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Background: Rapid identification of causative bacteria in treatment of acute otitis media (AOM) is of paramount importance for appropriate antibiotic use., Materials and Methods: This prospective observational study was conducted in 15 hospitals and clinics in Japan between 2018 and 2020. A new rapid antigen test kit (AOS-116), which simultaneously detects antigens for Streptococcus pneumoniae (Sp) and Haemophilus influenzae (Hi), was applied for middle ear fluids (MEFs) and nasopharyngeal secretions (NPSs) in patients with moderate to severe AOM. We investigated relationship between the results of rapid test, severity at initial visit, and clinical course., Results: Regarding performance accuracy based on culture results, AOS-116 showed 1) high (>80%) sensitivity, specificity, and negative predictive value (NPV) in MEFs for both antigens, 2) high sensitivity, specificity, and positive predictive value (PPV) in NPSs for Hi antigen, and 3) high specificity, and PPV in NPSs for Sp antigen. Regarding predictive value of nasopharyngeal culture and antigen detection for causative middle ear pathogens, similar results were observed between AOS-116 and culture, which was characterized with high sensitivity and NPV for both pathogens. MEFs/NPSs positive for Hi antigen were significantly associated with eardrum findings, and severity. MEFs/NPSs positive for pneumococcal antigen were significantly associated with severity of otalgia, fever, and otorrhea. Among patients with prior antimicrobial treatment, improvement tended to be slower in cases positive for Hi than in cases negative., Conclusion: The rapid antigen detection test is useful as a decision-making tool for prescribing antimicrobial agents and may play an important role in promoting appropriate antimicrobial use., Competing Interests: Declaration of competing interest The authors declare financial support from ASAHI KASEI Corporation. Takuji Miyata and Norikazu Okamura are employees of Asahi Kasei Corporation. The funder had no role in the experimental design, implementation of the study, interpretation of the data, or decision to submit the manuscript for publication. This does not alter the authors’ adherence to any publication policies on sharing data and materials., (Copyright © 2024 Japanese Society of Chemotherapy, Japanese Association for Infectious Diseases, and Japanese Society for Infection Prevention and Control. Published by Elsevier Ltd. All rights reserved.)

Background: The widespread administration of the Haemophilus influenzae type b vaccine has led to the predominance of non-typable H. influenzae (NTHi). However, the occurrence of invasive NTHi infection based on gynecologic diseases is still rare., Case Presentation: A 51-year-old Japanese woman with a history of adenomyoma presented with fever. Blood cultures and a vaginal discharge culture were positive with NTHi. With the high uptake in the uterus with 67 Ga scintigraphy, she was diagnosed with invasive NTHi infection. In addition to antibiotic administrations, a total hysterectomy was performed. The pathological analysis found microabscess formations in adenomyosis., Conclusions: Although NTHi bacteremia consequent to a microabscess in adenomyosis is rare, this case emphasizes the need to consider the uterus as a potential source of infection in patients with underlying gynecological diseases, including an invasive NTHi infection with no known primary focus.

Objectives: Fever of 39°C or higher and a white blood cell (WBC) count of 15,000/μL or greater are known predictors of occult bacteremia (OB). However, because of a decreasing prevalence of OB, WBC counts have become poor predictors of OB in populations of routinely immunized children. Thus, we aim to evaluate the clinical characteristics of OB in Japanese children and identify potential risk factors for OB., Methods: We conducted an observational study of children aged 3 to 36 months old with positive blood cultures for Streptococcus pneumoniae or Haemophilus influenzae at an emergency department in a tertiary care children's hospital between April 2002 and December 2015. Patients with significant underlying diseases, a proven source of infection, or toxic appearance, were excluded., Results: Positive blood cultures were recorded in 231 patients; of these, 110 were included in the study (S. pneumoniae, n = 102; H. influenzae, n = 8). Median age was 16 (3-34) months. Patients had a high median body temperature of 39.2 (interquartile range, 38.6-39.9) °C and median WBC of 21,120 (interquartile range, 16,408-24,242)/μL. A high rate of febrile seizures (58 patients, 53%) was observed, with complex febrile seizures accounting for 43% of the episodes. Frequency of febrile seizures was positively associated with age (P = 0.001)., Conclusions: Our study revealed a high rate of children presenting with febrile seizures, especially complex seizures, among children with OB in Japan. A further study is necessary to evaluate the role of febrile seizures as a predictor for OB.

Introduction. Recently, a Haemophilus influenzae clone with low susceptibility to quinolones emerged in paediatric patients in Japan. Isolates of this clone survived for a long time when exposed to the therapeutic concentration of quinolones, despite being classified as 'susceptible' under the criteria of the Clinical and Laboratory Standards Institute. In the present study, we report the first outbreak of this clone in paediatric patients in 2018. Aim. Our aim was to characterise the first outbreak of an H. influenzae clone with low susceptibility to quinolones. Methodology. All H. influenzae isolates ( n =62), collected at a Japanese teaching hospital in 2018, were characterized by both antimicrobial susceptibility tests and multilocus sequence typing. In addition, the similarity in genetic backgrounds was analysed by PFGE. Results. Among all the isolates ( n =62), quinolone low-susceptible isolates accounted for 19.4 % ( n =12). Seven out of 12 isolates were identified as sequence type 422 (ST422) and showed more than 90 % similarity to each other by PFGE analysis. All ST422 isolates exhibited identical amino acid substitutions in both quinolone resistance-determining regions in GyrA and ParC. In addition, all these isolates were from paediatric patients who had been referred by different primary care clinics and had no relationship to each other. Conclusion. In this study, we describe an outbreak of a quinolone low-susceptible ST422 clone in paediatric patients in Japan. Because ST422 isolates have already been reported in at least five other countries, it has the potential to spread worldwide.

Introduction . Certain nontypeable Haemophilus influenzae cannot be assigned a sequence type (ST) by Multilocus Sequence Typing (MLST) due to the lack of the fucK gene, one of seven MLST loci in H. influenzae , which encodes a fucose-operon enzyme. Aims . To confirm whether the loss of fucK is also found in the encapsulated strains, we analysed clinical isolates of H. influenzae serotype e (Hie). Methodology . We conducted MLST, PFGE, and antimicrobial susceptibility tests of 45 Hie strains; the majority ( n =43) were derived from respiratory samples of pediatric patients at Chiba Children's Hospital between 2000 and 2016. The two remaining strains were obtained from the blood of elderly patients with invasive H. influenzae diseases (IHiDs) between 2015 and 2016 at general hospitals. For the fucK -negative strains, PCR analysis for fucose operon was also performed. Results . Four STs (ST18, 122, 621 and 1758) were assigned to 13 strains, and remaining 32 (including one associated with IHiD) were fucK -negative, completely missing the fucose operon. The allelic profiles of six other loci were identical among 31 strains and to that of ST18, 122 and 621, and these strains were genetically closely related. Forty of 45 isolates were ampicillin-sensitive. Conclusions . The loss of fucK was frequently observed in clinical isolates of Hie from children. Moreover, fucK -negative Hie may be the cause of IHiD in adult patients. The majority of Hie, including fucK -negative strains, were shown to be clonally related and were ampicillin sensitive. This represents the first report examining fucK losses in encapsulated H. influenzae .

Acute otitis media (AOM) occurs commonly in pediatric populations. We examined resistance genotype, antibiotic susceptibility, quinolone (QL) resistance, and multilocus sequence type (MLST) among Haemophilus influenzae isolates causing AOM following introduction of pneumococcal conjugate vaccines in Japan. The AOM surveillance group included 69 participating otolaryngologists. Causative pathogens isolated from middle ear fluid (MEF) samples collected from 582 children with AOM were identified using both bacterial culture and real-time PCR. H. influenzae isolates among these pathogens were characterized by capsular type, resistance genotype, antibiotic susceptibility, QL resistance, and MLST. In 2016, H. influenzae was identified in 319 samples (54.8%), among which 72.4% (n = 231) tested positive by both culture and PCR; remaining H. influenzae cases were only PCR-positive. This proportion of H. influenzae positivity has increased significantly from 41.2% in 2006 (p < 0.001). Among culture-positive strains, genotypic β-lactamase-nonproducing ampicillin (AMP)-resistant (gBLNAR) strains were frequent (63.2%), with β-lactamase-nonproducing AMP-susceptible (gBLNAS) strains accounting for only 24.2%. Susceptibilities of gBLNAR to oral antimicrobials were best for tosufloxacin, followed by cefditoren and tebipenem; MIC 90 s were 0.031 μg/mL, 0.5 μg/mL, and 1 μg/mL, respectively. In 7 gBLNAR isolates (3.0%), QL susceptibility was low, owing to amino acid substitutions in GyrA and/or ParC. Sequence types identified numbered 107, including 28 that were new. Prevention of further increases in resistance to antimicrobial agents will require antibiotic selection based on characterization of causative pathogens in clinical practice., (Copyright © 2019 Japanese Society of Chemotherapy and The Japanese Association for Infectious Diseases. Published by Elsevier Ltd. All rights reserved.)

