In vitro exposure to macrolide antibiotics in <Emphasis Type="Italic">Helicobacter pylori</Emphasis> strains isolated from children

Clarithromycin resistance of Helicobacter pylori is a serious problem in eradication therapy. We investigated whether the use of maclorides (clarithromycin, erythromycin, and azithromycin) induces clarithromycin resistance in the organism. Twenty H. pylori strains were isolated from pediatric patients with gastrointestinal symptoms. The minimum inhibitory concentrations (MICs) of each macrolide antibiotic were determined by the Etest. Among these, 17 strains susceptible to macrolide antibiotics were used for the in vitro induction of drug resistance. In each of these 17 strains of H. pylori, 30-day exposure to clarithromycin in experiments for in vitro induction did not change the MIC of any antibiotic, nor did it induce either the A2143G or the A2144G mutation in the 23S rRNA gene. These results suggest that the use of macrolide antibiotics does not induce clarithromycin resistance in H. pylori by 23S rRNA gene mutation.     

Chronic <Emphasis Type="Italic">Helicobacter pylori</Emphasis> infection is associated with peripheral arterial disease

It is reported that Helicobacter pylori infection is associated with coronary atherosclerosis both epidemiologically and pathogenetically, but no conclusions have yet been reached. Therefore, we investigated the relationship between H. pylori infection and peripheral arterial disease (PAD). Sixty-nine patients with PAD attending Harasanshin General Hospital (Fukuoka, Japan) were compared with 143 controls (age-matched asymptomatic outpatients with hyperlipidemia). H. pylori infection was diagnosed by the detection of IgG antibodies, the (13)C-urea breath test, and histological examination. Multiple logistic regression analysis was used to assess the data. The 69 PAD patients and 143 controls were aged from 50 to 92 years. According to the Fontaine classification, 43/69 PAD patients (62.3%) were grade I, 25 (36.2%) were grade II, and 1 (0.14%) was grade III. The prevalence of H. pylori infection was higher in the PAD patients than in the controls (79.7% versus 44.8%; P < 0.01). Stepwise logistic regression analysis revealed that H. pylori infection and hypertension had a significant influence on the occurrence of PAD. Our results suggest that chronic H. pylori infection may be one of the risk factors for PAD.     

<Emphasis Type="Italic">Helicobacter pylori</Emphasis> infection as an environmental risk factor for migraine without aura

Helicobacter pylori (H. pylori) infection has recently been associated with various extraintestinal pathologies and migraine. The aim of this study was to investigate the correlation of the H. pylori infection with the pathogenesis of migraine without aura, especially in cases not affected by endogenous risk factors, like hereditary pattern or hormonal fluctuations. A total of 49 outpatients (37 females and 12 males; age range: 19–47 years; mean age: 31,±14 years) affected by migraine without aura was evaluated. We divided them in 2 subgroups: a) with positive familial history, and/or with menstrual type of migraine b) with negative familial history and with menstrual unrelated type of migraine. H. pylori infection was diagnosed by the 13 C- urea breath test (INFAI — test). Control subjects consisted of 51 patients without any primary headache history (38 females; mean age of 32,±14,4 years; range 21–49 years), who underwent upper gastrointestinal (GI) endoscopy for investigation of anaemia or non ulcer dyspepsia. H. pylori detection was based on the histologic analysis of gastric mucosa biopsy. The prevalence of H. pylori infection was significantly higher in the migraineurs without aura compared to controls (p=0.016). The prevalence of H. pylori infection was significantly high in the mixed and in the female group of our patients without other predisposing factors for migraine without aura (81 and 87% respectively), while in the same groups with predisposing factors (menstruation and/or family history) the prevalence was only 36 and 37% respectively (p=0,001 for the first group and p=0,002 for the second group). Our results seem to highlight the role of H. pylori infection as a probable independent environmental risk factor for migraine without aura, especially in patients that are not genetically or hormonally susceptible to migraine.     

