Kit using the thermal lysis method.The genomic DNA of 1 wild-type strain and 1 katG S315T ACC mutant strain were extracted with AxyPrep Bacterial Genomic DNA Miniprep Kit.The mutations were discriminated by the △Tm between the samples and wild type control in katG315 codon,inhA promoter -17 to -8 region,ahpC promoter region -44 to -30 and - 15 to 3,and inhA94 codon.A 10-fold dilution series of MTB DNA from 3 × 105 to 300 copies/ reaction obtained from a wild-type strain and a katG S315T ACC mutant strain,respectively,were prepared to determine the analytical sensitivity.Seven MTB INH-resistant strains with 9 predetermined mutations were used for the analytical specificity assay,and 5 mutants of which were used for the repeatability assay.The clinical detection performance of PMA assay were confirmed by the sequencing method in 833 samples.Results Results could be obtained within 3 hours from DNA extraction to PMA assay,including 46 samples in a standard 96-well real-time PCR instrument simultaneously.The analytical sensitivities of PMA were 300 copies/reaction for both the wild-type strain and katG S315T ACC mutant strain.Nine INH-resistant point mutations could be discriminated and 5 of which had standard deviations of melting temperature less than 0.5 ℃.Fully concordant results of mutant locus between PMA assay and sequencing were obtained in all 162 mutant samples.INH-resistant mutations in the four loci were found in 19.4％ (162/833) samples by PMA assay in Xiamen and Zhangzhou.Among the 14 lNH-resistant mutant types detected,katG S315T ( AGC→ACC),inhA promoter - 15C→T and katG S315N (AGC→AAC) accounted for 83.3％ (135/162) of the overall mutations.

Objective To explore a practical way for traceability and comparability of the value obtained from clinical biochemical examination in regional medical organizations by transferring the assigned value in fresh serum to daily serum calibrator.Methods Six local comprehensive hospitals with Grade III were chosen to analyze the effects of calibration by traceability and the comparability of examined results.The definite values of standard panel in matching analytical system were transferred to fresh sera for value assignment.Verified biochemical analyzers were used to examine the values of alaninetransaminase(ALT),total cholesterol(TC),urea and total bilirubin(TBil) in mixed fresh sera and intercomparison with target system was performed to analyze the effect of traceable calibration and the comparability among various analytical systems.Results Although all the applied instruments in the present study were in good conditions with high precision,the comparability of obtained results of ALT,TC,Urea and TBil between tested system and target system was low and the differences were significant(P0.05).Conclusion Traceable precision transfer by using fresh serum is a simple,practical,feasible way to realize the traceability and comparability of regional clinical biochemical examination.

Objective To inquire into the clinical acceptability of self-developed biochemical test system by comparing the results and bias estimation with the traceable assay system.Methods According to the requests of National Committee for Clinical Laboratory Standards (NCCLS) EP-9A, comparison test and bias estimation for self-developed test system and traceable Roche reference system (HITACHI 7180) were performed by using patient's sera. The self-developed system was set up as axle Y, the traceable Roche assay system as axle X and 1/2 CLIA (%) was used as the acceptable judge standard.Results The relationship of LDL-C between self-developed system and comparison system was inferior, so the results of LDL-C as well as the low values of HDL-C were of low acceptability. The comparabilities of results of ALT, urea, creatinine, TC, ApoA1 and ApoB were acceptable.Conclusion The self-developed test system must compare with traceable assay system, and estimate its clinical acceptability. The regression equation Y=bX+a can be used to correct the bias of test result.

Objective To investigate the strategy of diagnosis and treatment for non-functional parathyroid cysts.Methods The clinical data of non-functional parathyroid cysts patients undergoing resection between Jan.1991 and Jan.2013 in Peking Union Medical College Hospital were retrospectively analyzed.Results Non-functional parathyroid cysts were confirmed by intra-operation exploration and post-operation pathology in 22 patients.Ultrasound scan revealed cervical cystic lesions in all patients before operations,and concomitant lesions in thyroid were found in 10 patients.The serum calcium and phosphate level were in normal range in all patients.Local excisions were performed in 11 cases for isolated cyst.Subtotal thyroidectomy were performed in 10 cases for concomitant nodular goiter or close anatomy association cyst and thyroid.Radical resection of thyroid and cervical lymph node dissection were performed in one patient for concomitant thyroid carcinoma.No operation related complications was recorded in these series.No relapse presented in these patients after more than 7 months of follow-up.Conclusion Non-functional parathyroid cysts should be considered in the differential diagnosis of cervical mass even though it rarely occurs.Surgical resection was one promising treatment for non-functional parathyroid cysts.

