Recent Progress in Small Molecular Inhibitors of DNA Gyrase.

BACKGROUND: In the past few decades, with the abuse of antibiotics, bacterial resistance has enhanced constantly. More and more super species of bacteria, which are seriously threatening human health, have been discovered. Developing novel antibacterial agents to overcome the drug-resistance is an urgent duty. We all know that blocking the information-transfer of bacterial DNA and RNA is one of the effective ways to inhibit bacterial growth. Therefore, as the indispensable enzyme for DNA replication and transcription, DNA gyrase is one of the important targets for bacterial inhibitors. Accordingly, many inhibitors of DNA gyrase have also been developed. METHODS: In this review, to highlight the recent progress in DNA gyrase inhibitors, the study in this field over the past three years (2017-2019) was summarized and organized based on their backbones or core moieties. Both of the subunits of DNA gyrase were taken into consideration. RESULTS: These DNA gyrase inhibitors have been classified based on their backbones or core moieties. After the comparison of the divided 14 categories, we could achieve some clues for future modification. In particular, we found that benzodiazepines and naphthalene heterocycles were the most common structures in the drug design. On the other hand, isopropyl and cyclopropyl have also been used in drug design, which provides more inspiration for the investigations. Except for GSK2140944, which has entered the phase III clinical trial stage, other compounds here were not fully promulgated with their optimal pharmacokinetic activity. CONCLUSION: We briefly summed up the current situation and future challenges on this topic. Through the discussion of the design strategies and drug effect, we hope that this review can provide a focused direction for future researches.

Introducing ortho-methoxyl group as a fluorescence-enhancing and bathochromic-shift bi-functional strategy for typical cysteine sensors.

A practical strategy of introducing ortho-methoxyl group was explored to achieve the fluorescence-enhancing and bathochromic-shift bi-functional optimization. It was tested in the Cys sensing ISOPH-X series, thus the successful case, ISOPH-2, was obtained. It realized the optimization in a simple and compatible way. The corresponding strategy was basically established during the confirmation of checkpoints including applicable steadiness (over 24 h), wide pH range (7.0-9.0), rapid response (20 min), good biocompatibility, high sensitivity (LOD = 0.072 nm), high selectivity and biological monitoring of Cys in living cells as well as C. elegans. In this work, the o-methoxyl introduction strategy led to a 15 nm red shift and a near 4-fold fluorescence enhancement. This strategy could be combined with the double bond-introducing approach. Compared with reported strategies, by breaking the dilemma between red shift and strong fluorescent intensity, this strategy might offer beneficial information for exploiting better sensors with more fluorophores and mechanisms for their targets.

Reduced expression of annexin A1 promotes gemcitabine and 5-fluorouracil drug resistance of human pancreatic cancer.

Intrinsic chemoresistance is the main reason for the failure of human pancreatic ductal adenocarcinoma (PDAC) therapy. To identify the candidate protein, we compared the protein expression profiling of PDAC cells and its distinct surviving cells following primary treatment with gemcitabine (GEM) and 5-fluorouracil (5-FU) by two-dimensional electrophoresis combined with liquid chromatography-mass spectrometry or mass spectrometry. A total of 20 differentially expressed proteins were identified, and annexin A1 (ANXA1) was analyzed for further validation. The functional validation showed that the downregulation of ANXA1 contributes to GEM and 5-FU resistance in PDAC cells through protein kinase C/c-Jun N-terminal kinase/P-glycoprotein signaling pathway. Our findings provide a platform for the further elucidation of the underlying mechanisms of PDAC intrinsic chemoresistance and demonstrated that ANXA1 may be a valid marker for anticancer drug development.

An umbelliferone-derivated fluorescent sensor for selective detection of palladium(II) from palladium(0) in living cells.

Palladium (Pd) has drawn worldwide attentions because its connections to industry, chemistry, biological material and public health. Quantitative and selective detection tools for Pd and its ion forms are in urgent necessity. Here an umbelliferone derivative Umb-Pd2 was provided as a small, steady, safe and selective sensor for detecting Pd(II). It indicated advantages including sensitive (LOD 1.1 nM), wide pH tolerance (5.0-10.0), applicable linear range (0-1.8 equivalent) and low toxicity. The most attractive point was its explicit selectivity towards Pd(II) from Pd(0) in both independent and coexistence systems. This distinguishing ability was further utilized in imaging in living cells, raising this work as a rare and important example among all the published papers on palladium sensing. Thus, Umb-Pd2 supplied a potential approach for further improvement and applications in both daily chemistry and public health.

