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Background Degenerative lumbar spondylolisthesis (DLS) is a major cause of spinal canal stenosis and is often related to lower back pain. IL-20 is emerging as a potent angiogenic, chemotactic, and proinflammatory cytokine related to several chronic inflammatory bone disorders likes intervertebral disc herniation, rheumatoid arthritis (RA), osteoporosis, and bone fracture. IL-19 also acts as a proinflammatory cytokine in RA. The aim of the present study was to investigate whether IL-19 and IL-20 are involved in DLS and compare three different tissues including disc, facet joint, and ligamentum flavum of patients with DLS to verify which tissue is affected more by inflammation. Methods Disc, facet joint and ligamentum flavum from 13 patients with DLS was retrieved, and the expression pattern of IL-19, IL-20, IL-20R1, IL-20R2, TNF-α, IL-1β, and MCP-1 was evaluated using immunohistochemical staining with specific antibodies. The disc cells were isolated and incubated with IL-19 and IL-20 under CoCl2-mimicked hypoxic conditions to analyze the proinflammatory cytokine expression pattern using real-time quantitative PCR with specific primers. Results IL-19 and IL-20 were positively stained and accompanied by abundant expression of TNF-α, IL-1β, and MCP-1 in facet joints of DLS patients. IL-19 and IL-20’s receptors (IL-20R1 and IL-20R2) were expressed on chondrocytes and fibrocytes/fibroblasts in facet joint and ligamentum flavum tissues from patients with DLS. There was a significant correlation between the expression of IL-20 and IL-1β in facet joint. In vitro assay, IL-19 and IL-20 upregulated the expression of IL-1β, IL-6, TNF-α, IL-8, VEGF, and MCP-1 in primary cultured DLS disc cells under CoCl2-mimicked hypoxic conditions. Conclusions IL-19, IL-20, and their receptors as well as proinflammatory cytokines (TNF-α, IL-1β, and MCP-1) were expressed more in facet joints than the other tissues in patients with DLS; therefore, the etiology of inflammation might be more facet-centric. IL-19 and IL-20 induced proinflammatory cytokine expression in disc cells and might play a role in the pathogenesis of DLS.

Background/Purpose The aim of this study is to characterize antibiotic-nonsusceptible Salmonella isolates in Taiwan. Methods A total of 76 Salmonella isolates showing lower susceptibility to cephalosporins or quinolones were identified from 1416 clinical isolates from 1999 to 2008. Minimal inhibitory concentrations for selected antimicrobial agents were tested by the agar dilution method. Antibiotic resistance-related genes were determined by polymerase chain reaction (PCR) combined with sequencing. Southern blotting, conjugation tests, and transformation tests were used to characterize plasmid-mediated quinolone resistance (PMQR) determinants. Results The observed nonsusceptible phenotypes of 76 isolates were against cefoxitin (57.9%), cefotaxime (43.4%), ceftazidime (40.8%), ceftriaxone (42.1%), cefepime (5.3%), ciprofloxacin (80.3%), and levofloxacin (81.6%). Among 44 cephalosporin-resistant isolates, TEM-1, CMY-2, CMY-14, CTX-M-3-like and CTX-M-15-like determinants were present in 31 (70.5%), 32 (72.7%), 1 (2.3%), 1 (2.3%), and 1 (2.3%) of isolates, respectively. PCR screening for PMQR genes of 62 quinolone-nonsusceptible isolates revealed the presence of qnrS , qnrD , aac(6 ′ )-Ib-cr , and oqxAB in 3 (4.8%), 2 (3.2%), 1 (1.6%), and 10 (16.1%) isolates, respectively. Among 36 isolates showing high resistance to quinolones, S83F/D87N and S83F/D87G amino acid substitutions of GyrA were found in 29 (80.6%) and 6 (16.7%) isolates, respectively. Moreover, among quinolone highly resistant isolates, eight (22.2%) of isolates showed over-expression of the PAβN-sensitive efflux pump. Transformants and transconjugants harboring qnrD - or oqxAB -plasmids showed decreased susceptibility to quinolones. Conclusion GyrA mutations are the major mechanisms associated with quinolone-resistant Salmonella isolates in Taiwan. Overproduction of efflux pump genes and the presence of qnr and oqxAB play additional roles in reduced susceptibility to quinolones.

This study investigates the evolution of an Escherichia coli sequence type 219 clone in a patient with recurrent urinary tract infection, comparing isolate EC974 obtained prior to antibiotic treatment and isolate EC1515 recovered after exposure to several β-lactam antibiotics (ceftriaxone, cefixime, and imipenem). EC974 had a smooth colony morphology, while EC1515 had a rough colony morphology on sheep blood agar. RAPD-PCR analysis suggested that both isolates belonged to the same clone. Antimicrobial susceptibility tests showed that EC1515 was more resistant to piperacillin/tazobactam, cefepime, cefpirome, and ertapenem than EC974. Comparative genomic analysis was used to investigate the genetic changes of EC974 and EC1515 within the host, and showed three plasmids with replicons IncI1, P0111, and IncFII in both isolates. P0111-type plasmids pEC974-2 and pEC1515-2, contained the antibiotic resistance genes aadA2, tetA, and drfA12. IncFII-type plasmids pEC974-3 and pEC1515-3 contained the antibiotic resistance genes blaTEM−1, aadA1, aadA22, sul3, and inuF. Interestingly, blaCMY−111 and blaCMY−4 were found in very similar IncI1 plasmids that also contained aadA22 and aac(3)-IId, from isolates EC974 (pEC974-1) and EC1515 (pEC1515-1), respectively. The results showed in vivo amino acid substitutions converting blaCMY−111 to blaCMY−4 (R221W and A238V substitutions). Conjugation experiments showed a high frequency of IncI1 and IncFII plasmid co-transference. Transconjugants and DH5α cells harboring blaCMY-4 or blaCMY-111 showed higher levels of resistance to ampicillin, amoxicillin, cefazolin, cefuroxime, cefotaxime, cefixime, and ceftazidime, but not piperacillin/tazobactam, cefpime, or ertapenem. All known genes (outer membrane proteins and extended-spectrum AmpC β-lactamases) involved in ETP resistance in E. coli were identical between EC974 and EC1515. This is the first study to identify the evolution of an IncI1 plasmid within the host, and to characterize blaCMY−111 in E. coli.