Background: The use of non-β-lactam agents has increased in Japan due to the prevalence of β-lactam-resistant pathogens. This study aimed to clarify the recent trend of antimicrobial susceptibility and molecular epidemiological features in Haemophilus influenzae., Methods: Fifty-seven Haemophilus influenzae isolated from a Japanese teaching hospital in 2017 were characterised, and the data were compared with those of a previous study. The MICs were determined using the broth dilution method. Genetic backgrounds were compared by multilocus sequence typing. The bactericidal activity of tosufloxacin at, or near, the therapeutic Cmax was determined in vitro, with susceptible isolates and quinolone low-susceptible isolates by time-kill assay., Results: The results of the susceptibility tests showed that >90% of isolates were susceptible to cephalosporins and carbapenems, whereas ampicillin-susceptible and clarithromycin-susceptible isolates decreased. Regarding quinolones, low-susceptible isolates were noted in 2017, although all isolates were judged as susceptible. All low-susceptible isolates had an amino acid substitution in GyrA, and two isolates had an additional substitution in ParC. These isolates had different genetic backgrounds. Furthermore, the time-kill kinetic assay using the Cmax of tosufloxacin indicated that the low-susceptible isolates could persist for at least 8hours., Conclusions: This study revealed that Haemophilus influenzae has demonstrated multidrug low-susceptibility in recent years. The low-susceptible isolates had genetic diversity, meaning that resistance occurred independently., (Copyright © 2019 International Society for Chemotherapy of Infection and Cancer. Published by Elsevier Ltd. All rights reserved.)

Purpose: Haemophilus influenzae strains with low susceptibility to quinolones have recently emerged in the paediatric field in Japan. These strains are judged as 'susceptible' in routine susceptibility tests, although they may survive after quinolone treatment. Therefore, we aimed to construct a simple and cost-effective identification method for low-susceptibility strains using disc diffusion assays., Methodology: A total of 33 H. influenzae clinical isolates and a control strain were used. For the disc diffusion assay, levofloxacin, norfloxacin, nalidixic acid and pipemidic acid were employed. Correlations between the inhibition zone diameter and amino acid substitutions were evaluated., Results: All of the tested strains formed clear inhibition zones on both levofloxacin and norfloxacin discs. By contrast, none of the low-susceptibility strains showed inhibition zones against nalidixic acid, while the low-susceptibility strains with amino acid substitutions in both GyrA and ParC did not show inhibition zones against pipemidic acid discs, indicating that low-susceptibility strains can be detected with high sensitivity and specificity by the presence or absence of inhibition zones for earlier quinolones., Conclusion: A disc diffusion test combining results from nalidixic acid and pipemidic acid can detect low-susceptibility strains harbouring amino acid substitutions without the need for genetic analysis. This test can help reduce inappropriate and unnecessary fluoroquinolone use.

We report the case of a 36-year-old previously healthy woman who presented with fever and headache. Blood and cerebrospinal cultures and a bacterial analysis revealed the presence of β-lactamase non-producing ampicillin-resistant Haemophilus influenzae type f (Hif) with sequence type 124. Accordingly, the patient was diagnosed with bacterial meningitis with bacteremia caused by Hif. She had normal humoral immunity, and antibiotic therapy rapidly improved her condition. Our case indicates that serotype replacement can occur in Japan and suggests that a certain sequence type causes invasive Haemophilus influenzae disease, regardless of host immunity.

β-Lactam-resistant Haemophilus influenzae is a clinical concern. A high prevalence (>40%) of β-lactamase-negative high-level ampicillin-resistant H. influenzae (high-BLNAR) isolates in Japan has been reported. However, the reasons for the expansion are unknown. High-BLNAR strains possess an amino acid substitution, either Asn526Lys (group III) or Arg517His (group III-like) in addition to Ser385Thr, in penicillin-binding protein 3 (PBP3). To determine the current prevalence of high-BLNAR strains and the mechanisms behind their expansion in Japan, their prevalence, PBP3 types, multilocus sequence types, and susceptibilities to quinolones approved in Japan as alternatives were determined. Sixty percent of H. influenzae clinical isolates (62/104 isolates) were β-lactamase-negative ampicillin-resistant H. influenzae (BLNAR) strains. Among BLNAR isolates, 92% (57/62 isolates) were high-BLNAR strains. Most isolates were classified as belonging to group III, which contained many genotypes (11 PBP3 types and 25 sequence types). These results indicated that the expansion of high-BLNAR isolates was multiclonal and such strains are still predominant in Japanese clinical settings. One high-BLNAR isolate harbored the novel amino acid substitution Asn526Met in addition to Ser385Thr in PBP3, suggesting a new group (group IV). No quinolone-resistant H. influenzae isolates were identified. The MICs for the quinolones (moxifloxacin, garenoxacin, and tosufloxacin) were similar to that for levofloxacin, whereas sitafloxacin exhibited a lower MIC. However, we obtained 4 H. influenzae isolates with decreased quinolone susceptibility with the amino acid substitution Ser84Leu in GyrA, and 3 of those isolates were high-BLNAR isolates. In summary, this study shows that multiclonal high-BLNAR strains predominate in a Japanese university hospital. Isolates remain sensitive to quinolones, but vigilance is required to prevent the development of fluoroquinolone resistance in high-BLNAR strains., (Copyright © 2018 American Society for Microbiology.)

OBJECTIVETo describe the epidemiologic features of an outbreak of an acute respiratory tract infection (ARI) caused by β-lactamase-negative ampicillin-resistant (BLNAR) nontypeable Haemophilus influenzae (NTHi) in an acute-care ward.DESIGNCross-sectional case-control study.SETTINGAn acute-care ward (ward A) in a general hospital of Kochi in western Japan.METHODSPatients who shared a room with an index patient and all staff in ward A were screened and followed from July 1 to August 31, 2015. Sputum or throat swab samples were collected from participants and tested by culture and polymerase chain reaction (PCR). The association between detected pathogens and ARI development among all participants was examined. A case-control study was conducted to identify risk factors for disease.RESULTSIn total, 78 participants, including the index patient, were enrolled. Of all participants, 27 (34.6%) developed mild respiratory symptoms during a 3-week period: 24 were diagnosed as upper respiratory tract infections, and 3 were diagnosed as lower respiratory tract infections. The presence of BLNAR NTHi was confirmed in 13 participants, and multilocus sequence typing demonstrated that these isolates belonged to sequence type 159. All isolates showed identical pulsed-field gel electrophoresis patterns. The presence of BLNAR NTHi was strongly associated with ARI development, whereas viruses were not associated with the disease. Multivariate analyses demonstrated that a history of contact with the index patient was independently associated with ARI caused by BLNAR NTHi.CONCLUSIONSBLNAR NTHi has the potential to cause upper respiratory tract infections among adults and to spread rapidly in hospital settings.Infect Control Hosp Epidemiol 2018;39:652-659.

Background: Acute otitis media is a leading cause of childhood morbidity and antibiotic prescriptions. We examined etiologic changes in acute otitis media after introduction of 13-valent pneumococcal conjugate vaccine as routine immunization for Japanese children in 2014. Serotypes, resistance genotypes, antibiotic susceptibilities and multilocus sequence typing of pneumococcal isolates were also characterized., Methods: Otolaryngologists prospectively collected middle ear fluid from 582 children by tympanocentesis or sampling through a spontaneously ruptured tympanic membrane between June 2016 and January 2017. Causative pathogens were identified by bacterial culture and real-time polymerase chain reaction for bacteria. Serotypes, resistance genotypes, sequence types and susceptibilities to 14 antimicrobial agents were determined for pneumococcal isolates., Results: At least 1 bacterial pathogen was identified in 473 of the samples (81.3%). Nontypeable Haemophilus influenzae (54.8%) was detected most frequently, followed by Streptococcus pneumoniae (25.4%), Streptococcus pyogenes (2.9%) and others. Pneumococci of current vaccine serotypes have decreased dramatically from 82.1% in 2006 to 18.5% (P < 0.001). Commonest serotypes were 15A (14.8%), 3 (13.9%) and 35B (11.1%). Serotype 3 was significantly less frequent among children receiving 13-valent pneumococcal conjugate vaccine compared with 7-valent pneumococcal conjugate vaccine (P = 0.002). Genotypic penicillin-resistant S. pneumoniae accounted for 28.7%, slightly less than in 2006 (34.2%; P = 0.393); the penicillin-resistant serotypes 15A and 35B had increased. Serotypes 15A, 3 and 35B most often belonged to sequence types 63, 180 and 558., Conclusions: Our findings are expected to assist in development of future vaccines, and they underscore the need for appropriate clinical choice of oral agents based on testing of causative pathogens.