Objective To investigate the clinical characters of pancreatitis in patients with primary hyperparathyroidism (PHPT).Methods The clinical data of patients with PHPT undergoing parathyroidectomy from Jan.2009 to May.2014 in Peking Union Medical College Hospital were retrospectively analyzed for coexistent pancreatitis.Results 571 patients received parathyroidectomy due to PHPT during this period.Thirteen patients (2.3％)with PHPT were confirmed with coexistent pancreatitis by clinical manifestation and abdominal imaging evaluation,including acute pancreatitis in 9 patients and chronic pancreatitis in 4 patients.PHPT with pancreatitis was associated with preoperative serum intact parathyroid hormone(iPTH)higher than 500 pg/ml (P=0.025)and occurrence of hypercalcemic crisis (P=0.013).The age and sex of patients,preoperative serum calcium,gallstones,urinary calculi and bone fracture were not related with pancreatitis in this group.The average follow-up period for these patients was 34(ranging from 13 to 68)months.All patients were free from hyperparathyroidism recurrence.Only one young lady with chronic pancreatitis suffered from repeated pancreatitis after surgery.Conclusions Serum iPTH higher than 500 pg/ml and occurrence of hypercalcemic crisis were possible risk factor of pancreatitis in patients with PHPT.Parathyroidectomy can help to prevent the recurrence of pancreatitis in these patients.

Objective To investigate the clinical and pathological features of small cell carcinoma of the urinary bladder. Methods The pathological and clinical data of 9 cases of small cell carcinoma were analyzed retrospectively. There were 6 males and 3 females, ages 45 to 79 years (mean age, 62 years). Clinical manifestations of 7 cases included gross hematuria and dysuria, the other 2 cases experienced lower abdominal pain. The mean tumor size was 2.0 cm (ranged, 0.5 to 7.0 cm). Two cases had multiple tumors and 5 cases had single tumors. The growth pattern in 2 cases was diffuse growth in the whole bladder. In 4 cases tumor cells were found in urine cytology. All 9 patients underwent surgical treatment, including TURBt. Four patients were diagnosed as superficial tumors before operation. All the patients underwent regular theprubicine irrigation in the bladder. One case underwent additional intravenous chemotherapy for 3 cycles. Partial cystectomy was performed in 2 cases, with regular theprubicine irrigation in bladder and 1 case underwent intravenous chemotherapy for 2 cycles. Radical cystectomy was performed in 3 cases, with 2 cases undergoing intravenous chemotherapy after operation. Results Pathological findings showed that tumor cells were small and round in shape. These hyperchromatic nuclei showed limited cytoplasm with lack of nesting characters. CgA and NSE were positive in immunohistochemistry. The final diagnosis was small cell carcinoma, with 1 case accompanied with transitional cell carcinoma and 1 case accompanied with prostate cancer. One case showed high preoperative serum calcium (3.15 mmol/L) and low serum phosphate (0.61 mmol/L), which returned to normal 1 month after operation. Four cases who′s bladder was preserved were followed up, 3 cases were alive for 4, 9 and 25 months after operation. The 1 case who underwent intravenous chemotherapy was followed up for 24 months and there was no sign of relapse or metastasis. In all the 3 cases with radical cystectomy, 2 cases died 2 and 28 months postoperativly. Another case with adjuvant chemotherapy was followed up for 24 months without recurrence or metastasis. Conclusions Small cell carcinoma of the urinary bladder is highly malignant with poor prognosis. Radical cystectomy in combination with systemic chemotherapy has better efficacy. Retained bladder surgery with systemic chemotherapy is an alternative choice. The most important factors which influence the prognosis of the tumor are clinical stage and therapeutic methods.

Objective To explore the clinical and histopathological features of metanephric adenoma (MA). MethodsClinical and pathological data of 10 cases of MA were analyzed retrospectively.There were 4 males and 6 females,aged from 33 to 65 years,with an average of 45 years.2 patients had flank pain,4 patients had gross hematuria,and 4 patients were found by physical examination.The average diameter of tumor was 4.5 cm (2.5 - 8.0 cm).All patients were diagnosed as renal tumor by CT scan.9 patients underwent radical nephrectomy and 1 patient underwent partial nephrectomy. Results Pathological examination found that the tumors are composed of densely packed small uniform cells with regular nuclei that formed a tubular or adenoid pattern.Mitotic figures were absent or rare.4 patients were diagnosed as MA,2 cases were diagnosed as low-grade malignant MA,and 4 cases were diagnosed as MA with malignant component (2 cases of adenocarcinoma,1 case of chromophobe cell carcinoma,and 1 case of well differentiated papillary adenocarcinoma),7 cases were followed up for 22 months ( 10 to 34 months) without recurrence or metastasis. Conclusions MA is very rare benign renal tumor originating from epithelium,and a few are malignant,and some may contain malignant ingredients.Nephron-sparing surgery and radical nephrectomy are eligible for the treatment of MA.Considering the uncertainty of the biological behavior and cellular origin of MA,a long-term follow-up is necessary.

