Akihisa Togawa

Haemophilus influenzae and Haemophilus haemolyticus in tonsillar cultures of adults with acute pharyngotonsillitis

OBJECTIVE The aim of this study was to evaluate the clinical implication of Haemophilus haemolyticus, one of the closest relative of Haemophilus influenzae, on acute pharyngotonsillitis. METHODS We applied polymerase chain reaction (PCR) for 16S ribosomal DNA (rDNA) and IgA protease gene (iga) to distinguish H. haemolyticus and H. influenzae. RESULTS Among the 199 Haemophilus spp. isolated from 214 patients with acute pharyngotonsillitis, 52 (24.3%) H. influenzae strains and 23 (10.7%) H. haemolyticus strains were identified by polymerase chain reaction (PCR) for 16S rDNA and IgA protease gene (iga). All H. haemolyticus strains showed hemolysis on horse blood agar and there were no other Haemophilus spp., nonhemolytic H. haemolyticus and H. influenzae variant strains that had absent iga gene. H. hemolyticus showed close genetic relationship with H. influenzae evaluated by pulsed field gel electrophoresis (PFGE). The cases of acute pharyngotonsillitis showing WBC=7000/mm(3) or CRP=8 mg/dl were frequently found among cases with H. influenzae rather than cases with H. haemolyticus. CONCLUSION H. haemolyticus is a pharyngeal commensal that is isolated frequently from adults with acute pharyngotonsillitis.

Evaluation of a rapid immunochromatographic ODK-0901 test for detection of pneumococcal antigen in middle ear fluids and nasopharyngeal secretions.

Since the incidence of penicillin-resistant Streptococcus pneumoniae has been increasing at an astonishing rate throughout the world, the need for accurate and rapid identification of pneumococci has become increasingly important to determine the appropriate antimicrobial treatment. We have evaluated an immunochromatographic test (ODK-0901) that detects pneumococcal antigens using 264 middle ear fluids (MEFs) and 268 nasopharyngeal secretions (NPSs). A sample was defined to contain S. pneumoniae when optochin and bile sensitive alpha hemolytic streptococcal colonies were isolated by culture. The sensitivity and specificity of the ODK-0901 test were 81.4% and 80.5%, respectively, for MEFs from patients with acute otitis media (AOM). In addition, the sensitivity and specificity were 75.2% and 88.8%, respectively, for NPSs from patients with acute rhinosinusitis. The ODK-0901 test may provide a rapid and highly sensitive evaluation of the presence of S. pneumoniae and thus may be a promising method of identifying pneumococci in MEFs and NPSs.

A comparison of conventional and molecular microbiology in detecting differences in pneumococcal colonization in healthy children and children with upper respiratory illness

Conventional microbiology (CM) and real-time polymerase chain reaction (PCR) were used to determine rate and serotype of pneumococcal nasopharyngeal colonization in healthy children and children with upper respiratory illnesses (URI). One hundred and thirty-six healthy children and 79 children with URI were evaluated. Pneumococcal colonization was detected more often by real-time PCR than CM in healthy children (50% vs. 24%, p ≤ 0.001), while detection rates were comparable by CM and real-time PCR in children with URI (61% vs. 65%, NS). Pneumococcal serotypes were identified 2.3 times more often in healthy children by real-time PCR than CM, p ≤ 0.001 and 1.5 times more often in children with URI by PCR than CM, p = 0.01. Real-time PCR was also more sensitive in detecting multiple strains rather than CM in both healthy (p = 0.001) and children with URI (p ≤ 0.001). Overall real-time PCR proved superior to CM in detection and serotyping of Streptococcus pneumoniae. Future studies should incorporate real-time PCR technology along with CM to fully understand the epidemiology of colonization in health and illness.