We determined minimum inhibitory concentrations (MICs) of 41 antimicrobial agents for 73 clinical strains of Haemophilus influenzae isolated from the urethra of men with acute urethritis and/or epididymitis and examined the strains for the production of β-lactamase. We also compared their antimicrobial susceptibilities with those of H. influenzae strains from respiratory tract or otorhinolaryngological infections that were reported in Japan. The proportion of β-lactamase-nonproducing ampicillin-resistant strains from acute urethritis and/or epididymitis appeared to be lower, but that of β-lactamase-producing ampicillin-resistant strains appeared to be higher, compared with those from respiratory tract or otorhinolaryngological infections. However, their antimicrobial susceptibilities to a variety of other antimicrobial agents would be similar to those from respiratory tract or otorhinolaryngological infections. Almost all of the strains of H. influenzae from acute urethritis and/or epididymitis were susceptible to the agents, including ceftriaxone, quinolones, macrolides, and tetracyclines, commonly prescribed for treatment of acute urethritis based on the MIC breakpoints recommended by the Clinical and Laboratory Standards Institute. Ceftriaxone and quinolones could be effective on H. influenzae-induced urethritis. However, azithromycin treatment failures were reported in acute urethritis caused by H. influenzae strains considered susceptible to azithromycin. Further studies will be needed to determine MIC breakpoints of antimicrobial agents, which are recommended for treatment of urogenital infections, for H. influenzae strains causing these infections. Nevertheless, this study provides useful data regarding antimicrobial susceptibilities of H. influenzae strains isolated from the urogenital tract, which have rarely been studied., (Copyright © 2017 Japanese Society of Chemotherapy and The Japanese Association for Infectious Diseases. Published by Elsevier Ltd. All rights reserved.)

The nationwide surveillance on antimicrobial susceptibility of bacterial respiratory pathogens from the patients in Japan was conducted by Japanese Society of Chemotherapy, Japanese association for infectious diseases and Japanese society for Clinical Microbiology in 2012. The isolates were collected from clinical specimens obtained from well-diagnosed adult patients with respiratory tract infections during the period between January and December in 2012 by three societies. Antimicrobial susceptibility testing was conducted at the central reference laboratory according to the method recommended by Clinical Laboratory Standard Institutes. Susceptibility testing was evaluated in 1236 strains (232 Staphylococcus aureus, 225 Streptococcus pneumoniae, 16 Streptococcus pyogenes, 231 Haemophilus influenzae, 147 Moraxella catarrhalis, 167 Klebsiella pneumoniae and 218 Pseudomonas aeruginosa). Ratio of methicillin-resistant S. aureus was 51.3%, and those of penicillin-intermediate S. pneumoniae was 0.4%. Among H. influenzae, 5.6% of them were found to be β-lactamase-producing ampicillin-resistant strains, and 37.2% to be β-lactamase-non-producing ampicillin-resistant strains. Extended spectrum β-lactamase-producing K. pneumoniae and multi-drug resistant P. aeruginosa with metallo β-lactamase were 4.2% and 3.2%, respectively. Continuous national surveillance is important to determine the actual situation of the resistance shown by bacterial respiratory pathogens to antimicrobial agents., (Copyright © 2017 Japanese Society of Chemotherapy and The Japanese Association for Infectious Diseases. Published by Elsevier Ltd. All rights reserved.)

This study aimed to identify trends in frequency, serotype, and antimicrobial susceptibility of Streptococcus pneumoniae and Haemophilus influenzae isolated from middle ear fluid specimens of children aged≤15 years (mean, 2 years), before and after the introduction of the 7-valent pneumococcal conjugate vaccine (PCV7) and the H. influenzae type b vaccine, at a pediatric facility in Japan. Sixty-six S. pneumoniae and 88 H. influenzae strains were isolated from 820 middle ear fluid samples. Serotyping and antimicrobial susceptibility testing were performed. The study time-frame was divided into period 1 (2007-2010) and period 2 (2011-2014), according to the availability of vaccine public funding. The S. pneumoniae detection rate decreased from 9.6% in period 1-6.1% in period 2 (p = 0.042). PCV7 serotypes decreased from 56.8% to 9.1% (p = 0.0002). No significant change was observed for the 13-valent pneumococcal conjugate vaccine (PCV13) serotypes: 72.7% in period 1 and 59.1% in period 2. Penicillin-resistant strains (penicillin G-MIC ≥2 μg/mL) decreased from 25% to 4.5% (p = 0.038). Detection rates for H. influenzae did not change significantly: 10.3% in period 1 and 11.3% in period 2. Serotypes were mostly non-typeable: 97.9% in period 1 and 90.2% in period 2, and only one serotype b strain was isolated in each period. The frequency of ampicillin-resistant strains (MIC ≥4 μg/mL) did not change. These results show a preventative effect of PCV7 on otitis media due to S. pneumoniae. PCV7 was replaced with PCV13 in 2013 in Japan; therefore, a further decrease in pneumococcal otitis media is anticipated in the future., (Copyright © 2016 Japanese Society of Chemotherapy and The Japanese Association for Infectious Diseases. Published by Elsevier Ltd. All rights reserved.)

β-Lactamase-negative ampicillin-resistant (BLNAR) Haemophilus influenzae account for a large portion of H. influenzae clinical isolates in Japan. The aim of this study was to clarify the antimicrobial susceptibility of BLNAR H. influenzae clinical isolates as well as the annual changes in susceptibility. BLNAR H. influenzae isolates were collected from a tertiary care hospital from 2007 to 2012. Antimicrobial susceptibility testing was performed and resistance mechanisms were analysed. All of the isolates (n=304) had amino acid substitutions in penicillin-binding protein 3 (PBP3) and isolates were classified by these amino acid substitutions: R517H or N526K (class I); S385T and R517H (class II); and S385T and N526K (class III). Classes I, II and III represented 8.2% (n=25), 9.5% (n=29) and 81.6% (n=248) of the isolates, respectively; 2 isolates could not be classified because they had a PBP3 with a substantially mutated FtsI transpeptidase domain. All of the isolates were highly susceptible to fluoroquinolones and carbapenems. The number of clarithromycin (CAM)-non-susceptible [minimum inhibitory concentration (MIC) ≥16μg/mL] H. influenzae isolates increased significantly between 2010 and 2012. Moreover, CAM-non-susceptible H. influenzae isolates were prevalent among class II and class III BLNAR H. influenzae. Multilocus sequence typing (MLST) of the CAM-resistant (MIC ≥32μg/mL) H. influenzae isolates showed that they were not specific sequence types, suggesting that CAM resistance may occur in any isolates. These results raise concern regarding the occurrence of multidrug-resistant BLNAR H. influenzae., (Copyright © 2016 International Society for Chemotherapy of Infection and Cancer. Published by Elsevier Ltd. All rights reserved.)

Background: Nontypeable Haemophilus influenzae is a particularly important cause of acute otitis media (AOM). There is a high prevalence of β-lactamase-nonproducing ampicillin-resistant (BLNAR) strains in Japanese children, which is associated with recurrent AOM and prolonged treatment. The aim of this study was to investigate the antimicrobial susceptibility profile, mechanisms of ampicillin resistance and molecular epidemiology of ampicillin resistance in H. influenzae strains causing AOM in Japanese children., Methods: One hundred fifty-seven strains of H. influenzae isolated from the middle ear fluid of pediatric patients (aged 0-3 years) with AOM from various areas of Japan were studied. The antimicrobial susceptibility profile, genes encoding β-lactamase and alterations of penicillin-binding protein 3 were investigated. Genetic relatedness among ampicillin-resistant isolates was examined by multilocus sequence typing and pulsed-field gel electrophoresis., Results: Of 157 isolates, 108 (68.8%) demonstrated reduced susceptibility to ampicillin, including 95 (60.5%) of β-lactamase-nonproducing isolates and 13 (8.3%) of β-lactamase-producing isolates. All BLNAR (minimum inhibitory concentration of ampicillin ≥ 4 mg/L) isolates had amino acid substitutions related to ampicillin resistance. Multilocus sequence typing and pulsed-field gel electrophoresis demonstrated genetic diversity although there were 2 clusters of highly resistant isolates with identical STs (sequence types; ST161 and 549)., Conclusions: Alterations of penicillin-binding protein 3 represented the most prevalent mechanism of ampicillin resistance among H. influenzae isolates causing AOM in Japanese children. BLNAR isolates from children with AOM demonstrated genetic diversity. This study identified for the first time ST clones associated with BLNAR H. influenzae causing AOM in Japanese children.

Setting: Community-acquired pneumonia (CAP) is a common cause of morbidity and mortality worldwide. While interactions among bacterial pathogens in the respiratory tract have been studied, the impact of polybacterial aetiology on mortality has not been fully elucidated., Objective: To investigate the impact of polybacterial aetiology on mortality, prevalence, microbial patterns and clinical characteristics among CAP patients., Design: Retrospective data analysis., Results: Bacterial aetiology was established in 711 (46.0%) of 1544 enrolled hospitalised CAP patients. Of these, polybacterial aetiology was identified in 89 (12.5%): Streptococcus pneumoniae was the most frequently identified pathogen (n = 55, 61.8%). The most prevalent microbial combination was S. pneumoniae and Haemophilus influenzae (n = 19, 21.3%). Alcoholism and lack of previous antibiotic therapy were independent predictors of polybacterial aetiology. The proportion of patients with severe pneumonia was significantly higher in the polybacterial aetiology group than in the monobacterial group. Multivariate analysis showed that polybacterial aetiology was a predictor of 30-day mortality (OR 2.14, 95%CI 1.07-4.24, P = 0.030), independently of severe pneumonia status, pneumonia severity index, chronic obstructive pulmonary disease and inappropriate empirical therapy., Conclusion: Polybacterial CAP tends to advance in severity, and indicates adverse outcomes.

