Tohru Takata

Use of an Open-Reading Frame–Specific Campylobacter jejuni DNA Microarray as a New Genotyping Tool for Studying Epidemiologically Related Isolates

Findings from use of an open-reading frame-specific Campylobacter jejuni DNA microarray to investigate genetic diversity among clinical isolates associated with 5 independent clusters of infection were compared with data from random amplified polymeric DNA (RAPD) and Penner serotyping analyses. The DNA microarray provides a highly specific epidemiological typing tool for analysis of C. jejuni isolates and reveals both divergent and highly conserved gene classes among isolates.

Polymorphisms of Helicobacter pylori HP0638 Reflect Geographic Origin and Correlate with cagA Status

ABSTRACT Since the associations between Helicobacter pylori genotype and disease differ in Asia and the West, we investigated the correlation between HP0638, encoding an outer membrane protein, and potential markers of virulence (cagA, vacA, and iceA). For 109 strains from nine countries, the status of cagA, vacA, and iceA was determined by PCR and/or a line probe assay. We also studied 18 strains from 8 patients (parents and 6 daughters) from a Dutch family and paired strains collected on average 8 years apart from 11 patients. When the HP0638 signal sequences were amplified by PCR and DNA sequence determinations were performed, 89 (96%) of 93 cagA-positive strains had HP0638 in frame, versus none (0%) of 16 cagA-negative strains (P < 0.001). Among strains in which HP0638 was in frame, a six-CT dinucleotide repeat pattern was dominant in Western countries (23 of 33 strains [70%]), while a pattern of three CT repeats with another CT after four T’s (3 + 1-CT-repeat pattern) was dominant in East Asia (31 of 46 strains [67%]); however, specific CT repeat patterns did not correlate with clinical outcome. HP0638 phylogenetic trees also showed geographic characters. The HP0638 frame status and CT dinucleotide repeat patterns were identical for 9 of 11 pairs of strains obtained on average 8 years apart from individuals and the 15 strains obtained from the mother and all six daughters. Thus, HP0638 frame status and cagA status are strongly correlated. The CT dinucleotide repeat pattern in the putative HP0638 signal sequence has geographic characters and appears stable in particular patients and families over a period of years. Analysis of HP0638 CT polymorphisms may serve as a new typing system to discriminate H. pylori isolates for epidemiological purposes.

Helicobacter pylori Does Not Require Lewis X or Lewis Y Expression To Colonize C3H/HeJ mice

ABSTRACT Helicobacter pylori strains frequently express Lewis X (Lex) and/or Ley on their cell surfaces as constituents of the O antigens of their lipopolysaccharide molecules. To assess the effect of Lex and Ley expression on the ability of H. pylori to colonize the mouse stomach and to adhere to epithelial cells, isogenic mutants were created in which fucT1 alone or fucT1 and fucT2, which encode the fucosyl transferases necessary for Lex and Ley expression, were deleted. C3H/HeJ mice were experimentally challenged with either wild-type 26695 H. pylori or its isogenic mutants. All strains, whether passaged in the laboratory or recovered after mouse passage, colonized the mice well and without consistent differences. During colonization by the mutants, there was no reversion to wild type. Similarly, adherence to AGS and KatoIII cells was unaffected by the mutations. Together, these findings indicate that Le expression is not necessary for mouse gastric colonization or for H. pylori adherence to epithelial cells.

Regulation of the HpyII restriction–modification system of Helicobacter pylori by gene deletion and horizontal reconstitution