PspA Family Distribution, Antimicrobial Resistance and Serotype of Streptococcus pneumoniae Isolated from Upper Respiratory Tract Infections in Japan

Background The protection against pneumococcal infections provided by currently available pneumococcal polysaccharide conjugate vaccines are restricted to the limited number of the serotypes included in the vaccine. In the present study, we evaluated the distribution of the pneumococcal capsular type and surface protein A (PspA) family of pneumococcal isolates from upper respiratory tract infections in Japan. Methods A total of 251 S. pneumoniae isolates from patients seeking treatment for upper respiratory tract infections were characterized for PspA family, antibiotic resistance and capsular type. Results Among the 251 pneumococci studied, the majority (49.4%) was identified as belonging to PspA family 2, while most of the remaining isolates (44.6%) belonged to family 1. There were no significant differences between the distributions of PspA1 versus PspA2 isolates based on the age or gender of the patient, source of the isolates or the isolates’ susceptibilities to penicillin G. In contrast, the frequency of the mefA gene presence and of serotypes 15B and 19F were statistically more common among PspA2 strains. Conclusion The vast majority of pneumococci isolated from the middle ear fluids, nasal discharges/sinus aspirates or pharyngeal secretions represented PspA families 1 and 2. Capsular serotypes were generally not exclusively associated with certain PspA families, although some capsular types showed a much higher proportion of either PspA1 or PspA2. A PspA-containing vaccine would potentially provide high coverage against pneumococcal infectious diseases because it would be cross-protective versus invasive disease with the majority of pneumococci infecting children and adults.

The benefit of cochlear implantation in the Japanese elderly

The purpose of this study was to evaluate the benefit of cochlear implant for Japanese elderly people with profound hearing loss. A questionnaire was administered to the five Japanese elderly people over 65 years old with the Nucleus 22 channel device and their family. The questionnaire was designed to gain insight into the patients daily use and to evaluate the quality of life. Elderly patients who were implanted with the Nucleus device showed good ability to identify environmental sound and revealed no significant deterioration compared to younger patients. Implanted patients showed some difficulties in understanding speech in the noisy environment and conversation among several persons. Nevertheless, 80% of patients over 65 and their family reported communication benefits in the family and in social activities. In addition, 80% of the patients reported that they felt the better or improved quality of life after the implantation. All patients considered the implant was good for their life and would recommend a cochlear implant to a deaf friend. It is notable that 60% of patients and their family report that the recipients have become more cheerful. This study showed the cochlear implant contributes to the better quality of life in the Japanese elderly.

Distribution of fibronectin-binding protein genes ( prtF1 and prtF2 ) and streptococcal pyrogenic exotoxin genes ( spe ) among Streptococcus pyogenes in Japan

Two hundred and seventy-two strains of Streptococcus pyogenes isolated from patients with invasive and noninvasive infections in Japan were evaluated for the prevalence of fibronectin-binding protein genes (prtF1 and prtF2). The possible associations of the genes with streptococcal pyrogenic exotoxin genes, macrolide resistance genes, and emm types were also evaluated. Overall, about 50% of S. pyogenes isolates carried fibronectin-binding protein genes. The prevalence of the prtF1 gene was significantly higher among isolates from noninvasive infections (71.4%) than among isolates from invasive infections (30.8%; P = 0.0037). Strains possessing both the prtF1 and prtF2 genes were more likely to be isolates from noninvasive infections than isolates from invasive infections (50.6% vs 15.4%; P = 0.019). S. pyogenes isolates with streptococcus pyrogenic exotoxin genes (speA and speZ) were more common among isolates without fibronectin-binding protein genes. The speC gene was more frequently identified among isolates with fibronectin-binding protein genes (P = 0.05). Strains belonging to emm75 or emm12 types more frequently harbored macrolide resistance genes than other emm types (P = 0.0094 and P = 0.043, respectively). Strains carrying more than one repeat at the RD2 region of the prtF1 gene and the FBRD region of the prtF2 gene were more prevalent among strains with macrolide resistance genes than among strains negative for macrolide resistance genes. These genes (i.e., the prtF1, prtF2, and spe genes) may enable host-bacteria interaction, and internalization in the host cell, but may not enable infection complications such as invasive diseases.

