Surveillance of multi-drug resistance phenotypes in Staphylococcus aureus in Japan and correlation with whole-genome sequence findings.

BACKGROUND: Antimicrobial resistance in Staphylococcus aureus imposes a high disease burden. Both phenotypic and genotypic monitoring are key to understanding and containing emerging resistant strains. AIM: Phenotypic monitoring of emerging resistance in S. aureus and correlation of priority strain phenotypes with whole-genome sequencing (WGS) findings. METHODS: Antimicrobial susceptibility test results of >40,000 isolates from 213 participating hospitals from 2011 to 2019 were exported from the national Japan Nosocomial Infections Surveillance (JANIS) database. Longitudinal and geographic distribution and prevalence of distinct multi-drug resistance phenotypes ('resistance profiles') of S. aureus were examined among hospitals and prefectures. We further conducted a genome sequence analysis of strains with specific resistance profiles of concern. FINDINGS: The overall prevalence of meticillin-resistant S. aureus (MRSA) decreased from 40.3% to 35.1% from 2011 to 2019. However, among dozens of S. aureus resistance profiles, only one profile of a type of MRSA, exhibited a statistically significant increase in inpatient frequency, exceeding 10% during the nine years. This MRSA profile showed resistance to oxacillin, erythromycin and levofloxacin. Analysis of WGS results of S. aureus isolates with this phenotype revealed that most belonged to clonal complex 8, and all carried SCCmec IV, typical of community-acquired MRSA. CONCLUSION: Tracking distinct resistance profiles deepened our understanding of the overall decrease in MRSA and led to recognition of the emergence of a new resistance phenotype. This study provides a model for future epidemiological research on antimicrobial resistance correlating multi-drug resistance phenotypes with selective genome sequencing, which can be applied to other bacterial species.

Impact of the COVID-19 pandemic on the surveillance of antimicrobial resistance.

BACKGROUND: The impact of the coronavirus disease (COVID-19) pandemic on antimicrobial resistance (AMR) is a major concern. AIM: To compare the number of patients and isolation rate of antimicrobial-resistant bacteria before and after the beginning of the COVID-19 pandemic using the comprehensive national surveillance data. METHODS: We utilized comprehensive surveillance data, collected in the Japan Nosocomial Infections Surveillance programme, which included a total of 16.7 million samples of 5.9 million tested patients from >1300 hospitals. We compared the number of patients and isolation rate of five bacteria between 2019 and 2020, including antimicrobial-susceptible and -resistant bacteria of Staphylococcus aureus, Streptococcus pneumoniae, Escherichia coli, Klebsiella pneumoniae and Pseudomonas aeruginosa. FINDINGS: The number of patients and isolation rate of S. aureus and meticillin-resistant S. aureus decreased slightly; those of S. pneumoniae and penicillin-resistant S. pneumoniae decreased by 60%; and those of third-generation cephalosporin-resistant K. pneumoniae increased. The isolation rate of the remaining bacteria apparently increased, although the number of patients decreased. This was due to a substantial decrease in the total number of tested patients (the denominator of the isolation rate), which was larger than that of the number of patients (the numerator of the isolation rate). Consistent results were obtained when the same data were re-aggregated using the procedure of the World Health Organization Global Antimicrobial Resistance Surveillance System, demonstrating the general importance of this problem. CONCLUSION: Surveillance data during the COVID-19 pandemic must be carefully interpreted based on examination of the numerator, denominator and background factors that affect the denominator.

Reduction in chlorhexidine efficacy against multi-drug-resistant Acinetobacter baumannii international clone II.