Prokaryotic CRISPR-Cas systems limit the acquisition of genetic elements and provide immunity against invasive bacteriophage. The characteristics of CRISPR-Cas systems in clinical Klebsiella pneumoniae isolates are still unknown. Here, 97 K. pneumoniae genomes retrieved from the Integrated Microbial Genomes & Microbiomes genome database and 176 clinical isolates obtained from patients with bloodstream (BSI, n = 87) or urinary tract infections (UTI, n = 89) in Taiwan, were used for analysis. Forty out of ninety-seven genomes (41.2%) had CRISPR-Cas systems identified by the combination of CRISPRFinder and cas1 gene sequence alignment. The phylogenetic trees revealed that CRISPR-Cas systems in K. pneumoniae were divided into two types (type I-E, 23; subtype I-E∗, 17) based on the sequences of Cas1 and Cas3 proteins and their location in the chromosome. The distribution of type I-E and I-E∗ CRISPR-Cas systems was associated with the multilocus sequence typing and the pulsed-field gel electrophoresis results. Importantly, no CRISPR-Cas system was identified in published genomes of clonal complex 258 isolates (ST11 and ST258), which comprise the largest multi-drug resistant K. pneumoniae clonal group worldwide. PCR with cas-specific primers showed that 30.7% (54/176) of the clinical isolates had a CRISPR-Cas system. Among clinical isolates, more type I-E CRISPR-Cas systems were found in UTI isolates (BSI, 5.7%; UTI, 11.2%), and subtype I-E∗ CRISPR-Cas systems were dominant in BSI isolates (BSI, 28.7%; UTI, 15.7%) (p = 0.042). Isolates which had subtype I-E∗ CRISPR-Cas system were more susceptible to ampicillin-sulbactam (p = 0.009), cefazolin (p = 0.016), cefuroxime (p = 0.039), and gentamicin (p = 0.012), compared to the CRISPR-negative isolates. The strains containing subtype I-E∗ CRISPR-Cas systems had decreased numbers of plasmids, prophage regions, and acquired antibiotic resistance genes in their published genomes. Here, we first revealed subtype I-E∗ CRISPR-Cas system in K. pneumoniae potentially interfering with the acquisition of phages and plasmids harboring antibiotic resistance determinants, and thus maintained these isolates susceptible to antibiotics.

This study investigates the evolution of an Escherichia coli sequence type 219 clone in a patient with recurrent urinary tract infection, comparing isolate EC974 obtained prior to antibiotic treatment and isolate EC1515 recovered after exposure to several β-lactam antibiotics (ceftriaxone, cefixime, and imipenem). EC974 had a smooth colony morphology, while EC1515 had a rough colony morphology on sheep blood agar. RAPD-PCR analysis suggested that both isolates belonged to the same clone. Antimicrobial susceptibility tests showed that EC1515 was more resistant to piperacillin/tazobactam, cefepime, cefpirome, and ertapenem than EC974. Comparative genomic analysis was used to investigate the genetic changes of EC974 and EC1515 within the host, and showed three plasmids with replicons IncI1, P0111, and IncFII in both isolates. P0111-type plasmids pEC974-2 and pEC1515-2, contained the antibiotic resistance genes aadA2, tetA, and drfA12. IncFII-type plasmids pEC974-3 and pEC1515-3 contained the antibiotic resistance genes blaTEM−1, aadA1, aadA22, sul3, and inuF. Interestingly, blaCMY−111 and blaCMY−4 were found in very similar IncI1 plasmids that also contained aadA22 and aac(3)-IId, from isolates EC974 (pEC974-1) and EC1515 (pEC1515-1), respectively. The results showed in vivo amino acid substitutions converting blaCMY−111 to blaCMY−4 (R221W and A238V substitutions). Conjugation experiments showed a high frequency of IncI1 and IncFII plasmid co-transference. Transconjugants and DH5α cells harboring blaCMY-4 or blaCMY-111 showed higher levels of resistance to ampicillin, amoxicillin, cefazolin, cefuroxime, cefotaxime, cefixime, and ceftazidime, but not piperacillin/tazobactam, cefpime, or ertapenem. All known genes (outer membrane proteins and extended-spectrum AmpC β-lactamases) involved in ETP resistance in E. coli were identical between EC974 and EC1515. This is the first study to identify the evolution of an IncI1 plasmid within the host, and to characterize blaCMY−111 in E. coli.

The OmpK36 porin plays a role in carbapenem resistance and may contribute to bacterial virulence inKlebsiella pneumoniae. This study aimed to investigate the characteristics of different groups ofK. pneumoniaeseparated byompK36typing. Among 226 nonduplicateK. pneumoniaebloodstream isolates collected at a Taiwanese hospital in 2011, fourompK36types, designated types A, B, C, and D, were identified by PCR in 61, 28, 100, and 36 isolates, respectively; 1 isolate was untypeable. Statistical analysis showed significantly higher rates of antimicrobial resistance (all tested antibiotics except meropenem), extended-spectrum β-lactamases or DHA-1 (47.5% together), Qnr-type quinolone resistance determinants (50.8%), and IncFIIA-type plasmids (49.2%) in group A than in others. Seventeen isolates were identified as belonging to 3 international high-risk clones (4 sequence type 11 [ST11], 10 ST15, and 3 ST147 isolates); all isolates but 1 ST15 isolate were classified in group A. The significant characteristics of group C were hypermucoviscosity (62.0%) and a higher virulence gene content. This group included all serotype K1 (n= 30), K2 (n= 25), and K5 (n= 3) isolates, 6 of 7 K57 isolates, all isolates of major clones associated with pyogenic liver abscesses (29 ST23, 11 ST65, 5 ST86, 7 ST373, and 1 ST375 isolates), and 16 (94.1%) of 17 isolates causing bacteremic liver abscesses. Twelve (42.9%) of the group B isolates were responsible for bacteremic biliary tract infections. Group D was predominant (83.3%) among 12 K20 isolates. This study suggests that most clinicalK. pneumoniaeisolates can be allocated into four groups with distinct characteristics based onompK36types.

