Implementation of two novel schemes for patients on dialysis as a response to the COVID-19 surge in Tokyo.

The Japanese government recommended hospitalization of patients on dialysis once they tested positive because of their high COVID-19 mortality rate and definite need for periodic dialysis. However, after experiencing the Delta variant surge, strategic changes towards outpatient care for mild or asymptomatic cases, along with strengthening emergency preparedness were needed. Facing the Omicron surge, the Tokyo Metropolitan Government introduced two novel schemes: i) a temporary medical facility with a dialysis center for infected patients on hemodialysis, which started admitting patients on dialysis on January 20, 2022, to provide additional bed capacity and access to hemodialysis and ii) a transportation scheme for patients who need travel to maintenance dialysis facilities from their homes, which was introduced on February 5. The Tokyo Metropolitan Government, cooperating with some nephrology experts, announced these schemes and urged local dialysis facilities to change strategies, providing information regarding infection prevention measures and treatments in online seminars on February 3 and 7. Consequently, promoting outpatient care did not lead to an increase in the case fatality ratio (CFR) in patients on dialysis with COVID-19 in Tokyo during the first Omicron surge (January 7 to February 10, 8.2%; February 11 to March 31, 5.5%). Furthermore, after an additional online seminar on July 20, the CFR dramatically declined in the second Omicron surge (July 8 to September 8, 1.2%). Implementation of public health intervention and careful communication with local dialysis facilities were both crucial to the strategic changes. To maintain essential health services, emergency preparedness should be cultivated during regular times.

Photobiochemical mechanisms of biomolecules relevant to germicidal ultraviolet irradiation at 222 and 254 nm.

To inactivate viruses and microorganisms, ultraviolet light in the short wavelength region is a promising candidate for mitigating the infection of disease. Germicidal mercury lamps emitting at 254 nm and KrCl excimer lamps emitting at 222 nm have sterilisation properties. In this work, wavelength dependence of the photobiochemical mechanisms was investigated with 222- and 254-nm irradiation to analyze the underlying damage mechanisms of DNA/RNA and proteins, using Escherichia coli, a protease, an oligopeptide, amino acids, plasmid DNA and nucleosides. The photorepair of damaged DNA and the "dark" reversion of the hydrates of uracil phosphoramidite coupling blocks were also investigated.

Comparative study of bacterial flora in bronchoalveolar lavage fluid of pneumonia patients based on their pneumonia subtypes and comorbidities using 16S ribosomal RNA gene analysis.

INTRODUCTION: The culture method is the gold standard for identifying pathogenic bacteria in patients with pneumonia but often does not reflect the exact bacterial flora in pulmonary lesions of pneumonia, partly owing to easiness or difficulties in culturing certain bacterial species. We aimed to evaluate bacterial flora in bronchoalveolar lavage fluid (BALF) samples directly obtained from pneumonia lesions using 16S ribosomal RNA (rRNA) gene analysis to compare the results of the BALF culture method in each category of pneumonia. METHODS: Bacterial florae were detected by a combination of the culture method, and the clone library method using the 16S rRNA gene sequencing in BALF directly obtained from pneumonia lesions in pneumonia patients from April 2010 to March 2020 at the University of Occupational and Environmental Health, Japan, and affiliated hospitals. Clinical information of these patients was also collected, and lung microbiome was evaluated for each pneumonia category. RESULTS: Among 294 pneumonia patients (120 with community-acquired pneumonia (CAP), 101 with healthcare-associated pneumonia (HCAP), and 73 with hospital-acquired pneumonia (HAP)), significantly higher percentages of obligate anaerobes were detected in CAP than in HCAP and HAP patients by the clone library method. Corynebacterium species were significantly highly detected in HAP patients and patients with cerebrovascular diseases than in patients without, and Streptococcus pneumoniae was frequently detected in patients with diabetes mellitus. CONCLUSION: Obligate anaerobes may be underestimated in patients with CAP. Corynebacterium species should be regarded as the causative bacteria for pneumonia in patients with HAP and cerebrovascular diseases.

Risk factors associated with hospital transfer among mild or asymptomatic COVID-19 patients in isolation facilities in Tokyo: a case-control study.