BACKGROUND: Nosocomial infections caused by Acinetobacter baumannii international clone II (IC II) can cause severe clinical outcomes. AIM: Differential evaluation of bactericidal efficacy of chlorhexidine gluconate (CHX) and benzethonium chloride (BZT) disinfectants against IC II and non-IC II isolates. METHODS: Minimum inhibitory concentrations (MICs) of CHX and BZT were determined for 137 A. baumannii IC II, 99 non-IC II and 69 non-baumannii isolates, further classified according to MIC values into disinfectant-reduced susceptible (DRS) and disinfectant-susceptible (DS) groups. Time-kill curves and minimum bactericidal concentrations (MBCs) were evaluated for representative isolates in each group. RESULTS: CHX and BZT MIC90s for IC II isolates were 100 and 175mg/L, respectively, but those for non-IC II and non-baumannii isolates were <100mg/L. Nevertheless, time-kill curves indicated that CHX and BZT reduced live bacterial cell number by 5 log10 for IC II and non-IC II isolates within 30s when used at 1000mg/L, comparable to practical use concentrations. CHX MBC at 30s was 1000mg/L for IC II and non-IC II isolates, and was not influenced by addition of 3% bovine serum albumin (BSA); BZT MBC at 30s was 100mg/L without BSA and increased up to 500mg/L upon addition of BSA. No significant differences in BSA were found between DRS and DS isolates. CONCLUSION: CHX and BZT were effective against Acinetobacter spp. including IC II at a concentration of 1000mg/L and exposure for at least 30s, but their concentrations should be considered carefully to ensure sufficient effects in both clinical and healthcare settings.

Emergence of New Delhi metallo-beta-lactamase 1 and other carbapenemase-producing Acinetobacter calcoaceticus-baumannii complex among patients in hospitals in Ha Noi, Viet Nam.

Acinetobacter baumannii is an important cause of multidrug-resistant hospital acquired infections in the world. Here, we investigate the presence of NDM-1 and other carbapenemases among carbapenem-resistant A. baumannii isolated between August 2010 and December 2014 from three large hospitals in Hanoi, Vietnam. We identified 23/582 isolates (4 %) (11 from hospital A, five from hospital B, and seven from hospital C) that were NDM-1 positive, and among them 18 carried additional carbapenemase genes, including seven isolates carrying NDM-1, IMP-1, and OXA-58 with high MICs for carbapenems. Genotyping indicated that NDM-1 carrying A. baumannii have expanded clonally in these hospitals. Five new STs (ST1135, ST1136, ST1137, ST1138, and ST1139) were identified. One isolate carried NDM-1 on a plasmid belonging to the N-repA replicon type; no NDM-1-positive plasmids were identified in the other isolates. We have shown the extent of the carbapenem resistance and the local clonal spread of A. baumannii carrying NDM-1 in these hospitals; coexistence of NDM-1 and IMP-1 is reported for the first time from Vietnam here, and this will further seriously limit future therapeutic options.

Variation in patient position and impact on carbon-ion scanning beam distribution during prostate treatment.

OBJECTIVE: We assessed the impact of changes in patient position on carbon-ion scanning beam distribution during treatment for prostate cancer. METHODS: 68 patients were selected. Carbon-ion scanning dose was calculated. Two different planning target volumes (PTVs) were defined: PTV1 was the clinical target volume plus a set-up margin for the anterior/lateral sides and posterior side, while PTV2 was the same as PTV1 minus the posterior side. Total prescribed doses of 34.4 Gy [relative biological effectiveness (RBE)] and 17.2 Gy (RBE) were given to PTV1 and PTV2, respectively. To estimate the influence of geometric variations on dose distribution, the dose was recalculated on the rigidly shifted single planning CT based on two dimensional-three dimensional rigid registration of the orthogonal radiographs before and after treatment for the fraction of maximum positional changes. RESULTS: Intrafractional patient positional change values averaged over all patients throughout the treatment course were less than the target registration error = 2.00 mm and angular error = 1.27°. However, these maximum positional errors did not occur in all 12 treatment fractions. Even though large positional changes occurred during irradiation in all treatment fractions, lowest dose encompassing 95% of the target (D95)-PTV1 was >98% of the prescribed dose. CONCLUSION: Intrafractional patient positional changes occurred during treatment beam irradiation and degraded carbon-ion beam dose distribution. Our evaluation did not consider non-rigid deformations, however, dose distribution was still within clinically acceptable levels. ADVANCES IN KNOWLEDGE: Inter- and intrafractional changes did not affect carbon-ion beam prostate treatment accuracy.

