Spodiobacter cordis gen. nov. sp. nov., a member of the family Flavobacteriaceae isolated from patients with infective endocarditis.

Two gram-negative, catalase-negative, oxidase-positive strains (PAGU 1467T and PAGU 1468) isolated from patients with infective endocarditis were investigated to determine their taxonomic status. 16S rRNA gene sequence analysis indicated that the two strains were members of the Bergeyella-Chryseobacterium-Riemerella branch of the family Flavobacteriaceae. Strains PAGU 1467T and PAGU 1468 were highly related to each other (98.8% 16S rRNA gene sequence similarity). Phylogenetically closely-related species to PAGU 1467T comprised Bergeyella zoohelcum (95.0% 16S rRNA gene sequence similarity), Riemerella anatipestifer (94.3%) and Cloacibacterium normanense (94.3%). The major fatty acids of the two isolates were iso-C15:0 , iso-C17:0 3-OH and iso-C15:0 3-OH. The presence of C16:0 3-OH and iso-C15:0 2-OH allowed these isolates to be distinguished from B. zoohelcum. Menaquinone MK-6 was the only respiratory quinone in these organisms; this is a consistent characteristic of the family Flavobacteriaceae. The guanine-plus-cytosine content of the genomic DNA was 42.0%, which is higher than that of other close phylogenetic relatives. On the basis of their phenotypic properties and genetic distinctiveness, isolates PAGU 1467T and PAGU 1468 were classified within the novel genus Spodiobacter, as Spodiobacter cordis gen. nov., sp. nov., which is also the type species. The type strain of S. cordis is PAGU 1467T ( = CCUG 65564T = NBRC 109998T ).

Comparison of fluorescence optical imaging, ultrasonography and clinical examination with magnetic resonance imaging as a reference in active rheumatoid arthritis patients.

BACKGROUND: Fluorescence optical imaging with indocyanine-green enhancement (FOI) is a new imaging modality for the assessment of hand arthritis. The objective of this study was to compare performance profiles of clinical examination (CE), US and FOI using MRI as a reference in the same active rheumatoid arthritis (RA) patients. METHODS: CE, US, FOI and MRI were performed on six subjects with active RA. Each sequence of FOI was divided into three phases based on indocyanine-green dynamics and the joints were graded semi-quantitatively. Sensitivities and specificities of CE, US and FOI were calculated using the RAMRIS synovitis score >0 as a reference in a total of 30 joints (the second to fifth metacarpophalangeal (MCP) joints and the wrist of the clinically dominant hand). RESULTS: FOI showed sensitivities and specificities, respectively, of 85% and of 94% for Phase-1 and 69% and 94% for Phase-2. Sensitivities and specificities were 100% and 35% for CE (tender or swollen), 92% and 41% for gray scale US, and 77% and 100% for color-Doppler US. CONCLUSIONS: The performance characteristics of FOI in detection of synovitis in patients with active RA are comparable to those of US and more specific than CE. FOI has a potential as an assessment modality of RA.

Serial measurement of Wisteria floribunda agglutinin positive Mac-2-binding protein is useful for predicting liver fibrosis and the development of hepatocellular carcinoma in chronic hepatitis C patients treated with IFN-based and IFN-free therapy.

AIM: Wisteria floribunda agglutinin positive (WFA+) Mac-2-binding protein (M2BPGi) is a noninvasive glyco-marker for liver fibrosis. This study evaluated the utility of serial measurement of serum M2BPGi and total M2BP as a predictor of fibrosis and the development of hepatocellular carcinoma (HCC). METHODS: This study included 119 patients with chronic hepatitis C (CHC). Of these patients, 97 were treated with IFN-based therapy and 22 were treated with daclatasvir and asunaprevir. Serum M2BPGi values were measured prior to, at the end of, and at 24 weeks after the completion of treatment. As subanalysis, serum total M2BP levels were measured in patients treated with pegylated-interferon and ribavirin. RESULTS: In patients treated with IFN-based therapy, M2BPGi levels were elevated at the end of treatment but decreased afterwards. In contrast, M2BPGi levels in patients treated with IFN-free therapy decreased immediately after starting the treatment without transient elevation. Though pre-treatment M2BPGi levels significantly correlated with fibrosis in both patients with a sustained virological response (SVR) and non-SVR, post-treatment M2BPGi levels decreased regardless of the degree of fibrosis in patients with SVR. In multivariate analysis, non-SVR and HCC development were independent factors associated with M2BPGi level ≥2.2. In patients treated with pegylated-interferon and ribavirin, total M2BP levels were positively correlated with fibrosis and HCC development. CONCLUSION: Real-time monitoring of the serum M2BPGi level after antiviral therapy for CHC patients could be a helpful screening tool for assessing the risk of HCC. M2BP and its glycan structure could be associated together with hepatocarcinogenesis.