Developing potential Helicobacter pylori urease inhibitors from novel oxoindoline derivatives: Synthesis, biological evaluation and in silico study.

By recruiting the important moiety from Shikonin, a series of novel oxoindoline derivatives S1-S20 have been synthesized for inhibiting H. pylori urease. The most potent compound S18 displayed better activity (IC50 = 0.71 µM; MIC = 0.48 µM) than the positive controls AHA (IC50 = 17.2 µM) and Metronidazole (MIC = 31.3 µM). With low cytotoxicity, it showed considerable potential for further development. Docking simulation revealed the possible binding pattern of this series. 3D QSAR model was built to discuss SAR and give useful hints for future modification.

Investigation of Urination Disorder in Parkinson's disease.

BACKGROUND: Urination disorders are common in Parkinson's disease (PD) and respond poorly to medication. This study aimed to analyze the risk factors for urination disorders in PD. METHODS: Ninety-one patients with PD (aged 34-83 years old) were recruited. Patients were assessed with the Unified PD Rating Scale (UPDRS), Hoehn and Yahr stage, Pittsburgh Sleep Quality Index (PSQI), Hamilton Depression Rating Scale (HAMD), and Hamilton Anxiety Scale (HAMA). Micturition number was recorded, and Type B ultrasound was used to evaluate residual urine. Statistics was performed using binary logistic regression, bivariate correlations, and Chi-square and t-tests. RESULTS: Of 91 patients, urinary dysfunction occurred in 55.0%. Among these, 49.5% suffered with nocturia, 47.3% with pollakiuria. Nocturia number had a positive linear relationship with HAMA score (odds ratio [OR] = 0.340, P = 0.001), HAMD score (OR = 0.323, P = 0.002), duration of L-dopa medication (OR = 0.328, P = 0.001), dose of L-dopa (OR = 0.273, P = 0.009), UPDRS-II (OR = 0.402, P = 0.000), UPDRS-III score (OR = 0.291, P = 0.005), and PSQI score (OR = 0.249, P = 0.017). Micturition number over 24 h was positively associated with HAMA (OR = 0.303, P = 0.004) and UPDRS-II scores (OR = 0.306, P = 0.003). Of patients with residual urine, 79.3% had a volume of residual urine <50 ml. Residual urine was present in 44.4% of the patients with nocturia, 46.5% of the patients with pollakiuria, and 80.0% of the patients with dysuria. More men than women had residual urine (35.2% male vs. 13.3% female; P = 0.002). CONCLUSIONS: Nocturia and pollakiuria were common micturition symptoms in our participants with PD. Nocturia was associated with depression, anxiety, sleep problems, and severity of PD. Pollakiuria was associated with anxiety and severity of PD. Male patients were more prone to residual urine and pollakiuria.

Evaluation of anterior ethmoidal artery by 320-slice CT angiography with comparison to three-dimensional spin digital subtraction angiography: initial experiences.

OBJECTIVE: To explore the usefulness of 320-slice CT angiography (CTA) for evaluating the course of the anterior ethmoidal artery (AEA) and its relationship with adjacent structures by using three-dimensional (3D) spin digital subtraction angiography (DSA) as standard reference. MATERIALS AND METHODS: From December 2008 to December 2010, 32 patients with cerebrovascular disease, who underwent both cranial 3D spin DSA and 320-slice CTA within a 30 day period from each other, were retrospectively reviewed. AEA course in ethmoid was analyzed in DSA and CTA. In addition, adjacent bony landmarks (bony notch in medial orbital wall, anterior ethmoidal canal, and anterior ethmoidal sulcus) were evaluated with CTA using the MPR technique oriented along the axial, coronal and oblique coronal planes in all patients. The dose length product (DLP) for CTA and the dose-area product (DAP) for 3D spin DSA were recorded. Effective dose (ED) was calculated. RESULTS: The entire course of the AEA was seen in all 32 cases (100%) with 3D spine DSA and in 29 of 32 cases (90.1%) with 320-slice CTA, with no significant difference (p = 0.24). In three cases where AEA was not visualized on 320-slice CTA, two were due to the dominant posterior ethmoidal artery, while the remaining case was due to diminutive AEA. On MPR images of 320-slice CT, a bony notch in the orbital medial walls was detected in all cases (100%, 64 of 64); anterior ethmoidal canal was seen in 28 of 64 cases (43.8%), and the anterior ethmoidal sulcus was seen in 63 of 64 cases (98.4%). The mean effective dose in CTA was 0.6 ± 0.25 mSv, which was significantly lower than for 3D spin DSA (1.3 ± 0.01 mSv) (p < 0.001). CONCLUSION: 320-slice CTA has a similar detection rate for AEA to that of 3D spin DSA; however, it is noninvasive, and may be preferentially used for the evaluation of AEA and its adjacent bony variations and pathologic changes in preoperative patients with paranasal sinus diseases.