Objective To assess the feasibility and clinical efficiency of laparoscopic Madigan enucleation of the prostate for the treatment of benign prostatic hyperplasia (BPH) with large prostate.Methods From Nov.2008 to Jan.2010,24 eases of BPH patients with large prostate ( ＞90 g) were treated in our institute by laparoscopic Madigan enucleation.The average patient's age was 65.2 yrs,the average prostate weight were 132.3 ± 21.9 g,preoperative residual urine was 143.2 ± 23.2 ml,average IPSS was 28.6 ± 3.8,average QOL was 5.5 ± 2.4 and average MFR was 5.4 ± 2.3 ml/s.All patients accepted the laparoscopic Madigan enucleation of the prostate.The pre-surgery and three months post-operative IPSS,QOL,and MFR were compared and analyzed.Results All 24 patients successfully completed the surgery.The resected prostate tissue weight was 104.7 ± 23.3 g,blood loss was 112.5 ± 47.8 ml,postoperative hospital stay was 3.5 ± 0.8 d,bladder irrigation time was 1.3 ± 0.9 d,drainage time was 2.3 ± 0.5 d and catheterization time was 3.1 ±0.4 d.Three months after surgery,patient's IPSS was 4.5 ± 1.8,QOL was 19.9 ±3.1,and MFR was 18.5 ±2.9 ml/s.All the parameters significantly improved compared with the pre-surgery data ( P ＜ 0.05 ).Conclusions Laparoscopic Madigan enucleation of the prostate is a safe and effective method in the treatment of BPH with large prostate volume.

Objective To investigate the clinical features,diagnosis and treatment of prostate sarcoma. Methods19 cases of prostate sarcoma had been treated in our hospital from Oct 1995 to Sep 2010,aged from 16 to 48 years (mean,36 years).They were hospitalized due to different degrees of difficulty in urination,6 cases with bladder irritation,3 cases with gross hematuria,3 cases with blood clots in urination,3 cases with difficult defecation,2 cases with urinary tract infection,and 1 case with low back pain.Digital rectal examination showed prostate volume increased significantly in 16 cases,of which 6 cases prominent into bladder,and 3 cases touched hard nodules and tenderness.B ultrasound examination showed prostate neoplasms with internal echo uneven.Pelvic MRI in 12 cases indicated irregular soft tissue density in prostate with unclear edge,a high-low mixed signal inside,and mass protruding outward,among which 7 cases with significantly enlarged prostates and unclear bladder boundaries,6 cases with unclear boundaries with rectum,and 1 case with solid and cystic change. Pelvic CT in 10 cases showed irregular prostate tumors,among which 5 cases with tumor violation of bilateral seminal vesicle,2 cases with tumor invasion of unilateral seminal vesicle,and 3 cases with lymph node enlargement.Chest X-ray in 2 cases showed widespread metastatic lung. Results Different treatments were used according to different conditions:6 cases with full cystoprostatectomy,8 cases with radical prostatectomy,1 case with skin routine ureter fistulization and rectal fistulization,1 case with bladder fistulization,1 case with chemotherapy after prostate biopsy,and 2 cases with prostate biopsy alone. Pathological diagnosis showed 6 cases of leiomyosarcoma,6 cases of rhabdomyosarcoma,2 cases of malignant mesenchymal tumor,1 case of phyllodes sarcoma,1 case of granulocytic sarcoma,1 case of phyllodes sarcoma,1 case of malignant fibrous tissue sarcoma,and 1 case of poorly differentiated muscle induced sarcoma. Immunohistochemical examination in 19 cases of paraffin specimens stained with SP showed vimentin was positive,smooth muscle actin and myoglobin in leiomyosarcoma and rhabdomyosarcoma were positive respectively,and desmin in two kinds of afore-mentioned sarcoma expressed in different level.CD117 and S-100 were weakly positive in 1 patient with undifferentiated component and 1 patient with liposarcoma of the malignant mesenchymoma.Peroxidase was positive in granulocytic sarcoma,p63 was positive in lobulated sarcoma,and CD34 was positive in malignant fibrous histiocytoma.14 patients were given chemotherapy,among which 6 cases were combined with radiotherapy.19 cases were followed up for 4 -30 months (mean 13 months).4 patients survived for 24 -30 months without recurrence,while 15 cases died within 18 months after diagnosis. Conclusions The prostate sarcoma is a clinically rare malignant tumor with high degree of malignancy and unspecific clinical manifestations.The diagnosis relies on biopsy and immunohistochemistry,and early diagnosis and radical surgical excision are helpful for increasing survival rate.