Lin-Li Chang

Characterisation of drug resistance gene cassettes associated with class 1 integrons in clinical isolates of Escherichia coli from Taiwan, ROC

The presence of class 1 integrons in clinical isolates of Escherichia coli was detected by PCR. Of 104 E. coli isolates from Kaohsiung, 54 (52%) carried class 1 integrons, with inserted DNA regions of 1-3 kb. These integrons were located on plasmids, as demonstrated by Southern hybridisation. DNA sequencing was used to identify the genetic content of the integron-variable regions. Different class 1 integrons contained various numbers, kinds and combinations of gene cassettes within their variable regions. These gene cassettes included those encoding resistance to trimethoprim (dfrIa, dfrV, dfr12 and dfr17), aminoglycosides (aadA1a, aadA2, aadA4 and aadB), chloramphenicol (cmlA), erythromycin (ereA2) and beta-lactams (blaP1). An integron carrying three inserted cassettes - dfr12-orJF-aadA2 - was present in 33 (61%) of the 54 isolates with class 1 integrons. Gene cassettes encoding resistance were expressed phenotypically. The results indicate that class 1 integrons are widespread in clinical E. coli isolates in Taiwan. The types, combinations and frequency of the gene cassettes in integrons may reflect the specific selective pressures to which the isolates were exposed and could provide useful surveillance data for relation to antibiotic usage information.

Increase sensitivity in detecting superficial, low grade bladder cancer by combination analysis of hypermethylation of E-cadherin, p16, p14, RASSF1A genes in urine.

OBJECTIVES To identify a better set of DNA methylation markers to detect superficial, low grade cancer cell in urine sediment for improving cancer treatment, morbidity, and mortality. MATERIALS AND METHODS Methylation-specific PCR (MSP) assay was used to detect promoter hypermethylation in 4 genes (E-cadherin, p16, p14, and RASSF1A) to identify reliable biomarkers for bladder cancer diagnosis in primary tumor DNA and urine sediment DNA from 57 bladder cancer patients. Urine DNA was compared with 20 healthy controls. RESULTS Fifty-one (90%) tumor DNA and 47 urine DNA (83%) samples from bladder cancer patients revealed hypermethylation in at least 1 of the 4 analyzed genes, whereas all urine samples from normal controls were negative. The sensitivity of MSP assay for detecting E-cadherin, p16, p14 and RASSF1A in tumor cells in voided urine was 35%, 35%, 33%, and 65%, respectively. Diagnostic sensitivity was 75% for combining RASSF1A and p14, and 83% for RASSF1A, p14 and E-cadherin. Urine cytology, however, detect only 13 (28%) cases of cancer or suspicious cancer. For detecting superficial and invasive bladder tumor, urine cytology revealed a sensitivity of 23% (6/26) and 35% (7/20), respectively. In contrast, MSP detected hypermethylation in the urine of 80% (37/46) bladder cancer patients. Moreover, hypermethylation analysis of E-cadherin, p14 or RASSF1A genes in urine sediment DNA detected in 85% (22/26) of superficial, 85% (11/13) of low grade, 75% (15/20) of invasive and 79% (26/33) of high grade bladder cancers. Importantly, hypermethylation was detected in the urine DNA of 90% (18/20) superficial tumors with negative or atypia cytology. CONCLUSIONS Hypermethylation of E-cadherin, p14 or RASSF1A in urine sediment DNA is a potential biomarker for detecting superficial, low grade cancer. Besides, hypermethylation of these 3 genes is a valuable adjunct diagnostic marker to urine cytology, which can enhance the diagnostic accuracy and follow-up treatment of bladder cancer patients.

