Epidemiological study of respiratory syncytial virus infection in adults during the pandemic of COVID-19.

INTRODUCTION: While respiratory syncytial virus (RSV) is one of the most common pathogens in adults admitted to the ICU due to respiratory diseases, no reports regarding the occurrence rate of RSV infections in adults in Japan during the COVID-19 pandemic exist. PATIENTS AND METHODS: We conducted this retrospective study to examine the exact occurrence rate of RSV infections in adults. We reviewed all patients (≥18 years) with any respiratory symptoms who received quantitative polymerase chain reaction (PCR) using nasopharyngeal samples for respiratory viruses by GeneLEAD at the Aichi Medical University Hospital between November 2022 and November 2023. RESULTS: A total of 541 adult patients who underwent PCR test were enrolled in this study. RSV was identified in 18 cases (3.3 %); 8 (1.5 %) upper and 10 (1.8 %) lower respiratory tract infections. Influenza A and SARS-CoV-2 were found in 10 (1.8 %) and 61 (11.3 %), respectively. Patients with RSV infections and COVID-19 had more comorbidities than those with Influenza virus infections. As for RSV-associated with lower respiratory tract infection cases, 10 developed acute respiratory failure, resulting in 1 fatal case due to pneumonia and 1 died of septic shock due to ileus. The 30-, 90-day mortality rates were 1 (6 %) and 2 (11 %) respectively. CONCLUSION: About 3 % of adults had RSV infections during the COVID-19 pandemic. The outcomes of RSV infections in adults were similar to those by COVID-19. Those with comorbidities should have a preventive method against RSV infections, the same as for COVID-19.

Clinical characteristics and antimicrobial susceptibility of Fusobacterium species isolated over 10 years at a Japanese university hospital.

PURPOSE: Anaerobic bacteria, existing on human skin and mucous membranes, can cause severe infections with complications or mortality. We examined the clinical characteristics of patients infected with Fusobacterium spp. and assessed their antibiotic susceptibility. METHODS: Clinical data were collated from patients diagnosed with Fusobacterium infections in a Japanese university hospital between 2014 and 2023. Antibiotic susceptibility tests were conducted following the Clinical and Laboratory Standards Institute guidelines. RESULTS: We identified 299 Fusobacterium isolates. The median age was 61 years (range, 14-95 years), with females constituting 43.1% of the patients. Most infections were community-acquired (84.6%, 253/299). Multiple bacterial strains were isolated simultaneously in 74.6% of cases. One-fourth of the patients had solid organ malignancies (25.4%, 76/299), and 14.5% (11/76) of those had colorectal cancer. The 30-day mortality rate was 1.3%. Fusobacterium species were isolated from blood cultures in 6% (18/299) of the patients. Patients, aged 75 years or older, with cerebrovascular disease or hematologic malignancy exhibited significantly higher prevalence of blood culture isolates in univariate analysis. Each Fusobacterium species had its characteristic infection site. Approximately 5% F. nucleatum and F. necrophorum isolates showed penicillin G resistance. Moxifloxacin resistance was observed in varying degrees across strains, ranging from 4.6 to 100% of isolates. All isolates were sensitive to ß-lactam/ß-lactamase inhibitors, carbapenems, and metronidazole. CONCLUSION: We show a link between Fusobacterium species and solid organ malignancies. We observed resistance to penicillin, cefmetazole, clindamycin, and moxifloxacin, warranting caution in their clinical use. This study offers valuable insights for managing Fusobacterium infections and guiding empirical treatments.

Three Successfully Treated Cases of Lodderomyces elongisporus Fungemia: Case Reports and a Review of the Literature.