Helicobacter pylori, Gram‐negative, curved bacteria colonizing the human stomach, possess strain‐specific complements of functional restriction–modification (R–M) systems. Restriction–modification systems have been identified in most bacterial species studied and are believed to have evolved to protect the host genome from invasion by foreign DNA. The large number of R–Ms homologous to those in other bacterial species and their strain‐specificity suggest that H. pylori may have horizontally acquired these genes. A type IIs restriction–modification system, hpyIIRM, was active in two out of the six H. pylori strains studied. We demonstrate now that in most strains lacking M.HpyII function, there is complete absence of the R–M system. Direct DNA repeats of 80 bp flanking the hpyIIRM system allow its deletion, resulting in an ‘empty‐site’ genotype. We show that strains possessing this empty‐site genotype and strains with a full but inactive hpyIIRM can reacquire the hpyIIRM cassette and functional activity through natural transformation by DNA from the parental R–M+ strain. Identical isolates divergent for the presence of an active HpyII R–M pose different restriction barriers to transformation by foreign DNA. That H. pylori can lose HpyII R–M function through deletion or mutation, and can horizontally reacquire the hpyIIRM cassette, is, in composite, a novel mechanism for R–M regulation, supporting the general hypothesis that H. pylori populations use mutation and transformation to regulate gene function.

Antibiotic susceptibility survey of blood-borne MRSA isolates in Japan from 2008 through 2011

We conducted an antibiotic susceptibility survey of 830 blood-borne methicillin resistant Staphylococcus aureus collected from nationwide hospitals in Japan over a three-year period from January 2008 through May 2011. Antibiotic susceptibility was judged according to the criteria recommended by the Clinical Laboratory Standard Institute. Over 99% of the MRSA showed to be susceptible to teicoplanin, linezolid, sulfamethoxazole/trimethoprim and vancomycin, and over 97% of them were susceptible to daptomycin, arbekacin and rifampin. The majority of the MRSA strains showed resistant to minocycline, meropenem, imipenem, clindamycin, ciprofloxacin, cefoxitin, and oxacillin in the rates of 56.6, 72.9, 73.7, 78.7, 89.0, 99.5, and 99.9%, respectively. Among the MRSA strains, 72 showed reduced susceptibility to vancomycin, including 8 strains (0.96%) of vancomycin-intermediate S. aureus (VISA), 54 (6.51%) of heterogeneous vancomycin-intermediate S. aureus (hVISA), and 55 (5.63%) of β-lactam antibiotics-induced vancomycin resistant S. aureus (BIVR). Unexpectedly, among the 54 hVISA and 55 BIVR, 45 isolates (83.3% and 81.8%, respectively) showed both hVISA and BIVR phenotypes. A new trend of vancomycin resistance found in this study was that VISA strains were still prevalent among the bacteremic specimens. The high rates of the hVISA/BIVR two-phenotypic vancomycin resistance, and the prevalence of VISA in the bloodborne MRSA call attention in the MRSA epidemiology in Japan.

A patient with acute myeloid leukemia who developed fatal pneumonia caused by carbapenem-resistant Bacillus cereus

Bacillus cereus is known as a serious bacterial pathogen in neutropenic patients. B. cereus is often resistant to β-lactams, including penicillins and cephalosporins. We report a case of fatal pneumonia caused by B. cereus in a patient with newly diagnosed acute myeloid leukemia (AML) during remission induction therapy. Cefepime was started for febrile neutropenia (FN) initially and was switched to panipem/betamipron, when fulminant pneumonia supervened. The isolated strain was resistant not only to cefepime but also to panipenem/betamipron. This is the first report of fulminant infection caused by carbapenem-resistant B. cereus in a neutropenic patient. B. cereus should be kept in mind as a target of empirical treatment when neutropenic patients develop pneumonia

Presence of both heterogeneous vancomycin-intermediate resistance and β-lactam antibiotic-induced vancomycin resistance phenotypes is associated with the outcome in methicillin-resistant Staphylococcus aureus bloodstream infection