Common isolation of New Delhi metallo-beta-lactamase 1-producing Enterobacteriaceae in a large surgical hospital in Vietnam.

This study sought to monitor the presence of carbapenem-resistant Enterobacteriaceae (CRE) and the proportion New Delhi metallo-beta-lactamase 1 (NDM-1)-producing bacteria between August 2010 and December 2012 in a surgical hospital in Vietnam. We identified 47 CRE strains from a total of 4,096 Enterobacteriaceae isolates (1.1 %) that were NDM-1-positive from 45 patients admitted to 11 different departments, with the majority being from the urology department. The NDM-1 gene was found in seven different species. Genotyping revealed limited clonality of NDM-1-positive isolates. Most of the isolates carried the NDM-1 gene on a plasmid and 17.8 % (8/45) of those were readily transferable. We found five patients at admission and one patient at discharge with NDM-1-positive bacteria in their stool. From 200 screening environmental hospital samples, five were confirmed to be NDM-1-positive and included Acinetobacter species (n = 3) and Enterobacter aerogenes (n = 2). The results reveal that NDM-1-producing Enterobacteriaceae are commonly isolated in patients admitted to a Vietnamese surgical hospital and are also detected in the hospital environment.

Laboratory-based surveillance of pertussis using multitarget real-time PCR in Japan: evidence for Bordetella pertussis infection in preteens and teens.

Between January 2013 and December 2014, we conducted laboratory-based surveillance of pertussis using multitarget real-time PCR, which discriminates among Bordetella pertussis, Bordetella parapertussis, Bordetella holmesii and Mycoplasma pneumoniae. Of 355 patients clinically diagnosed with pertussis in Japan, B. pertussis, B. parapertussis and M. pneumoniae were detected in 26% (n = 94), 1.1% (n = 4) and 0.6% (n = 2), respectively, whereas B. holmesii was not detected. It was confirmed that B. parapertussis and M. pneumoniae are also responsible for causing pertussis-like illness. The positive rates for B. pertussis ranged from 16% to 49%, depending on age. Infants aged ≤ 3 months had the highest rate (49%), and children aged 1 to 4 years had the lowest rate (16%, p < 0.01 vs. infants aged ≤ 3 months). Persons aged 10 to 14 and 15 to 19 years also showed high positive rates (29% each); the positive rates were not statistically significant compared with that of infants aged ≤ 3 months (p ≥ 0.06). Our observations indicate that similar to infants, preteens and teens are at high risk of B. pertussis infection.

Immunohistochemical localization of elastin, fibrillins and microfibril-associated glycoprotein-1 in the developing periodontal ligament of the rat molar.

BACKGROUND AND OBJECTIVE: Elastic system fibers are a major component of the periodontal ligament, but little information is available about their detailed composition or the mechanism of elastogenesis in the developing periodontal ligament. The purpose of this study was to investigate immunolocalization of elastin, fibrillins and microfibril-associated glycoprotein-1 (MAGP-1) in the developing periodontal ligament of the rat molar. MATERIAL AND METHODS: Frozen sections of demineralized as well as non-demineralized periodontal ligament of Wistar rats of various ages from 19 days to 7 weeks were incubated with anti-elastin, anti-fibrillin-1 and -2 and anti-MAGP-1 antibodies followed by peroxidase-conjugated secondary antibodies. After incubation with diaminobenzidine solution, immunoreaction products were observed with a light microscope. RESULTS: In the developing periodontal ligament of 19-day-old rats, fibers immunopositive to elastin were not present, but fibers positively stained for fibrillin-2 and MAGP-1 were widely distributed throughout the ligament. The latter fibers were arranged in the apico-occlusal direction along with blood vessels. In 3-week-old rats, fibers stained for elastin were observed for the first time in the apical region of the ligament. The number and distribution pattern of these elastin-positive fibers was basically the same as those in rats aged 5 and 7 weeks. In contrast, fibrillin-2- and MAGP-1-positive fibers were more extensively distributed in the ligament, and their pattern of distribution was comparable to that of reported oxytalan fibers. Fibrillin-1 was, however, not detected either in demineralized sections or in non-demineralized sections, indicating its absence in periodontal ligament. CONCLUSION: Elastin expressed in the periodontal ligament assembled into elaunin fibers in the vicinity of blood vessels. Both fibrillin-2 and MAGP-1 are structural components not only of the elastin-associated microfibrils but also of elastin-free microfibrils, with possible roles in elastogenesis and in periodontal ligament homeostasis.