In Japan, publicly subsidized Haemophilus influenzae serotype b vaccines became available in 2011; consequently, the incidence of invasive H. influenzae infection in paediatric patients of less than 5 years of age decreased dramatically. In 2013, the first case of H. influenzae serotype f (Hif) meningitis in a Japanese infant was reported, and another case of Hif meningitis in a Japanese infant was observed in 2013. We experienced a fatal paediatric case of Hif bacteraemia in 2004; therefore, we conducted an analysis of the three Hif strains isolated from these three Japanese children with invasive Hif infections. All three strains were β-lactamase-non-producing, ampicillin-sensitive strains, with MICs of 1 µg ml(-1) or less. However, one of the three strains showed slightly elevated MICs for ampicillin (1 µg ml(-1)), cefotaxime (0.25 µg ml(-1)) and meropenem (0.13 µg ml(-1)). A molecular analysis by multilocus sequence typing identified all three strains as sequence type (ST) 124, which is a predominant invasive Hif strain in many countries. SmaI-digested PFGE showed variable DNA fragmentation patterns among the strains, suggesting that some highly virulent strains have originated from a single ST124 clone and caused invasive Hif infections in Japan. Additional studies are needed to determine the factors that have led to the clonal expansion of virulent ST124 strains., (© 2015 The Authors.)

We performed a postmarketing surveillance study to determine the efficacy and safety of the oral quinolone antibacterial agent, garenoxacin (Geninax(®) Tablets 200 mg), against bacterial pneumonia. Between October 2009 and March 2011, patients with community-acquired pneumonia visited 174 facilities in Japan; we collected survey forms from 739 patients of these patients who were suspected with bacterial pneumonia on the basis of factors, e.g., the presence of purulent sputum or suspected presence of bacterial pathogens in clinical specimens. We examined the safety in 730 patients and the efficacy in 535 patients. The efficacy rate of garenoxacin for bacterial pneumonia was 92.8% (479/516 patients). The eradication rates for Streptococcus pneumoniae and Haemophilus influenzae, the major pathogens of bacterial pneumonia, were 98.5% (65/66 strains) and 100% (65/65 strains), respectively. The incidence of adverse drug reactions (including abnormal laboratory tests) was 7.9% (58/730 patients). Among the main adverse drug reactions, abnormal laboratory tests were observed in 2.1% patients (15/730), hepatobiliary disorders were observed in 1.8% patients (13/730), and skin and subcutaneous tissue disorders were observed in 1.6% patients (12/730). In conclusion, garenoxacin showed an efficacy rate of greater than 90% for bacterial pneumonia and is considered to be useful in daily practice., (Copyright © 2014 Japanese Society of Chemotherapy and The Japanese Association for Infectious Diseases. Published by Elsevier Ltd. All rights reserved.)

The 7-valent pneumococcal conjugate vaccine (PCV7) and Haemophilus influenzae type b (Hib) vaccine reduce nasopharyngeal carriage of vaccine-type bacteria, which may in turn influence the presence of other nasopharyngeal bacterial pathogens. To investigate this possibility, nasopharyngeal carriage of potential pathogens was examined before and after official financial support was provided to offer the PCV7 and Hib vaccines in healthy children attending a day care centre in Japan during 2011-2012. Despite a virtual disappearance of PCV7 serotypes over time, the overall pneumococcal carriage rate remained unchanged. Although others have reported an increase in PCV13 serotypes following PCV7 vaccination, only non-PCV13 serotypes were observed to have increased in this study. The majority of H. influenzae isolates were non-typeable and Hib was not found. Our data identified an unexpected pattern of pneumococcal serotype replacement following PCV7. Continuous monitoring of pneumococcal carriage is important for decisions regarding the future of national vaccination policy in Japan., (Copyright © 2013 Japanese Society of Chemotherapy and the Japanese Association for Infectious Diseases. Published by Elsevier Ltd. All rights reserved.)

Before the introduction of vaccines, the incidence of bacterial meningitis among children aged 28 days to 5 years was 8.48, Haemophilus influenzae type-b meningitis was 5.65 and Streptococcus pneumoniae meningitis was 1.85 per 100,000 person-years in Hokkaido, Japan. The incidence of bacteremia caused by S. pneumoniae was 60.15 and H. influenzae was 18.80.

We examined the antimicrobial susceptibility of 1,208 Haemophilus influenzae isolates obtained at a pediatric facility between 2009 and 2012. The percentage distribution of beta-lactamase-non-producing ampicillin (ABPC)-sensitive (BLNAS) strains was 38.2%, that of beta-lactamase-non-producing ABPC-intermediately resistant (BLNAI) strains was 13.9%, that of beta-lactamase-non-producing ABPC-resistant (BLNAR) strains was 38.2%, that of beta-lactamase-producing ABPC-resistant (BLPAR) strains was 5.2%, and that of beta-lactamase-producing clavulanic acid/amoxicillin-resistant (BLPACR) strains was 4.5%. Although the percentage of BLNAR strains increased dramatically from 13.9% (2000-2003; period I) to 32.7% (2004-2008; period II), it increased more slowly from period II to the present period (2009-2012). However, the percentage of BLNAI strains, which had decreased from 10.6% (period I) to 8.9% (period II), began to increase during the present period. Tosufloxacin (< or = 0.06 microg/mL) and tazobactam/piperacillin (< or = 0.13 microg/mL) exhibited a low 90% minimum inhibitory concentration for H. influenzae, as well as for BLNAR strains. A decreased susceptibility to cephems was also observed throughout all 3 periods. Serotype b strains (Hib) were observed in 54 of the 1,208 isolates (4.5%); their distribution decreased since period II (6.3%). In Hib, the percentage distribution of strains from patients less than 2 years of age, who are recommended to be vaccinated against Hib, decreased from 56.8% to 29.6%, and this reduction seems to have been achieved by increasing the percentage of Hib vaccine inoculations in Japan. Non-serotype b-capsulated strains were identified in 21 isolates (1.7%), and 11 of them were serotyped as type e, whereas the other 10 were serotyped as type f.

Background: Acute otitis media (AOM) is one of the most common forms of bacterial infection and cause for clinic visits in children. The incidence of AOM was 0.9-1.2 episodes per person-year during the first 2 years of life in previous reports conducted before 2000. The aim of this study was to 1) evaluate the latest AOM incidence in pediatric outpatients and 2) identify the bacterial pathogens from these patients and ascertain their serotypes and resistance., Methods: The study was conducted in a closed population, involving all pediatricians and otolaryngologists in Sado Island allowing accurate determination of AOM incidence. In each month, one week was assigned as "surveillance week", and all outpatients with acute illness aged 0-18 years examined during the surveillance weeks were enrolled. AOM was diagnosed on the basis of otoscopic findings and clinical symptoms were recorded. Specimens were collected from the nasopharynx or middle ear cavity of AOM patients and examined for bacteria. Antimicrobial susceptibilities, serotypes, and molecular typing for resistance were determined among Streptococcus pneumoniae and Haemophilus influenzae., Results: In total, 8,283 clinic visits were conducted, and 354 episodes (4.3%, 95% CI: 3.9-4.7%) among 312 children were diagnosed as AOM. The incidence of AOM was highest in children of 1 year of age (0.54 episodes/child/year, 95% CI: 0.44-0.64). Serotype coverage of 7- and 13-valent pneumococcal conjugate vaccines in this study were 38.0% (95% CI: 29.3-47.3) and 62.8% (95% CI: 53.6-71.4), respectively. Of 122 H.influenzae isolates available for typing, 120 were nontypeable and 2 were type b. A high proportion of S. pneumoniae isolates (46%) showed resistance to penicillin. Approximately half of H. influenzae isolates had genetic markers for beta-lactamase-negative ampicillin-resistance., Conclusions: Approximately 4-5% of pediatric outpatients, even without AOM-related symptoms, had AOM in our study. Pediatricians as well as otolaryngologists should check the tympanic membrane findings of all pediatric outpatients.

Background: The first step in a bacterial disease is the establishment of nasopharyngeal carriage., Methods: We conducted a birth cohort study to identify factors associated with colonization in healthy children and evaluate the serotype distributions and resistances of Streptococcus pneumoniae/Haemophilus influenzae. Nasopharyngeal cultures were obtained from 349 subjects at 5 time points coinciding with health checkups (4, 7, 10, 18 and 36 months)., Results: A total of 551 S. pneumoniae (penicillin resistance rate: 46.3%) and 301 H. influenzae (ampicillin resistance rate: 44.5%) isolates were obtained from 1654 samples. In this study, 47.5% and 60.9% of S. pneumoniae isolates were included in the serotypes of 7- and 13-valent pneumococcal conjugate vaccines, respectively. Analyzing by Cox proportional hazards models, cohabiting older sibling(s) attending day-care (hazard ratios: 2.064-3.518, P < 0.001) and an early start of day-care attendance by the subjects themselves (2.259-2.439, P < 0.001) were associated with a higher risk of early colonization regardless of their susceptibility. Recent exposure to antimicrobials was also significantly associated with increased risk of colonization (odds ratios: 2.032-2.999, P < 0.001) but not with resistance rates. This data indicated that introduction of appropriate antimicrobial usage in areas of overuse of antimicrobials could contribute to lower colonization of S. pneumoniae/H. influenzae, resulting in a decrease in the absolute number of resistant isolates., Conclusions: Strategies to control transmission at day-care centers or from older sibling(s) as well as appropriate use of antimicrobials are essential for reducing colonization and the absolute number of resistant isolates.

