Implementable assay for monitoring minimum residual disease after radical treatment for colorectal cancer.

Considering the cost and invasiveness of monitoring postoperative minimal residual disease (MRD) of colorectal cancer (CRC) after adjuvant chemoradiotherapy (ACT), we developed a favorable approach based on methylated circulating tumor DNA to detect MRD after radical resection. Analyzing the public database, we identified the methylated promoter regions of the genes FGD5, GPC6, and MSC. Using digital polymerase chain reaction (dPCR), we termed the "amplicon of methylated sites using a specific enzyme" assay as "AMUSE." We examined 180 and 114 pre- and postoperative serial plasma samples from 28 recurrent and 19 recurrence-free pathological stage III CRC patients, respectively. The results showed 22 AMUSE-positive of 28 recurrent patients (sensitivity, 78.6%) and 17 AMUSE-negative of 19 recurrence-free patients (specificity, 89.5%). AMUSE predicted recurrence 208 days before conventional diagnosis using radiological imaging. Regarding ACT evaluation by the reactive response, 19 AMUSE-positive patients during their second or third blood samples showed a significantly poorer prognosis than the other patients (p = 9E-04). The AMUSE assay stratified four groups by the altered patterns of tumor burden postoperatively. Interestingly, only 34.8% of cases tested AMUSE-negative during ACT treatment, indicating eligibility for ACT. The AMUSE assay addresses the clinical need for accurate MRD monitoring with universal applicability, minimal invasiveness, and cost-effectiveness, thereby enabling the timely detection of recurrences. This assay can effectively evaluate the efficacy of ACT in patients with stage III CRC following curative resection. Our study strongly recommends reevaluating the clinical application of ACT using the AMUSE assay.

Single-tube one-step gel-based RT-RPA/PCR for highly sensitive molecular detection of HIV.

We developed a single-tube one-step gel-based reverse transcription-recombinase polymerase amplification (RT-RPA)/polymerase chain reaction (PCR) (termed "SOG RT-RPA/PCR") to detect the human immunodeficiency virus (HIV). To improve the assay sensitivity, the RNA template is pre-amplified by RT-RPA prior to PCR. To simplify the detection process and shorten the assay time, we embedded PCR reagents into agarose gel, constructing it to physically separate the reagents from the RT-RPA reaction solution in a single tube. Due to the thermodynamic properties of agarose, the RT-RPA reaction first occurs independently on top of the PCR gel at a low temperature (e.g., 39 °C) during the SOG RT-RPA/PCR assay. Then, the RPA amplicons directly serve as the template for the second PCR amplification reaction, which begins when the PCR agarose dissolves due to the elevated reaction temperature, eliminating the need for multiple manual operations and amplicon transfer. With our SOG RT-RPA/PCR assay, we could detect 6.3 copies of HIV RNA per test, which is a 10-fold higher sensitivity than that of standalone real-time RT-PCR and RT-RPA. In addition, due to the high amplification efficiency of RPA, the SOG RT-RPA/PCR assay shows stronger fluorescence detection signals and a shorter detection time compared to the standalone real-time RT-PCR assay. Furthermore, we detected HIV viral RNA in clinical plasma samples and validated the superior performance of our assay. Thus, the SOG RT-RPA/PCR assay offers a powerful method for simple, rapid, and highly sensitive nucleic acid-based molecular detection of infectious diseases.

Flow cytometry assay for the detection of single-copy DNA in human lymphocytes.

Specific nucleic acid sequences can be detected in individual cells by in situ hybridization. However, when very few copies of a target sequence are present per cell, its signal is undetectable by flow cytometry. Although various approaches have been developed to increase fluorescence signals for in situ hybridization, flow cytometric detection of specific genomic DNA sequences has not been established. Here, we present a flow cytometry assay for detection of single-copy genomic sequences in human lymphocytes using in situ PCR with universal energy transfer-labelled primers.

Evaluation of a novel rapid TRC assay for the detection of influenza using nasopharyngeal swabs and gargle samples.