Objectives: The Tokyo Metropolitan Government has been implementing facility-based isolation of asymptomatic/mild coronavirus disease (COVID-19) patients to facilitate timely hospital referral. However, there are only a few published studies in prehospital settings, and the factors associated with hospital transfer are unclear. Our study identified the factors associated with COVID-19 deterioration in a prehospital setting. Methods: This case-control study assessed the risk factors for hospital transfer from isolation facilities and the need for ambulance transport due to deterioration among COVID-19 patients, using multivariate logistic regression analysis. Results: In total, 10 590 patients (median age 34 years), with male predominance (61.1%), were included. 367 (3.5%) were transferred to hospital, of whom 44 (12.0%) required ambulance transport. Hypertension, diabetes, and bronchial asthma were prevalent in 704 (6.6%), 195 (1.8%), and 305 (2.9%) patients, respectively. After adjustment, older age, male sex, higher body mass index (BMI), and comorbidities (including diabetes, inflammatory bowel disease, and bronchial asthma) were associated with hospital transfer. Older age, male sex, and higher BMI significantly increased the risk of transfer by ambulance. Conclusions: Our results may be beneficial for the development of intervention measures for probable future COVID-19 waves.

Reporting on the implementation to set up a "care and isolation facility" for mild COVID-19 cases in Tokyo.

The increasing number of COVID-19 cases has placed pressure on medical facilities. Against this backdrop, the Tokyo Metropolitan Government established a facility for mild and asymptomatic COVID-19 cases by using existing hotels. These kinds of facilities were established in several countries, and represented a spectrum from hotel-like to hospital-like care. In this article, we focused on implementation and related strategies for establishing such a facility in Tokyo as implementation research, while ensuring patient and staff safety. This facility had three functions: care, isolation, and buffering. For the implementation strategy, we used several strategies from the Expert Recommendations for Implementing Change (ERIC) to implement functions similar to an ordinary hospital, but using fewer inputs. This experience can be applied to other resource-limited settings such as that in less developed countries.

Spontaneous pulmonary emphysema in mice lacking all three nitric oxide synthase isoforms.

The roles of endogenous nitric oxide (NO) derived from the entire NO synthases (NOSs) system have yet to be fully elucidated. We addressed this issue in mice in which all three NOS isoforms were deleted. Under basal conditions, the triple n/i/eNOSs-/- mice displayed significantly longer mean alveolar linear intercept length, increased alveolar destructive index, reduced lung elastic fiber content, lower lung field computed tomographic value, and greater end-expiratory lung volume as compared with wild-type (WT) mice. None of single NOS-/- or double NOSs-/- genotypes showed such features. These findings were observed in the triple n/i/eNOSs-/- mice as early as 4 weeks after birth. Cyclopaedic and quantitative comparisons of mRNA expression levels between the lungs of WT and triple n/i/eNOSs-/- mice by cap analysis of gene expression (CAGE) revealed that mRNA expression levels of three Wnt ligands and ten Wnt/ß-catenin signaling components were significantly reduced in the lungs of triple n/i/eNOSs-/- mice. These results provide the first direct evidence that complete disruption of all three NOS genes results in spontaneous pulmonary emphysema in juvenile mice in vivo possibly through down-regulation of the Wnt/ß-catenin signaling pathway, demonstrating a novel preventive role of the endogenous NO/NOS system in the occurrence of pulmonary emphysema.

Effects of sublingual immunotherapy on nasal symptoms and sleep disturbance in patients with Japanese cedar pollinosis.

OBJECTIVE: Japanese cedar (JC) pollinosis is the most common seasonal allergic rhinitis (AR) in Japan. AR reduces the quality of life not only because of nasal symptoms but also because of sleep disturbance. In the present study, we investigated the effects of sublingual immunotherapy (SLIT) with a standardized JC pollen extract on nasal symptoms and AR-related sleep disturbance in patients with JC pollinosis. METHODS: In the present non-randomized controlled study, we assigned thirty-one patients with JC pollinosis who received SLIT into the SLIT group, and another thirty-eight patients with JC pollinosis who visited our hospital without treatment into the untreated group. We evaluated nasal symptoms and sleep disturbance using the classification of the severity of AR symptoms and the Athens Insomnia Scale, respectively. RESULTS: The nasal symptom scores and the Athens Insomnia Scale scores of patients in the SLIT group were both significantly lower than those of patients in the untreated group. There was a significant correlation between total nasal symptom scores and the Athens Insomnia Scale scores. CONCLUSIONS: These findings suggested that SLIT with JC pollen extract suppressed nasal symptoms in patients with JC pollinosis, leading to improvements in AR-related sleep disturbance and daytime troubles with daily life.