Background Matrix-assisted laser desorption/ionization–time of flight mass spectrometry (MALDI-TOF MS) is a valuable method for rapid identification of blood stream infection (BSI) pathogens. Integration of MALDI-TOF MS and blood culture system can speed the identification of causative BSI microorganisms. Materials and methods We investigated the minimal microorganism concentrations of common BSI pathogens required for positive blood culture using BACTEC FX and for positive identification using MALDI-TOF MS. The time to detection with positive BACTEC FX and minimal incubation time with positive MALDI-TOF MS identification were determined for earlier identification of common BSI pathogens. Results The minimal microorganism concentrations required for positive blood culture using BACTEC FX were >10 7 –10 8 colony forming units/mL for most of the BSI pathogens. The minimal microorganism concentrations required for identification using MALDI-TOF MS were > 10 7 colony forming units/mL. Using simulated BSI models, one can obtain enough bacterial concentration from blood culture bottles for successful identification of five common Gram-positive and Gram-negative bacteria using MALDI-TOF MS 1.7–2.3 hours earlier than the usual time to detection in blood culture systems. Conclusion This study provides an approach to earlier identification of BSI pathogens prior to the detection of a positive signal in the blood culture system using MALDI-TOF MS, compared to current methods. It can speed the time for identification of BSI pathogens and may have benefits of earlier therapy choice and on patient outcome.

Objectives: Neurogenic pulmonary edema (NPE) is a fatal complication in children with enterovirus 71 (EV71) encephalitis. Endothelin 1 (ET-1), a potent vasoconstrictor, can induce pulmonary edema in rats via intrathecal injections. Thus, it was hypothesized that ET-1 in the central nervous system may correlate with NPE in children with EV71 encephalitis. Methods: Clinical data and ET-1 in the cerebrospinal fluid (CSF) were compared between three groups: (1) EV71 encephalitis with NPE; (2) EV71 encephalitis without NPE; and (3) non-EV71 aseptic meningitis. ET-1 immunostaining was performed on the brainstem of autopsy patients. Results: The EV71 with NPE group showed significantly increased CSF levels of ET-1 compared to the EV71 without NPE and the non-EV71 aseptic meningitis groups (both p

Carbapenem-resistant Klebsiella pneumoniae strain 1756 was isolated from a pus specimen from a Taiwanese patient. Here, the complete genome sequence of strain 1756 is presented.

The transcriptional network of the SRY (sex determining region Y)-box 17 (SOX17) and the prognostic impact of SOX17 protein expression in human cancers remain largely unclear. In this study, we evaluated the prognostic effect of low SOX17 protein expression and its dysregulation of transcriptional network in esophageal squamous cell carcinoma (ESCC). Low SOX17 protein expression was found in 47.4% (73 of 154) of ESCC patients with predicted poor prognosis. Re-expression of SOX17 in ESCC cells caused reduced foci formation, cell motility, decreased ESCC xenograft growth and metastasis in animals. Knockdown of SOX17 increased foci formation in ESCC and normal esophageal cells. Notably, 489 significantly differential genes involved in cell growth and motility controls were identified by expression array upon SOX17 overexpression and 47 genes contained putative SRY element in their promoters. Using quantitative chromatin immunoprecipitation-PCR and promoter activity assays, we confirmed that MACC1, MALAT1, NBN, NFAT5, CSNK1A1, FN1 and SERBP1 genes were suppressed by SOX17 via the SRY binding-mediated transcriptional regulation. Overexpression of FN1 and MACC1 abolished SOX17-mediated migration and invasion suppression. The inverse correlation between SOX17 and FN1 protein expression in ESCC clinical samples further strengthened our conclusion that FN1 is a transcriptional repression target gene of SOX17. This study provides compelling clinical evidence that low SOX17 protein expression is a prognostic biomarker and novel cell and animal data of SOX17-mediated suppression of ESCC metastasis. We establish the first transcriptional network and identify new suppressive downstream genes of SOX17 which can be potential therapeutic targets for ESCC.