We evaluated a novel transcription-reverse transcription concerted reaction (TRC) assay that can detect influenza A and B within 15 min using nasopharyngeal swab and gargle samples obtained from patients with influenza-like illness, between January and March 2018 and between January and March 2019. Based on the combined RT-PCR and sequencing results, in the nasal swabs, the sensitivity and specificity of TRC for detecting influenza were calculated as 1.000 and 1.000, respectively. In the gargle samples, the sensitivity and specificity of TRC were 0.946 and 1.000, respectively. The TRC assay showed comparable performance to RT-PCR in the detection of influenza viruses.

Rapid detection and surveillance of cfiA-positive Bacteroides fragilis using matrix-assisted laser desorption ionization time-of-flight mass spectrometry.

OBJECTIVES: To perform surveillance of cfiA-positive Bacteroides fragilis using new subtyping software module, MALDI Biotyper Subtyping Module (MBT Subtyping Module), on MALDI-TOF MS system, and to evaluate the detection ability of the module. METHODS: cfiA-positive strains were presumed using the module against B. fragilis isolated between 2006 and 2019. The cfiA gene was confirmed using PCR. In cfiA-positive B. fragilis, the insertion sequence (IS) elements were examined and the MBT STAR-BL assay was performed to examine meropenem hydrolysis activity. RESULTS: Of the 396 B. fragilis strains included, the MBT Subtyping Module detected 33 presumptive cfiA-positive strains (8.3%), of which 32 harbored the cfiA gene. The sensitivity and specificity of the MBT Subtyping Module for detecting cfiA-positive B. fragilis were 100.0% and 99.7%, respectively. Of the 32 strains harboring the cfiA gene, seven strains possessed IS elements, which were thought to induce high cfiA expression. Meropenem hydrolysis was detected in all seven strains that were positive for both cfiA and IS elements, and they exhibited resistance to meropenem and imipenem. The overall non-susceptibility rates to meropenem and imipenem were 84.8% and 36.4%, respectively, in the 33 presumptive cfiA-positive strains. CONCLUSION: The MBT Subtyping Module can detect cfiA-positive B. fragilis rapidly and accurately, supporting its use for surveillance of cfiA-positive B. fragilis in clinical settings.

A novel macrolide, solithromycin suppresses mucin overexpression induced by Pseudomonas aeruginosa LPS in airway epithelial cells.

Some macrolides such as 14- and 15-membered macrolides have immunomodulatory effects such as suppression of mucin overproduction. Because a novel macrolide, solithromycin, was developed, we examined whether it suppresses the overexpression of mucin in vitro. A human airway epithelial cell line NCI-H292 was stimulated by Pseudomonas aeruginosa lipopolysaccharides to induce the overproduction of a major mucin, MUC5AC. Treatment with 10 µg/mL of solithromycin significantly inhibited LPS-induced MUC5AC in both mRNA and protein levels as well as a 15-membered macrolide, azithromycin. These findings support that solithromycin has a potential immunomodulatory effect.

Effect of probiotics on gut microbiome in patients with administration of surgical antibiotic prophylaxis: A randomized controlled study.

Surgical antibiotic prophylaxis (SAP) is recommended for the prevention of surgical site infections. However, there is a concern about adverse effects of SAP, such as antibiotic-associated diarrhea (AAD). To prevent AAD, administration of probiotics has been investigated. Although recent advances in next-generation sequencing makes it possible to analyze the gut microbiome, the effect of probiotics on the gut microbiome in the patients with SAP remains unknown. To test a hypothesis that SAP influences the gut microbiome and probiotics prevent the influence, a randomized controlled study was conducted with patients who underwent spinal surgery at Nagasaki University Hospital. After obtaining informed consent, the patients were automatically classified into the non-probiotics group and the probiotics group. In the probiotics group, the patients took 1 g of Enterococcus faecium 129 BIO 3B-R, 3 times a day on postoperative days (PODs) 1-5. The feces of all patients were sampled before administration of SAP and on PODs 5 and 10. We compared alpha and beta diversity and differential abundance analysis of the gut microbiome before and after SAP. During the study period, a total of 33 patients were evaluated, comprising 17 patients in the non-probiotics group and 16 in the probiotics group. There was no significant difference between the groups regarding patient characteristics. In alpha and beta diversity, there were no significant differences among all combinations. In differential abundance analysis at operational taxonomic unit level, Streptococcus gallolyticus and Roseburia were significantly increased in the non-probiotics group and significantly decreased in the probiotics group.

