Strategic Retesting to Reduce False Positive SARS-CoV-2 PCR Test Results.

BACKGROUND: Nucleic acid amplification testing is the gold standard for SARS-CoV-2 diagnostics, although it may produce a certain number of false positive results. There has not been much published about the characteristics of false positive results. In this study, based on retesting, specimens that initially tested positive for SARS-CoV-2 were classified as true or false positive groups to characterize the distribution of cycle threshold (CT) values for N1 and N2 targets and number of targets detected for each group. METHODS: Specimens that were positive for N-gene on retesting and accompanied with S-gene were identified as true positives (true positive based on retesting, rTP), while specimens that retested negative were classified as false positives (false positive based on retesting, rFP). RESULTS: Of the specimens retested, 85/127 (66.9%) were rFP, 16/47 (34.0%) specimens with both N1 and N2 targets initially detected were rFP, and the CT values for each target was higher in rFP than in rTP. ROC curve analysis showed that optimal cutoff values of CT to differentiate between rTP and rFP were 34.8 for N1 and 33.0 for N2. With the optimal cutoff values of CT for each target, out of the 24 specimens that were positive for both N1 and N2 targets and classified as rTP, 23 (95.8%) were correctly identified as true positives. rFP specimens had a single N1 target in 52/61 (85.2%) and a single N2 target in 17/19 (89.5%). Notably, no true positive results were obtained from any specimens with only N2 target detected. CONCLUSIONS: These results suggest that retesting should be performed for positive results with a CT value greater than optimal cutoff value for each target or with a single N1 target amplified, considering the possibility of a false positive. This may provide guidance on indications to perform retesting to minimize the number of false positives.

First report of New Delhi metallo-ß-lactamase-1-producing Acinetobacter soli in Japan.

New Delhi metallo-ß-lactamase (NDM)-producing gram-negative rods, including Acinetobacter species, are a global problem but have rarely been isolated in Japan. To our knowledge, this is the first study to isolate an NDM-1-producing Acinetobacter soli strain, KUH106, in Japan. We analyzed this strain using next-generation sequencing to examine the plasmid carrying NDM-1. This plasmid, named pKUH106_NDM1, is 41,135 bp in length and contains genetic contexts with the structure ISAba14-aph(3')-VI-ISAba125-blaNDM-1ble-MBL. Comparative analysis of the plasmid revealed that it resembled the plasmids of Acinetobacter detected in various countries, such as the A. soli isolate from Taiwan and the Acinetobacter baumannii isolate from a healthcare facility in Osaka Prefecture, Japan. These results suggest that blaNDM-1 may spread via this plasmid in Acinetobacter species. This phenomenon needs to be confirmed through the genetic analysis of A. baumannii and other carbapenem-resistant Acinetobacter species. In particular, blaNDM-1 and other resistance genes must be investigated, and the spread of these genes in the community must be cautioned.

Nationwide surveillance of bacterial respiratory pathogens conducted by the surveillance committee of the Japanese Society of Chemotherapy, the Japanese Association for Infectious Diseases, and the Japanese Society for Clinical Microbiology in 2019-2020: General view of the pathogens' antibacterial susceptibility.

The trends and prevalence of antimicrobial susceptibility of pathogens vary by country, region, and time. Long-term regular surveillance is required to investigate trends in the antimicrobial resistance of various isolated bacterial pathogens. We report the results of a nationwide surveillance on the antimicrobial susceptibility of bacterial respiratory pathogens in Japan conducted by the Japanese Society of Chemotherapy, the Japanese Association for Infectious Diseases, and the Japanese Society for Clinical Microbiology. The isolates were collected from clinical specimens obtained from adult patients who visited a collaborating medical facility between June 2019 and December 2020 and were diagnosed with respiratory tract infections by a physician. Antimicrobial susceptibility testing was performed in a centralized laboratory according to the methods recommended by the Clinical and Laboratory Standards Institute. Susceptibility testing was performed for 932 strains (201 Staphylococcus aureus, 158 Streptococcus pneumoniae, 6 S. pyogenes, 136 Haemophilus influenzae, 127 Moraxella catarrhalis, 141 Klebsiella pneumoniae, and 163 Pseudomonas aeruginosa) collected from 32 facilities in Japan. The proportions of methicillin-resistant S. aureus and penicillin-resistant S. pneumoniae were 35.3% and 0%, respectively. In H. influenzae, 16.2% and 16.9% were ß-lactamase-producing ampicillin resistant and ß-lactamase-negative ampicillin resistant, respectively. Extended-spectrum ß-lactamase-producing K. pneumoniae accounted for 5.0% of all K. pneumoniae infections. Carbapenemase-producing K. pneumoniae and multi-drug-resistant P. aeruginosa with metallo-ß-lactamase were not detected in this study. This surveillance will be a useful reference for treating respiratory infections in Japan and will provide evidence to enhance the appropriate use of antimicrobial agents.