Fungemia is a fatal systemic infection that can occur in immunocompromised patients. Despite that, antifungal stewardship is spreading widely, but the mortality rate is extremely high, showing 40-60%. Loderomyces elongiporus is a newly morphologically detected pathogen, first described in 1994, followed by isolation in humans in 2008. It has been misrecognized as Candida parapsilosis. Recently, fever attributable to L. elongisporus fungemia cases has been reported, and the etiology and clinical features are still unknown. Here, we present three successfully treated L. elongisporus fungemia cases by echinocandin. In total, 11 cases were reviewed, including ours. Six of the eleven cases (55%) had external devices. All cases had some immunocompromised conditions or underlying diseases, such as diabetes mellitus, lung cancer, etc. Six patients survived, and the remaining five died. Seven patients who had received echinocandin initially survived. Risk factors for L. elongiporus fungemia overlap with those of candidemia. Even though there is no breakpoint for L. elongiporus, echinocandin can be a helpful treatment regimen for L. elongiporus fungemia.

Comparative study of SmartAmp assay and reverse transcription-polymerase chain reaction by saliva specimen for the diagnosing COVID-19.

INTRODUCTION: The pandemic of a novel coronavirus disease 2019 (COVID-19) caused by a severe acute respiratory coronavirus 2 (SARS-CoV-2) infection has been problematic worldwide. A new SARS-CoV-2 diagnostic test (SmartAmp) was licensed in Japan in July 2021. This method, which enables us to diagnose COVID-19 as well as a gene mutation on the virus, is promising to reduce medical costs and staff labor. PATIENTS AND METHODS: To analyze the diagnostic accuracy of the SmartAmp assay for diagnosing COVID-19, we performed this retrospective study at our institute during April and May 2021. We compared the results of the SmartAmp assay and real-time reverse transcription-polymerase chain reaction (rRT-PCR) using a saliva sample from individuals suspected as having COVID-19. RESULTS: Out of 70 samples tested, the SmartAmp assay had 50 (71%) positive and 20 (29%) negative results. Using rRT-PCR as a reference, the diagnostic accuracy displayed a sensitivity of 84%, a specificity of 95%, a positive predictive value of 97.7%, and a negative predictive value of 70.4%. On the other hand, false-negative cases were found in 7 (10%), and there was no significant difference of Ct-value between true positive and false negative cases (Mean Ct-value 25.2 vs. 27.5 cycles, p = 0.226 by Mann-Whitney U test). CONCLUSION: The SmartAmp assay is a valuable method to diagnose COVID-19 rapidly. However, the negative predictive value is not high enough to diagnose the disease, so that negative results should be considered for rRT-PCR testing if patients are suspected of having COVID-19.

Efficacy and validity of automated quantitative chemiluminescent enzyme immunoassay for SARS-CoV-2 antigen test from saliva specimen in the diagnosis of COVID-19.

INTRODUCTION: The pandemic of a novel coronavirus disease 2019 (COVID-19) caused by a severe acute respiratory coronavirus 2 (SARS-CoV-2) infection has been problematic worldwide. A new SARS-CoV-2 antigen test (LUMIPULSEⓇ) was licensed and widely used in Japan since May 2020. We conducted this study intending to whether the automated quantitative CLEIA antigen test using a saliva sample is effective and valid for the diagnosis of COVID-19. PATIENTS AND METHODS: We analyzed and compared the diagnostic accuracy of both the automated quantitative CLEIA antigen test and real-time RT-PCR (rRT-PCR) using a saliva sample from individuals suspected as having COVID-19. RESULTS: A total of 305 samples were collected and tested in Aichi Medical University Hospital and affiliated facilities from December 2020 until January 2021 at our institute. Using reverse-transcription PCR as a reference, the AUROC of the automated quantitative CLEIA antigen test was 0.903 (95% confidential interval 0.845-0.962, p < 0.001). The appropriate cut-off antigen level was 4.0 pg/mL and had a sensitivity of 77.8%, a specificity of 99.6%, a positive predictive value of 98%, and a negative predictive value of 94.5%. On the other hand, the diagnostic accuracy of the antigen test decreased among patients among patients with COVID-19 with threshold cycle (Ct-value)≥27, which shows the AUROC was 0.795 (95%CI 0.687-0.907, p < 0.001). CONCLUSION: While the automated quantitative CLEIA antigen test from saliva specimen could be one of the most useful diagnostic tests for the diagnosis of COVID-19 in general practice, clinicians should know the limitations of the antigen test.