Comparison of standard- and low-tube voltage 320-detector row volume CT angiography in detection of intracranial aneurysms with digital subtraction angiography as gold standard.

RATIONALE AND OBJECTIVES: The aim of this study was to prospectively assess the effect of low-tube voltage (80 kVp) 320-detector row volume computed tomographic (CT) angiography (L-VCTA) in the detection of intracranial aneurysms, with three-dimensional (3D) spin digital subtraction angiography (DSA) as the gold standard. MATERIALS AND METHODS: Forty-eight patients with clinically suspected subarachnoid hemorrhages were divided into two groups. One group underwent L-VCTA and DSA, while the other group underwent conventional-tube voltage (120 kVp) volume CT angiography (C-VCTA) and DSA. Vascular enhancement, image quality, detection accuracy of aneurysms, and radiation dose were compared between the two groups. RESULTS: For objective image quality, the L-VCTA group had higher mean vessel attenuation, correlated with higher image noise and lower signal-to-noise ratio, than the C-VCTA group. For subjective image quality, there were no significant differences between the two groups regarding scores for arterial enhancement, depiction of small arterial detail, interference of venous structures, and overall image quality scores. The mean effective dose for the L-VCTA group was significantly lower than for the C-VCTA group (0.56 ± 0.25 vs 1.84 ± 0.002 mSv), with a reduction of radiation dose of 69.73%. With 3D DSA as the reference standard, the sensitivity, specificity, and accuracy in the L-VCTA and C-VCTA groups were 94.12%, 100%, 94.4% and 100%, 100%, and 100%, respectively. In both groups, there were significant correlations for maximum aneurysm diameter measurements between volume CT angiography and 3D DSA; no statistical difference in the mean maximum diameter of each aneurysm was measured between volume CT angiography and 3D DSA. CONCLUSIONS: L-VCTA is helpful in detecting intracranial aneurysms, with results similar to those of 3D DSA, but at a lower radiation dose than C-VCTA.

[Value of protein array in the diagnosis of Helicobactor pylori infection in children].

OBJECTIVE: To study the value of multiple Helicobacter pylori (H.pylori) antibody detection by protein array in the diagnosis of H.pylori infection in children. METHODS: Biopsy specimens obtained by gastroscopy from 120 children with digestive system symptoms were detected by rapid urease test (RUT) and modified Giemsa staining. Positivity in both RUT and Giemsa staining was the "gold criterion" of H.pylori infection. Serum samples of these patients were obtained and the antibodies against cytotoxin associated gene A protein (CagA), vacuolating toxin A (VacA), urease, heat shock protein 60 (Hsp60) and RdxA (nitroreductase) were detected by protein array technique. RESULTS: H.pylori infection was identified according to the "gold criterion" in 60 children. Compared with the "gold criterion", the goodness of fit and the coefficient of contingency in the diagnosis of H.pylori infection of the following four groups antibody detection were all statistically significant (P<0.001): anti-Ure antibody alone, anti-Ure antibody combined with anti-CagA antibody, anti-Ure antibody combined with anti-VacA antibody and anti-Ure antibody combined with anti-CagA and anti-VacA antibody. The sensitivity, specificity and accuracy of the detection of anti-Ure antibody combined with anti-CagA antibody for the diagnosis of H.pylori infection were 81.7%, 91.7% and 86.7%, respectively. The antibody detection showed a high positive predictive value (90.7%) and a high negative predictive value (83.3%). CONCLUSIONS: The antibody detection by protein array, especially the detection of anti-Ure antibody combined with anti-CagA antibody, is valuable in the diagnosis of H.pylori infection.