Effects of corticosteroid on mRNA levels of histamine H1 receptor in nasal mucosa of healthy participants and HeLa cells.

The purpose of this study is to examine the effect of intranasal corticosteroid (INCS) administration on histamine H1 receptor (H1R) gene expression in the nasal mucosa of healthy participants and the effects of dexamethasone on basal and histamine-induced H1R mRNA expression, and histamine-induced phosphorylation of extracellular signal-regulated kinase (ERK) in HeLa cells. Sixteen healthy participants were given INCS once daily for a week. After pretreatment of dexamethasone, HeLa cells were treated with histamine. Levels of H1R mRNA and phosphorylation of ERK were measured using real time PCR and immunoblot analysis, respectively. Levels of H1R mRNA in the nasal mucosa of healthy participants receiving INCS was significantly decreased. Dexamethasone suppressed basal levels of H1R mRNA, and histamine-induced up-regulation of H1R mRNA and ERK phosphorylation in HeLa cells. These data suggested that corticosteroid inhibited both basal transcription and histamine-induced transcriptional activation of H1R through its suppression of ERK phosphorylation in the signaling pathway involved in H1R gene transcription. It is further suggested that pre-seasonal prophylactic administration of INCS suppresses both basal and pollen-induced upregulation of H1R gene expression in the nasal mucosa of patients with pollinosis, leading to prevention of the exacerbation of nasal symptoms during peak pollen season. J. Med. Invest. 67 : 311-314, August, 2020.

Coinfection With Multiple Nontuberculous Mycobacteria as a Possible Exacerbating Factor in Pulmonary Nontuberculous Mycobacteriosis: Clone Library Analysis Using the 16S Ribosomal RNA Gene.

BACKGROUND: Mycobacterial culture is the gold standard for the diagnosis of nontuberculous Mycobacterium (NTM) infections. However, this method is not suitable for detection of coinfection with different NTMs. RESEARCH QUESTION: The goal of this study was to determine if clone library analysis of BAL fluid (BALF) was useful for detection of NTM phylotypes, including multiple NTM phylotypes, in pulmonary NTM infections. STUDY DESIGN AND METHODS: BALF samples obtained from 120 patients with suspected pulmonary NTM infections were retrospectively evaluated by using the mycobacterial culture and clone library methods between July 2010 and August 2016. RESULTS: In total, 55 (45.8%) patients were diagnosed as NTM positive according to results of mycobacterial culture, and 52 patients were NTM positive as determined by using the clone library method. Furthermore, 45 (86.5%) and seven (13.5%) patients exhibited a single phylotype (mono-phylotype group) and multiple phylotypes of NTM (multi-phylotype group), respectively. Compared with the mono-phylotype group, the multi-phylotype group had a significantly higher incidence of adverse chest CT findings (P = .048). In addition, 11 patients who were NTM negative according to results of BALF mycobacterial culture were determined to be NTM positive according to the clone library method. Six of these 11 patients were eventually diagnosed as NTM positive by using mycobacterial culture results within 6.2 ± 2.1 months following the initial sample collection. INTERPRETATION: Coinfection multiple phylotypes could be associated with adverse clinical findings. In addition, patients who test positive for NTM genes but negative for mycobacterial culture may be diagnosed with NTM lung infection within 1 year of the initial sample collection. Further follow-up of these patients may facilitate early detection of NTM species.

222-nm UVC inactivates a wide spectrum of microbial pathogens.