A case of gray platelet syndrome masked by immune thrombocytopenia at presentation.

We report a case of gray platelet syndrome (GPS) associated with immune thrombocytopenia (ITP) at presentation. A 22-year-old male patient presenting with petechiae on his limbs was diagnosed with ITP due to a gradual decrease of his platelet count to a minimum of 26 × 10(9) /liter and an elevated platelet-associated IgG (PA-IgG) level in the absence of any other specific cause of thrombocytopenia. Administration of prednisolone increased his platelet count, but this dropped again to approximately 50 × 10(9) /liter as the dose was tapered, and remained at the same level after the treatment was terminated. Thirteen years later, we reassessed the cause of the thrombocytopenia because the PA-IgG level was found to be within the normal range. There were large hypogranular platelets on the blood film and a deficit of α-granules in the platelets on electron microscopy. On this basis, we diagnosed his thrombocytopenia as GPS. To our knowledge, this is the first report of a GPS case associated with ITP at presentation. This case illustrates the importance of carefully reviewing blood film results in the differential diagnosis of thrombocytopenia.

Catheter-related bloodstream infection by Tsukamurella inchonensis in an immunocompromised patient.

We report a case of catheter-related bloodstream infection by Tsukamurella inchonensis, identified using 16S rRNA gene sequencing, in a patient with myelofibrosis who underwent a bone marrow transplant. Tsukamurella species infections are rare. To our knowledge, this is the first case of T. inchonensis bloodstream infection in an immunocompromised patient.

First report of KPC-2 Carbapenemase-producing Klebsiella pneumoniae in Japan.

We investigated a novel Japanese isolate of sequence type 11 (ST11), the Klebsiella pneumoniae carbapenemase-2 (KPC-2)-producing K. pneumoniae strain Kp3018, which was previously obtained from a patient treated at a Brazilian hospital. This strain was resistant to various antibiotic classes, including carbapenems, and harbored the gene blaKPC-2, which was present on the transferable plasmid of ca. 190 kb, in addition to the blaCTX-M-15 gene. Furthermore, the ca. 2.3-kb sequences (ISKpn8-blaKPC-2-ISKpn6-like), encompassing blaKPC-2, were found to be similar to those of K. pneumoniae strains from China.

[Pulmonary function testing: history up to the present].

The main roles of pulmonary function testing are: aiding in the diagnosis, assessing disease severity, and monitoring the progress and effects of treatment. The spirometric measurements of highest clinical value are the vital capacity(VC) and forced expiratory volume in one second(FEV1). Measurements of VC were reported in both healthy subjects and patients with respiratory disease during the mid-19th century. The VC, however, was little used in clinical medicine until the second half of the 20th century. The FEV1, FEV1/VC ratio, and/or FEV1/forced vital capacity (FVC) ratio all came into existence after 1950. Pulmonary function testing evolved over time into a more complex set of lung function measurements using various techniques and devices. If the variability of the results can be minimized and the measurement accuracy can be improved, abnormalities will be more easily detected. In 2005, the American Thoracic Society (ATS) and European Respiratory Society (ERS) together published an updated set of standards for pulmonary function testing that could be applied more widely. These standards are structured to cover definitions, equipment, and patient-related procedures commonly applied for many methods of lung function testing. In 2004, the Japanese Respiratory Society issued an initial set of recommendations for the standardized measurement of VC, FVC, and the single-breath carbon monoxide-diffusing capacity. These recommendations have not been updated, and the prediction equations for pulmonary function testing have not been fully established. The statements need to be periodically updated in accordance with new developments. (Review).