Genes responsible for dextran-dependent aggregation of Streptococcus sobrinus strain 6715.

INTRODUCTION: Streptococcus sobrinus exhibits more significant dextran-dependent aggregation mediated by glucan-binding proteins than Streptococcus mutans. We have identified four glucan-binding protein C gene (gbpC) homologues designated as gbpC1, gbpC2, dblA and dblB in S. sobrinus in contrast to the single gene gbpC in S. mutans. We attempted to determine which gene is most responsible for the dextran-dependent aggregation of S. sobrinus. METHODS: We introduced mutation with a chemical mutagen, 1-methyl-3-nitro-1-nitrosoguanidine, into S. sobrinus strain 6715 and analysed the four gbpC homologous gene sequences in the parental strain 6715 and an obtained aggregation-negative mutant NUM-Ssg99. We also examined the localization of proteins encoded by these genes in the mutant NUM-Ssg99. RESULTS: The nucleotide sequences of the gbpC1, gbpC2 and dblA genes in NUM-Ssg99 were 100% identical to the homologous genes in parental strain 6715. In contrast, a truncated mutation was detected in the dblB gene and the mutant protein devoid of the LPXTG motif was confirmed by Western blot analysis to be released into the extracellular milieu. CONCLUSION: We conclude that the dblB gene among the four GbpC homologous protein genes is most responsible for aggregation in strain 6715.

One of two gbpC gene homologues is involved in dextran-dependent aggregation of Streptococcus sobrinus.

INTRODUCTION: Streptococcus sobrinus exhibits marked dextran-dependent aggregation mediated by glucan-binding proteins (GBPs). In contrast to Streptococcus mutans, in which the gbpC gene responsible for dextran-dependent aggregation of this organism has been characterized, genes encoding the S. sobrinus GBPs have not yet been identified. METHODS: Recently, we identified the gbpC gene homologue from Streptococcus macacae using polymerase chain reaction primers based on the conserved regions of the gbpC sequence exhibiting intraspecies variations. This method was applied to amplify a S. sobrinus homologue. RESULTS: Unexpectedly, two gbpC gene homologues were identified in S. sobrinus strain 100-4. One homologue, named gbpC, was more similar to the S. mutans gbpC gene than the other and was approximately half the molecular size of its homologue with similar regions interrupted by several non-similar stretches. However, the dextran-binding activity of the protein expressed from gbpC in Escherichia coli was not detected in contrast to the other homologue, a protein designated as Dbl, expressing this activity. The gbpC gene was shown to be intact on the chromosome of strain OMZ176, which does not exhibit dextran-dependent aggregation, while the dbl gene of this strain contained a single adenine nucleotide insertion at approximately one-third the distance from the 5'-end. The insertion mutation in the dbl gene resulted in translation of a premature protein missing its LPXTG sequence signature sequence of the wall-anchored proteins. CONCLUSION: These results suggest that the dbl gene is very likely responsible for S. sobrinus dextran-dependent aggregation.

Protective effect against 17beta-estradiol on neuronal apoptosis in hippocampus tissue following transient ischemia/recirculation in mongolian gerbils via down-regulation of tissue transglutaminase activity.