Microbial substitution of <Emphasis Type="Italic">Mycobacterium avium-intracellulare</Emphasis> to <Emphasis Type="Italic">Mycobacterium abscessus</Emphasis> during clinical course

Mycobacterium avium-intracellulare (MAI) is, among acid-fast bacilli, the most common cause of nontuberculous pulmonary diseases, and MAI infections are often treated according to the guidelines of the American Thoracic Society. However, despite the use of multiple drugs, patients sometimes do not recover with the initial round of treatment. Other kinds of nontuberculous mycobacteria are sometimes found in patients' respiratory samples, even during such treatment for MAI pulmonary disease. We experienced three patients with pulmonary disease due to Mycobacterium abscessus (MA) in whom the disease was difficult to treat because of resistance to all the antituberculous agents used. MA infection had occurred in these patients after long-term treatment with multiple drugs for previous MAI pulmonary disease. We considered that the MA infection in these patients appeared to be the result of insufficient efficacy of the drugs used for MAI and insufficiently aggressive use of antituberculous agents. It thus appeared that MA infection was the result of microbial substitution, and that, if this is the case, the guidelines for the treatment of MAI may need to be modified to eliminate microbial substitution.     

First isolation of <Emphasis Type="Italic">bla</Emphasis><Subscript>IMP-7</Subscript> in a <Emphasis Type="Italic">Pseudomonas aeruginosa</Emphasis> in Japan

We report here the first isolation in Japan of a carbapenem-resistant Pseudomonas aeruginosa strain that carries the metallo-β-lactamase gene bla _IMP-7. This isolate revealed high-level resistance to all of the tested antibiotics except for piperacillin, showing a multidrug-resistant phenotype.     

Potent antibacterial activity of Y-754, a novel benzimidazole compound with selective action against <Emphasis Type="Italic">Helicobacter pylori</Emphasis>

Y-754, a novel benzimidazole compound, was investigated for in vitro and in vivo antibacterial activity. Unlike amoxicillin, clarithromycin, and metronidazole, the compound had no activity against common aerobic and anaerobic bacteria other than Helicobacter pylori. The minimum inhibitory concentration of Y-754 against H. pylori, at 0.025µg/ml, was nearly equal to that of amoxicillin and clarithromycin. The respective concentrations of Y-754, amoxicillin, clarithromycin, and metronidazole required to inhibit 90% of 39 isolates of H. pylori were 0.05, 0.39, 6.25, and 25µg/ml, indicating the potent activity of Y-754, including activity against clarithromycin- and metronidazole-resistant strains. The anti-H. pylori activity of Y-754 was potent even at pH 5.5 and was bactericidal at concentrations of 0.1µg/ml and above. Exposure of H. pylori to Y-754 did not result in the induction of drug-resistant mutation. Oral administration (10mg/kg twice a day for 7 days) to Mongolian gerbils infected with strain ATCC 43504 demonstrated that Y-754 was effective in H. pylori eradication and that its eradication efficacy increased in line with the progress of damage to the gastric mucosa caused by H. pylori infection. Y-754 was also efficacious in the treatment of infection by the clarithromycin-resistant strain OIT-36. The results obtained lead to the expectation that the new benzimidazole Y-754 will, in the near future, be used for H. pylori eradication therapy in peptic ulcer patients.     

Prevalence of β-lactamase-nonproducing ampicillin-resistant <Emphasis Type="Italic">Haemophilus influenzae</Emphasis> and <Emphasis Type="Italic">Haemophilus influenzae</Emphasis> type b strains obtained from children with lower respiratory tract infections