Molecular epidemiology of Clostridioides difficile and risk factors for the detection of toxin gene-positive strains.

In this study, we investigated all Clostridioides difficile strains isolated from stool samples in Nagasaki University Hospital between January 2012 and December 2014. Toxin genes (tcdA, tcdB and cdtA/cdtB) were analyzed for multiplex PCR in a total of 213 strains. In the toxin gene-positive strain, PCR ribotyping was conducted using capillary gel electrophoresis-based PCR and the Webribo database. Patients' backgrounds were analyzed by departments, disorders, antimicrobials, and clinical dates. The positive rates of tcdA, tcdB, and cdtA/cdtB genes were 62.9%, 63.4%, and 2.8%, respectively. The most frequent PCR ribotype was 047 (14.1%), followed by 014/0 (11.1%) and 002/0 (8.2%). In univariate analysis, the risk factors for the detection of toxin gene-positive strains in patients were older age (p = 0.0036), over ≥ 65 years old (p = 0.0175), the patients hospitalized at Department of Digestive Surgery (P = 0.0059), higher CRP level (P = 0.0395), and lower albumin level (p = 0.0014). In the multivariate analysis, the risk factor for detection of toxin gene-positive strains was the patients hospitalized at Department of Digestive Surgery (OR; 4.62, 95% CI; 1.18-18.0, p = 0.0274). In this study, the percentage of toxin gene-positive and cdtA/cdtB gene-positive strains was almost the same as that reported in previous studies, but the ribotype was different. In addition, we revealed that the risk factor associated with the detection of toxin gene-positive strains was the patients hospitalized at Department of digestive surgery.

Molecular and epidemiological analysis of IMP-1 metallo-ß-lactamase-producing Klebsiella pneumoniae in a tertiary care hospital in Japan.

This study investigated the molecular and phenotypic characteristics of carbapenemase-producing Klebsiella pneumoniae, and identified the risk factors underlying its acquisition. We evaluated K. pneumoniae isolated in Nagasaki University Hospital between January 2009 and June 2015. The presence of carbapenemase genes and plasmid characteristics were investigated. We performed multilocus sequence typing (MLST), and generated a dendrogram based on the results of pulsed-field gel electrophoresis (PFGE) for carbapenemase-producing strains. We also performed a case-control study of patients. Of the 88 K. pneumoniae strains that showed minimum inhibitory concentration ≥1 µg/mL for imipenem and/or meropenem, and that were available from our bacterial collection, 18 had the IMP-type carbapenemase gene, all of which were IMP-1 according to sequencing analysis. Strains included seven different sequence types (STs), of which the most common was ST1471. A dendrogram showed the significant similarity of some strains with relationships in PFGE patterns, STs, and the wards in which they were isolated. Plasmid incompatibility group was similar among the IMP-1 producers. Regarding risk factors, multivariate analysis showed that liver disease and previous uses of carbapenems and anti-fungal drugs were significant factors for the acquisition of IMP-1-producing strains. Our results demonstrate that IMP-1 is a major carbapenemase produced by K. pneumoniae. The PFGE results indicated the possibility of transmission in the hospital. The identified risk factors should be considered for appropriate antibiotic therapy and infection-control measures.

In Vitro Activity of Lascufloxacin against Streptococcus pneumoniae with Mutations in the Quinolone Resistance-Determining Regions.