Variable Gene Cassette Patterns of Class 1 Integron-Associated Drug-Resistant Escherichia Coli in Taiwan

This study characterized class 1 integrons in Escherichia coli in Taiwan. The stability and changes in gene cassettes inserted into integrons were also evaluated. The study included 436 clinical strains of E. coli isolated in 2002. Class 1 integrons were characterized by polymerase chain reaction and direct sequencing. Genetic localization of class 1 integrons was determined by conjugal transfer and Southern hybridization. The results indicated that 64% of E. coli isolates carried class 1 integrons. Molecular analysis revealed that the class 1 integrons harbored 13 different antimicrobial resistance gene cassettes and two unknown gene cassettes; the predominant cassettes were aadA and dfrA. Novel gene cassettes first recovered from E. coli were aacA4 and linF. Cassette arrays orfD‐aacA4‐catB8 and aadA1‐linF were also observed. Gene cassette dfrA12‐orfF‐aadA2 was stable. The class 1 integron and dfrA17‐aadA5 gene cassette were located on the same transferable plasmids and were capable of transmission. Therefore, the increased drug resistance of clinical isolates may be explained by antibiotic selective pressure and widespread presence of integrons. Under antibiotic selective pressure, gene cassette‐mediated resistance may not be easily lost. The potential role of integrons in the uptake and dissemination of resistance genes by plasmid between species of bacteria may decrease the therapeutic effectiveness of antibiotics.

Integron types, gene cassettes, antimicrobial resistance genes and plasmids of Shigella sonnei isolates from outbreaks and sporadic cases in Taiwan

This study analysed the presence, location and transferability of integrons and antibiotic resistance genes in 103 Shigella sonnei outbreak isolates and in 32 sporadic isolates from Taiwan. Multiple antimicrobial resistance was common in both outbreak (95 %) and sporadic (97 %) isolates. Class 1 integrons were present in 34 outbreak isolates (33 %) and in six sporadic isolates (19 %). This study is the first, to our knowledge, to identify an atypical sul3-associated class 1 integron carrying the estX-psp-aadA2-cmlA-aadA1-qacH cassette array in Shigella. Class 2 integrons carrying the dfr1-sat2-aadA1 cassette array were predominant in outbreak isolates (90 %) but were not present in sporadic isolates. Other antimicrobial resistance genes not associated with integrons were found to encode resistance to ampicillin (bla(TEM)), chloramphenicol (cat1), sulfonamide (sul2) and tetracycline (tetA and tetB). The most common plasmid size was 130 kb (observed in 43 and 97 % of 1998 outbreak and sporadic isolates, respectively). In conclusion, the plasmid location of resistance genes and horizontal plasmid transfer promote the spread of multiple resistance genes in outbreak and sporadic isolates of S. sonnei.

The associations among eNOS G894T gene polymorphism, erectile dysfunction and related risk factors

To investigate the possible correlations among eNOS G894T polymorphism, erectile dysfunction (ED) and related risk factors in a Taiwanese population.

Amoxicillin resistance with β‐lactamase production in Helicobacter pylori

Background  Amoxicillin‐resistant Helicobacter pylori with minimal inhibitory concentration (MIC) ≥ 256 mg L−1 was isolated from a gastritis patient. The aims were to investigate the mechanism of high‐level amoxicillin resistance in H. pylori.

Persistent mechanical allodynia positively correlates with an increase in activated microglia and increased P‐p38 mitogen‐activated protein kinase activation in streptozotocin‐induced diabetic rats

In experimental early painful diabetic neuropathy, persistent hyperglycaemia induces dys‐regulated sodium channel (Navs) expression in the dorsal root ganglion (DRG) and activates microglia in the spinal dorsal horn (SDH). However, information on diabetes‐induced chronic neuropathic pain is limited. Therefore, we investigated abnormal Navs in the DRG and activated glial cells in the SDH of diabetic rats with chronic neuropathic pain.

Impaired dendritic cell maturation and IL-10 production following H. pylori stimulation in gastric cancer patients

The current study was to investigate the interaction between Helicobacter pylori and human dendritic cells (DCs). Whether impaired DC function can influence the outcome of H. pylori infections. Human monocyte-derived DCs (MDDCs) from five gastric cancer patients and nine healthy controls were stimulated with H. pylori. Maturation markers of MDDC were examined by flow cytometry. IL-10 and TNF-α released by MDDCs and IL-17 produced by T cells were measured by ELISA. Regulatory signaling pathways of IL-10 were examined by ELISA, western blotting, and chromatin immunoprecipitation assay. The results showed that as compared with healthy individuals, the maturation marker CD40 in MDDCs, IL-17A expression from T cells, and IL-10 expression from MDDCs were significantly lower in gastric cancer patients. Blocking DC-SIGN, TLR2, and TLR4 could reverse H. pylori-associated IL-10 production. Activation of the p38 MAPK and NF-kB signaling pathways concomitant with decreased tri-methylated H3K9 and increased acetylated H3 accounted for the effect of H. pylori on IL-10 expression. Furthermore, upregulated IL-10 expression was significantly suppressed in H. pylori-pulsed MDDCs by histone acetyltransferase and methyltransferase inhibitors. Taken together, impaired DC function contributes to the less effective innate and adaptive immune responses against H. pylori seen in gastric cancer patients. H. pylori can regulate IL-10 production through Toll-like and DC-SIGN receptors, activates p-p38 MAPK signaling and the transcription factors NF-kB, and modulates histone modification.