BACKGROUND: UVC has been used to inactivate several pathogens. Unlike the conventional 254-nm UVC, 222-nm UVC is harmless to mammalian cells. AIM: To investigate the disinfection efficacy of 222-nm UVC against human pathogens which are commonly found in the environment and healthcare facilities. METHODOLOGY: Staphylococcus aureus, Pseudomonas aeruginosa, Escherichia coli, Salmonella enterica subsp. serovar Typhimurium, Campylobacter jejuni, Bacillus cereus (vegetative cells and endospores), Clostridium sporogenes (vegetative cells and endospores), Clostoridioides difficile (endospores), Candida albicans (yeast), Aspergillus niger (hyphae and spores), Trichophyton rubrum (hyphae and spores), feline calicivirus and influenza A virus were irradiated with 222-nm UVC at various doses. The remaining live bacterial and fungal cells, and the viral infectivity were evaluated. The efficiency of 222-nm UVC germicidal effect was compared to that of the conventional 254-nm UVC. RESULTS: The 222-nm UVC showed potent germicidal effect to vegetative bacterial cells, yeast and viruses as efficient as the 245-nm UVC. The 222-nm UVC exhibited more potent germicidal effect to bacterial endospores, compared with the 254-nm UVC. The fungicidal effect of 222-nm UVC against the fungal spores and hyphae was weaker than that of 254-nm UVC. CONCLUSIONS: The 222-nm UVC is able to inactivate a wide spectrum of microbial pathogens. In comparison with the conventional 254-nm UVC, the germicidal effect of 222-nm UVC to the fungal hyphae and spores is low, but the 222-nm UVC exhibits strong germicidal effect to the bacterial endospores.

[Microbiology of Aspiration Pneumonia].

The number of patients with pneumonia has been increasing as the population ages, and most fatal pneumonia cases are the elderly with aspiration pneumonia. Although aspiration pneumonia leads to poor short- and long-term prognosis, there have been no practical ways to diagnose it precisely. Persistent subclinical aspiration without any subjective symptoms is problematic in clinical practice in patients with aspiration pneumonia, and physicians can only use aspiration risks such as brain infarction to diagnose aspiration pneumonia. Anaerobes have been believed to be major causative pathogens in aspiration pneumonia, based on data from the 1970's. In relation to these data, Marik insisted that there is a possible overestimation of anaerobes because 1) the sampling of microbiologic specimens was in the late phase in the course of the illness, especially frequently after developing complications such as abscesses, necrotizing pneumonia, or empyema thoracis; 2) the organisms recovered by percutaneous transtracheal aspiration (PTA) sampling could have been contaminated by the aspiration of oropharyngeal flora during the PTA procedure or colonized in the trachea; and 3) many of the patients had chronic alcoholism or were under general anesthesia. In addition, 4) oral care was not common in the 1970s, and 5) the patients in these reports were relatively young. Molecular biological approaches using the 16S ribosomal RNA (rRNA) gene have recently been used, and have enabled us to detect more exact pathogens compared to conventional bacterial culture. Using the method with the detection of the 16S rRNA gene, we evaluated the bacterial phylotypes in bronchoalveolar lavage fluid in patients with aspiration pneumonia and found that oral streptococci were the most detected phylotypes (31.0%), while anaerobes were only 6.0%. Our results suggest that oral streptococci are important, and anaerobes may have been overestimated as causative pathogens in patients with aspiration pneumonia.

Induction of plasmid-mediated AmpC ß-lactamase DHA-1 by piperacillin/tazobactam and other ß-lactams in Enterobacteriaceae.

Chromosomal AmpC ß-lactamase induction by several types of ß-lactams has been reported, but not enough data are available on DHA-1 ß-lactamase, a plasmid-mediated AmpC ß-lactamase. Therefore, we evaluated the DHA-1 ß-lactamase induction by various antibiotics including piperacillin/tazobactam (PIP/TZB) in this study. Six strains (Enterobacter cloacae 2 strains, Citrobacter freundii 1 strain, Serratia marcescens 2 strain, and Morganella morganii 1 strain) possessing chromosomal inducible AmpC ß-lactamase were used as controls. Four strains (Escherichia coli 2 strains, Klebsiella pneumoniae 1 strain, and C. koseri 1 strain) possessing DHA-1 ß-lactamase were used. The ß-lactamase activities were determined by a spectrophotometer using nitrocefin. ß-lactamase induction by PIP, PIP/TZB was not observed in any strains and ß-lactamase induction by third- and fourth-generation cephems was not observed in most strains. The induction ratios of the chromosomal AmpC ß-lactamase in the reference group by PIP/TZB were <1.51, and those of the DHA-1 ß-lactamase were <1.36, except for K. pneumoniae Rkp2004 (2.22). The ß-lactamase induction by first- and second-generation cephems, flomoxef, and carbapenem differed in each strain. Cefmetazole (CMZ) strongly induced ß-lactamase. This study demonstrated that the induction of DHA-1 ß-lactamase was similar to that of chromosomal AmpC using various Enterobacteriaceae, although the induction of ß-lactamase in both groups by PIP/TZB was low. We also reported that the induction of PIP/TZB, a ß-lactamase inhibitor combination antibiotic, against various AmpC-producing Enterobacteriaceae, including DHA-1 producers, was low.