The Drug-Resistant Pathogen Surveillance Group in Pediatric Infectious Disease conducted national surveillance for Haemophilus influenzae in 2007 (phase 3) and 2010 (phase 4), following the previous surveillance conducted from 2000 to 2001 (phase 1) and in 2004 (phase 2). We examined the antimicrobial susceptibility for H. influenzae derived from clinical specimens of pediatric patients collected nationwide from 27 institutions during phases 3 (386 strains) and 4 (484 strains). The frequency of β-lactamase-nonproducing ampicillin (ABPC)-resistant (BLNAR) strains, which rapidly increased from 11.4 % in phase 1 to 43.4 % in phase 2, has gradually decreased from 38.3 % in phase 3 to 37.8 % in phase 4. In contrast, On the other hand, the frequency of β-lactamase-producing strains, which continuously decreased from 8.3 % in phase 1 to 4.4 % in phase 3, has increased to 8.7 % in phase 4. Prevalence of β-lactamase-producing clavulanic acid/amoxicillin-resistant (BLPACR) strains, especially, has increased from 1.6 % in phase 3 to 4.8 % in phase 4. The oral antimicrobial agents with the lowest MIC90 were levofloxacin in both phases, and tosufloxacin in phase 4 (≤0.063 μg/ml), whereas for intravenous use the corresponding agent was tazobactam/piperacillin in both phases (0.125 μg/ml). There was no increase in the MIC90 of most β-lactams between phase 3 and phase 4. In relationship to sex, age, presence of siblings, attendance at a daycare center, siblings' attendance at a daycare center, and prior administration of antimicrobial agents within 1 month, the frequency of β-lactamase-nonproducing ABPC-intermediately resistant (BLNAI) strains + BLNAR strains was high (P = 0.005) in cases with prior administration of antimicrobial agents in phase 3.

We investigated the susceptibility to antibacterial agents of 197 strains of Haemophilus influenzae isolated from pediatric patients at medical facilities in Gifu and Aichi prefectures between 2009 and 2010. Those strains were also examined for the mutations of ftsI coding for penicillin-binding protein 3, presence of bla TEM-1, serotype and beta-lactamase producing ability. Among the 197 strains, the most prevalent serotype was non-typeable (89.8%), followed by serotype b (8.1%), e (1.5%) and f (0.5%). Based on the susceptibility among the 197 strains to antibacterial agents, beta-lactamase nonproducing ampicillin-susceptible H. influenzae (BLNAS) accounted for 27.4%, beta-lactamase nonproducing ampicillin-resistant H. influenzae (BLNAR) for 62.4%, beta-lactamase producing ampicillin-resistant H. influenzae (BLPAR) for 6.1% and beta-lactamase producing amoxicillin/ clavulanic acid-resistant H. influenzae (BLPACR) for 4.1%. According to PCR-based genotyping, the strains were classified into 6 categories: gBLNAS, gLow-BLNAR, gBLNAR, gBLPAR, gBLPACR-I and gBLPACR-II. The incidences of each resistant class were 17.3% for gBLNAS, 6.6% for gLow-BLNAR, 66.0% for gBLNAR, 5.6% for gBLPAR and 4.6% for gBLPACR-II. The combined incidence of gLow-BLNAR and gBLNAR was 72.6%, which was higher than that of BLNAR (62.4%). The MIC90s of antibacterial agents against the 197 strains were as follows; 0.0156 microg/mL for tosufloxacin and garenoxacin, 0.0313 microg/mL for levofloxacin and pazufloxacin, 0.0625 microg/mL for norfloxacin, 0.25 microg/mL for tazobactam/piperacillin (TAZ/PIPC) and ceftriaxone, 0.5 microg/mL for TAZ/PIPC (1:8) and cefditoren, 1 microg/mL for piperacillin, cefteram, cefotaxime, meropenem, tebipenem and minocycline, 2 microg/mL for doripenem, 4 microg/mL for cefcapene, imipenem and azithromycin, 8 microg/mL for sulbactam/ampicillin, clavulanic acid/amoxicillin (1:2, CVA/AMPC) and cefdinir, 16 microg/mL for CVA/AMPC (1:14), flomoxef and clarithromycin, 32 microg/mL for ampicillin. Although there was no rapid increase in the antibacterial resistance, the prevalence of BLNAR was still over 50%. In order to ensure the appropriate chemotherapy, it is important to continue the surveillance of susceptibility among H. influenzae.

A survey on adult community-acquired pneumonia was conducted jointly by multiple centers nationwide to verify the Japanese Respiratory Society Guidelines for the Management of Community-Acquired Pneumonia in Adults (JRS2005). The efficacy and safety of piperacillin (PIPC) were investigated at the same time. PIPC is recommended as the initial treatment for patients with suspected bacterial pneumonia and pneumococcal pneumonia in JRS2005. Overall, 552 and 333 patients were registered for safety and efficacy analysis in this study, respectively. The majority of the cases in which PIPC was used had moderate disease (63.7 %), and the most common daily dosage was 4 g (73.6 %). The efficacy rate was 83.5 % overall, 81.1 % in patients with suspected bacterial pneumonia, and 92.8 % in patients with pneumococcal pneumonia. The efficacy rate with a daily dosage of 4 g was 84.9 %, and the efficacy rates achieved with a daily dosage of 4 g in patients who had mild and moderate suspected bacterial pneumonia were 90.0 and 82.6 %, respectively. The most commonly isolated causative organisms were Streptococcus pneumoniae (S. pneumoniae) and Haemophilus influenzae (H. influenzae), and the bacterial eradication rates were high (97.2 and 100 %, respectively). The incidence of adverse drug reactions was 5.62 %, among which the main events were hepatic dysfunction and decreased white blood cell count. In conclusion, this study showed that PIPC is safe and effective at 4 g/day for mild-to-moderate adult community-acquired pneumonia.

This report presents the cases of three patients with fatal pneumonia that was highly suspected to be Pneumocystis pneumonia (PCP) based on serological diagnosis. Their chest radiographs showed bilateral pneumonia and each had presented with severe respiratory failure requiring mechanical ventilation when they arrived at the hospital. Although bronchoscopical sampling could not be performed, their chest computed tomography imaging and a marked elevation of serum KL-6 and β-D-glucan levels were characteristic of Pneumocystis pneumonia. All three were found to have been treated with temozolomide after surgery for malignant glioma. Temozolomide can cause Pneumocystis pneumonia. The three patients did not receive prophylactic medication against Pneumocystis pneumonia during treatment with temozolomide, and their histories suggested that all had delayed seeking treatment. It may be difficult to diagnose Pneumocystis pneumonia because the symptoms are not specific for Pneumocystis pneumonia and they tend to be similar to those of common respiratory infectious diseases. Therefore, patients who receive temozolomide therapy have the potential to develop fatal pneumonia and should be carefully observed. The patients should also be adequately informed about Pneumocystis pneumonia, and prophylaxis against Pneumocystis pneumonia should be considered proactively before treatment with temozolomide is initiated.

Objective: Pneumonia is reported to be associated with high morbidity in elderly and compromised individuals, with poor oral health demonstrated to be a significant risk factor for pneumonia. Several opportunistic pathogens, such as Streptococcus pneumoniae, Klebsiella pneumoniae, Escherichia coli and Staphylococcus aureus, have been detected in patients with pneumonia. We investigated the prevalence of opportunistic pathogens in the oral cavity of healthy independent living Japanese elderly subjects and analysed factors related to harbouring those pathogens., Materials and Methods: We studied 265 subjects, each of whom received a dental examination, during which specimens were collected with a tongue swab and examined for the presence of 10 oral opportunistic pathogens using single or multiple selective media. Furthermore, the presence of occult blood in saliva was examined using test paper strips., Results: Oral opportunistic pathogens were detected in 13.6% of the subjects. Those positive for occult blood in saliva had a significantly higher rate of harbouring the pathogens (p < 0.05). In addition, age was a significant factor for the presence of pathogenic microbes in the oral cavity (p = 0.033)., Conclusion: Positive findings of occult blood in saliva and older age are suggested to be significant factors for harbouring opportunistic pathogens in the oral cavity., (© 2010 The Gerodontology Society and John Wiley & Sons A/S.)

Nontypeable Haemophilus influenzae (NTHi) commonly colonizes the upper respiratory tract of children and causes otitis media, sinusitis, and bronchitis. Invasive NTHi diseases such as meningitis and septicemia have rarely been reported, especially in children with underlying predisposing conditions such as head trauma and immune compromise. However, we report a previously healthy 2-year-old girl who developed meningitis and septicemia caused by NTHi biotype ΙΙΙ. She was treated with dexamethasone, meropenem, and ceftriaxone, and recovered uneventfully. We wish to emphasize that NTHi should be borne in mind as a potential pathogen that can cause meningitis and septicemia, even in previously healthy children.