The prevalence of strains with ampicillin (ABPC) resistance among Haemophilus influenzae strains isolated from the nasopharynx of children with lower respiratory tract infections has increased significantly during the 6 years from 2000, when it was 41.9%, to 2005, when it reached 60.1%. From 2002, the prevalence exceeded 50%, and the prevalence of β-lactamase-nonproducing ABPC-resistant (BLNAR) strains with a minimum inhibitory concentration (MIC) of ABPC of over 4 μg/ml doubled, from 28.2% in 2002 to 54.7% in 2005. In H. influenzae strains obtained from the nasopharynx of children with lower respiratory tract infections between April 2004 and March 2006, identification of serotype b was defined, using the slide agglutination method. The frequency of isolation of H. influenzae type b (Hib) strains was then measured and the ABPC resistance conditions of the Hib strains were also evaluated. The frequency of the Hib strains was found to be 30 out of 479 strains, 6.3%. Of these 30 strains, BLNAR accounted for 53.3% (16 strains), approximately the same frequency of isolation as that of the BLNAR isolated from all H. influenzae strains during the same period. In Japan, the prevalence of BLNAR strains among clinically isolated H. influenzae strains has continued to increase, and the frequency of isolation of BLNAR strains among Hib strains has also continued to rise. As a countermeasure, attempts at improving resistance have been made through judicious antibiotic use, but concern that the choice of antibiotics for Hib meningitis may become complicated has sparked a keen interest in the introduction of Hib conjugate vaccine. Part of this report was presented at the fifty-fourth general meeting of the Japanese Society of Chemotherapy (Kyoto, May 2006).     

Characterization of oxacillin-susceptible <Emphasis Type="Italic">mecA</Emphasis>-positive <Emphasis Type="Italic">Staphylococcus aureus</Emphasis>: a new type of MRSA

Methicillin-resistant Staphylococcus aureus (MRSA) has been defined as S. aureus having the mecA gene or showing a minimum inhibitory concentration (MIC) of oxacillin higher than 4 mg/l. However, some clinical isolates are mecA-positive and oxacillin-susceptible. Therefore, we surveyed the occurrence of S. aureus having the mecA gene and an MIC of oxacillin of less than 2 mg/l (oxacillin-susceptible MRSA; OS-MRSA) in a total of 480 strains of S. aureus collected from 11 hospitals in different location in Japan isolated from 2003 through 2005. We found 6 strains matching the criteria for OS-MRSA. All 6 strains were staphylococcal cassette chromosome (SCC) mec-positive, without exception, and 4 strains showed the SCCmec type III-variant, which is unique in Japan. These OS-MRSAs were least resistant to oxacillin among the MRSAs tested and they were within the susceptible range to seven other beta-lactam antibiotics tested. Thus, OS-MRSA may become a high-resistant MRSA upon the treatment of patients with beta-lactam antibiotics. To characterize whether these OS-MRSAs were hospital-acquired or community-acquired MRSAs, we tested for the presence of the genes encoding toxins. Genes encoding hemolysin, exfoliative toxin, enterotoxin, toxic shock syndrome toxin-1, and Panton-Valentine leukocidin were found in 6, 4, 0, 0, and 0 strains, respectively. These results revealed that OS-MRSAs could be classified as a new type of MRSA that exhibits properties distinguishable from either hospital- or community-acquired MRSA. Coagulase typing of the OS-MRSAs supported the above conclusion. In this study, the occurrence of OS-MRSA at a certain frequency was noted; precautions are called for in the classification of oxacillin-resistant S. aureus and in the treatment of OS-MRSA infection.     

Dissemination of extended-spectrum β-lactamase-producing <Emphasis Type="Italic">Klebsiella pneumoniae</Emphasis> and <Emphasis Type="Italic">Escherichia coli</Emphasis> in Thai hospitals

Fifty clinical isolates of Klebsiella pneumoniae and Escherichia coli with reduced susceptibility to third-generation cephalosporins, collected from 11 hospitals in Thailand, were studied. All isolates were found to produce extended-spectrum β-lactamase (ESBL), as judged by double-disk synergy and combination disk methods. Most ESBL-producing K. pneumoniae isolates were resistant to ceftazidime (94%) and aztreonam (90%). In contrast, most ESBL-producing E. coli isolates were resistant to ceftriaxone (95%) and cefotaxime (74%). Plasmid DNA was isolated and β-lactamase genes were identified by PCR and sequencing. We found that SHV-12 and CTX-M-14 were the main ESBLs responsible for resistance in K. pneumoniae and E. coli, respectively. SHV-27, SHV-28, and CTX-M-14 were detected in three, two, and four K. pneumoniae isolates, respectively. A high genetic diversity among ESBL-producing K. pneumoniae and E. coli isolates was observed. In addition, the finding of a few isolates that produced identical restriction patterns on pulsed field gel electrophoresis (PFGE) suggests the clonal spread of resistant bacteria within the hospital.     