Evaluation of the Analytical Performance of the Fully Automated Gene Analyzer µTASWako g1 for the Detection of SARS-CoV-2.

BACKGROUND: The worldwide spread of coronavirus disease 2019 (COVID-19) has led to an urgent need for nucleic acid amplification test (NAAT) for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Because NAAT has many manual processes, results may vary depending on the operator. Therefore, it has been required to develop a fully automated testing device and reagent that detects genetic material from SARS-CoV-2. The µTASWako g1 system (FUJIFILM Wako Pure Chemical Corporation, Osaka, Japan), a genetic analyzer, provides results in 75 minutes by performing a fully automated PCR process. METHODS: We evaluated the analytical and clinical performance of the µTASWako g1 system for the detection of SARS-CoV-2 RNA. RESULTS: The µTASWako g1 system had the limit of detection at 2,000 copies/mL using a known concentration of RNA. In clinical samples, the µTASWako g1 system had a sensitivity of 88.0% and 100% specificity compared to conventional RT-PCR. The µTAS Wako g1 system could detect three variants of concern carrying spike mutations including N501Y, E484K, and L452R. CONCLUSIONS: As the assay on the µTASWako g1 system is highly accurate for the detection of SARS-CoV-2 regardless of the experience of operator, it can be widely applicable in clinical laboratories.

[Antimicrobial Stewardship and Education, the Role of Pharmacists in the University Hospital].

Over the last decade, many public institutions have emphasized the importance of antimicrobial stewardship (AS) in reducing the spread of antimicrobial resistance. In our facility, we have tackled AS that has adapted to clinical practices since 2015. In many facilities, especially university hospitals, multidrug-resistant bacteria, such as methicillin-resistant Staphylococcus aureus and extended-spectrum ß-lactamase-producing bacteria, may cause infections. Considering this, we recommend prompt treatment with suitable antibiotics and dosage. In AS teams, pharmacists are responsible for administering pharmacotherapy, including optimal dosage for each patient. Therefore, they should assess their patients carefully, implement thorough therapeutic drug monitoring, and utilize the information obtained from these assessments to administer optimal pharmacotherapy. However, optimal pharmacotherapy also requires a correct diagnosis. Although diagnostic stewardship is not a pharmacist's work, it is a great opportunity for pharmacists to learn how expert physicians think. Based on the type of situation above, we train younger pharmacists on the job.

The Use of Fusion Images as a Diagnostic and Neurosurgical Planning Tool in Microvascular Decompression.

Preoperative assessment of surgery using high-quality images can help surgeons to achieve best result of treatment. With the advances in computer technology, interactive multimodality fusion images have been developed. The use of fusion images as a preoperative planning tool is described with its examples in illustrative cases of trigeminal neuralgia and hemifacial spasm microvascular decompression (MVD). Interactive computer graphics such as multimodality fusion method is a useful tool to preoperatively predict the need of bone exposure and configuration of blood vessels with its correlation to cranial nerves in MVD.

Dynamic modelling of cold-hardiness in tea buds by imitating past temperature memory.