Abstract Background: Although the individual expression of heterogeneous vancomycin-intermediate resistance (hVISA) and β-lactam antibiotic-induced vancomycin resistance (BIVR) phenotypes has been associated with treatment failure and recurrence in methicillin-resistant Staphylococcus aureus (MRSA) infections, the effect of the co-expression of these phenotypic profiles on clinical outcome has not been fully elucidated. The aim of this study was to determine the impact of the combination of hVISA and BIVR phenotypes on the clinical outcome in MRSA bacteremia. Methods: One hundred and sixty-two MRSA blood isolates from a 21-y period, 1987–2007, were randomly selected. Screening for hVISA was done by the macromethod Etest and confirmed by population analysis profiles. BIVR was identified using Mu3 agar containing 4 μg/ml of vancomycin. Results: Thirty (18.5%) and 39 (24.1%) of the 162 MRSA blood isolates were positive for the hVISA and BIVR phenotypes, respectively. Eighteen (11.1%) isolates possessed both hVISA and BIVR phenotypes (hVISA(+)/BIVR(+)). In a subset of patients who received initial treatment with glycopeptides, only the patients whose isolates were hVISA(+)/BIVR(+) displayed a significantly higher mortality rate in comparison to those with non-hVISA(+)/BIVR(+) (80.0% vs 31.3%, p = 0.004). The presence of both hVISA and BIVR phenotypes was a predictor of mortality using a logistic regression analysis (p = 0.025). Conclusions: The combined phenotype of hVISA and BIVR was associated with a higher probability of mortality in patients with MRSA bacteremia. Further prospective studies are warranted to delineate the clinical significance of the combined phenotype of hVISA and BIVR.

Acute hemorrhagic gastropathy with multiple shallow ulcers and duodenitis caused by a laboratory infection of Helicobacter pylori

It is now widely accepted that Helicobacter pylori (H. pylori) is the major cause of chronic active gastritis and an important pathogen of peptic ulcer disease, gastric cancer, lymphoma of mucosaassociated lymphoid tissue (MALT).1-4 It has previously been thought that the acute phase of H. pylori infection usually manifests only trivial symptoms and thus goes unnoticed by the patient.5 Even in the case of accompanying acute gastric illness caused by H. pylori infection owing to either experimental or accidental oral ingestion, the pathologic lesions remained microscopic abnormalities in most cases.6-8 However, acute H. pylori infection occasionally results in gross lesions such as erosive gastritis,9 purulent gastritis,10 gastric ulceration,11 and lesions that mimic either gastric cancer or lymphoma.12,13 We herein report a case of acute hemorrhagic gastropathy with multiple shallow ulcers caused by an accidental infection of H. pylori which occurred in a laboratory without any knowledge of how it was acquired. This case confirms that an H. pylori infection is etiologically important in the pathogenesis of acute hemorrhagic gastropathy and duodenitis.

A simple high‐performance liquid chromatography for the determination of linezolid in human plasma and saliva

Linezolid is an antimicrobial agent for the treatment of multiresistant Gram-positive infections. A practical high-performance liquid chromatography method was developed for the determination of linezolid in human plasma and saliva. Linezolid and an internal standard (o-ethoxybenzamide) were extracted from plasma and saliva with ethyl acetate and analyzed on a Capcell Pak C18 MG column with UV detection at 254 nm. The calibration curve was linear through the range 0.5-50 µg/mL using a 200 μL sample volume. The intra- and interday precisions were all <6.44% for plasma and 5.60% for saliva. The accuracies ranged from 98.8 to 110% for both matrices. The mean recoveries of linezolid were 80.8% for plasma and 79.0% for saliva. This method was used to determine the plasma and saliva concentrations of linezolid in healthy volunteers who were orally administered a 600 mg dose of linezolid. Our liquid-liquid extraction procedure is easy and requires a small volume of plasma or saliva (200 μL). This small volume can be advantageous in clinical pharmacokinetic studies, especially if children participate.

Genome Sequences of Multidrug-Resistant Acinetobacter baumannii Strains from Nosocomial Outbreaks in Japan

ABSTRACT Acinetobacter baumannii has emerged worldwide as an important nosocomial pathogen in medical institutions. Here, we present the draft genome sequences of A. baumannii strains MRY09-0642, MRY10-0558, and MRY12-0277 that were isolated from nosocomial outbreaks in Japan between 2008 and 2012 and that are resistant to antimicrobial agents, including carbapenems, fluoroquinolones, and aminoglycosides.