Background: Since intrinsic PD-1 receptor functions promote tumor growth was reported, we will investigate the role of PD-L1 in lung adenocarcinoma cells, and their impact on clinical outcome. Materials and methods: Lung adenocarcinoma CL1-5 cells, derived from CL1-0 cells have more invasive ability. We prepared PDL1-overexpression human lung adenocarcinoma cell line, derived from CL1-0 cells (CL1-0-PD1) and from PC14PE6 cells (PC14PE6-PD-L1). We observed the cellular morphology, and invasiveness; epithelial-mesenchymal transition (EMT) markers and regulators were also evaluated. To explore interaction between PD-1 and PD-L1, we treated the CL1-0, CL1-5, CL1-0-PDL1 cells, PC14PE6 cells, and PC14PE6-PD-L1 cells with anti-PD-1 antibody, and then evaluated cellular invasion. We also suppressed PD-1 to test whether PD-1/PDL-1 interaction contributed to the EMT change. Further, we evaluated cellular proliferation and chemosensitivity by MTT assay and colony formation assay. Finally, we correlated PD-L1 expression with clinical outcome in patients’ tumor specimen. Results: CL1-5 cells possessed higher PD-L1 expression than parental CL1-0 cells. CL1-0 cells with PD-L1 overexpression had more expression of EMT regulators and mesenchymal markers. Overexpression of PD-L1 in another lung adenocarcinoma PC14PE cells (PC14PE6-PD-L1 cells) become more aggressive. On the contrary, down-regulation of PD-L1 in CL1-5 cells and PC14-PE6-PD-L1 cells became indolent. Besides through the observation from confocal microscopy, PD-L1 expression was more co-localized with vimentin and slug, individually in CL1-5, CL1-0-PD-L1 and PC14PE6-PD-L1 cells. Therefore, PD-L1 promoted EMT in human lung adenocarcinoma cells. After adding anti-PD-1 antibody in CL1-5, CL1-0, and CL1-0-PDL1 cells, migration and invasion ability were decreased. Similar phenomenon was found when PD-1 antibody treatment in PC14PE6 cells, and PC14PE6-PD-L1 cells. Silencing PD-1 by siRNA also disrupt the cellular aggressivenss in CL1-0-PD-L1 and PC14PE6-PD-L1 cells. These results indicated PD-1/PD-L1 axis regulated cancer cells migration and invasiveness. PD-L1 expression also decreased cellular proliferation and had little influence on chemsensitivity. Finally, we found that higher PD-L1 expression was correlated with lymph node metastasis in clinical specimen. Conclusion: Lung adenocarcinoma cells with higher PD-L1 expression promote EMT via PD-1 receptor. PD-L1 expression lowers proliferation rate, but has little impact on chemosensitivity. Lung cancer patients with high PD-L1 expression in tumor have higher lymph node metastasis. Citation Format: Wen-Pin Su, Li-Chan Chang, Jing-Jou Yan, Wu-Chou Su. PD-1/PD-L1 axis in cancer cells contributes to cellular EMT in lung adenocarcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 4599.

The aim of this study was to characterize fluoroquinolone (FQ)-resistant Escherichia coli isolates from bacteremia in Taiwan in 2001–2015. During the study period, 248 (21.2%) of 1171 isolates were identified as levofloxacin-resistant. The results of phylogenetic group analysis showed that 38.7% of the FQ-resistant isolates belonged to phylogenetic group B2, 23.4% to group B1, 22.6% to groupA, 14.9% to group D and 0.4% belonged to group F. FQ-resistant isolates were highly susceptible to cefepime (91.5%), imipenem (96.0%), meropenem (98.8%), amikacin (98.0%) and fosfomycin (99.6%), as determined by the agar dilution method. β-lactamases, including blaTEM (66.1%), blaCMY-2 (16.5%), blaCTX-M (5.2%), blaDHA-1 (1.6%) and blaSHV-12 (1.6%), were found in FQ-resistant isolates. The results of PCR and direct sequencing showed that 37 isolates (14.9%) harbored plasmid-mediated quinolone resistance (PMQR) genes. qnrB2, qnrB4, qnrS1, coexistence of qnrB4 and qnrS1, oqxAB and aac(6′)-Ib-cr were found in 1, 4, 4, 1, 15 and 14 isolates, respectively. PMQR genes were successfully transfered for 11 (29.7%) of the 37 PMQR-harboring isolates by conjugation to E. coli C600. These findings indicate that qnr genes remained rare in E. coli but demonstrate the potential spread of oqxAB and aac(6′)-Ib-c in Taiwan.

The Abbott RealTime MTB assay, launched in June 2014, has been shown to have a competitive performance in the detection of the Mycobacterium tuberculosis (MTB) complex in respiratory specimens. The present study was conducted to investigate the usefulness of the Abbott MTB Realtime assay in the detection of MTB in formalin-fixed paraffin-embedded (FFPE) tissues.A total of 96 FFPE specimens obtained from microbiologically proven MTB cases (N=60) and nontuberculous Mycobacterium cases (N=36) were analyzed. The performance of the Abbott MTB Realtime assay was compared with that of the Roche Cobas TaqMan MTB assay.The overall sensitivity and specificity of the Abbott assay were 63.3% and 97.2%, respectively, compared with 11.7% and 100% for the Cobas assay. The detection rate of the Abbott assay was much higher among 37 acid-fast-positive specimens than among 23 acid-fast-negative specimens (89.3% versus 21.7%, respectively). The detection rate of the assay was higher among 29 resection specimens than among 31 small biopsy specimens (86.2% versus 41.9%, respectively).Our results suggest that the Abbott RealTime MTB assay can be used to differentiate MTB from nontuberculous mycobacterial infections in acid-fast-positive FFPE tissues.

Background The emergence of imipenem-resistant Pseudomonas aeruginosa (IRPA) has become a great concern worldwide. The aim of this study was to investigate resistance mechanisms associated with bloodstream isolated IRPA strains in Taiwan. Results A total of 78 non-duplicated IRPA isolates were isolated from patients with bloodstream infection. The average prevalence of imipenem-resistance in those isolates was 5.9 % during a 10-year longitudinal surveillance in Taiwan. PFGE results showed high clonal diversity among the 78 isolates. VIM-2, VIM-3, OXA-10, and OXA-17 β-lactamases were identified in 2 (2.6 %), 3 (3.8 %), 2 (2.6 %), and 1 (1.3 %) isolates, respectively. Active efflux pumps, AmpC β-lactamase overproduction, and extended-spectrum AmpC cephalosporinases (ESACs) were found in 58 (74.4 %), 25 (32.1 %) and 15 (19.2 %) of IRPA isolates, respectively. oprD mutations with amino acid substitution, shortened putative loop L7, premature stop codon caused by point mutation, frameshift by nucleotide insertion or deletion, and interruption by insertion sequence were found in 19 (24.4 %), 18 (23.1 %), 15 (19.2 %), 14 (17.9 %), and 10 (12.8 %) of isolates, respectively. Conclusions This study suggests that alterations in the OprD protein and having an active efflux pump are the main mechanisms associated with bloodstream isolated IRPA. Overproduction of AmpC, ESACs, and the presence of VIM- and OXA-type β-lactamases play additional roles in reduced susceptibility to imipenem in P. aeruginosa isolates in Taiwan. Electronic supplementary material The online version of this article (doi:10.1186/s12866-016-0719-2) contains supplementary material, which is available to authorized users.