[Efficacy of biofeedback therapy in children with functional constipation].

OBJECTIVE: To evaluate the benefits of biofeedback therapy in children with functional constipation caused by pelvic floor dysfunction (PFD). METHOD: Anorectal manometry (PC Polygraf HR, Medtronic) was performed in children with functional constipation according to the diagnostic criteria of Rome III. Among them, 47 cases with PFD were trained with biofeedback therapy (PC Polygraf HR, Medtronic) according to the degree of cooperation. They received the treatment twice every week, and those in whom the therapy was performed at least three times were enrolled to evaluate the efficacy in this study. RESULT: A total of 27 cases (20 male and 7 female) in whom the procedure was performed at least three times were eligible for inclusion into this study. The mean age of them was (6.7 + or - 2.2) years (range 4 to 12 years), and the mean duration of symptoms was (3.0 + or - 2.3) years with a range of 6 months to 8 years. Among them, 16 cases received the biofeedback training more than 5 times, while 8 cases more than 7 times. The rectal maximum contraction pressure during defecation was increased significantly with the number of biofeedback training before treatment and after 3, 5, and 7 times of treatment was (36.2 + or - 10.4), (45.1 + or - 9.5), (47.6 + or - 9.1), and (47.8 + or - 8.8) mm Hg (1 mm Hg = 0.133 kPa) respectively (P < 0.01). The potential of electromyography of external anal sphincter decreased progressively, but had not reached significance. Follow-up continued for 3 to 12 months by telephone, the clinical response to biofeedback treatment was evaluated as excellent (complete resolution of constipation), good (improvement of constipation), partial or poor (no improvement of constipation). The response was excellent in 13 cases (48.1%), good in 8 (29.6%), partial in 3 (11.1%), and 2 (7.5%) cases had no improvement, and 1 case was lost to follow-up, and the rate of success was 88.9% (24/27). CONCLUSION: Biofeedback therapy is a safe and effective treatment option for functional constipation due to PFD in children probably by increasing the rectal maximum contraction pressure during defecation.

[Role of oxidative stress in the pathogenesis of esophageal mucosal injury in children with reflux esophagitis].

OBJECTIVE: To investigate the role of oxidative stress in the pathogenesis of esophageal mucosa injury in children with reflux esophagitis (RE). METHODS: Esophageal mucosal samples from 36 children with RE (7 months to 16 years of age) were obtained by gastroscopy. The parameters of oxidative stress, including the contents of malondialdehyde (MDA), glutathione (GSH) and nitric oxide (NO) and total superoxide dismutase (T-SOD) activity in the esophageal mucosa as well as the protein content of the esophageal mucosa, were measured. Twenty children (3 to 16 years of age) without esophageal mucosal injury by gastroscopy served as controls. RESULTS: There was no significant difference in the protein content of the esophageal mucosa between the RE and the control groups. The content of MDA in the RE group (15.36+/- 16.67 nmol/mg) was significantly higher than that in the control group (7.51+/- 6.17 nmol/mg) (P<0.01). The activity of T-SOD in the RE group (30.43+/- 35.09 U/mg) was statistically lower than that in the control group (56.34+/- 51.73 U/mg) (P<0.05). No significant differences were observed in GSH and NO contents between the two groups. CONCLUSIONS: The MDA content increases and the SOD content decreases in the esophageal mucosa in children with RE. This suggests that oxidative stress seems to be an important mediator in generation of esophageal mucosal injury.

[Rapid detection of clarithromycin resistant Helicobacter pylori from children's gastric biopsy specimens by polymerase chain reaction-restriction fragment length polymorphism].