Pulmonary Nocardiosis Caused by Nocardia exalbida in a Patient with Lung Cancer and Radiation Pneumonitis: A Case Report and Literature Review.

We report a case of Nocardia exalbida (N.exalbida)-induced pneumonia in a 70-year old Japanese man with lung cancer and radiation pneumonitis. He initially received doripenem (1.5 g/day) for pneumonia treatment, and N.exalbida was identified by a clone library analysis of bronchoalveolar lavage fluid obtained from the pneumonia lesion. The doripenem dosage was therefore increased to 3.0 g/day with adjunctive trimethoprim/sulfamethoxazole, and his pneumonia improved. N. exalbida is susceptible to antibiotics; thus, in nocardiosis, N. exalbida infection might be associated with a good response to treatment, although its clinical findings are non-specific and similar to those of other Nocardia infections.

Systemic bone loss, impaired osteogenic activity and type I muscle fiber atrophy in mice with elastase-induced pulmonary emphysema: Establishment of a COPD-related osteoporosis mouse model.

Although it is suggested that chronic obstructive pulmonary disease (COPD) and bone are related, almost all of the pathological mechanisms of COPD-related osteoporosis remain unknown. There is a mouse model showing a deterioration of bone quality after cigarette smoke exposure; however, in smoking exposure models, various factors exist that affect bone metabolism, such as smoking and body weight loss (muscle and fat mass loss). We considered it appropriate to use an elastase-induced emphysema model to exclude factors influencing bone metabolism and to investigate the influence of pulmonary emphysema on bone metabolism. The purpose of this study was to establish a COPD/emphysema-related osteoporosis mouse model by using the elastase-induced emphysema model. The lumbar vertebrae and femurs/tibiae exhibited trabecular bone loss and impaired osteogenic activity in 24-week-old male elastase-induced emphysema model mice. In addition, the model mice showed atrophy of type I muscle fibers without atrophy of type II muscle fibers. We believe that the mice described in this experimental protocol will be accepted as a COPD/emphysema-related osteoporosis mouse model and contribute to further investigations.

Utility of the Quick Sequential Organ Failure Assessment in Japanese patients with nursing- and healthcare-associated pneumonia.

AIM: To clarify the utility of sepsis evaluation using the Quick Sequential Organ Failure Assessment (qSOFA) tool in addition to the Pneumonia Severity Index (PSI); age, dehydration, respiration, orientation and blood pressure (A-DROP) index; and immunodeficiency, respiration, orientation, age and dehydration (I-ROAD) scoring systems, and risk factor evaluation of potentially drug-resistant (PDR) pathogens are suggested in the 2017 guidelines for pneumonia of the Japanese Respiratory Society in nursing- and healthcare-associated pneumonia patients. METHODS: We included 289 hospitalized nursing- and healthcare-associated pneumonia patients between April 2016 and March 2017, and investigated the ability of PSI, A-DROP, I-ROAD and qSOFA to predict pneumonia-related mortality. We also evaluated the associations among the risk factors for PDR pathogens, the detection ratio of PDR pathogens and pneumonia-related mortality. RESULTS: The mortality rate of pneumonia during hospitalization was 6.9% (20/289). The area under the curve for pneumonia-related mortality predicted using PSI, A-DROP, I-ROAD and qSOFA was 0.697 (95% confidence interval [CI] 0.59-0.80), 0.63 (95% CI 0.51-0.76), 0.61 (95% CI 0.52-0.70) and 0.701 (95% CI 0.59-0.81), respectively. In addition, higher areas under the curve were observed for pneumonia-related mortality predicted according to a combination of PSI and hypoalbuminemia (<2.5 g/dL) (0.75, 95% CI 0.64-0.86), and qSOFA and hypoalbuminemia (0.74, 95% CI 0.62-0.86) than for PSI and qSOFA alone. No significant associations were observed among the risk factors for PDR pathogens, the detection ratios of PDR pathogens and pneumonia-related mortality. CONCLUSIONS: qSOFA and the combination of qSOFA and hypoalbuminemia might be simple and useful evaluation tools for predicting pneumonia-related mortality in nursing- and healthcare-associated pneumonia patients. Geriatr Gerontol Int 2019; 19: 177-183.