Oligonucleotide primers used in this study. (DOC 106 kb)

The objectives of the study were to investigate the distribution of cphA-related genes (cphA) encoding a CphA metallo-β-lactamase (MBL) among 51 consecutive Aeromonas blood isolates and to compare different phenotypic methods for detecting CphA. The presence of cphA was detected by PCR. Four phenotypic methods, the imipenem-EDTA combined disc test, imipenem-EDTA MBL Etest, agar dilution test and modified Hodge test (MHT), were used to detect imipenem susceptibility and MBL production. The results showed that 35 (69%) blood isolates had cphA. All (100%) of 16 Aeromonas aquariorum isolates and 12 Aeromonas veronii isolates, and 4 (80%) of 5 Aeromonas hydrophila isolates, carried cphA, but none of 15 Aeromonas caviae isolates did. With the standard inocula, irrespective of the presence or absence of cphA, all but one (50, 98%) isolates were susceptible to imipenem tested by disc diffusion, Etest and agar dilution (10(4) c.f.u. spot inocula), and did not exhibit MBL production by the imipenem-EDTA combined disc test and MBL Etest. By the agar dilution test using large inocula (10(7) c.f.u.), 34 (97%) of 35 cphA(+) isolates had imipenem MICs of ≥16 µg ml(-1), higher than the susceptible breakpoint (4 µg ml(-1)), and demonstrated positive results for the MHT, while one cphA(+) and all 17 cphA(-) isolates had imipenem MICs of ≤4 µg ml(-1). In conclusion, the distribution of cphA among aeromonads is species-specific, found in A. aquariorum, A. veronii and A. hydrophila, and the MHT may be a phenotypic screening test for CphA production.

IL-20 is a proinflammatory cytokine involved in rheumatoid arthritis, atherosclerosis, and stroke. However, little is known about its role in breast cancer. We explored the function of IL-20 in tumor growth and metastasis, as well as in clinical outcome. Tumor expression of IL-20 was assessed by immunohistochemical staining among 198 patients with invasive ductal carcinoma of the breast, using available clinical and survival data. IL-20 expression was associated with advanced tumor stage, greater tumor metastasis, and worse survival. Reverse transcription quantitative polymerase chain reaction showed that clinical breast tumor tissue expressed higher levels of IL-20 and its receptors than did nontumorous breast tissue. IL-20 was also highly expressed in breast cancer bone-metastasis tissue. In vitro, IL-20 upregulated matrix metalloproteinase-9, matrix metalloproteinase-12, cathepsin K, and cathepsin G, and enhanced proliferation and migration of breast cancer cells, which were inhibited by anti–IL-20 mAb 7E. In vivo, we generated murine models to evaluate the therapeutic potential of 7E, using luminescence intensity, radiological scans, and micro-computed tomography. 7E reduced tumor growth, suppressed bone colonization, diminished tumor-mediated osteolysis, and lessened bone density decrement in mice injected with breast cancer cells. In conclusion, our results suggest that IL-20 plays pivotal roles in the tumor progression of breast cancer. IL-20 expression in breast cancer tissue is associated with a poor clinical outcome. Anti–IL-20 mAb 7E suppressed bone colonization and decreased osteolytic bone lesions. Therefore, IL-20 may be a novel target in treating breast tumor-induced osteolysis.

Matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS) and conventional standard methods were compared for time to pathogen identification and impact on clinical outcomes in peritoneal dialysis-related peritonitis patients. The MALDI-TOF MS method identified the causative microorganisms earlier (average time saved, 64 h for all pathogens), and patients had a shorter hospital stay (mean ± standard deviation, 5.2 ± 4.8 days versus 8.2 ± 4.5 days, P = 0.001).

Although extended-spectrum-β-lactamase (ESBL)-producing aeromonads have been increasingly reported in recent years, most of them were isolates from case reports or environmental isolates. To investigate the prevalence of ESBL producers among Aeromonas blood isolates and the genes encoding ESBLs, consecutive nonduplicate Aeromonas blood isolates collected at a medical center in southern Taiwan from March 2004 to December 2008 were studied. The ESBL phenotypes were examined by clavulanate combination disk test and the cefepime-clavulanate ESBL Etest. The presence of ESBL-encoding genes, including bla TEM , bla PER , bla CTX-M , and bla SHV genes, was evaluated by PCR and sequence analysis. The results showed that 4 (2.6%) of 156 Aeromonas blood isolates, 1 Aeromonas hydrophila isolate and 3 Aeromonas caviae isolates, expressed an ESBL-producing phenotype. The ESBL gene in two A. caviae isolates was bla PER-3 , which was located in both chromosomes and plasmids, as demonstrated by Southern hybridization. Of four patients with ESBL-producing Aeromonas bacteremia, two presented with catheter-related phlebitis and the other two with primary bacteremia. Three patients had been treated with initial noncarbapenem β-lactams for 5 to 10 days, and all survived. In conclusion, ESBL producers exist among Aeromonas blood isolates, and clinical suspicion of ESBL production should be raised in treating infections due to cefotaxime-resistant Aeromonas isolates.

Liao I-C, Chen H-M, Wu J-J, Tsai P-F, Wang L-R, Yan J-J. Metallo-β-lactamase-producing Enterobacteriaceae isolates at a Taiwanese hospital: lack of distinctive phenotypes for screening. APMIS 2011; 119: 543–50. We investigated the prevalence of metallo-β-lactamases (MBLs) among 1,827 Enterobacteriaceae isolates collected in 2006 and evaluated the VITEK 2 microbiology system, modified Hodge test, and 2 combined disk tests as the screening tools for MBLs by using these isolates and 77 previously characterized IMP-8 producers. The IMP-8 MBL was identified in 18 isolates of 2006, and the IMP-8-positive isolates represented 0.2%, 1.1%, and 5.0% of all Escherichia coli, Klebsiella pneumoniae, and Enterobacter cloacae isolates, respectively. Only one-third of all MBL producers could be recognized by either VITEK 2 or the Hodge test. MBL production could be identified in 38 (40%) of the 95 IMP-8-producing isolates by the combined disk test using meropenem disks supplemented by phenylboronic acid and EDTA, and only 2 (2.1%) isolates gave positive results in the combined disk test using meropenem disks supplemented with dipicolinic acid. Of all IMP-8 producers, 37.9%, 50.5%, and 32.6% were nonsusceptible to tigecycline, fluoroquinolones, and both, respectively. In conclusion, this study demonstrated the lack of distinct phenotypes that could be easily identified among the IMP-8-producing Enterobacteriaceae isolates at a Taiwanese hospital. Continuous surveillance and monitoring are needed because the widespread of tigecycline- and fluoroquinolone-coresistant MBL producers may become a serious therapeutic problem.