Background and Objective: More than 100000 Japanese die of pneumonia every year. The number of people residing in nursing homes is increasing with the ageing of the population. In 2005, the American Thoracic Society/Infectious Diseases Society of America (ATS/IDSA) published important guidelines for the management of healthcare-associated pneumonia (HCAP). In Japan, however, the optimum strategy for management of HCAP is still unclear. The purpose of this study was to clarify the clinical features of patients with HCAP., Methods: Patients (n = 202) who were consecutively admitted with a diagnosis of acute pneumonia between October 2007 and September 2009 were retrospectively evaluated. Using the ATS/IDSA guidelines, patients were divided into three groups: a community-acquired pneumonia (CAP) group (n = 123), a nursing home-acquired pneumonia (NHAP) group (n = 46) and a HCAP other than NHAP (O-HCAP) group (n = 33). These groups were then compared with respect to laboratory data, microbiological findings and mortality., Results: Thirty-day mortality in the NHAP group (10.9%) tended to be higher than that in the CAP group (3.3%) or the O-HCAP group (0%). The pathogens most frequently identified were Streptococcus pneumoniae and Haemophilus influenzae in the CAP group, methicillin-resistant Staphylococcus aureus and Klebsiella pneumoniae in the NHAP group, and S. pneumoniae and K. pneumoniae in the O-HCAP group., Conclusions: The NHAP group was clinically different from the O-HCAP group, based on bacteriological examination and mortality rates. In order to accurately diagnose, and formulate optimum treatment strategies for Japanese patients, the categories of HCAP, as specified in the ATS/IDSA guidelines, should not be applied directly either to patients with NHAP or those with O-HCAP., (© 2011 The Authors; Respirology © 2011 Asian Pacific Society of Respirology.)

Objectives: We sought to identify predictors for hearing loss in Japanese children with meningitis., Methods: We analyzed 155 cases of pediatric meningitis without other entities causing hearing loss in children admitted to Saitama Children's Medical Center between 1990 and 2005 for potential risk factors for hearing loss, using multiple logistic regression. Auditory brain stem response tests were performed to evaluate hearing loss., Results: Of 155 children, 35 (23%) developed hearing loss (21 unilaterally and 14 bilaterally). Profound hearing loss (greater than 90 dB normal hearing level) occurred in 15 patients (9.7%; 4 unilaterally and 11 bilaterally). Of 112 patients with positive cerebrospinal fluid cultures, 27 (24%) developed hearing loss and 13 (12%) showed profound loss. Of 22 patients with Streptococcus pneumoniae meningitis, 11 (50%) developed hearing loss and 7 (32%) showed profound loss. Of 54 patients with Haemophilus influenzae meningitis, 11 (20%) developed hearing loss and 4 (7.4%) showed profound loss. High serum C-reactive protein levels and cerebrospinal fluid cultures positive for Streptococcus pneumoniae were identified as significant risk factors for hearing loss., Conclusions: A high serum C-reactive protein level was first identified as a risk factor for hearing impairment after pediatric meningitis.

In Japan, the increasing incidence of β-lactum-resistant Haemophilus influenzae infections is of growing concern. We retrospectively studied whether the prevalence of β-lactamase-negative ampicillin-resistant strains of H. influenzae was influenced by chronic lung diseases. H. influenzae isolates, obtained from patients who were diagnosed with acute or chronic bronchitis, or acute exacerbation of chronic bronchitis in 2005, were studied. In addition to susceptibility testing, polymerase chain reaction (PCR) was performed for the detection of TEM-1 β-lactamase, and Asn526-Lys and Ser385-Thr amino acid substitutions in the ftsI gene encoding penicillin-binding protein-3 (PBP-3). The minimum inhibitory concentration values of β-lactams were found to be increased in isolates from patients with chronic bronchitis who had been repeatedly administered antibiotics. Genetic analysis using PCR suggested that this might be associated with a high frequency of β-lactamase-negative strains with mutations in PBP-3. The presence of β-lactum-resistant strains needs to be considered for patients with chronic bronchitis in whom H. influenzae is isolated as a causative pathogen.

Background: Streptococcus pneumoniae and Haemophilus influenzae infections in children with influenza have been noted because of the severity of co-infection. In Japan, vaccination against S. pneumoniae and H. influenzae infections has been listed in the vaccine program in 2008, but the characteristics of the two organisms, colonizing at the initial stage of influenza infection, have not been investigated in detail., Methods: Nasopharyngeal swabs from children with influenza (flu(+)) (n= 236; mean age, 6.2 years) were examined for bacterial pathogens, including S. pneumoniae and H. influenzae. They were then examined for serotypes, drug susceptibilities, and resistance genes (or gene mutations). As a reference, children with upper respiratory tract infection (URTI(+), flu(-); n = 189; mean age, 6.2 years) were also examined., Results: S. pneumoniae, beta-streptococci (groups A, B, and G), methicillin-susceptible and -resistant S. aureus, Moraxella catarrhalis, and H. influenzae were isolated. For S. pneumoniae, nine serotypes were detected with prevalent types of 3, 6, 19 and 23. Penicillin resistance was detected in types 19 and 23, while resistance to macrolide and clindamycin was found in various types. For H. influenzae, only b serotype was detected, with marked ampicillin resistance. The majority was non-typeable. Very similar results were obtained even in URTI(+) (flu(-)) cases., Conclusion: Multiple drug-resistant S. pneumoniae with major serotypes, for example, 19 and 23 and H. influenzae with serotype b were already present at the initial stage of influenza infection, similar to URTI(+) flu(-) cases. They could be prevented by current vaccines, but drug-resistant non-typeable H. influenzae is troubling.

We examined the frequency of ampicillin (ABPC) resistance of 3784 strains of Haemophilus influenzae isolated from the infants aged from 1 month after birth to 12 years who had been treated in the Pediatric Department of our hospital for 9 years and 6 months from July 1999 to December 2008. We determined the drug susceptibility by Etest, and judged the resistance by the standard of Clinical and Laboratory Standard Institute. The rate of strains of MIC of less than 2 microg/ml to ABPC was 87.1% in 1999, but the ratio was decreasing year after year significantly up to 28.7% in 2008 (p < 0.001). beta-lactamase negative ABPC resistant (BLNAR) strains whose MIC to ABPC was more than 4 microg/ml was 2.9% in 1999, but increased significantly to 43.3% in 2008 (p < 0.001). MIC90 was 2 microg/ml in 1999, but increased to 16 microg/ml in 2008.

We studied changes in the drug resistance of 606 strains of Haemophilus influenzae (H. influenzae) and 502 strains of Streptococcus pneumoniae (S. pneumoniae) isolated from our patients between 1997 and 2006. The incidence of beta-lactamase nonproducing ampicillin-susceptible H. influenzae (BLNAS) in 1997-1998 and 1999-2000 was 72.0 and 69.6%, respectively. In 2005-2006, the incidence of BLNAS decreased to 31.0%, while that of beta-lactamase nonproducing ampicillin-resistant H. influenzae (BLNAR) and intermediate-resistant H. influenzae increased to 65.5%. Remarkably early development of BLNAR and intermediate-resistant H. influenzae was found in patients younger than 3 years, as compared to patients older than 3 years. The proportion of penicillin-susceptible S. pneumoniae (PSSP) in 1999-2000 was 18.4%. In 2005-2006, the proportions of penicillin-resistant S. pneumoniae (PRSP) and penicillin-intermediate S. pneumoniae (PISP) were lower, while that of PSSP increased to 38.2%. An early increase in the proportion of PSSP was found in patients older than 3 years, as compared to patients younger than 3 years. The difference between age groups may be attributed to entrance into nursery school, frequent administration of antibiotics, and the immature immunological state of patients younger than 3 years. Therefore, changes in the drug resistance of H. influenzae and S. pneumoniae should be investigated separately, depending on the age of the patients. The minimum inhibitory concentrations of antibiotics, including cefditoren and cefcapene, against BLNAR and PRSP did not increase. The marked increase in intermediate-resistant H. influenzae and BLNAR mandates a re-evaluation of the directions for antibacterial agents.

Four major causative bacteria (Streptococcus pneumoniae, Haemophilus influenzae, Moraxella catarrhalis, Streptococcus pyogenes) of pediatric respiratory infections, 295 clinical isolates in total, were isolated at general practitioners' clinics in Sendai city, and evaluated the drug susceptibility to seven antibiotics for pediatric. Penicillin-resistant S. pneumoniae and penicillin-intermediate S. pneumoniae (PRSP-PISP) were 55.8% of all S. pneumoniae isolates. The MIC90 of penicillin and cephalosporin antibiotics in S. pneumoniae were good, 0.5-1 microg/mL, however, macrolide-resistant strains remarkably increased. As for H. influenzae, 50.0% of all isolates were ampicillin-intermediate and -resistant (MIC: > or = 2 microg/mL), the MIC90s of cephalosporin antibiotics had large differences between 0.5 to 8 microg/mL, and generally less susceptibility was shown to other antibiotics. M. catarrhalis showed less susceptibility to amoxicillin which behaved unstably to penicillinase, on the other hand, the MIC90s of other antibiotics were relatively good, 0.25-1 microg/mL. S. pyogenes remarkably tend to be resistant to macrolide antibiotics, however, the MIC90s of penicillin and cephalosporin antibiotics were very good, 0.03-0.06 microg/mL. Pediatric respiratory infections are required a treatment which results in inhibition of drug-resistant bacteria. Based on the results of drug susceptibility testing, we should make a proper selection of antibiotics by reference to disposition such as drug concentration in serum and transfer into cells.