Influence of daily low-dose 14-membered-ring macrolide therapy on <Emphasis Type="Italic">Helicobacter pylori</Emphasis> infection in patients with chronic inflammatory disease of the airway

Our aim was to evaluate the effects of eradication and the incidence of secondary resistance by long-term low-dose daily 14-membered-ring macrolide therapy on Helicobacter pylori (H. pylori) infection in patients with chronic lower respiratory tract inflammatory disease. In a retrospective analysis, we studied the seroprevalence of H. pylori IgG in 90 patients with inflammation of the lower respiratory tract (68 had been treated with macrolide and 22 served as controls). Then, in a prospective analysis, we evaluated the eradication effect of macrolide therapy by the decline of IgG values and the ¹³C-urea breath test. Only long-term macrolide use significantly affected the seroprevalence of H. pylori IgG. However, macrolide therapy did not reduce the H. pylori IgG values in 24 patients and did not eradicate H. pylori in ¹³C-urea breath tests. Chemosensitivity testing was performed on three H. pylori strains obtained by gastric biopsy from patients in whom the disease could not be eradicated. Only one strain demonstrated a resistant character. Daily long-term low-dose 14-membered-ring macrolide therapy for patients with lower respiratory inflammatory disease may not be sufficient to eradicate H. pylori, but some strains do not acquire a resistant nature.     

Remarkably high prevalence of <Emphasis Type="Italic">fts I</Emphasis> gene mutations in <Emphasis Type="Italic">Haemophilus influenzae</Emphasis> isolates from upper respiratory tract infections in children of the Sapporo district,Japan

Recently, the frequency of isolation of beta-lactamase-negative ampicillin resistant (BLNAR) strains of Haemophilus influenzae in Japanese children has been increasing rapidly. Drug resistance in BLNAR strains is associated with mutations of the fts I gene, which encodes penicillin-binding protein 3. In the otolaryngological field, only a few reports have been available concerning fts I gene mutations in BLNAR. We investigated the prevalence of fts I gene mutations, by polymerase chain reaction (PCR) genotyping, in H. influenzae isolates from the upper respiratory tracts of children in the Sapporo district, Japan. When the isolates were classified according to PCR genotyping, 34 (44.2%) of 77 isolates were beta-lactamase-negative ampicillin-sensitive (g-BLNAS), 8 (10.4%) were g-low-BLNAR, 30 (39.0%) were g-high-BLNAR, 2 (2.6%) were beta-lactamase-positive ampicillin-resistant (g-BLPAR), and 3 (3.9%) were beta-lactamase-positive ampicillin/clavulanic acid-resistant (g-high-BLPACR). Mutations in the fts I gene were generally parallel to ampicillin susceptibility, and were frequently observed in children who were 7 years or younger. Of the beta-lactams tested, cefditoren showed the strongest inhibition of H. influenzae isolates, and it inhibited g-BLNAR and g-BLPACR. This study revealed a remarkably high prevalence of fts I gene mutations (g-BLNAR and g-BLPACR) in our district. Furthermore, a regional difference in the prevalence of fts I gene mutations was observed even at the district level.     

Multifocal osteomyelitis due to <Emphasis Type="Italic">Bartonella henselae</Emphasis> in a child without focal pain

We describe a case of an 11-year-old girl who presented with osteomyelitis of the vertebrae and right femur due to Bartonella henselae. Her only symptom was prolonged fever without focal pain. Magnetic resonance imaging (MRI) and nested polymerase chain reaction (PCR) were useful for the diagnosis. Osteomyelitis due to B. henselae should be considered in cases of prolonged fever of unknown origin.     