Lascufloxacin showed potent activity against Streptococcus pneumoniae with a GyrA or ParC mutation (first-step mutant). The frequency of selecting resistant strains tended to be lower for lascufloxacin than for levofloxacin and garenoxacin after drug exposure in first-step mutants but was similar in the comparison between lascufloxacin and moxifloxacin. The increase in MIC was smaller for lascufloxacin than for levofloxacin, garenoxacin, and moxifloxacin when clinical strains with only ParC mutations were exposed to the corresponding drug.

Evaluation of FilmArray respiratory panel multiplex polymerase chain reaction assay for detection of pathogens in adult outpatients with acute respiratory tract infection.

Although viruses are the major pathogen that causes upper respiratory tract infection (URTI) and acute bronchitis, antibiotics have been prescribed. This was a prospective observational study in influenza epidemics that enrolled adult outpatients who visited a hospital with respiratory tract infection symptoms. In this study, we evaluated the usefulness of FilmArray respiratory panel (RP). Fifty patients were enrolled. FilmArray RP detected the pathogens in 28 patients. The common pathogens were influenza virus (n = 14), respiratory syncytial virus (n = 6), and human rhinovirus (n = 6). Of the 14 patients with influenza virus, 6 were negative for the antigen test. The physicians diagnosed and treated the patients without the result of FilmArray in this study. Of the patients with positive FilmArray RP, 9 were treated with antibiotics; however, bacteria were detected in only 3 patients. By implementing FilmArray RP, URTI and acute bronchitis would be precisely diagnosed, and inappropriate use of antibiotics can be reduced.

Efficacy and Pharmacokinetics of the Combination of OP0595 and Cefepime in a Mouse Model of Pneumonia Caused by Extended-Spectrum-Beta-Lactamase-Producing Klebsiella pneumoniae.

OP0595 (RG6080) is a novel diazabicyclooctane that inhibits class A and C serine beta-lactamases. Although the combination of OP0595 and cefepime (FEP) showed good in vitro activity against extended-spectrum-beta-lactamase (ESBL)-producing pathogens, the effect of the combination therapy against severe infections, such as pneumonia or bacteremia, remains unknown in vivo In this study, we investigated the efficacy and pharmacokinetics of the combination therapy of OP0595 and FEP in a mouse model of pneumonia caused by Klebsiella pneumoniae harboring SHV- and CTX-M-9-type ESBLs. The infected BALB/c mice were intraperitoneally administered saline (control), 100 mg/kg of body weight of FEP, 20 mg/kg of OP0595, or both FEP and OP0595, twice a day. The MIC of FEP against the bacteria was 8 mg/liter and markedly improved to 0.06 mg/liter with the addition of 0.5 mg/ml of OP0595. In the survival study, the combination of FEP and OP0595 significantly improved the survival rate compared with that reported with either OP0595 or FEP alone (P < 0.001). The number of bacteria in the lungs and blood significantly decreased in the combination therapy group compared to that reported for the monotherapy groups (P < 0.001). In addition, the in vivo effect depended on the dose of FEP. However, pharmacokinetic analysis revealed that the percentage of time above MIC remained constant when increasing the dose of FEP in combination with 20 mg/kg of OP0595. The results of our study demonstrated the in vivo effectiveness of the combination of OP0595 and FEP.

Evaluation of Sensitivity and Specificity Performance of Elecsys HTLV-I/II Assay in a Multicenter Study in Europe and Japan.