Many human lung cancer cell lines express apolipoprotein E (ApoE), especially cells derived from malignant pleural effusions (MPE) in patients with lung adenocarcinoma. This study aimed to investigate the influence of ApoE expression on lung cancer. In lung cancer tissues, ApoE expression was more frequently found in malignant pleural effusions (MPE)-associated lung adenocarcinoma than in lung adenocarcinoma or squamous cell carcinoma without MPE (P

Background/PurposeIdentifying the pathogens present in blood stream infections is crucial to initiate appropriate antimicrobial therapy and avoid morbidity and mortality. The aim of this study was to evaluate the effect of overnight storage of aerobic and anaerobic BACTEC 9240 blood culture bottles on the detection time for common pathogens.MethodsFrom November 2007 to July 2008, a total of 2,105 isolates were positively detected using the BACTEC 9240 system. The time to positive detection (TTD) was calculated by subtracting the time of receipt in the laboratory from the time required to detect a positive culture. The mean TTD values were calculated using the TTD value of the first positive culture bottle only. Overnight delay at the National Cheng Kung University Hospital, Taiwan was 15 hours (from 5 pm to 8 am).ResultsOf the 2,105 total isolates, 972 (46.1%) were Gram-positive bacteria, 1,024 (48.6%) were Gram-negative bacteria and 109 (5.1%) were fungi. Among the top 10 pathogens, 24.7% grew only in the aerobic bottle and 15.1% in the anaerobic bottle, including Staphylococcus spp., Enterococcus faecium, Enterobacteriaceae, and Gram-positive bacilli. Due to the overnight delay in loading a blood culture bottle into the instrument, for most of the pathogens (including Staphylococcus spp. and Enterobacteriaceae), a decrease in TTD by £ 4.4 hours was observed. An increase in TTD by 20.8 hours was observed for Gram-positive bacilli. We also found that the difference between TTD in aerobic versus anaerobic bottles during the day was higher in coagulase-negative staphylococcus (12 hours) and lower in Escherichia coli and Staphylococcus aureus (< 2 hours). TTD was longer than 72 hours in 20.5% of Gram-positive bacilli and 7.3% of Candida albicans.ConclusionNo difference in the TTD of major pathogens was observed in bottles processed during the day and after overnight delay, suggesting that the delayed entry of the blood culture bottle into the instrument may affect the detection time. Since high numbers of facultative anaerobes were detected in anaerobic bottles only, use of a single aerobic bottle might have a detrimental effect on the clinical therapy outcome.

Background/Purpose An increase in incidence of Clostridium difficile infection (CDI) among Western countries has been noted in recent years. Epidemiological data of CDI are scarce in Taiwan. This study is intended to depict the clinical features of CDI at a medical center in Southern Taiwan. Methods From January 1, 2007 to March 31, 2008, hospitalized patients with CDI (defined as the presence of gastrointestinal symptoms and fecal C. difficile toxin) were identified. Their medical records were reviewed for further evaluation. Results A total of 86 cases of CDI were identified in the study period. The incidence was 42.6 cases per 100,000 patient-days, or 3.4 cases per 1,000 discharges, and was highest in intensive care units (110.6 cases per 100,000 patient-days). Variable incidence rates were noted in different wards, and prevalence was higher in the infectious ward. Diarrhea, fever, and abdominal distension were common in 82 (95.3%), 47 (54.7%), and 29 (33.7%) patients, respectively. Metronidazole was the initial therapeutic regimen for 83 (96.5%) patients. Prolonged diarrhea was noted in 31 (36.4%) patients, especially in those on hemodialysis therapy. Recurrence was noted in 7 (8.1%) patients. Fecal carriage of vancomycin-resistant Enterococcus colonization was found in three patients after therapy for CDI. All-cause mortality rate of patients with CDI at 30 days was 23.3%. Conclusion CDI is increasingly being recognized within the medical departments, and should be considered in hospitalized adults with diarrhea, fever, or abdominal distension alone, or in combination.

Background/Purpose: The emergence of multidrug-resistant Neisseria gonorrhoeae is a great challenge in controlling gonorrhea. This study was conducted to survey the prevalence of molecular mechanisms of antimicrobial resistance among 45 clinical isolates of N. gonorrhoeae collected at a university hospital in Southern Taiwan during 1999-2004. Methods: Mutations in mtrR loci and quinolone-resistance-determining regions (QRDRs) were examined by gene sequencing. Polymerase chain reactions with specific primers were performed to detect ermA, ermB, ermC, and ermF. Serogroups and serovars were determined by commercial kits. Results: The percentage of multidrug resistance, that is, resistance to penicillin, tetracycline, erythromycin, and ciprofloxacin, among the 45 isolates was 40%. Ceftriaxone and spectinomycin were active against all isolates in vitro. The frequency of mutations in the QRDR and mtrR promoter was 82.2% and 93.3%, respectively. Eighty-two percent of the isolates carried mutations both in the QRDR and mtrR loci. Of nine mutation profiles with QRDR mutations (n=37), gyrA-Ser91Phe/gyrA-Asp95Gly/parC-Ser87Arg was the most common type (56.8%). Acquired genes for rRNA methylase were detected in 11 isolates (10 ermB and 1 ermA). Twenty-seven serovars were identified and all belonged to serogroup B, which suggested that multiple clones of N. gonorrhoeae were circulating in the community in the Tainan area. Conclusion: The high prevalence of multidrug resistance caused by varied resistance mechanisms in N. gonorrhoeae limits the drug choice. Ongoing surveillance of antimicrobial resistance and discovery of new effective antibiotic therapy are warranted in endemic areas.