We analyzed the protective effect of 17beta-estradiol (17beta-ED) injection against delayed neuronal death in the hippocampus tissue of the brain in Mongolian gerbils after transient ischemia/recirculation treatment, especially in relation with bcl-2 gene expression and enzymatic activity changes of caspase-3 and tissue transglutaminase (tTGase). Daily intraperitoneal injection of 17beta-ED to the animal after the ischemia stimulated the expression of an apoptosis suppressor gene, bcl-2, in the hippocampal tissue for a week. The gradually increasing apoptotic enzyme activity of caspase-3 and increased number of TUNEL positive fragmented neuronal nuclei caused by ischemic attack in the gerbil brain were clearly suppressed by 17beta-ED administration. The reduced activity and enzyme protein of tTGase, a neurodegenerative marker of apoptosis in the hippocampus after ischemia, were also restored to nearly normal levels by 17beta-ED injection. These results suggest that daily 17beta-ED administration to the gerbil after transient ischemic insult with progressing neuronal deteriorative changes in hippocampus tissue can effectively prevent apoptotic changes through a molecular cascade involving gene expression regulation.

Plasma brain natriuretic peptide concentrations in patients undergoing pulmonary vein isolation.

OBJECTIVE: To examine whether raised plasma brain natriuretic peptide (BNP) concentrations decrease after successful pulmonary vein isolation (PVI) in patients with atrial fibrillation (AF). METHODS: 53 patients (mean age 53 years) with drug-refractory, paroxysmal lone AF underwent segmental ostial PVI. Blood samples were collected before and after PVI. BNP concentrations were determined by immunoassays. RESULTS: Median plasma BNP concentrations were significantly higher in patients with lone AF than in controls (patients with supraventricular tachyarrhythmias, n = 21) (64.6 (71.9) v 13.9 (7.8) pg/ml, p < 0.01). AF recurred in 21 patients after the initial PVI procedure (recurrent AF group), and the others were free from AF without antiarrhythmic drugs (non-recurrent AF group). BNP concentrations were significantly decreased by PVI in the non-recurrent AF group (38.9 (39.1) to 18.3 (16.1) pg/ml, p < 0.01) but not in the recurrent AF group. CONCLUSIONS: Raised plasma BNP concentrations decreased after successful segmental ostial PVI in patients with AF.

Identification of a glucan-binding protein C gene homologue in Streptococcus macacae.

BACKGROUND/AIMS: The past few decades have seen the isolation of certain glucosyltransferases and a number of proteins from mutans streptococci. Some of these proteins have been shown to possess glucan-binding capabilities which confer an important virulence property on mutans streptococci for the role of these bacteria play in dental caries. Among these proteins is glucan-binding protein C, which is encoded by the gbpC gene, and which we have identified as being involved in the dextran-dependent aggregation of Streptococcus mutans. However, gbpC homologues have yet to be identified in other mutans streptococci. METHODS: We carried out polymerase chain reaction amplification of Streptococcus macacae using primers that were designed based on conserved sequences of S. mutans gbpC and identified a gbpC gene homologue. The gene of that homologue was then characterized. RESULTS: Nucleotide sequencing of the S. macacae gbpC homologue revealed a 1854 bp open reading frame encoding a protein with an N-terminal signal peptide. The molecular mass of the processed protein was calculated to be 67 kDa. We also found an LPxTG motif, the consensus sequence for gram-positive cocci cell wall-anchored surface proteins, which was followed by a characteristic sequence at the carboxal terminal region of the putative protein. This suggests that the S. macacae GbpC homologue protein was tethered to the cell wall. CONCLUSION: Based on these results, together with the demonstrated glucan-binding ability of the S. macacae GbpC homologue protein, we suggest that S. macacae cells are capable of binding dextran via the GbpC homologue protein, which is similar to the S. mutans GbpC protein. In addition, Southern hybridization analysis using the S. macacae gbpC homologue as a probe showed a distribution of gbpC homologues throughout the mutans streptococci.

Active membrane transport and receptor proteins from bacteria.