Flow cytometric analysis of Notch1 and Jagged1 expression in normal blood cells and leukemia cells.

Notch1 and its ligand Jagged1 are proteins with important roles in the growth of leukemia cells. Although the detection of Notch1 protein in acute lymphoblastic leukemia cells using immunoblot analysis has been previously reported, the expression patterns of Notch1 and Jagged1 detected by flow cytometry (FCM) in normal blood cells and various leukemia cells have not been well-characterised. In the present study, we examined the expression patterns of Notch1 and Jagged1 in 10 normal blood samples, 8 bone marrow samples, 11 leukemia/lymphoma cell lines and leukemia cells from 22 patients with acute myeloid leukemia (AML), mature T-cell neoplasms or B-cell chronic lymphocytic leukemia (B-CLL) using FCM. The results showed that Notch1 expression is relatively strong in monocytes and granulocytes but weak in lymphocytes. The expression of Notch1 is stronger in bone marrow cells than in the equivalent cells in blood. All the cell lines examined strongly expressed Notch1, and eight cell lines expressed Jagged1. In leukemia cells from patients, four AML samples expressed Notch1 and/or Jagged1. However, three samples expressed neither Notch1 and/or Jagged1 and none of the mature T-cell neoplasm samples expressed either protein. However, all B-CLL samples expressed high levels of both Notch1 and Jagged1. We found that the expression of Notch1 and Jagged1 is detected in various hematological malignancies by FCM. The examination of these proteins is likely to be useful in the characterisation of diseases and individual cases. Examination of these proteins may also be useful in the selection of patients most likely to benefit from novel molecular-targeted therapies using Notch inhibitors in the future.

[Pulmonary function testing in Japan: present status and new developments].

In 2004, the Japanese Respiratory Society issued an initial set of recommendations on the standardized measurement of the most frequently used tests for pulmonary function, i.e., tests to assess slow vital capacity, forced vital capacity, and single-breath carbon monoxide diffusing capacity. This statement has not been updated, and the prediction equations for pulmonary function testing are not fully established. Thus, the guidelines will need to be periodically updated in accordance with new developments in this rapidly evolving field. Nitric oxide (NO) is now recognized as a biological mediator in animals and humans. The human lung produces NO and exhales it in breath. The fractional nitric oxide (NO) concentration in exhaled breath (FE(NO)) can be quantitatively measured by a simple, safe, and noninvasive procedure as a complementary tool for assessing airway inflammation in airway diseases such as asthma. While the measurement of exhaled NO is standardized for clinical use, FE(NO) measurement is not approved or covered under the public health insurance system in Japan.

First report of sepsis caused by Rhodococcus corynebacterioides in a patient with myelodysplastic syndrome.

We report a case of sepsis caused by Rhodococcus corynebacterioides, identified using 16S rRNA gene sequencing, in a myelodysplastic syndrome patient who had undergone hematopoietic stem cell transplantation. This is the first report of R. corynebacterioides infection in a human.

Point-of-care testing system enabling 30 min detection of influenza genes.

We developed a portable and easy-to-use nucleic acid amplification test (NAT) system for use in point-of-care testing (POCT). The system shows sensitivity that is sufficiently higher than that of the currently available rapid diagnostic kit and is comparable to that of real-time reverse transcription polymerase chain reaction (RT-PCR) for influenza testing.

Symptoms of atrial fibrillation in patients with and without subsequent permanent atrial fibrillation based on a retrospective questionnaire survey.