A total of 317 Klebsiella pneumoniae and 291 Escherichia coli nonduplicate isolates were tested by the VITEK 2 system to evaluate its capability to detect extended-spectrum beta-lactamases (ESBLs) among putative ESBL-producing isolates, in particular those with the coproduction of AmpC enzymes. beta-lactamases produced by the test isolates had been characterised. The sensitivity and specificity for ESBLs were 98.9% and 98.5%, respectively. Ninety of the isolates were AmpC (CMY-2, CMY-8 or DHA-1) and ESBL (SHV and/or CTX-M) coproducers, and 74 isolates (82.2%) of them were flagged as ESBL producers. Our study indicates that the VITEK 2 system is an acceptable tool for ESBL detection among K. pneumoniae and E. coli isolates for laboratories where both imported AmpC and ESBLs are prevalent.

Due to reduced antibiotic consumption in Taiwan, erythromycin resistance rate had decreased in Streptococcus pyogenes, but it increased in Streptococcus pneumoniae. The objectives of the present study were (1) to determine the erythromycin and clindamycin resistance rate and minimum inhibitory concentration (MIC) of the group B streptococcus (GBS) clinical isolates, and (2) to investigate the mechanism responsible for the macrolide, lincosamide, and group B streptogramin (MLS(B)) resistance. A total of 1,395 GBS isolates were collected from June 2001 to April 2007. Forty-four percent of the GBS isolates were resistant to erythromycin, and 39% were resistant to clindamycin. The annual erythromycin resistance rate increased from 32% in 2001 to 51% in 2004; a significant decrease was observed in 2005 (47%), 2006 (42%), and 2007 (38%). Percentage of erythromycin-resistant isolates with erm(B) gene significantly increased from 72% in 2001 to 90% in 2007. We found that the plasmid encoded zeta gene was present in 13% of the resistant isolates, along with erm(B). When compared to our previous study (1991 to May 2001), the overall erythromycin resistance rate increased from 30% to 44%. erm(B) was the major resistant determinant, and zeta toxin encoding plasmid has a limited role in mediating erythromycin resistance unlike in GAS isolates as reported earlier.

Background/Purpose: To understand the resistance patterns of rapidly growing mycobacteria (RGM) in Taiwan, antimicrobial resistance of clinical isolates was determined as part of the SMART (Surveillance from Multicenter Antimicrobial Resistance in Taiwan) program. Methods: During the period from January 2002 to December 2003, clinical isolates were collected from eight hospitals located on the west side of Taiwan and one reference laboratory. Broth microdilution minimum inhibitory concentrations of 11 antimicrobial agents were determined for 312 clinical isolates of RGM, including the Mycobacterium fortuitum group (110 isolates), Mycobacterium abscessus group (168 isolates), and Mycobacterium chelonae group (34 isolates). Results: Nearly all of the RGM were susceptible to amikacin and ofloxacin (≥90%) and resistant to doxycycline (＜3% susceptible). Tobramycin showed similar in vitro activity against the M. fortuitum and M. chelonae (77%) groups, but was less active against the M. abscessus group (58%). Ciprofloxacin was active mainly against M. fortuitum (95%). Nearly all RGM were resistant to erythromycin and doxycycline. However, around half of the RGM isolates remained susceptible to minocycline (50-54%). Clarithromycin was active against the M. abscessus group (53% susceptible), with a high rate of resistance in the M. chelonae (38% susceptible) and M. fortuitum (15% susceptible) group. Cefoxitin was more active against the M. fortuitum group (65%) than the other two RGM (40-44%), and ＜40% of the RGM isolates remained susceptible to imipenem (21-38%). Conclusion: The resistance of RGM in Taiwan is not as high as previously reported (notably for tobramycin, ciprofloxacin and cefoxitin), but reduction in the susceptibility rates of clarithromycin and imipenem for the M. fortuitum and M. abscessus groups demonstrates the importance of in vitro susceptibility testing of clinically important isolates, as susceptibility may differ in different geographical areas, even regionally, and over time. [J Formos Med Assoc 2008; 107(4):281-287]

A 269-kilobase conjugative plasmid, pK29, from a Klebsiella pneumoniae strain was sequenced. The plasmid harbors multiple antimicrobial resistance genes, including those encoding CMY-8 AmpC-type and CTX-M-3 extended-spectrum β-lactamases in the common backbone of IncHI2 plasmids. Mechanisms for dissemination of the resistance genes are highlighted in comparative genomic analyses.

Resume Nous rapportons l'observation d'un jeune garcon âge de cinq ans et huit mois, atteint d'une histiocytose a cellules de Langerhans (HCL) vertebrale, qui s'etait revelee par des douleurs dorsales et abdominales, associees a un abaissement du moignon de l'epaule et mimant une hemivertebre T10 avec scoliose sur les radiographies. L'aspect radiologique de la vertebre atteinte par l'HCL n'etait pas celui de la classique vertebra plana , jusqu'a ce que survienne six mois plus tard un tassement vertebral complet, contemporain de la disparition de la scoliose. Le diagnostic d'HCL fut porte par la biopsie, mais avec un retard diagnostique de six mois. Une atteinte polyostotique etait alors mise en evidence, avec atteinte de C6, T5, T9–T12 et L2. Dans cette observation, la presentation de l'HCL est inhabituelle et trompeuse, faisant evoquer une scoliose douloureuse. Il semble que nous rapportions le premier cas posant un tel diagnostic differentiel.