Screening of blood for human T-cell lymphotropic virus type 1 and type 2 (HTLV-1 and -2, respectively) is important to diagnose and prevent infection and ensure the safety of blood supplies. The Elecsys HTLV-I/II assay is a newly developed, electrochemiluminescence screening assay for the detection of HTLV-1/2 infection. The sensitivity and specificity of the Elecsys HTLV-I/II assay were determined using well-characterized HTLV-1/2-positive serum and plasma samples and routine diagnostic and blood donor samples expected to be HTLV negative, respectively. These results were compared with those for at least one of the following CE-marked assays at seven independent laboratories and the Roche Diagnostics facility in Penzberg, Germany: Abbott Architect rHTLV-I/II, Ortho Avioq HTLV-I/II Microelisa system, Abbott Prism HTLV-I/HTLV-II, and DiaSorin Murex HTLV I+II. Fujirebio INNO-LIA HTLV-I/II Score was used as a confirmatory assay. The Elecsys HTLV-I/II, Abbott Architect rHTLV-I/II, and Abbott Prism HTLV-I/HTLV-II assays detected all HTLV-1/2-positive samples (sensitivity, 100%). Sensitivity for Ortho Avioq HTLV-I/II was 98.63%. The Elecsys HTLV-I/II assay had a specificity of 99.95% in blood donor samples, which was comparable to results for the other assays (range, 99.91 to 100%). In routine diagnostic samples, the specificity of the Elecsys HTLV-I/II assay was 99.83%, compared with 99.70% for Abbott Architect rHTLV-I/II. Specificity for the Elecsys HTLV-I/II assay in potentially cross-reactive samples was 100%, compared with 99.0% for Ortho Avioq HTLV-I/II and 99.2% for DiaSorin Murex HTLV I+II. The Elecsys HTLV-I/II assay has the sensitivity and specificity to support its use as a routine screening assay for detecting HTLV infection.

Efficacy and pharmacokinetics of ME1100, a novel optimized formulation of arbekacin for inhalation, compared with amikacin in a murine model of ventilator-associated pneumonia caused by Pseudomonas aeruginosa.

Background: Arbekacin is an aminoglycoside that shows strong antimicrobial activity against Gram-positive bacteria, including MRSA, as well as Pseudomonas aeruginosa . The therapeutic effectiveness of arbekacin is directly related to C max at the infection site. To maximize drug delivery to the respiratory tract and minimize the systemic toxicity, arbekacin optimized for inhalation, ME1100, is under development. In this study, we investigated the efficacy and pharmacokinetics of ME1100 in a murine model of ventilator-associated pneumonia caused by P. aeruginosa by using a customized investigational nebulizer system. Methods: The mice were treated for 5 min, once daily, with placebo, 3, 10 or 30 mg/mL ME1100 or 30 mg/mL amikacin. Results: In the survival study, the survival rate was significantly improved in the 10 and 30 mg/mL ME1100 treatment groups compared with that in the placebo group. The number of bacteria in the lungs was significantly lower in the 30 mg/mL ME1100 treatment group at 6 h after the initial treatment, compared with all other groups. In the pharmacokinetic study, the C max in the 30 mg/mL ME1100 treatment group in the epithelial lining fluid (ELF) and plasma was 31.1 and 1.2 mg/L, respectively. Furthermore, we compared the efficacy of ME1100 with that of amikacin. Although there were no significant differences in ELF and plasma concentrations between 30 mg/mL of ME1100 and 30 mg/mL of amikacin, ME1100 significantly improved the survival rate compared with amikacin. Conclusions: The results of our study demonstrated the in vivo effectiveness of ME1100 and its superiority to amikacin.

TNF-α inhibits the growth of Legionella pneumophila in airway epithelial cells by inducing apoptosis.

BACKGROUND: TNF-α plays an important role in the pathogenesis of Legionella pneumophila (Lp)-induced pneumonia. Patients undergoing anti-TNF-α therapy are at an increased risk of Lp infection. Lp infects both phagocytic and non-phagocytic cells such as airway epithelial cells; however, the role of TNF-α in airway epithelial cells is unknown. METHODS: Human airway epithelial cell line NCI-H292 was infected with Lp NUL1 strain. After infection, both intracellular growth of Lp and cell death were evaluated after treating the cells with or without TNF-α. Apoptosis was examined by performing activated caspase-3/7 staining and by using a pan-caspase inhibitor. RESULTS: Lp infected and replicated in NCI-H292 cells in a time-dependent manner, and TNF-α treatment of Lp-infected NCI-H292 cells inhibited Lp replication. Inhibitory effects of TNF-α on Lp replication were suppressed after treatment with a TNF-α-neutralizing antibody. Lp infection increased extracellular lactate dehydrogenase levels and decreased the number of living cells. Increased number of Lp-infected NCI-H292 cells showed caspase-3/7 activation, indicating they underwent apoptosis. TNF-α treatment inhibited Lp replication by increasing the apoptosis of NCI-H292 cells. CONCLUSIONS: Thus, our results suggested that airway epithelial cells were involved in the pathogenesis of Lp infection and that TNF-α played a protective role by inhibiting the intracellular replication of Lp and by increasing the apoptosis of Lp-infected airway epithelial cells. However, Lp infection should be investigated further in patients undergoing anti-TNF-α therapy who develop pneumonia.