BACKGROUND AND AIMS: Most perennial plants memorize cold stress for a certain period and retrieve the memories for cold acclimation and deacclimation, which leads to seasonal changes in cold-hardiness. Therefore, a model for evaluating cold stress memories is required for predicting cold-hardiness and for future frost risk assessments under warming climates. In this study we develop a new dynamic model of cold-hardiness by introducing a function imitating past temperature memory in the processes of cold acclimation and deacclimation. METHODS: We formulated the past temperature memory for plants using thermal time weighted by a forgetting function, and thereby proposed a dynamic model of cold-hardiness. We used the buds of tea plants (Camellia sinensis) from two cultivars, 'Yabukita' and 'Yutakamidori', to calibrate and validate this model based on 10 years of observed cold-hardiness data. KEY RESULTS: The model captured more than 90 % of the observed variation in cold-hardiness and predicted accurate values for both cultivars, with root mean square errors of ~1.0 °C. The optimized forgetting function indicated that the tea buds memorized both short-term (recent days) and long-term (previous months) temperatures. The memories can drive short-term processes such as increasing/decreasing the content of carbohydrates, proteins and antioxidants in the buds, as well as long-term processes such as determining the bud phenological stage, both of which vary with cold-hardiness. CONCLUSIONS: The use of a forgetting function is an effective means of understanding temperature memories in plants and will aid in developing reliable predictions of cold-hardiness for various plant species under global climate warming.

Nationwide surveillance of bacterial respiratory pathogens conducted by the surveillance committee of japanese society of chemotherapy, the japanese association for infectious diseases, and the japanese society for clinical microbiology in 2016: General view of the pathogens' antibacterial susceptibility.

The nationwide surveillance on antimicrobial susceptibility of bacterial respiratory pathogens from the patients in Japan was conducted by the Japanese Society of Chemotherapy, the Japanese Association for Infectious Diseases, and the Japanese Society for Clinical Microbiology in 2016. The isolates were collected from clinical specimens obtained from well-diagnosed adult patients with respiratory tract infections during the period between February 2016 and August 2016 by three societies. Antimicrobial susceptibility testing was conducted at the central reference laboratory according to the method recommended by Clinical Laboratory Standards Institute. Susceptibility testing was evaluated in 1062 strains (143 Staphylococcus aureus, 210 Streptococcus pneumoniae, 17 Streptococcus pyogenes, 248 Haemophilus influenzae, 151 Moraxella catarrhalis, 134 Klebsiella pneumoniae, and 159 Pseudomonas aeruginosa). Ratio of methicillin-resistant S. aureus was 48.3%, and those of penicillin-susceptible S. pneumoniae was 99.5%. Among H. influenzae, 14.1% of them were found to be ß-lactamase-producing ampicillin-resistant strains, and 41.1% to be ß-lactamase-non-producing ampicillin-resistant strains. Extended spectrum ß-lactamase-producing K. pneumoniae and multi-drug resistant P. aeruginosa with metallo ß-lactamase were 4.5% and 0.6%, respectively.

[Frontal Lobe Signs Caused by Migration of a Burr Hole Cap into the Brain after Deep Brain Stimulation Surgery:A Case Report].

We report a rare complication in a patient with Parkinson's disease who underwent deep brain stimulation(DBS)surgery. The patient was a 60-year-old woman who presented with frontal lobe signs, including ataxic gait and memory disturbance, that were caused by the unexpected migration of a burr hole cap into the brain three to four months after surgery. The patient had no incidence of a head injury prior to development of symptoms. The patient underwent surgery to extract the migrated cap from the frontal lobe, and her symptoms improved several months after the operation. The cap serves to fix the DBS lead to the skull using an adjunctive burr hole ring. It was intraoperatively confirmed that only the cap detached from the ring, and no cap or ring defects were detected in a postoperative quality check by the manufacturer. We have previously utilized a burr hole ring and cap, which are packaged along with the DBS electrode, when employing the product made by Medtronic Inc. No previous report has described the cap packed in the official DBS kit to have migrated into the intracranial space. It seems unlikely that the cap migration into the intracranial space would occur without the cap and/or ring breaking through either traumatic injury or from manufacturing defects. It is important to consider the migration of a burr hole cap into the intracranial space in the absence of head injury as a possible device complication after DBS surgery.

Essential human resources for antimicrobial stewardship teams in Japan: Estimates from a nationwide survey conducted by the Japanese Society of Chemotherapy.