Comparative evaluation of nasopharyngeal swab and saliva specimens for the molecular detection of SARS-CoV-2 RNA in Japanese patients with COVID-19.

Considering the issues of shortage of medical resources and the invasiveness and infection risk involved in the collection of nasopharyngeal swab specimens, there is a need for an effective alternative test specimen for SARS-CoV-2 RNA detection. Here, we investigated suitability of saliva as a non-invasively obtained specimen for molecular detection of SARS-CoV-2 RNA in Japanese patients with COVID-19. In total, 28 paired clinical specimens of saliva and nasopharyngeal swabs were collected from 12 patients at various time points after symptom onset. Each specimen was assayed using reverse transcription real-time polymerase chain reaction (rRT-PCR) on the BD MAX open system using primers and probes targeting the N-gene. The saliva and nasopharyngeal swab specimens showed 19 and 15 positive results, respectively. No invalid (PCR inhibition) result was observed for any specimen. The qualitative results of each specimen obtained in the period immediately after symptom onset were similar. Three convalescent patients presented saliva-positive results, whereas their nasopharyngeal swabs were negative at four different time points, suggesting that saliva may be superior to nasopharyngeal swabs in terms of obtaining stable assay result of SARS-CoV-2. In conclusion, our results suggest that saliva can potentially serve as an alternative to nasopharyngeal swabs as a specimen for SARS-CoV-2 rRT-PCR. As saliva can be collected by patients themselves, it may be an effective way to overcome the shortage of personal protective equipment and specimen sampling tools.

Could threshold cycle value correctly reflect the severity of novel coronavirus disease 2019 (COVID-19)?

The novel coronavirus disease 2019 (COVID-19) is diagnosed by positive result of reverse transcription polymerase chain reaction (RT-PCR) for the novel coronavirus. We concluded that cycle threshold value (Ct-value) of real-time RT-PCR (rRT-PCR) assay could decrease as patients recover. Results of rRT-PCR assay could remain positive among asymptomatic patients for longer than 2 weeks. The discharge criteria of COVID-19 patients using a negative result of rRT-PCR should be reconsidered.

[Performance Evaluation of "BinaxNOW™ Streptococcus pneumoniae" Using Alere™ Reader].

Streptococcus pneumoniae is one of the major bacterial pathogens of community-acquired pneumonia. Immunochromatographic assay tests are used to detect pneumococcal capsular antigen. In many cases, They can be read visually. The Alere™ reader (Reader), which was developed in October 2018 by Alere Medical Co., Ltd. (currently, Abbott Diagnostics Medical Co., Ltd.) for interpreting BinaxNOW™ Streptococcus pneumoniae test (BinaxNOW™), quickly displays the results of the immunochromatographic tests, objectively and accurately, as it was launched for the purpose of streamlining laboratory workflow. The performance of the reader was evaluated by using urine samples from 100 patients, who were ordered pneumococcal urine antigen test from September 2018 to February 2019 at our hospital. Of the 100 samples, 14 were visually positive and 19 were reader positive. All visually positive samples generated reader positive result. Because 1 of the 5 cases which indicated a negative visual determination and positive reader determination was a sample with strong viscosity and turbidity, it was retested after centrifugation at 3,000×g for 10 min, resulting in negative reader determination. In 2 cases, S. pneumoniae were detected in sputum gram stains and culture tests. 5 discrepant samples were all visually and reader positive after concentration by centrifugal ultrafiltration. A questionnaire about visual interpretation was conducted among 31 individuals, by using urine from day 0 to day 4 collected from the patients whose test result was visually negative, reader positive and sputum culture positive at day 0. As a result, the number of operators who determined visually positive was 0 on day 0 (0%), 16 on day 1 (51.6%), 13 on day 2 (41.9%), 2 on day 3 (6.5%), and 0 on day 4 (0%). There were individual differences in ability to interpret low level positive result visually. On the other hand, reader can remove individual differences among operators from the interpretation of BinaxNOW™ and interpret positive result earlier than visual interpretation. Therefore reader was considered to be useful tool in clinical settings.