Performance evaluation of the Verigene®Clostridium difficile nucleic acid test, an automated multiplex molecular testing system for detection of C. difficile toxin.

The Verigene®Clostridium difficile nucleic acid test (Verigene® CDF test) is an automatic and rapid detection system for the genes encoding tcdA, tcdB, binary toxin, and the single nucleotide deletion at base pair 117 in the tcdC based on microarray and PCR amplification. We compared the performance of the Verigene® CDF test to that of two enzyme immunoassays, C. DIFF QUIK CHEK COMPLETE and X/Pect Toxin A/B, using 118 specimens. We found overall concordance rates of 81.4% and 78.8% between C. DIFF QUIK CHEK COMPLETE and Verigene® CDF test, and X/Pect Toxin A/B and Verigene® CDF test. The Verigene® CDF test showed the highest sensitivity (93.9%) and had a specificity of 96.5%. The sensitivity and specificity were respectively 45.5 and 94.1% for C. DIFF QUIK CHEK COMPLETE and 27.3 and 100.0% for X/Pect Toxin A/B. These results indicated that the Verigene® CDF test was highly accurate for the detection of C. difficile toxin in fecal specimens and supported its use in daily diagnostic practice.

Tedizolid inhibits MUC5AC production induced by methicillin-resistant Staphylococcus aureus in human airway epithelial cells.

The innate immune system plays an important role in early immunity against respiratory tract infection. Although airway epithelial cells produce mucus to eliminate pathogens and irritants, hypersecretion of mucus is harmful for the host as it may cause airway obstruction and inhibit influx of antimicrobial agents. It has been reported that several antimicrobial agents have an immunomodulatory effect in vitro and in vivo, but little is known about whether tedizolid, a novel oxazolidinone, can modulate immune responses. In this study, we evaluated whether tedizolid can suppress MUC5AC production in human airway epithelial cells stimulated by methicillin-resistant Staphylococcus aureus (MRSA). Compared with the control, tedizolid significantly inhibited MUC5AC protein production and mRNA overexpression at concentrations of both 2 and 10 µg/mL (representative of trough and peak concentrations in human epithelial lining fluid). Among the mitogen-activated protein kinase inhibitors tested, only extracellular signal-regulated protein kinase 1/2 (ERK1/2) phosphorylation was inhibited by tedizolid as indicated by western blot analysis. These results indicate that tedizolid inhibits the overproduction of MUC5AC protein by inhibiting phosphorylation of ERK1/2. This study revealed that tedizolid suppresses excessive mucin production in human airway epithelial cells. The immunomodulatory effect of tedizolid may improve outcomes in patients with severe respiratory infectious diseases caused by MRSA.

Induction of apoptosis by HBI-8000 in adult T-cell leukemia/lymphoma is associated with activation of Bim and NLRP3.

Adult T-cell leukemia/lymphoma (ATL) is an aggressive T-cell malignancy caused by human T-cell lymphotropic virus 1. Treatment options for acute ATL patients include chemotherapy, stem cell transplantation, and recently the anti-chemokine (C-C motif) receptor 4 antibody, although most patients still have a poor prognosis and there is a clear need for additional options. HBI-8000 is a novel oral histone deacetylase inhibitor with proven efficacy for treatment of T-cell lymphomas that recently received approval in China. In the present study, we evaluated the effects of HBI-8000 on ATL-derived cell lines and primary cells obtained from Japanese ATL patients. In most cases HBI-8000 induced apoptosis in both primary ATL cells and cell lines. In addition, findings obtained with DNA microarray suggested Bim activation and, interestingly, the contribution of the NLR family, pyrin domain containing 3 (NLRP3) inflammasome pathway in HBI-8000-induced ATL cell death. Further investigations using siRNAs confirmed that Bim contributes to HBI-8000-induced apoptosis. Our results provide a rationale for a clinical investigation of the efficacy of HBI-8000 in patients with ATL. Although the role of NLRP3 inflammasome activation in ATL cell death remains to be verified, HBI-8000 may be part of a novel therapeutic strategy for cancer based on the NLRP3 pathway.