Occurrence of vancomycin-intermediate-resistant <Emphasis Type="Italic">Staphylococcus aureus</Emphasis> in Japan

The Clinical and Laboratory Standards Institute (CLSI) amended the criteria for vancomycin susceptibility and resistance of Staphylococcus aureus in 2006. The earlier criteria had established that S. aureus with minimum inhibitory concentrations (MICs) of vancomycin of < or =4 microg/ml, 8 to 16 microg/ml, and > or =32 microg/ml were vancomycin-susceptible, -intermediate-resistant and -resistant, respectively. The revised recommendation states that bacteria showing vancomycin MICs of < or =2 microg/ml, 4 to 8 microg/ml, and > or =16 microg/ml are -susceptible, -intermediate-resistant, and -resistant, respectively. We examined, based on these new criteria, the vancomycin susceptibility of methicillin-resistant S. aureus (MRSA) strains isolated in Japan from 1978 through 2005 at 17 general hospitals. The results showed that, among 2446 MRSA isolates tested, 8 were classified as intermediate-vancomycin-resistant (VISA). Re-examination of vancomycin susceptibility in these 8 strains in 2006 revealed that 6 strains showed a vancomycin MIC of 4 microg/ml, as tested by the agar dilution method, broth dilution methods, and E-test; the 2 other strains had lost the vancomycin resistance. Pulsed-field gel electrophoresis (PFGE) of the chromosomal DNA of these strains exhibited five unique profiles; 2 strains isolated from the same hospital were identical. These results revealed that at least five different types of VISA strains could be identified in Japan so far according to the new CLSI criteria. All these VISA strains had type II staphylococcal cassette chromosome, mec. This study revealed, for the first time in Japan, the presence of intermediate vancomycin-resistant MRSA in this country.     

Splenic abscess in an infant caused by <Emphasis Type="Italic">Streptococcus intermedius</Emphasis>

We report a 20-month-old girl with splenic abscess. The patient was admitted to our hospital because of persistent high fever and abdominal pain. Laboratory data showed leucocytosis and elevated C-reactive protein levels. Abdominal computed tomography showed multiple low-density lesions in the spleen. These findings were consistent with a diagnosis of splenic abscess. She was successfully treated with ultrasonographically guided percutaneous drainage for 11 days and intravenous antibiotic for 17 days. On culture, aspirated fluid from the abscess grew Streptococcus intermedius. This case illustrates that the differential diagnosis of unknown-focus infection in infants should include splenic abscess. We recommend conservative therapy (antibiotics and drainage) as first-line therapy for splenic abscess in pediatric patients, based on the importance of the immunological functions of the spleen.     

Molecular epidemiology of uropathogenic <Emphasis Type="Italic">Escherichia coli</Emphasis>

Escherichia coli is the most common cause of complicated as well as uncomplicated urinary tract infections (UTIs). Most of these uropathogenic E. coli (UPEC) strains exhibit certain virulence factors (VFs), including adhesins, iron uptake systems, synthesis of cytotoxins, and specific O:K:H serotypes. Molecular epidemiological studies of UPEC have contributed to the discovery of uropathogenic VFs, to an understanding of the pathogenesis of UTIs as ascending infections, and to the clarification of genetic linkages between different virulence genes such as pathogenicity islands (PAIs), which are one of the mechanisms for horizontal VF gene transfers between the same or related species. Uropathogenic VFs not only play an important role individually but also work cooperatively in a fine-tuned manner with coordinated regulation and expression.     

Comparison of the efficacy of 250 mg and 500 mg clarithromycin used with lansoprazole and amoxicillin in eradication regimens for<Emphasis Type="Italic">Helicobacter pylori</Emphasis> infection