A novel disk-based detection method with superior sensitivity for ß-lactamase production in third-generation cephalosporin-resistant Enterobacteriaceae.

OBJECTIVE: Current phenotypic methods for extended-spectrum ß-lactamase (ESBL), AmpC ß-lactamase (AmpC), and carbapenemases fail to detect isolates that co-produce other classes of ß-lactamases. In this study, we have developed a novel assay (Applied Multiplex Disk Method: AMU-DM) for the phenotypic detection and identification of ß-lactamases produced by Enterobacteriaceae. METHODS: We evaluated the performance of the method by comparison with PCR results for 78 Enterobacteriaceae clinical isolates that were positive by the ESBL screening test and negative by the ESBL confirmation test. Additionally, one NCTC strain and four ATCC strains were also included in the test population for the study as reference. RESULTS: For 79/83 (95%) isolates tested, the AMU-DM results matched those obtained by PCR. The concordance rates were 31/31 (100%), 11/11 (100%), 3/3 (100%), 0/1 (0%), 15/15 (100%), 16/19 (84%), and 3/3 (100%) for AmpC, ESBL and AmpC co-production, Klebsiella pneumoniae carbapenemase (KPC), KPC and ESBL co-production, metallo ß-lactamase (MBL), MBL and ESBL co-production, and MBL and AmpC co-production, respectively. CONCLUSION: The AMU-DM is convenient to perform, economical, and highly sensitive in identifying ESBLs, AmpCs, and carbapenemases. Our method may be useful in clinical settings for the implementation of relevant infection control measures and for surveillance purposes.

Effect of sodium mercaptoacetic acid on different antimicrobial disks in the sodium mercaptoacetic acid double disk synergy test for detection of IMP-1 metallo-ß-lactamase-producing Pseudomonas aeruginosa isolates in Japan.

We determined the optimal antimicrobial in the sodium mercaptoacetic acid double disk synergy test (SMA-DDST) for the detection of IMP-1-producing Pseudomonas aeruginosa isolates in Japan and evaluated the performance of the test. Fifty-four P. aeruginosa clinical isolates were tested, including 39 IMP-1 producers and 15 non-metallo-ß-lactamase (MBL)-producing carbapenem- and ceftazidime (CAZ)-resistant isolates. The SMA-DDST was performed with CAZ, cefepime (CFPM), imipenem (IPM), meropenem (MEPM), doripenem (DRPM), or biapenem (BIPM)-containing disks. The sensitivity of the SMA-DDST with CAZ, CFPM, IPM, MEPM, DRPM, and BIPM was 39/39 (100%), 36/39 (92%), 18/39 (46%), 8/39 (21%), 19/39 (49%), and 36/39 (92%), respectively. The specificity was 15/15 (100%) for all SMA-DDSTs. This suggests that the isolates may have a resistance mechanism other than MBL production for IPM, MEPM, or DRPM. Since the CAZ resistance mechanism in P. aeruginosa is the same as that of CFPM, but differs from that of carbapenems, we conclude that combining CAZ with BIPM SMA-DDSTs can prevent any failure in the detection of IMP-1-producing P. aeruginosa.

[Is BC-GN applicable for analysis of strains and urine/sputum specimens?]