TX99 Is a Neutralizing Monoclonal Antibody Against Mouse TIGIT.

T cell immunoglobulin and ITIM domains (TIGIT) is an inhibitory immunoreceptor expressed on NK cells, effector and memory T cells, and regulatory T cells (Tregs). The ligands for TIGIT are CD155 (PVR) and CD112 (PVRL2, nectin-2), which are broadly expressed on hematopoietic cells and nonhematopoietic cells. TIGIT negatively regulates antitumor responses, but promotes autoimmune reaction. Although neutralizing anti-human TIGIT mAbs are under clinical trials for cancers, how the blockade of TIGIT interaction with the ligands shows tumor immunity still remains unclear. Although analyses of mouse tumor model using a neutralizing anti-mouse TIGIT (mTIGIT) mAbs should be useful to address this issue, there are limitations to this type of studies due to unavailability of neutralizing anti-mTIGIT mAbs. In this study, we generated five clones of anti-mTIGIT mAbs, designated TX99, TX100, TX103, TX104, and TX105. We show that TX99 and TX100 showed the strongest binding to TIGIT. We also show that TX99 interfered with the interaction between TIGIT and CD155 and increased NK cell-mediated cytotoxicity against CD155-expressing RMA-S cells. Thus, TX99 is a unique neutralizing mAb that can be used for studies of mTIGIT functions.

Protective Role of Myelocytic Nitric Oxide Synthases against Hypoxic Pulmonary Hypertension in Mice.

RATIONALE: Nitric oxide (NO), synthesized by NOSs (NO synthases), plays a role in the development of pulmonary hypertension (PH). However, the role of NO/NOSs in bone marrow (BM) cells in PH remains elusive. OBJECTIVES: To determine the role of NOSs in BM cells in PH. METHODS: Experiments were performed on 36 patients with idiopathic pulmonary fibrosis and on wild-type (WT), nNOS (neuronal NOS)-/-, iNOS (inducible NOS)-/-, eNOS (endothelial NOS)-/-, and n/i/eNOSs-/- mice. MEASUREMENTS AND MAIN RESULTS: In the patients, there was a significant correlation between higher pulmonary artery systolic pressure and lower nitrite plus nitrate levels in the BAL fluid. In the mice, hypoxia-induced PH deteriorated significantly in the n/i/eNOSs-/- genotype and, to a lesser extent, in the eNOS-/- genotype as compared with the WT genotype. In the n/i/eNOSs-/- genotype exposed to hypoxia, the number of circulating BM-derived vascular smooth muscle progenitor cells was significantly larger, and transplantation of green fluorescent protein-transgenic BM cells revealed the contribution of BM cells to pulmonary vascular remodeling. Importantly, n/i/eNOSs-/--BM transplantation significantly aggravated hypoxia-induced PH in the WT genotype, and WT-BM transplantation significantly ameliorated hypoxia-induced PH in the n/i/eNOSs-/- genotype. A total of 69 and 49 mRNAs related to immunity and inflammation, respectively, were significantly upregulated in the lungs of WT genotype mice transplanted with n/i/eNOSs-/--BM compared with those with WT-BM, suggesting the involvement of immune and inflammatory mechanisms in the exacerbation of hypoxia-induced PH caused by n/i/eNOSs-/--BM transplantation. CONCLUSIONS: These results demonstrate that myelocytic n/i/eNOSs play an important protective role in the pathogenesis of PH.

Frequency of detection of Chlamydophila pneumoniae using bronchoalveolar lavage fluid in patients with community-onset pneumonia.

BACKGROUND: Chlamydophila pneumoniae is a causative pathogen of lower respiratory tract infection, which generally infects healthy, young people. However, it is often difficult to evaluate acute C. pneumoniae infection using upper respiratory tract specimens and/or sputum samples due to its persistent infection or colonization. The interpretation of frequency of detection of C. pneumoniae seems to be insufficient in community-onset pneumonia. The aim of this study was to evaluate the presence of C. pneumoniae using bronchoalveolar lavage fluid (BALF) samples. METHODS: BALF samples from 147 patients with pneumonia were retrospectively evaluated using C. pneumoniae-specific polymerase chain reaction (PCR) primers. RESULTS: None of the samples had positive PCR results for C. pneumoniae using two different sets of specific primers. Single and paired serological analyses were performed in 54 (36.7%) and 37 (25.2%) patients, respectively. These analyses revealed that 1 of 37 (2.7%) patients had a presumptive acute infection with C. pneumoniae, 8 of the 54 (14.8%) patients were suspected of having a C. pneumoniae infection, and 7 of the 37 (18.9%) patients were suspected of having past C. pneumoniae infection. In addition, cultivation and/or 16S rRNA gene sequencing detected Haemophilus influenzae in the presumptive case using the serological method. CONCLUSIONS: The results of the present study revealed that C. pneumoniae might be a minor causative agent of community-onset pneumonia according to an evaluation of specimens obtained from the lower respiratory tract.