The activity of telithromycin and comparator antibacterials was examined in isolates of Streptococcus pneumoniae and Haemophilus influenzae isolated from patients with community-acquired pneumonia (CAP), acute exacerbations of chronic bronchitis (AECB), or sinusitis during year 5 (2003-2004) of the Prospective Resistant Organism Tracking and Epidemiology for the Ketolide Telithromycin global resistance surveillance study. Among S. pneumoniae, penicillin nonsusceptibility and erythromycin resistance were 35.7% and 36.0%, respectively. beta-Lactamase was produced by 12.3% of H. influenzae isolates. beta-Lactamase-negative ampicillin-resistant strains, mainly from Japan, comprised 5.2% of global H. influenzae isolates. Telithromycin and levofloxacin were the most active agents tested against S. pneumoniae and H. influenzae (>99% of isolates susceptible) isolated from patients with CAP, AECB, or bacterial sinusitis. Amoxicillin-clavulanate, levofloxacin, and telithromycin were the most active agents against multidrug-resistant S. pneumoniae.

We investigated the susceptibility to antibacterials of 194 strains of Haemophilus influenzae isolated from medical facilities in Gifu prefecture between 2005 and 2006, and compared these results with those of 280 strains of H. influenzae isolated between 1999 and 2000. Additionally, the strains that had been separated between 2005 and 2006 were examined for beta-lactamase (BL) production, the mutation of ftsI gene coding for PBP3, the bla gene coding for TEM type of BL and the serotype. Referring to the CLSI breakpoint, H. influenzae strains were classified into the following categories: (1) beta-lactamase-negative ampicillin-susceptible (BLNAS) strains, which showed BL negative, ampicillin (ABPC) and ampicillin/sulbactam (ABPC/SBT)-MIC < or = microg/ml, (2) beta-lactamase producing ampicillin-resistant (BLPAR) strains, which showed BL producing and ABPC/SBT-MIC < or =2 microg/ml, (3) beta-lactamase-negative ampicillin-resistant (BLNAR) strains, which showed BL negative, ABPC and ABPC/SBT-MIC > or =2 microg/ml, (4) beta-lactamase-producing amoxicillin/clavulanic acid-resistant (BLPACR) strains, which showed BL producing and ABPC/SBT-MIC > or =4 microg/ml. The prevalence of each resistance class were 71.8% for BLNAS, 7.9% for BLPAR, 19.6% for BLNAR and 0.7% for BLPACR in strains isolated between 1999 and 2000. But they were 38.1% for BLNAS, 4.6% for BLPAR, 54.6% for BLNAR and 2.6% for BLPACR in strains isolated between 2005 and 2006, indicating that the percentage of BLNAS and BLPAR decreased and that of BLNAR and BLPACR increased from 1999-2000 to 2005-2006. On the basis of ftsI substitutions and having bla gene, the strains isolated between 2005 and 2006 were classified into the following distribution: 24.2% for gBLNAS, 4.1% for gBLPAR, 10.8% for gLow-BLNAR, 57.7% for gBLNAR, and 3.1% for gBLPACR-II. Ratio of BLNAR belonging to gBLNAR and gLow-BLNAR based on the ftsI substitutions and having bla gene was higher than that based on the susceptibility pattern. The MIC50 and MIC90 for those strains isolated between 2005 and 2006 were as follows; 0.0039, 0.0156 microg/ml for garenoxacin, 0.0078, 0.0156 microg/ml for tosufloxacin and ciprofloxacin, 0.0156, 0.0313 microg/ml for levofloxacin, 0.0313, 0.0625 microg/ml for norfloxacin, 0.0625, 0.25 microg/ml for piperacillin/ tazobactam, 0.0625, 0.5 microg/ml for piperacillin, 0.125, 0.25 microg/ml for ceftriaxone and cefditoren, 0.5, 1 microg/ml for cefteram, chloramphenicol and tetracycline, 0.5, 2 microg/ml for cefotaxime, 2, 8 microg/ml for ampicillin, ampicillin/sulbactam and cefdinir. In comparison with the values for the strains isolated between 1999 and 2000, the MIC50s of beta-lactam for the strains isolated between 2005 and 2006 increased over 4 times.

We analyzed the incidence of beta-lactamase non-producing ampicillin-resistance (BLNAR) Haemophilus influenzae in Showa University Hospital from June 2004 to March 2005. The ratio of BLNAR in total H. influenzae isolates was 44.5%. BLNAR accounted for 89.6% of the ampicillin non-susceptible strains displaying a minimal inhibitory concentration(MIC) of ampicillin of more than 2 microg/ml, which is the break point advocated by the Clinical and Laboratory Standards Institute. Of the total BLNARs, 71.4% were isolated from children of six and under. We analyzed mutations of the SSN and KTG motif of penicillin binding protein 3 encoded by the ftsI gene, since mutations in these regions are considered to be responsible for the drug-resistance of BLNAR. Mutations in SSN and KTG motifs were identified in all BLNAR isolates. We also detected mutations in the ampicillin susceptible strains displaying an ampicillin MIC of 0.5 or microg/ml. When a noticeable MIC increase of ABPC or other beta-lactams in the routine clinical laboratory practice, gene analysis of ftsI gene of the isolates displaying increased MIC is required to measure the spreading of BLNAR.

We surveyed pediatric bacterial meningitis epidemiology from January 2005 to December 2006 in Japan, with the following results. Bacterial meningitis cases numbered 246 -138 boys and 108 girls-, equivalent to 1.7-1.72 children of 1,000 hospitalized in pediatrics per year. The age distribution for infection was highest in those under 1 year of age and decreased with increasing age, Haemophilus influenzae was the most common infection causing the pathogen, followed by Streptococcus pneumoniae, group B streptococcus, and Escherichia coli. The relationship between causative pathogens and age distribution was as follows: group B streptococcus and E. coli were major pathogens in patients under 4 months old and H. influenzae and S. pneumoniae in those over 4 months old. Susceptibility tests at individual facilities showed 59.3% of H. influen- zae isolates and 69.3% of S. pneumoniae isolates in 2004 to be drug-resistant. Ampicillin and cephem antibiotics are effective against GBS, E. coli, and Listeria, so combined of ampicillin and cephem antibiotics are used as first-line antibiotics in many facilities in patients under 4 month old and combined of carbapenem antibiotics effective against PRSP and cephem effective against H. influenzae were the first choice against childhood bacterial meningitis in patients over 4 month old.

Any increase in beta-lactam-resistant Haemophilus influenzae is a serious problem in respiratory and otolaryngology medicine. In this study, we examined the antibiotic susceptibility and genotype of 457 clinical Haemophilus influenzae strains isolated in Hokkaido Prefecture, Japan. Strains with beta-lactam-resistant mutations in gene encoding penicillin-binding protein 3 were more frequently found in lower respiratory tract specimens (sputa) than in upper respiratory tract specimens, such as rhinorrhea. The existence of the TEM-1 beta-lactamase gene occurred more frequently in adult patients than in pediatric patients. The results suggest that beta-lactam-resistant or nonsusceptible strains are more prevalent in adult patients with respiratory diseases. We observed only a very few strains which were nonsuscpetible to third-generation cephalosporins (CEPs) and carbapenems. However, 12%-13% of the strains were shown to be resistant to penicillins and second-generation CEPs, and approximately 4% of the strains were shown to be nonsusceptible to fourth-generation CEPs. In addition, we identified tetracycline-resistant (2.8%), chloramphenicol-resistant (0.6%), clarithromycin-resistant (2.6%), and fluoroquinolone-nonsusceptible (approximately 2%) H. influenzae strains.

We conducted a prospective bacteriological survey to investigate antibiotic resistance-related genetic characteristics and the turnover of nasopharyngeal Haemophilus influenzae carriage in healthy children in day-care centers (DCCs). A total of 363 nasopharyngeal mucus samples were collected from children aged 0 to 6 years attending two DCCs in the summer of 2004 (n = 184) and the following winter (n = 179). We obtained 172 H. influenzae isolates and analyzed them by antimicrobial susceptibility testing, PCR for bla(TEM-1) and the penicillin-binding protein (PBP) gene, and pulsed-field gel electrophoresis (PFGE). The overall carriage rate was 47.4% (172/363), and 37.2% of the isolates (64/172) were ampicillin (AMP) resistant. All the resistant isolates had a PBP mutation(s), while only three isolates had TEM-1. The carriage rate was significantly higher in the winter than in the summer (56.4% and 38.6%, respectively), owing to the increase in the numbers of AMP-susceptible H. influenzae isolates in the winter. Children aged < or = 3 years showed a higher rate of carriage of H. influenzae isolates with an AMP resistance gene(s) than those aged > or = 4 years (21.9% and 12.6%, respectively). Forty-two strains with different PFGE patterns were obtained from among the 172 isolates. Only five strains were observed in both seasons. None of the strains isolated in the summer was isolated from the same carrier in the winter. Twenty-seven strains (64.3%) were isolated from two or more children, and 25 of these were each isolated from children belonging to the same DCC. These results indicate the spread of H. influenzae, particularly those with a PBP mutation(s), and the highly vigorous genetic turnover and substantial horizontal transmission of this pathogen in healthy children attending DCCs in Japan.