OBJECTIVE: Infection with clarithromycin-resistant Helicobacter pylori (Hp) is often predictive of treatment failure. Susceptibility testing for Hp could guide therapy of Hp infections. However, agar dilution approved by the Clinical and Laboratory Standards Institute (CLSI) to test for antimicrobial susceptibility of Hp is time consuming (results are often not available in a week or more). So a more expeditious method is necessary. The purpose of this study was to evaluate polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) test performed directly on gastric biopsy specimen from children to detect 23S rRNA mutations (A2143G and A2144G) indicating clarithromycin resistance. METHODS: All biopsy specimens were derived from patients presenting with upper gastrointestinal symptoms, submitted to endoscopy in the Affiliated Children's Hospital, Zhejiang University School of Medicine from September 2006 to February 2007. No patients had undergone eradication therapy. Thirty-nine samples randomly selected from positive specimens by rapid urease test, were homogenized in 500 microl brucella broth with 30% glycerol. The 200 microl homogenized fluid was used to purify genomic DNA with the kit according to the instructions provided by manufacturer, and the rest was used to isolate Hp strains by culturing. All the Hp isolates were tested for clarithromycin susceptibility with the agar dilution and classified as resistant if the minimum inhibitory concentrations (MIC) exceeded 1 microg/ml. Simultaneously, PCR-RFLP analysis was performed in order to identify 23S rRNA mutations (A2143G and A2144G). Finally, the two methods were compared by statistics. The agar dilution was used as a standard to determine the sensitivity and specificity of the PCR-RFLP assay. RESULTS: Of the 39 samples, agar dilution and PCR-RFLP method respectively detected 13 (33.3%) and 14 (35.9%) clarithromycin-resistant gastric specimens. The sensitivity and specificity of PCR-RFLP for the detection of Hp in biopsy specimens were both 92%. The positive and negative predictive value was 85.7% and 96% respectively. No statistically significant difference was found between the two methods (chi2=0.06, P>0.05). The rate of Hp resistance to clarithromycin significantly increased compared with a previous report from the authors' hospital in 2004 (chi2=6.20, P<0.05). CONCLUSIONS: Rising clarithromycin resistance rates were observed in children who visited the authors' hospital. PCR-RFLP test is reliable and rapid for detection of clarithromycin resistance directly on gastric biopsy specimen from children and may help choose appropriate antibiotic in Hp eradication therapy.

[Percutaneous kyphoplasty with double or single balloon in treatment of osteoporotic vertebral body compressive fracture: a clinical controlled study].

OBJECTIVE: To evaluate the clinical efficacy of percutaneous bipediculary kyphoplasty with double or single balloon I treatment of osteoporosis vertebral compressive fractures. METHODS: Fifty-one patients with painful osteoporotic vertebral compressive fracture involving 69 vertebrae. 19 males and 32 females, aged 72.5 (63 - 85), underwent percutaneous kyphoplasty with double inflatable balloon (29 cases, 38 vertebrae) or single inflatable balloon (22 cases, 31 vertebrae) under X-ray fluoroscopy monitoring. The fractured vertebral bodies were punctuated, balloon was put into the subsided areas and then distended, and bone cement was injected into the cavity. The vertebral height and Cobb angle, preoperative and postoperative, were measured by radiography. Follow-up was conducted for 6 - 12 months. RESULTS: All patients tolerated the procedure well with dramatic pain relief within 96 hours after the procedure. No clinical complication was found. Visual analog scale score was improved from 7.8 preoperatively to 2.6 postoperatively (P < 0.01). Oswestry disability index was decreased from 73% preoperatively to 26% postoperatively. In the double balloon group, the heights loss of the anterior and middle portions of the vertebral body reduced from 14.7 mm and 10.5 mm to 10.1 mm and 5.5 mm respectively (both P < 0.01), and the Cobb angle was corrected from 22.4 degrees to 12.3 degrees (P < 0.01). In the single balloon group, the heights loss of the anterior and middle portions of the vertebral body reduced from 14.7 mm and 10.6 mm to 10.4 mm and 6.5 mm respectively (both P < 0.01), and the. Cobb angle was corrected averagely from 21.2 degrees to 11.6 degrees (P < 0.01). The mean vertebral height restoration rates were 72.8% and 70.1% in the double and single balloon groups respectively. The mean Cobb angle correct degrees were 10.1 degrees and 9.5 degrees in double and single balloon groups respectively. There were no significant differences in the average height restoration rate and Cobb angle correction between the double and single balloon groups (72.8% vs 70.1%, and 10.0 degrees vs 9.5 degrees both P > 0.05). The pain relief and functional recovery were substantial and maintained to the last follow up. CONCLUSION: Percutaneous bipediculary kyphoplasty with double or single balloon for painful osteoporotic vertebral body compressive fractures is effective and safe.