Genetic characteristics of Haemophilus influenzae and Streptococcus pneumoniae isolated from children with conjunctivitis-otitis media syndrome

Acute conjunctivitis is the most common ocular disorders among children and frequently concomitant with acute otitis media (AOM) as conjunctivitis-otitis syndrome. In this study, we evaluated prevalence of causative pathogens and PCR-based genotypes of Haemophilus influenzae and Streptococcus pneumoniae among children with conjunctivitis-otitis media syndrome. Nontypeable H. influenzae (NTHi) is identified most often at 61.8% in conjunctiva exudates followed by S. pneumoniae at 28.2% and Moraxella catarrhalis at 19.1%. Genetic β-lactamase nonproducing ampicillin resistant (gBLNAR) strains of NTHi and genetic penicillin resistant S. pneumoniae (gPRSP) were identified at 72.1% and at 74.2% among conjunctiva isolates by polymerase chain reaction (PCR), respectively. Pneumococcal strains having either ermB or mefE genes were identified at 93.5% among conjunctiva isolates. The restriction fragment of patterns of 89.7% pairs of H. influenzae isolates and 100% pairs of pneumococcal isolates from conjunctiva exudates, middle ear fluids (MEFs) and nasopharyngeal swabs were identical. In contrast to the previous reports, most prevalent strains from conjunctivitis-otitis media syndrome was BLNAR H. influenzae in this study. The causative pathogen responsible for acute conjunctivitis will be originated from the nasopharynx. In the absence of MEFs one can possibly rely on the nasopharyngeal culture to guide an appropriate treatment.

Primary laryngeal cryptococcosis resembling laryngeal carcinoma

A case of an 82-year-old female with primary laryngeal cryptococcosis who had undergone long-term corticosteroid therapy for chronic obstructive pulmonary disease and rheumatoid arthritis is reported. She complained hoarseness with swallowing pain and irritability of the larynx for over a month. Endoscopic examination revealed a white, exudative irregular region on right arytenoid that mimicked a laryngeal carcinoma. Histological examination showed pseudoepitheliomatous hyperplasia and severe submucosal inflammation with ovoid budding yeasts by Grocotts stain. A serological study indicated a high titer of cryptococcal antigen. After treating with oral fluconazole for 3 months, her primary lesion of larynx turned to be clear. We implicate a long-term use of steroids as the significant risk factor in developing cryptococcosis of the larynx.

Amount of Haemophilus influenzae Genomic DNA in Middle Ear Fluids ofPediatric Acute Otitis Media

Background: Nontypeable Haemophilus influenzae (NTHi) is a leading causative pathogen responsible for acute otitis media (AOM). Objective: In this study, we applied real-time PCR to quantify the amount of genomic NTHi DNA in MEEs and to evaluate the influence of the amount of bacteria in MEEs on the clinical outcome of AOM. Methods: Thirty-two children with severe AOM were evaluated for the quantification of the amount of NTHi genomic DNA in middle ear effusions (MEEs) by real-time PCR. Results: The improvement of tympanic membrane abnormalities at the second visits was significantly worse in cases with MEEs containing dense NTHi genomic DNA rather than in cases with MEEs containing sparse NTHi genomic DNA. The amount of NTHi DNA genome in MEEs were significantly higher in cases of which improvement ratio of tympanic membrane abnormalities at the second visit was less than 50% than in cases of which improvement ratio of tympanic membrane abnormalities above 50%. Discussion: It is important to predict the clinical futures of AOM for appropriate treatments. The quantity of infected pathogens will be an important factor for poor clinical outcomes of AOM. Conclusion: The current findings suggest that severity of tympanic membrane abnormalities evaluated scoring system proposed by Japanese AOM guideline reflect the amount of NTHi in MEEs and that predict the poor improvement of AOM.

An application of outer membrane protein p6-specific enzyme-linked immunosorbent assay for detection of haemophilus influenzae in middle ear fluids and nasopharyngeal secretions.

An enzyme-linked immunosorbent assay specific to outer membrane protein P6 (P6-ELISA) was applied for detecting Haemophilus influenzae in middle ear fluids (MEFs) from acute otitis media (AOM) patients and in nasopharyngeal secretions (NPSs) from acute rhinosinusitis patients. P6-ELISA had a sensitivity of 83.3% for MEFs and 71.5% for NPSs and a specificity of 85.6% for MEFs and 92.5% for NPSs, respectively. Real-time PCR exhibited significant differences in the number of ompP1 gene copies among samples determined by P6-ELISA to be positive and negative for H. influenzae. However, because the P6-ELISA test has the reactivity in Haemophilus species include two commensals H. haemolyticus and H. parainfluenzae, it is thus a weak method in order to detect only NTHi correctly. Consequently, diagnosis using the P6-ELISA should be based on an overall evaluation, including the results of other related examinations and clinical symptoms to prevent misleading conclusions in clinical setting.