We screened 457 Haemophilus influenzae strains isolated in Japan during 2002 to 2004 and identified 12 fluoroquinolone-resistant strains (2.6%). The resistant strains were divided into three genotypes (eight, three, and one of each type). These were isolated from patients over 58 years of age. Several fluoroquinolone-resistant clones appeared to have invaded the population of elderly patients in a particular area, Sapporo city.

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 beta-lactamase-nonproducing ABPC-resistant (BLNAR) strains with a minimum inhibitory concentration (MIC) of ABPC of over 4 microg/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.

We evaluated the recent prevalence of antimicrobial-resistant Haemophilus influenzae isolated from the upper respiratory tracts (URT) of patients in Japan. Mutations in the ftsI gene, which encodes penicillin binding protein 3 (PBP3), and the clonal dissemination of the resistant strains were also investigated. A total of 264 H. influenzae isolates were collected from patients with URT infections. According to the criteria of the Clinical and Laboratory Standards Institute for the susceptibility of H. influenzae to ampicillin (AMP), the isolates were distributed as follows: 161 (61.0%) susceptible strains (MIC < or = 1 microg/ml), 37 (14.0%) intermediately resistant strains (MIC = 2 microg/ml), and 66 (25.0%) resistant strains (MIC > or = 4 microg/ml). According to PCR-based genotyping, 172 (65.1%) of the isolates had mutations in the ftsI gene and were negative for the beta-lactamase (bla) gene. These 172 isolates were thus defined as genetically beta-lactamase-negative ampicillin-resistant (gBLNAR) strains. The ftsI mutant group included 98 (37.1%) strains with group I/II mutations in the variable mutated region (group I/II gBLNAR) and 74 (28.0%) strains with group III mutations in the highly mutated region (group III gBLNAR). Eighty-seven (33.0%) of the isolates were genetically beta-lactamase-negative ampicillin-susceptible (gBLNAS) strains. The group III gBLNAR strains showed resistance to beta-lactams. Only five strains (1.9%) were positive for a bla gene encoding TEM-type beta-lactamase. The three clusters consisting of 16 strains found among the 61 BLNAR strains (MIC > or = 4 microg/ml and without the bla gene) showed identical or closely related DNA restriction fragment patterns. Those isolates were frequently identified among strains with a MIC to AMP of 16 microg/ml. The current study demonstrates the apparent dissemination and spread of a resistant clone of H. influenzae among medical centers in Japan. The gBLNAR strains show a remarkable prevalence among H. influenzae isolates, with the prevalence increasing with time. This fact should be taken into account when treating URT infections.

Background: The clinical management of community-acquired respiratory tract infections (RTIs) is complicated by the increasing worldwide prevalence of antibacterial resistance, in particular, beta-lactam and macrolide resistance, among the most common causative bacterial pathogens. This study aimed to determine the mechanisms and molecular- and sero-epidemiology of antibacterial resistance among the key paediatric respiratory pathogens in Japan., Methods: Isolates were collected at 18 centres in Japan during 2002 and 2003 from children with RTIs as part of the PROTEKT surveillance programme. A proportion of Haemophilus influenzae isolates was subjected to sequencing analysis of the ftsI gene; phylogenetic relatedness was assessed using multilocus sequence typing. Streptococcus pneumoniae isolates were screened for macrolide-resistance genotype by polymerase chain reaction and serotyped using the capsular swelling method. Susceptibility of isolates to selected antibacterials was performed using CLSI methodology., Results and Discussion: Of the 557 H. influenzae isolates collected, 30 (5.4%) were beta-lactamase-positive [BL+], 115 (20.6%) were BL-nonproducing ampicillin-resistant (BLNAR; MIC >or= 4 mg/L) and 79 (14.2%) were BL-nonproducing ampicillin-intermediate (BLNAI; MIC 2 mg/L). Dabernat Group III penicillin binding protein 3 (PBP3) amino acid substitutions in the ftsI gene were closely correlated with BLNAR status but phylogenetic analysis indicated marked clonal diversity. PBP mutations were also found among BL+ and BL-nonproducing ampicillin-sensitive isolates. Of the antibacterials tested, azithromycin and telithromycin were the most active against H. influenzae (100% and 99.3% susceptibility, respectively). A large proportion (75.2%) of the 468 S. pneumoniae isolates exhibited macrolide resistance (erythromycin MIC >or= 1 mg/L); erm(B) was the most common macrolide resistance genotype (58.8%), followed by mef(A) (37.2%). The most common pneumococcal serotypes were 6B (19.7%), 19F (13.7%), 23F (13.5%) and 6A (12.8%). Telithromycin and amoxicillin-clavulanate were the most active antibacterials against S. pneumoniae (99.8% and 99.6% susceptibility, respectively)., Conclusion: Approximately one-third of H. influenzae isolates from paediatric patients in Japan are BLNAI/BLNAR, mainly as a result of clonally diverse PBP3 mutations. Together with the continued high prevalence of pneumococcal macrolide resistance, these results may have implications for the clinical management of paediatric RTIs in Japan.

The minimum inhibitory concentration (MIC) of five antibiotics and the presence of resistance genes was determined in 163 Haemophilus influenzae isolates collected over 13 years (1987-2000) in four two-yearly sampling periods from patients with respiratory tract infections. The prevalence of beta-lactamase-negative ampicillin-susceptible strains was approximately 80% over the sampling period although fewer strains (65.9%) were recovered in the period 1995-1997. TEM-1 type beta-lactamase-producing strains were less frequent starting at 15.6% and declining to 2.2% in the final sampling period. Low-beta-lactamase-negative ampicillin-resistant (BLNAR) strains were uncommon in 1987-1989 (2.2%), peaked to 19.5% in 1995-1997, but fell back to 11.1% by 2000. Fully BLNAR strains were not detected until the last sampling period (6.7%). The MICs of ampicillin, levofloxacin, cefditoren and ceftriaxone remained stable but there was an eight-fold increase in the MIC of cefdinir over the sampling period. Pulsed-field gel electrophoresis of DNA digests showed that three representative BLNAR strains were genetically distinct and 11 DNA profiles were identified among 17 low-BLNAR strains. These data suggest that the number of genetically altered BLNAR and low-BLNAR strains are increasing in Japan.

Infants at day care centers tend to contract repetitive upper respiratory infections and prolonged otitis media. The increase in antimicrobial-resistant bacteria, particularly in infants, has given rise to a stubborn therapeutic problem. We studied the nasopharyngeal carriage and drug resistance to Haemophilus influenzae (H. influenzae) and Streptococcus pneumoniae (S. pneumoniae), the most common pathogens of upper respiratory infections, in infants at day care centers. Nasopharyngeal cultures of infants between the ages of 0 and 6 years were conducted at two day care centers in July 2004 ("summer"; n=183), and in February 2005 ("winter"; n=182). Isolated H. influenzae and S. pneumoniae were subjected to antibiotic susceptibility tests by broth microdilution. We also conducted an otolaryngological examination and a survey on past and life histories. H. influenzae in summer (38.3%) increased significantly in winter (57.7%). Beta-lactamase-negative and positive ampicillin-resistant H. influenzae (BLNAR+ BLPAR) in summer decreased significantly in winter. S. pneumoniae did not differ in summer (42.1%) or in winter (43.4%). Penicillin-resistant and intermediate S. pneumoniae (PRSP+PISP) was 41.3% in summer and decreased significantly to 19.0% in winter. BLNAR + BLPAR and PRSP + PISP differed with the day care center. In otolaryngological examination, rhinosinusitis was commonest (28.4% in summer and 30.8% in winter), followed by allergic rhinitis (8.7% in summer and 6.0% in winter) and otitis media (8.2% in summer and 6.0% in winter). Tonsillitis was minor (0.5% in both seasons). Rhinosinusitis in winter was significantly higher in carriers of H. influenzae and/or S. pneumoniae than in non carriers (36.4% versus 16.0%). Breast-fed infants tended to have less otitis media than bottle-fed infants (38.2% versus 52.9%). H. influenzae and/or S. pneumoniae plateaued (75-80%) after 12 months in day care centers. These results suggest that infants attending day care centers are immediately colonized by H. influenzae and S. pneumoniae in the nasopharynx after entering the centers. Nasopharyngeal drug-resistant H. influenzae and S. pneumoniae varied during the seasons and between day care centers. Further prospective studies are needed to determine upper respiratory tract infection in infants at day care centers and to evaluate carriage, epidemiology, and the drug-resistance rates of these pathogens.