[Clinical manifestations and endoscopic features of abdominal type Henoch purpura in children].

OBJECTIVE: To investigate the clinical manifestations and endoscopic features of abdominal type Henoch purpura in children and improve the diagnostic level. METHODS: Retrospective review was made on the clinical, endoscopic and histopathological features of 57 cases of children with Henoch purpura abdominal type who had been hospitalized from Jan. 2002 to May 2007. Upper gastrointestinal endoscopy was performed in all cases. All the cases had various digestive system symptoms without cutaneous purpura before endoscopy. Mucosal specimens were taken from sinus ventriculi and duodenum for histopathological analysis. Helicobacter pylori (H.pylori) infection was identified by rapid urease test and histology, and diagnosis of H.pylori infection was made when both were positive. RESULTS: The common gastrointestinal symptoms of Henoch purpura were abdominal pain (46 cases), vomiting (32 cases), hematochezia (11 cases), diarrhea (4 cases) and abdominal distention (1 case). Three cases had arthralgia and joint swelling. The main laboratory findings were increased peripheral white blood cells (33 cases, 57.9%), 1/5 of cases had elevated C reactive protein (CRP), low serum albumin and seroperitoneum. Endoscopy demonstrated the damages to the mucosa, which varied from congestion, edema, petechia and ecchymosis (37 cases, 64.9%) to erosive and multiple ulcers (14 cases, 24.6%), granulation of mucosa in descendent duodenum (4 cases, 7.0%), and diffuse hemorrhage of mucosa (2 cases, 3.5%). The upper gastrointestinal endoscopy showed that the commonest and most serious position involved was the descendent duodenum (55 cases, 96.5%), followed by duodenal bulb (32 cases, 56.1%) and stomach (18 cases, 36.1%), esophagus was less involved (1 case, 1.8%). Histological manifestations showed swollen vascular endothelial cells of capillary vessels and small blood vessels, fibrotic necrosis of small vessels and bleeding, diffuse perivascular lymphocytic and neutrophilic infiltration and nuclear debris in mucosa and submucosa. Three cases (5.3%) were found infected with H. pylori. In 49 cases (86.0%) cutaneous purpura appeared 1 - 7 days after endoscopy. Eight cases had no cutaneous purpura until they left hospital. Two cases were lost to follow up and 6 cases (10.5%) remained free from cutaneous purpura were followed up until now (1 - 5 years). CONCLUSION: Descending duodenum is the commonest and most serious position of upper gastrointestinal tract involved in Henoch purpura. Upper gastrointestinal endoscopy with the mucosal biopsy are useful for the early diagnosis of Henoch purpura.

[Prevalence of resistance to metronidazole in Helicobacter pylori from children and induction of resistance in vitro].

OBJECTIVE: To investigate the prevalence of resistance of Helicobacter pylori (H. pylori) to metronidazole (MTZ) and the distribution change of minimal inhibitory concentrations (MICs) in H. pylori from local children, evaluate the applicability of E-test for MIC determination, and display the propensity of acquired-resistance to MTZ after induction of resistance in vitro. METHODS: One group of 44 H. pylori isolates obtained from Oct. 2002 to Nov. 2003 and another 83 H. pylori isolates obtained from Dec. 2004 to Jul. 2005 from the local children who underwent gastroscopy in the Children's Hospital affiliated to Zhejiang University Medical School and were diagnosed as H. pylori-associated gastritis or peptic ulcer were studied. Susceptibility was tested by agar dilution method or E-test method. In 11 randomly selected metronidazole-sensitive isolates (MTZ(S)), resistance was induced in vitro with MTZ. RESULTS: The resistance rate was 31.8% (14/44) in the 44-islates obtained from Oct. 2002 to Nov. 2003 and 51.8% (43/83) (chi(2) = 4.64, P < 0.05) in 83-isolates obtained from Dec. 2004 to Jul. 2005, respectively. The distribution of MICs were < 0.125 - 128 mg/L and 0.25- > 256 mg/L, in which, the MIC(50) was 0.5 mg/L and 16 mg/L, the MIC(90) was 128 mg/L, respectively. Comparing to agar dilution method which is recommended by National Committee for Clinical Laboratory Standards (NCCLS) for MIC, E-test was significantly associated with agar dilution method (chi(2) = 32.38, P < 0.001). The sensitivity, specificity, agreement rate of E-test were 73.08%, 100%, 87.27%, respectively, while there were factors of 2(2) to 2(6) difference in MICs between the results obtained by E-test and agar dilution. For all the 11 MTZ(S) isolates inducted resistance in vitro with MTZ, 16 MICs were achieved through 7 - 9 (7.2 +/- 0.6) passages of induction in vitro, and 100% acquired-resistance to MTZ through 8 - 10 generations; as a result, 10 of 11 MTZ(S) isolates achieved stable high-level resistance (256 mg/L for 2 and > 256 mg/L for 8) and 1 stable 64 mg/L resistance to MTZ. CONCLUSIONS: The prevalence of resistance to MTZ seems to be increasing in H. pylori from local children. To avoid missed diagnosis of H. pylori resistant to MTZ (MTZ(R)), agar dilution method was needed when detecting susceptibility of H. pylori to MTZ. Resistance to MTZ of H. pylori from children is readily induced in vitro.