AIMS AND METHODS: The aim of this study was to compare the efficacy of 250 mg and 500 mg clarithromycin used with lansoprazole and amoxicillin in eradication of H. pylori infection. 235 patients with H. pylori infections and non-ulcer dyspepsia were randomly assigned to one of the following regimens: lansoprazole 30 mg, amoxicillin 1000 mg, clarithromycin 250 mg (LAC250) and lansoprazole 30 mg, amoxicillin 1000 mg, clarithromycin 500 mg (LAC500). All drugs were given twice daily for 7 days. The patients were assessed for prevalence of H. pylori with the CLO test. Gastric biopsy samples obtained during upper gastrointestinal endoscopy before randomization and 4-6 weeks after completion of therapy were used for histology and culture. Bacterial sensitivity to clarithromycin and amoxicillin was determined with the E-test. RESULTS: 101 patients in the LAC250 mg group and 102 in the LAC500 group completed the study. On intention-to-treat analysis, eradication rates were 81% with LAC250 and 82% with LAC500 (p=0.88). On per-protocol analysis, eradication rates were 92% with LAC250 and 96% with LAC500 (p=0.23). Among the 203 patients (86% of the entire study group) for whom H. pylori antibiotic-sensitivity testing was technically feasible, primary resistance to clarithromycin was found in 9% and to amoxicillin in 0%. Eradication of clarithromycin sensitive/resistant strains was 94%/38% for LAC250 (p < 0.001) and 93%/40% for LAC500 (p < 0.001). CONCLUSIONS: The cure rates for the two regimens were similar, although adverse effects were more frequent with the LAC500 regimen, suggesting that 250 mg of clarithromycin b.d. may be sufficient in our patient population.     

Fusion of <Emphasis Type="Italic">Gaussia</Emphasis> Luciferase to an Engineered Anti-carcinoembryonic Antigen (CEA) Antibody for <Emphasis Type="Italic">In Vivo</Emphasis> Optical Imaging

The bioluminescent protein Gaussia luciferase (GLuc) was fused to an anti-carcinoembryonic antigen (CEA) antibody fragment, the diabody, for in vivo optical tumor imaging. A 15-amino acid N-terminal truncation (GLΔ15) resulted in a brighter protein. Fusions of the anti-CEA diabody to full-length GLuc and GLΔ15 retained high affinity for the antigen, emitted light, and exhibited excellent enzymatic stability. In vivo optical imaging of tumor-bearing mice demonstrated specific targeting of diabody-GLΔ15 to CEA-positive xenografts, with a tumor/background ratio of 3.8 ± 0.4 at four hours after tail-vein injection, compared to antigen-negative tumors at 1.3 ± 0.1 (p = 0.001). MicroPET imaging using ¹²⁴I-diabody-GLΔ15 demonstrated specific uptake in the CEA-positive tumor (2.6% ID [injected dose]/g) compared to the CEA-negative tumor (0.4% ID/g) at 21 hours. Although further optimization of this fusion protein may be needed to improve in vivo performance, the diabody-GLΔ15 is a promising optical imaging probe for tumor detection in vivo.     

<Emphasis Type="Italic">Mycobacterium peregrinum</Emphasis>: bactericidal activity of antibiotics alone and in combination

Mycobacterium peregrinum is a rapidly growing mycobacterium that is occasionally associated with disease at different locations. At present, little information is available on antibiotic activity against this microorganism. For this reason, we have carried out a study on the in vitro activity of 15 antibiotics, alone and in combination, against M. peregrinum. Our study shows that the new fluoroquinolones with a C8-methoxy group, especially moxifloxacin, exhibit greater activity against this species. These data should be evaluated in clinical assays or animal models in order to confirm their clinical significance.     

Successfully treated nosocomial <Emphasis Type="Italic">Stenotrophomonas maltophilia</Emphasis> bacteremia following desensitization to trimethoprim-sulfamethoxazole

Stenotrophomonas maltophilia has emerged as an important cause of morbidity and mortality in hospitalized patients. Because trimethoprim-sulfamethoxazole (TMP-SMX) remains the most effective drug for the treatment of S. maltophilia infections, desensitization should be considered in patients with hypersensitivity to TMP-SMX.     

Surgical-site infection with toxin A-nonproducing and toxin B-producing <Emphasis Type="Italic">Clostridium difficile</Emphasis>

To date, few cases of extraintestinal infection with Clostridium difficile have been reported. We describe a case of surgical-site infection with C. difficile following a colonic operation. Administration of metronidazole was considered to be effective for treatment of the infection. The isolate was a toxin A-nonproducing and toxin B-producing strain.