A general strategy for the expression of bacterial membrane transport and receptor genes in Escherichia coli is described. Expression is amplified so that the encoded proteins comprise 5-35% of E. coli inner membrane protein. Depending upon their topology, proteins are produced with RGSH6 or a Strep tag at the C-terminus. These enable purification in mg quantities for crystallization and NMR studies. Examples of one nutrient uptake and one multidrug extrusion protein from Helicobacter pylori are described. This strategy is successful for membrane proteins from H. pylori, E. coli, Enterococcus faecalis, Bacillus subtilis, Staphylococcus aureus, Microbacterium liquefaciens, Brucella abortus, Brucella melitensis, Campylobacter jejuni, Neisseria meningitides, Streptomyces coelicolor and Rhodobacter sphaeroides.

Polymorphism of the polyalanine tract of thyroid transcription factor-2 gene in patients with thyroid dysgenesis.

OBJECTIVE: One of the thyroid-specific transcription factors, thyroid transcription factor-2 (TTF-2), performs a crucial role in the development of the thyroid gland. We performed genetic analysis of the TITF2 gene (encoding TTF-2) in patients with thyroid dysgenesis. METHODS: By direct sequencing of the PCR products of TITF2, we screened the genomic DNA from 46 patients with thyroid dysgenesis (five had agenesis, six had hypoplasia, 15 had ectopy, and 20 were undetermined). We also studied the transcriptional activities of TITF2 by co-expressing the luciferase gene directed by the human thyroglobulin gene promoter. RESULTS: Human TITF2 consists of a forkhead domain, a polyalanine tract, and unique C-terminal residues. In one of the patients with an ectopic sublingual thyroid, we found a polyalanine tract of 11 alanine residues on one chromosome instead of the 14 alanine residues found in normal controls. In one patient with hypoplasia, the polyalanine tract consisted of 12 heterozygous alanine residues. The reduced polyalanine tracts were not detected in 101 normal individuals. However, the expression study showed that the transcriptional activities of TITF2 with reduced polyalanine-tract lengths were equal to that of TITF2 with an unreduced polyalanine tract. CONCLUSION: These results suggest that the polymorphism of the polyalanine tract of TITF2 is not a frequent cause of developmental defects of the human thyroid gland.

Oxidation mechanism of 7-hydroxy-delta 8-tetrahydrocannabinol and 8-hydroxy-delta 9-tetrahydrocannabinol to the corresponding ketones by CYP3A11.

A cDNA isolated from a C57BL/6 mouse liver cDNA library had the identical nucleotide sequence in coding region with the mouse CYP3A11, and the NH(2)-terminal sequence was also identical to that of cytochrome P450 (P450) MDX-B, a microsomal alcohol oxygenase. The COS-7 cells transfected with the CYP3A11 expression vector formed 7-oxo-Delta(8)-tetrahydrocannabinol (7-oxo-Delta(8)-THC) from 7alpha- and 7beta-hydroxy-Delta(8)-THC. An immunologically related protein with P450 MDX-B was expressed in the COS-7 cell microsomes. The cell microsomes expressed CYP3A11; COS-3A11 catalyzed the oxidation of 7-hydroxy-Delta(8)-THC and 8-hydroxy-Delta(9)-THC to 7-oxo-Delta(8)-THC and 8-oxo-Delta(9)-THC, respectively, in a reconstituted system. (18)O derived from atmospheric oxygen was incorporated into about 30% of the corresponding ketones formed from 7alpha-hydroxy-Delta(8)-THC and 8beta-hydroxy-Delta(9)-THC by mouse hepatic microsomes, P450 MDX-B, and COS-3A11, although incorporation of the stable isotope into the oxidized metabolites from 7beta-hydroxy-Delta(8)-THC and 8alpha-hydroxy-Delta(9)-THC was negligible. (18)O, however, was not incorporated into 7-oxo-Delta(8)-THC formed from 7alpha-hydroxy-Delta(8)-THC by using cumene hydroperoxide instead of NADPH under (18)O(2). When (18)O-labeled 7alpha-hydroxy-Delta(8)-THC and 8beta-hydroxy-Delta(9)-THC were incubated with above enzymes under air, about 30% of the ketones formed released (18)O from a hydroxy group at the 7 and 8 positions in the course of the oxidation. These results suggest that 7alpha-hydroxy-Delta(8)-THC and 8beta-hydroxy-Delta(9)-THC may be oxidized to the corresponding ketones by CYP3A11 via a gem-diol pathway. 7beta-Hydroxy-Delta(8)-THC and 8alpha-hydroxy-Delta(9)-THC may be also converted to the ketones through a stereoselective dehydration of an enzyme-bound gem-diol rather than through a direct hydrogen extraction as a peroxy form of the enzyme.