Antimicrobial and immunomodulatory effects of tedizolid against methicillin-resistant Staphylococcus aureus in a murine model of hematogenous pulmonary infection.

Tedizolid (TZD) is a second-generation oxazolidinone and demonstrates potent in-vitro activity against multidrug-resistant Gram-positive bacteria. Phase III studies in patients with acute bacterial skin and skin structure infections (ABSSSI) have demonstrated the non-inferiority of TZD to linezolid (LZD). However, there are only a few studies that show the effect of TZD in pulmonary infections. In this study, we investigated the effect of TZD in a murine model of hematogenous pulmonary infection caused by methicillin-resistant Staphylococcus aureus (MRSA). The mice were treated either twice daily with saline (control), 25mg/kg of vancomycin (low-VAN), 110mg/kg of vancomycin (high-VAN), 120mg/kg of LZD or once daily with 20mg/kg of TZD. As compared to the control, the low- and high-VAN treatment groups, LZD and TZD significantly improved the survival rate, reduced the bacterial count in the lungs. Furthermore, TZD decreased the area of central bacterial colony zone (CBCZ) at 36h post-inoculation, compared with the control. In addition, we investigated the immunomodulatory effect of TZD by evaluating the plasma concentrations of the inflammatory cytokines. Although there were no significant differences in the bacterial count in the lungs amongst the drugs at 26h post-inoculation, TZD and LZD significantly improved the plasma concentrations of TNF-alpha, IL-6 and MIP-2, in comparison with the control. In this study, both TZD and LZD demonstrated antimicrobial and immunomodulatory efficacy in a murine model of hematogenous pulmonary infection caused by MRSA.

Antimicrobial susceptibility and molecular characteristics of methicillin-resistant Staphylococcus aureus in a Japanese secondary care facility.

Methicillin-resistant Staphylococcus aureus (MRSA) is prevalent in Japan, and the Staphylococcus cassette chromosome mec (SCCmec) type II is common among hospital-acquired MRSA isolates. Information pertaining to MRSA characteristics is limited, including SCCmec types, in primary or secondary care facilities. A total of 128 MRSA isolates (90 skin and soft tissue isolates and 38 blood isolates) were collected at a secondary care facility, Kawatana Medical Center, from 2005 to 2011. Antimicrobial susceptibility testing for anti-MRSA antibiotics and molecular testing for SCCmec and virulence genes (tst, sec, etb, lukS/F-PV) were performed. Strains positive for lukS/F-PV were analyzed by multilocus sequence typing and phage open-reading frame typing. SCCmec typing in skin and soft tissue isolates revealed that 65.6% had type IV, 22.2% had type II, 8.9% had type I, and 3.3% had type III. In blood isolates, 50.0% had type IV, 47.4% had type II, and 2.6% had type III. Minimum inhibitory concentrations, MIC(50)/MIC(90), against vancomycin, teicoplanin, linezolid, and arbekacin increased slightly in SCCmec II isolates from skin and soft tissue. MICs against daptomycin were similar between sites of isolation. SCCmec type II isolates possess tst and sec genes at a greater frequently than SCCmec type IV isolates. Four lukS/F-PV-positive isolates were divided into two clonal patterns and USA300 was not included. In conclusion, SCCmec type IV was dominant in blood, skin, and soft tissue isolates in a secondary care facility in Japan. Because antimicrobial susceptibility varies with the SCCmec type, SCCmec typing of clinical isolates should be monitored in primary or secondary care facilities.