Implementation of antimicrobial stewardship programs (ASPs) with multidisciplinary antimicrobial stewardship teams (ASTs) is critical for appropriate antimicrobial use at healthcare facilities. Although the Japanese medical reimbursement system was revised to allow fees for ASP implementation, several concerns remain, including understaffing and enforcement of the recommendations on ASTs and ASPs in practice. Furthermore, there are no recommendations on full-time equivalents (FTEs) of the core members in ASTs in Japan. This committee report presents our recommendations on ASTs based on an analysis of the nationwide survey on implemented ASPs and staff FTEs at 1358 healthcare facilities conducted by the Japanese Society of Chemotherapy. Our report provides a directive for structural and financial support of ASTs and should aid in planning for the enhancement of AST practices and the organization of new ASTs.

The incidence of endophthalmitis or macular involvement and the necessity of a routine ophthalmic examination in patients with candidemia.

BACKGROUND: The incidence of ocular candidiasis (OC) in patients with candidemia varies across different reports, and the issue of whether routine ophthalmoscopy improves outcomes has been raised. This study investigated the incidence of OC and evaluate whether the extent of OC impacts the clinical outcomes. METHODS: This retrospective study included non-neutropenic patients with candidemia who underwent treatment at one of 15 medical centers between 2010 and 2016. Chorioretinitis without other possible causes for the ocular lesions and endophthalmitis was classified as a probable OC. If signs of chorioretinitis were observed in patients with a systemic disease that causes similar ocular lesions, they were classified as a possible OC. RESULTS: In total, 781 of 1089 patients with candidemia underwent an ophthalmic examination. The prevalence of OC was 19.5%. The time from the collection of a positive blood culture to the initial ophthalmic examination was 5.0 ± 3.9 days in patients with OC. The leading isolate was Candida albicans (77.9%). Possible OC was associated with unsuccessful treatments (resolution of ocular findings) (odds ratio: 0.354, 95% confidence interval: 0.141-0.887), indicating an overdiagnosis in patients with a possible OC. If these patients were excluded, the incidence fell to 12.8%. Endophthalmitis and/or macular involvement, both of which require aggressive therapy, were detected in 43.1% of patients; a significantly higher incidence of visual symptoms was observed in these patients. CONCLUSION: Even when early routine ophthalmic examinations were performed, a high incidence of advanced ocular lesions was observed. These results suggest that routine ophthalmic examinations are still warranted in patients with candidemia.

Thermophilic ethanol fermentation from lignocellulose hydrolysate by genetically engineered Moorella thermoacetica.

A transformant of Moorella thermoacetica was constructed for thermophilic ethanol production from lignocellulosic biomass by deleting two phosphotransacetylase genes, pdul1 and pdul2, and introducing the native aldehyde dehydrogenase gene (aldh) controlled by the promoter from glyceraldehyde-3-phosphate dehydrogenase. The transformant showed tolerance to 540mM and fermented sugars including fructose, glucose, galactose and xylose to mainly ethanol. In a mixed-sugar medium of glucose and xylose, all of the sugars were consumed to produce ethanol at the yield of 1.9mol/mol-sugar. The transformant successfully fermented sugars in hydrolysate prepared through the acid hydrolysis of lignocellulose to ethanol, suggesting that this transformant can be used to ferment the sugars in lignocellulosic biomass for ethanol production.

Homolactic Acid Fermentation by the Genetically Engineered Thermophilic Homoacetogen Moorella thermoacetica ATCC 39073.