Bacteriological incidence in pneumonia patients with pulmonary emphysema: a bacterial floral analysis using the 16S ribosomal RNA gene in bronchoalveolar lavage fluid.

Pulmonary emphysema is an important radiological finding in chronic obstructive pulmonary disease patients, but bacteriological differences in pneumonia patients according to the severity of emphysematous changes have not been reported. Therefore, we evaluated the bacteriological incidence in the bronchoalveolar lavage fluid (BALF) of pneumonia patients using cultivation and a culture-independent molecular method. Japanese patients with community-acquired pneumonia (83) and healthcare-associated pneumonia (94) between April 2010 and February 2014 were evaluated. The BALF obtained from pneumonia lesions was evaluated by both cultivation and a molecular method. In the molecular method, ~600 base pairs of bacterial 16S ribosomal RNA genes in the BALF were amplified by polymerase chain reaction, and clone libraries were constructed. The nucleotide sequences of 96 randomly selected colonies were determined, and a homology search was performed to identify the bacterial species. A qualitative radiological evaluation of pulmonary emphysema based on chest computed tomography (CT) images was performed using the Goddard classification. The severity of pulmonary emphysema based on the Goddard classification was none in 47.4% (84/177), mild in 36.2% (64/177), moderate in 10.2% (18/177), and severe in 6.2% (11/177). Using the culture-independent molecular method, Moraxella catarrhalis was significantly more frequently detected in moderate or severe emphysema patients than in patients with no or mild emphysematous changes. The detection rates of Haemophilus influenzae and Pseudomonas aeruginosa were unrelated to the severity of pulmonary emphysematous changes, and Streptococcus species - except for the S. anginosus group and S. pneumoniae - were detected more frequently using the molecular method we used for the BALF of patients with pneumonia than using culture methods. Our findings suggest that M. catarrhalis is more frequently detected in pneumonia patients with moderate or severe emphysema than in those with no or mild emphysematous changes on chest CT. M. catarrhalis may play a major role in patients with pneumonia complicating severe pulmonary emphysema.

Determining the Possible Etiology of Hospital-Acquired Pneumonia Using a Clone Library Analysis in Japan.

Obtaining precise etiological information regarding causative bacteria is important for the proper use of antimicrobials in hospital-acquired pneumonia (HAP), which is associated with a high rate of mortality. The aim of this study was to comparatively investigate the bacterial diversity in bronchoalveolar lavage fluid (BALF) in Japanese patients with HAP by the clone library method using the 16S rRNA gene. This study included Japanese patients with HAP who were treated at our hospital and referring hospitals. BALF specimens were obtained from pneumonia lesions identified on chest radiographs and/or computed tomography. Sputum specimens were also evaluated in patients with sputum production. Sixty-eight patients were ultimately enrolled. BALF cultivation revealed bacterial positivity in 53 of 68 (77.9%) patients, and Staphylococcus aureus (30.9%) was the most frequently isolated, followed by Pseudomonas aeruginosa (16.2%), and Escherichia coli (10.3%). In contrast, the clone library analysis identified the presence of some bacterial phenotype in 65 of 68 (95.6%) patients, and streptococci (16.2%), Corynebacterium species (11.8%), anaerobes (10.3%) were frequently detected as the predominant phylotypes. Both methods tended to detect S. aureus, Klebsiella pneumoniae, and E. coli in patients with late-onset pneumonia. In addition, the cases that phylotypes of S. aureus and P. aeruginosa were found to account for > 5% of the bacterial flora of each case were 42.9% and 72.7%, respectively. These results indicate that attention should be paid to the roles of gram-positive bacilli such as streptococci, Corynebacterium species and anaerobes, in addition to Gram-negative bacilli, in the pathogenesis of HAP.