[Evaluation of clinical value of anorectal manometry for diagnosis of Hirschsprung's disease in neonate].

OBJECTIVE: Hirschsprung's disease (HD), one of the most common causes resulting in lower intestinal obstruction in children, is prone to be misdiagnosed or to be missed from diagnosis because of its atypical clinical symptoms and inconspicuous morphological findings by barium enema X-ray. Recently, this situation has been largely ameliorated by increased comprehension of anorectal kinetics and improvement of instrument for measurement of anorectal pressure. By now, anorectal manometry (ARMM) has been regarded as a routine means for functional assessment and diagnosis for anorectal disease. Nevertheless, the accuracy rate of diagnosis of HD in neonate by ARMM remains to be elucidated. In this study the clinical evaluation of anorectal manometry as an early diagnostic method for neonates with Hirschsprung's disease was appraised. METHODS: Forty-two HD patients defined by pathological study of rectal tissue obtained via rectal mucous membrane biopsy or operation were recruited in this study. ARMM was performed in liquid transmission using PC polygraph high rate gastrointestinal dynamical detection system (PC Polygraf HR, CTD-synectics, Sweden), with 4-lumen catheter with which a small 5-cm-long balloon was connected at the terminus. All children were positioned on their left side or back during the procedure and the pressure transducers were placed in the mid-axillary line level. The results of ARMM performed before operation or biopsy were compared with the results of barium enema X-ray testing. The decrease of internal anal sphincter pressure as rectoanal inhibitory reflex (RAIR) was measured based on the fluctuation curve of pressure detected. HD was defined when no decrease of anal catheter pressure was detected after insufflation (RAIR positive), and suspected HD state was assessed with the presentation of incomplete relaxation or positive/negative results coexisted (RAIR abnormal) in canal. RESULTS: Thirty patients (71.43%) were diagnosed as HD by ARMM including 18 patients who showed negative response to RAIR and 12 patients whose response was abnormal. While barium enema examinations were carried out in all the 45 patients, the results showed 5 HD patients and 14 suspected HD patients, giving an overall diagnostic accuracy of 45.24%. There were also 16 patients with positive ARMM response and negative barium enema findings together, and 5 patients with negative ARMM results and positive barium enema findings at the same time. There was a significant difference between the two diagnostic methods (chi(m)(2) = 4.76, P < 0.05). CONCLUSION: Anorectal manometry seems to be a more reliable method for diagnosis of Hirschsprung's disease in neonate than barium enema X-ray. Because ARMM is a simple, safe and non-invasive method, it can be used as a screening test of choice in neonates with clinically suspected HD. But for final diagnosis, it is reasonable to combine ARMM with other diagnostic methods in HD patients.

[Prevalence of Helicobacter pylori resistant to clarithromycin, amoxicillin and metronidazole in children].