As the reagent that can simultaneously detect bacterial nucleic acid/drug-resistant genes from the culture-positive liquid of blood cultures, Verigene® system includes the Verigene® Gram-Positive Blood Culture test (BC-GP) and the Verigene® Gram Negative Blood Culture test (BC-GN). This study used BC-GN to identify the names of bacteria from stock strains, urine samples, and sputum specimens and detect drug-resistant genes. The stock strains included 28 clinical isolates, 9 urine samples in which the target bacterial strain grew to 106CFU/ml or more in culture, and 9 sputum specimens in which the target bacterial strain grew to 105CFU/ml or more in culture. The bacterial identification and detection of drug-resistant genes used quality Matrix Assisted Laser Desorption/Ionization Time of Flight Mass Spectrometry (MALDI-TOF MS) analysis and conventional PCR method, respectively, followed in comparison with the results of Verigene®. As a result, the measurement results of Verigene® for the stock strains and urine samples had a high concordance rate with MALDI-TOF MS analysis and PCR method. On the other hand, the concordance rate of the sputum specimens with the Verigene® measurement results was only 40% (4 out of 10 specimens). These results suggest that BC‒GN can be an effective tool for AMR rapid diagnosis if the measurement target includes not only bacterial strains in the culture-positive liquid of blood cultures, but also other bacterial strains and urine.

[Development of a simplified assay for detection of van gene harbored enterococci using the automated BD MAX platform].

We developed and evaluated of multiplex real-time PCR assay for detection of vancomycin-resistant genes (vanA, vanB, vanC1 and vanC2/C3) using the new, fully automated BD MAX platform. Ct value analyses of real-time PCR simultaneous repeatability test have showed the usefulness; coefficient of variation: CV (%) were determined 2.09%, 1.72%, 1.41% and 1.52% with vanA, vanB, vanC1 and vanC2/C3, respectively. We also evaluated with 43 strains of enterococci were characterized by conventional PCR method; 4/4 for vanA-positive, 14/14 for vanB-positive, 1/1 for vanB plus vanC1-positive, 6/6 for vanC1-positive, 4/4 for vanC2/C3- positive and 14/14 for all-van gene-negative strains were identified correctly. This assay was automatically performing before and after PCR operations previously done manually by operator, such as DNA extraction, sample dispensing and gel electrophoresis or the ethidium bromide dyeing. As a result, work burden and the risk of the contamination were largely reduced and were shortened to about half for measurement time. We conclude that this assay could greatly contribute to efficient and rapid detection of vancomycin-resistant genes.

[Multicenter surveillance of Pseudomonas aeruginosa strains for antimicrobials in Aichi prefecture in 2009].

We investigated the susceptibility to antimicrobials of 204 Pseudomonas aeruginosa strains isolated from 21 hospitals in Aichi prefecture from September to November 2009. MIC distributions of various antimicrobials were analyzed in terms of geographic region of isolation, patient status (outpatient or inpatient), and type of specimens that the strain was isolated from. The results were as follows. 1. Although more than 90% of strains were susceptible to all aminoglycosides and colistin, 80-90% of them were susceptible to beta-lactams and fluoroquinolones. MIC distributions of all antimicrobials measured were not significantly different between regions. 2. Only 1 strain (0.5%) was multi-drug resistant Pseudomonas aeruginosa (MDRP). Thirteen strains (6.4%) showed imipenem MIC > or = 16 microg/mL, and 16 strains (7.8%) showed ciprofloxacin MIC > or = 4 microg/mL. These strains tended to be more isolated from urine, respiratory tract specimens, or surgical specimens. 3. The MICs of tazobactam/piperacillin, panipenem, meropenem, doripenem, biapenem, sulbactam/cefoperazone, cefepime, and aztreonam were significantly higher in strains isolated from inpatients than in those from outpatients. MIC distributions of antimicrobials other than beta-lactams were not significantly different between situations where strains were isolated. 4. MIC distributions of piperacillin, all carbapenems, cefepime, gentamicin, and all fluoroquinolones were significantly different among samples from which strains were isolated. The strains isolated from blood showed lower MICs against all antimicrobials than those from other samples. No difference was found in MIC distributions when categorized according to bacteremic origin. The MICs were apparently elevated against beta-lactams, fluoroquinolones, and gentamicin in strains isolated from respiratory tract specimens, and against beta-lactams, and fluoroquinolones in strains isolated from urine. It was suggested that in P. aeruginosa surveillance, the results should be reported by stratifying with patient status, and type of specimens that the strain was isolated from and that regional surveillance should be useful with such stratification to establish antibiograms for empirical antimicrobial choice.