Apoptotic signaling pathway activated by Helicobacter pylori infection and increase of apoptosis-inducing activity under serum-starved conditions.

The enhanced gastric epithelial cell apoptosis observed during infection with Helicobacter pylori has been suggested to be of significance in the etiology of gastritis, peptic ulcers, and neoplasia. To investigate the cell death signaling induced by H. pylori infection, human gastric epithelial cells were incubated with H. pylori for up to 72 h. H. pylori infection induced the activation of caspase -8, -9, and -3 and the expression of the proapoptotic Bcl-2 family proteins Bad and Bid. The peak of the activity of the caspases occurred at 24 h. At this time, the inhibition of caspase-8 or -9 almost completely suppressed H. pylori-induced apoptosis. Inhibition of caspase-8 suppressed the expression of Bad and Bid and the subsequent activation of caspase-9 and -3. These observations indicate that H. pylori induces apoptosis through a pathway involving the sequential induction of apical caspase-8 activity, the proapoptotic proteins Bad and Bid, caspase-9 activity, and effector caspase-3 activity. Activation of the pathway was independent of CagA or vacuolating toxin. A membrane fraction of H. pylori was sufficient to activate this pathway, and treatment with proteinase K eliminated the activity. Apoptotic activity of the membrane fraction was significantly increased by incubating the bacteria under serum-starved conditions for 24 h. These observations suggest that environmental conditions in the human stomach could induce H. pylori-mediated pathogenesis, leading to a variety of clinical outcomes.

Trends in antimicrobial-drug resistance in Japan.

Multidrug resistance in gram-positive bacteria has become common worldwide. In Japan until recently, gram-negative bacteria such as Pseudomonas aeruginosa, Klebsiella pneumoniae, and Serratia marcescens were controlled by carbapenems, fluoroquinolones, and aminoglycosides. However, several of these microorganisms have recently developed resistance against many antimicrobial drugs.

Synthesis and cytokinin activity of fluorescent 7-Phenylethynylimidazo[4,5-b]pyridine and its riboside.

7-Phenylethynylimidazo[4,5-b]pyridine and its riboside have been newly developed as fluorescent carbon-substituted cytokinin analogues. Palladium-catalyzed coupling of 7-iodo-3-(tri-O-acetyl-beta-D-ribofuranosyl)imidazo[4,5-b]pyridine with phenylacetylene followed by ammonolysis afforded the 7-phenylethynyl riboside via its tri-O-acetate. Acid hydrolysis of the riboside provided its free base, which showed a marked enhancement in fluorescence intensity in an aqueous alkaline solution. The free base and its riboside were more active than the corresponding 6-phenylethynylpurine and its riboside, respectively, in Amaranthus betacyanin and tobacco callus bioassays. Surprisingly, the imidazo[4,5-b]pyridine base exhibited strong cytokinin activity comparable to that of N(6)-benzyladenine in the tobacco callus bioassay. This compound would be useful for studying localization and transport of cytokinins in cells or tissues of plants.

A new SHV-derived extended-spectrum beta-lactamase (SHV-24) that hydrolyzes ceftazidime through a single-amino-acid substitution (D179G) in the -loop.

A new SHV-derived extended-spectrum beta-lactamase (SHV-24) conferring high-level resistance to ceftazidime but not cefotaxime and cefazolin was identified in Japan. This enzyme was encoded by a transferable 150-kb plasmid from an Escherichia coli clinical isolate. The pI and K(m) for CAZ of this enzyme were 7.5 and 30 microM, respectively. SHV-24 was found to have a D179G substitution in the Omega-loop of the enzyme.