STUDY DESIGN Prospective analysis. OBJECTIVE To investigate biologic influences of recombinant human bone morphogenetic protein (rhBMP)-2 on intervertebral discs after anular tears. SUMMARY OF BACKGROUND DATA Treatments for intervertebral disc injury or degeneration are unsatisfactory. rhBMP-2, a high-potency osteoinductive and chondroinductive substance, is approved for use in anterior lumbar interbody fusions. rhBMP-2 stimulates the proliferation of rat disc cells and the secretion of extracellular matrix in vitro. In vivo responses in the intervertebral disc after anular tears are rarely studied. METHODS Twenty New Zealand white rabbits received full-thickness anular tears and intradiscal injections of saline (control) and rhBMP-2 0.1 mg with and without coral grafts at L2-L3, L3-L4, and L4-L5, respectively. Three died or had infection. Therefore, 17 underwent radiography and sacrifice at 12 weeks. Spinal sections were stained with hematoxylin and eosin to examine responses to rhBMP-2. RESULTS Radiographs revealed degenerative changes, such as disc space narrowing and irregularity, subchondral sclerosis, osteophyte formation, and hypertrophy of vertebral endplates in all groups. Degeneration was more frequent and severe with rhBMP-2 with (P < 0.01) and without (P < 0.05) coral than with saline. Two rabbits receiving rhBMP-2 and coral achieved solid interbody bony fusion. New bone formation was noted in 2 controls, in 3 animals treated with rhBMP-2, and in 4 treated with rhBMP-2 and coral. Vascularity and fibroblast proliferation increased with rhBMP-2 (n = 14) and rhBMP-2 with coral (n = 9) compared with control (n = 3; P < 0.01 and P = 0.03, respectively). Inflammatory infiltrates increased with rhBMP-2 (n = 8) compared with control (n = 2; P = 0.03). CONCLUSIONS Degenerative changes were more frequent and severe in the groups treated with rhBMP-2 with or without coral in radiographic findings. In histopathologic findings, rhBMP-2 promoted hypervascularity and fibroblast proliferation of the intervertebral disc after an anular tear.

The prevalence of three plasmid-mediated quinolone resistance determinants, QnrA, QnrB, and QnrS, among 526 nonreplicate clinical isolates of Enterobacter cloacae collected at a Taiwanese university hospital in 2004 was determined by PCR and colony hybridization, and the association of Qnr with the IMP-8 metallo-β-lactamase was investigated. Eighty-six (16.3%) of all isolates were qnr positive, and the qnrA1 -like, qnrB2 -like, and qnrS1 -like genes were detected alone or in combination in 3 (0.6%), 53 (10.1%), and 34 (6.5%) isolates, respectively. Among 149 putative extended-spectrum-β-lactamase-producing isolates, 59 (39.6%) isolates, all of which were SHV-12 producers, harbored qnrA (0.7%; 1 isolate), qnrB (28.9%; 43 isolates), or qnrS (12.1%; 18 isolates). Forty-four (78.6%) of 56 IMP-8 producers carried qnrB (58.9%; 33 isolates), qnrS (25.0%; 14 isolates), or both. PCR and sequence analysis revealed that qnrA1 was located in a complex sul1 -type integron that contains dhr15 , aadA2 , qacE Δ 1 , sul1 , orf513 , qnrA1 , ampR , and qacE Δ 1 . Conjugation experiments revealed the coexistence of qnrB and bla IMP-8 on the transferred plasmids and the absence of β-lactamase content on the transferred qnrS -positive plasmids. The transferred bla IMP-8 -positive plasmids with and without qnrB had very similar restriction patterns, suggesting the horizontal mobility of qnrB . Pulsed-field gel electrophoresis showed six major patterns among the 44 qnr -positive IMP-8-producing isolates. Thus, the extremely high prevalence of qnr among the metallo-β-lactamase-producing E. cloacae isolates in the hospital may be due mainly to the intrahospital spread of a few clones and the dissemination of plasmids containing both qnrB and bla IMP-8 .

A plasmid containing the qnrS quinolone resistance determinant and the gene encoding the SHV-2 β-lactamase has been discovered from a clinical Klebsiella pneumoniae strain isolated in Taiwan. The complete 98-kb sequence of this plasmid, designated pK245, was determined by using a whole-genome shotgun approach. Transfer of pK245 conferred low-level resistance to fluoroquinolones in electroporant Escherichia coli epi300. The sequence of the immediate region surrounding qnrS in pK245 is nearly identical (>99% identity) to those of pAH0376 from Shigella flexneri and pINF5 from Salmonella enterica serovar Infantis, the two other qnr S-carrying plasmids reported to date, indicating a potential common origin. Other genes conferring resistance to aminoglycosides ( aacC2 , strA , and strB ), chloramphenicol ( catA2 ), sulfonamides ( sul 2), tetracycline ( tetD ), and trimethoprim ( dfrA14 ) were also detected in pK245. The dfrA14 gene is carried on a class I integron. Several features of this plasmid, including three separate regions containing putative replicons, a partitioning-control system, and a type II restriction modification system, suggest that it may be able to replicate and adapt in a variety of hosts. Although no critical conjugative genes were detected, multiple insertion sequence elements were found scattered throughout pK245, and these may facilitate the dissemination of the antimicrobial resistance determinants. We conclude that pK245 is a chimera which acquired its multiple antimicrobial resistance determinants horizontally from different sources. The identification of pK245 plasmid expands the repertoire of the coexistence of quinolone and extended-spectrum-β-lactam resistance determinants in plasmids carried by various species of the family Enterobacteriaceae in different countries.

Production of extended-spectrum β-lactamases and plasmid-mediated AmpC enzymes was investigated among 291 Escherichia coli and 282 Klebsiella pneumoniae isolates that showed decreased susceptibilities to extended-spectrum cephalosporins from seven Taiwanese medical centers. CTX-M-type and SHV-type enzymes were the most prevalent extended-spectrum β-lactamases. CMY-2-like and DHA-1-like β-lactamases were the most prevalent AmpC-type enzymes.