For the efficient production of target metabolites from carbohydrates, syngas, or H2-CO2 by genetically engineered Moorella thermoacetica, the control of acetate production (a main metabolite of M. thermoacetica) is desired. Although propanediol utilization protein (PduL) was predicted to be a phosphotransacetylase (PTA) involved in acetate production in M. thermoacetica, this has not been confirmed. Our findings described herein directly demonstrate that two putative PduL proteins, encoded by Moth_0864 (pduL1) and Moth_1181 (pduL2), are involved in acetate formation as PTAs. To disrupt these genes, we replaced each gene with a lactate dehydrogenase gene from Thermoanaerobacter pseudethanolicus ATCC 33223 (T-ldh). The acetate production from fructose as the sole carbon source by the pduL1 deletion mutant was not deficient, whereas the disruption of pduL2 significantly decreased the acetate yield to approximately one-third that of the wild-type strain. The double-deletion (both pduL genes) mutant did not produce acetate but produced only lactate as the end product from fructose. These results suggest that both pduL genes are associated with acetate formation via acetyl-coenzyme A (acetyl-CoA) and that their disruption enables a shift in the homoacetic pathway to the genetically synthesized homolactic pathway via pyruvate.IMPORTANCE This is the first report, to our knowledge, on the experimental identification of PTA genes in M. thermoacetica and the shift of the native homoacetic pathway to the genetically synthesized homolactic pathway by their disruption on a sugar platform.

Cerebellopontine angle epidermoid cysts: clinical presentations and surgical outcome.

Epidermoid cysts constitute less than 1% of intracranial tumors with the majority of them involving cerebellopontine angle (CPA). Although several mechanisms for cranial nerve dysfunction due to these tumors have been proposed, no direct evaluation for hyper- or hypoactive dysfunction has been done. In this case series, pathophysiology of cranial nerve dysfunction in CPA epidermoid cysts was evaluated with special attention to a new mechanism of capsule strangulation caused by stratified tumor capsule. Twenty-two cases with epidermoid cysts of CPA micro-neurosurgically treated in our departments since 2005 were reviewed. Clinical status of the patients before the surgery and post-operative functional outcome were recorded. Available data from the English literature were summarized for comparison. Mass reduction of cyst contents in most cases was usually associated with prompt and marked improvement of the symptoms suggesting neuroapraxia caused by compression of the tumor content and/or mild ischemia. Among them, two cases showed strangulation of the affected nerves by the tumor capsule whose preoperative dysfunction did not improve after surgery in spite of meticulous microsurgical removal of the lesion. Involved facial and abducent nerves in these two cases showed distortion of nerve axis and nerve atrophy distal to the strangulation site. We report the first direct evidence of etiology of cranial nerve dysfunction caused by cerebellopontine angle epidermoid tumors. Young age and rapidly progressive neurological deficit might be the characteristics for strangulation of the affected nerves by the cyst capsule. Even though the number of cases might be limited, immediate decompression and release of the strangulating band might be urged in such patients to prevent irreversible deficits.

[A CASE OF PULMONARY TUBERCULOSIS WITH PANCREATIC LESION].

A 52-year-old woman was referred to our hospital presenting with epigastric pain and weight loss. A contrast- enhanced abdominal computed tomography (CT) scan showed a low-density mass in the body of the pancreas, indicative of a malignancy. Endoscopic ultrasound-guided fine needle aspiration of the pancreatic mass was performed three times and showed no specific findings. A distal pancreatectomy was performed, and a pathological examination revealed epitheli- oid cell granulomas and necrosis. Ziehl-Neelsen staining did not reveal acid-fast bacilli in the pancreatic mass. A diagnosis of tuberculosis or sarcoidosis of the pancreas was con- sidered; however, the patient chose to undergo a follow-up examination without therapeutic intervention because the pancreatic mass had been removed completely and she had recovered well. Four months after the operation, the patient was readmitted to our hospital for insulin therapy for pancreatic diabetes. She presented with a fever and a productive cough, and a chest CT scan showed multiple nodules in both upper lobes. A bronchoscopy was performed and bronchoalveolar lavage fluid cultures for Mycobacterium tuberculosis were positive. The patient received antitubercular quadri-therapy and showed symptomatic and radiologic improvement. At the initial examination, we had been unable to establish the correct diagnosis; however, the detection of pulmonary lesions led to the time-delayed diagnosis of pancreatic tuber- culosis. Owing to its rarity, it is difficult to diagnose pancreatic tuberculosis using clinical symptoms and radiological imaging modalities; thus, pathologic and bacteriologic confirmation is essential. To avoid performing an unnecessary laparotomy in patients with pancreatic tuberculosis, increased vigilance and an accurate diagnostic approach are required.