Clinical and Molecular Characteristics of Invasive and Noninvasive Skin and Soft-Tissue Infections Caused by Group A Streptococcus

ABSTRACT The severity of skin and soft tissue infections caused by group A Streptococcus (GAS) is variable, and there are only a limited number of studies evaluating the characteristics of these infections in the literature. From May 2005 to November 2007, 73 patients with skin and soft tissue infections caused by group A Streptococcus were included in this study. Among these patients, 34 (46.6%) had invasive diseases. Diabetes mellitus, alcoholism, and hypertension were the most common underlying disorders. The overall mortality rate was 6.8%, and the elderly were predisposed to invasive infections (P < 0.001). Neutrophil percentages of ≥80, serum creatinine levels of ≥2 mg/dl, and high serum C-reactive protein levels were noted more frequently in patients with invasive infections than in patients with noninvasive infections, as were bacteremia and a high mortality rate. Of the 73 isolates, 93.2%, 97.3%, and 37% exhibited susceptibility to erythromycin, clindamycin, and tetracycline, respectively. The five most prevalent emm types were emm106 (24.7%), emm11 (12.3%), emm102 (9.6%), emm4 (8.2%), and emm12 (8.2%). Compared to other types, the emm106 type was significantly more likely to be associated with invasive diseases (P = 0.012). Dendrogram analysis showed a unique SmaI-digested pulsed-field gel electrophoresis pattern of the emm106 type that was particularly prone to cause invasive skin and soft tissue infections (P < 0.001). The results of this study suggest that isolates with the emm106 gene may be an emerging group A Streptococcus strain that causes invasive skin and soft tissue infections. Further surveillance study to understand the significance of this invasive strain is critical.

Suppressor of cytokine signaling 1 gene expression and polymorphisms in systemic lupus erythematosus

With the aim of investigating the role of suppressor of cytokine signaling 1 (SOCS1) in the pathogenesis of systemic lupus erythematosus, 107 patients with systemic lupus erythematosus, 101 healthy controls, and 151 patients with ankylosing spondylitis were enrolled in this study. SOCS1 mRNA level was measured by the method of quantitative real-time polymerase chain reaction. SOCS1 polymorphisms were detected by the polymerase chain reaction/restriction fragment length polymorphisms method. Systemic lupus erythematosus disease activity was evaluated with the SLEDAI. This study showed that the SOCS1 mRNA expression was significantly higher in the patients with systemic lupus erythematosus than in the healthy controls (p = 0.0014). Patients with active systemic lupus erythematosus had a higher expression of SOCS1 mRNA than the patients with inactive systemic lupus erythematosus (p = 0.035). There was no significant difference in the frequencies of the SOCS1-1478CA/del polymorphisms among the patients with systemic lupus erythematosus, healthy controls, and patients with ankylosing spondylitis. The genotype frequency of the SOCS1-1478 polymorphisms in the dominant model (CA/del+del/del versus CA/CA) was significantly decreased in the patients with thrombocytopenia compared with those without thrombocytopenia (pc = 0.035). Moreover, the allele frequency of SOCS1-1478del was also significantly lower in the patients with thrombocytopenia than in those without thrombocytopenia (p c = 0.02). In conclusion, this study demonstrated that the expression of SOCS1 mRNA was significantly increased in patients with systemic lupus erythematosus. Moreover, SOCS1 mRNA levels in patients with active systemic lupus erythematosus were significantly higher than those in the inactive patients. We also found that the systemic lupus erythematosus patients with thrombocytopenia have a lower frequency of SOCS1-1478del compared with patients without thrombocytopenia. Lupus (2010) 19, 696—702.