OBJECTIVE: To investigate the prevalence of Helicobacter pylori (Hp) resistant to clarithromycin, amoxicillin and metronidazole in children. METHODS: A total of 44 Hp clinical isolates were cultured from children patients with Hp gastritis and peptic ulcer who underwent gastroscopy during the period from October 2002 to November 2003 in Children's Hospital affiliated to Medical College of Zhejiang University. Nineteen of the patients were girls and 25 boys. The mean age was 8.66 +/- 2.66 years (3 approximately 14 years). The susceptibilities of the 44 Hp strains to clarithromycin, amoxicillin and metronidazole were tested by agar dilution test to determine minimum inhibitory concentration (MIC)(50), MIC(90) and resistance rates were also calculated. RESULTS: Among the 44 Hp strains isolated from children, 8, 4, and 14 strains were resistant to clarithromycin, amoxicillin and metronidazole and the resistant rate was 18.2% (8/44), 9.1% (4/44) and 31.8% (14/44), with MICs for all strains ranging from < 0.125 microg/ml to 64 microg/ml, < 0.125 microg/ml to 64 microg/ml and < 0.125 microg/ml to 128 microg/ml, respectively. MIC(50) and MIC(90) of clarithromycin, amoxicillin and metronidazole were < 0.125 microg/ml, 8 microg/ml; < 0.125 microg/ml, 0.5 microg/ml; 0.5 microg/ml, 128 microg/ml, respectively. The prevalence of Hp resistant to clarithromycin was higher in children than that in adults, while to metronidazole was lower than that in adults with high MICs (32 - 128 microg/ml) and MIC(90) (128 microg/ml) of metronidazole for resistant isolates. Three multidrug resistant strains (6.8%) were simultaneously resistant to these three antibiotics. CONCLUSIONS: The prevalence of Helicobacter pylori resistant to metronidazole was high in children, and the rate of resistance to clarithromycin was higher than that isolated from adults. Amoxicillin-resistant strains were now found, and multidrug resistant strains which resistant to the three antibiotics also emerged.

[Prevalence and mechanism of Helicobacter pylori resistance to clarithromycin in children].

OBJECTIVE: To investigate the prevalence of Helicobacter pylori (H. pylori) resistance to clarithromycin (CLM) in children and to demonstrate the correlation of 23S rRNA gene mutation to clarithromycin resistance of Helicobacter pylori isolates. METHODS: Totally 108 clinical strains of H. pylori were isolated from gastric biopsy specimens obtained from children who underwent endoscopy during the period from October 2002 to January 2004 in Children's Hospital Affiliated to Medical College of Zhejiang University. H. pylori was identified by morphology and biochemical tests after culture. Clarithromycin susceptibility of H. pylori isolates was determined by both E-test and two-fold agar dilution method. A strain was considered resistant when the MIC was defined as >or= 1 microg/ml. Genome DNAs of the 108 isolates were extracted and prepared for PCR to detect the corresponding gene in the V domain of the 23S rRNA. The amplified fragments were recognized and analyzed by restriction fragment length polymorphism (RFLP) when an additional restriction site is created by the mutation. The PCR products of all sensitive and resistant strains were digested with restriction enzyme BbsI and BsaI and were analyzed on a 1.5% agarose gel to discriminate different kinds of mutant genotype. RESULTS: Sixteen of 108 isolates of H. pylori were resistant to clarithromycin by the agar dilution method and E-test method in clinical isolates from children, and the CLM resistance rate was 14.8% (16/108) with MICs ranging from 1 microg/ml to 128 microg/ml. Comparison of results of the two methods showed that these two methods were quite consistent in determination of susceptibility and resistance. The target fragment 425 bp in length containing 23S rRNA corresponding gene was successfully amplified. An A2144G mutation digested with BsaI was detected in 13 resistant isolates, but an A2143G mutation digested with BbsI in only 3 among all 16 clarithromycin resistant strains. None of the sensitive isolates was cleaved by either BsaI or BbsI enzyme, indicating that there was no mutation on them. It was also found that all the fragments from the resistant strains were not completely digested, and 425 bp uncut fragments were also visible and showed three bands indicating that they were heterozygotic strains with a mixture of wild-types and A-->G genotypes. In addition, in this study, no statistically significant difference between mutations at positions 2143 and 2144 with respect to the MIC was observed (r = 0.035, P > 0.05). CONCLUSION: A high prevalence of clarithromycin-resistant H. pylori strains were detected among strains isolated from Chinese children studied. The 23S rRNA gene mutation at positions A2143G and A2144G plays an important role in clarithromycin resistance of H. pylori and A2144G mutation is the predominant finding among the resistant strains.