The aim of the present study was to determine whether symptoms of atrial fibrillation (AF) differ between patients with and without subsequent permanent AF. Sixty-four patients (68 +/- 10 years old, 45 males) were recruited. AF follow-up was started at the age of 61 +/- 10 years and accomplished in a median period of 4.9 years (396 person-years). Permanent AF, defined as lasting > 180 days, developed in 17 patients (14 males) (43 per 1000 person-years). The AF follow-up period was longer in the permanent AF group than in the non-permanent AF group (median, 9.8 versus 4.2 years, P < 0.001). For baseline characteristics, hypertension was less frequent in the permanent AF group than in the nonpermanent AF group (18% versus 45%, P < 0.05). A retrospective questionnaire survey regarding initial AF symptoms was conducted. The severity of AF symptoms by a 4-grade scale was significantly milder in the permanent AF group than in the nonpermanent AF group (P < 0.05). Cox proportional hazards model analysis revealed that the severity of initial AF symptoms was related to the subsequent development of permanent AF (hazard ratio 0.46 per grade, 95% confidence interval 0.23 - 0.93, P < 0.05), but age, gender, hypertension, diabetes mellitus, organic heart disease, and left atrial dimension were not. The permanent AF-free rate was significantly lower in 33 patients with mild symptoms than in 31 patients with severe symptoms (log-rank test, P < 0.05). These results point to an inconspicuous feature in the development of permanent AF.

[False-positive reaction of urinary ketone bodies caused by bucillamine].

In many patients taking bucillamine, an anti-rheumatic drug, urinary examinations show false-positive findings for ketone on urine dipsticks, because of the SH radicals in bucillamine. Our group examined the frequency of false-positive ketone testing and the causative factors in 179 outpatients being treated by the Department of Rheumatology in our hospital. The samples that remained purple even after boiling were determined to be false-positives, because boiling volatilizes urinary ketones while leaving bucillamine unchanged. Forty-six of 49 patients taking bucillamine tested false-positive for ketone, whereas there were no false-positive reactions among patients not receiving bucillamine. The patients showing false-positive reactions had significantly lower serum albumin levels, higher specific gravity of urine and more acidic urine. Moreover, we could predict whether the reaction was true-positive or bucillamine-induced false-positive by observing differences between the colors and levels of discoloration of the urine test paper. In the evaluation of ketone findings on urinary dipstick, the physician must ascertain whether the patient is taking bucillamine.

Recombinant human fibrinogen expressed in the yeast Pichia pastoris was assembled and biologically active.

Fibrinogen is a large plasma glycoprotein with a molecular mass of 340kDa that plays a critical role in the final stage of blood coagulation. Human plasma fibrinogen is a dimeric molecule comprising two sets of three different polypeptides (Aalpha, 66kDa; Bbeta, 55kDa; gamma, 48kDa). To express recombinant human fibrinogen in the methylotrophic yeast Pichia pastoris, we constructed an expression vector containing three individual fibrinogen chain cDNAs under the control of the mutated AOX2 (mAOX2) promoter. First, P. pastoris GTS115 was transformed with the vector, but the expressed recombinant fibrinogen suffered severe degradation by yeast-derived proteases under conventional nutrient culture conditions. Fibrinogen degradation was prevented by using the protease A-deficient strain SMD1168 as a host strain and regulating the pH of the culture to between 5.5 and 7.0. Western blot analysis revealed that the Aalpha, Bbeta and gamma chains of recombinant fibrinogen were assembled and secreted as a complete molecule. The Bbeta chain of the recombinant fibrinogen was N-glycosylated but the Aalpha chain, as in plasma fibrinogen, was not. The gamma chains however were heterologous, one being N-glycosylated and the other not. The recombinant fibrinogen was capable of forming a thrombin-induced clot in the presence of factor XIIIa and both the glycosylated and the non-glycosylated gamma chains were involved in the formation of cross-linking fibrin. The present study indicates that the recombinant fibrinogen expressed in P. pastoris, although different from plasma fibrinogen in post-translational modification, is correctly assembled and biologically active.

[Problems of lung function testing in the laboratory].

Spirometry is indispensable for the screening test of general respiratory function, and measurements of lung volume and diffusing capacity play an important role in the assessment of disease severity, functional disability, disease activity and response to treatment. Pulmonary function testing requires cooperation between the subjects and the examiner, and the results obtained depend on technical as well as personal factors. In order to diminish the variability of results and improve measurement accuracy, the Japan Respiratory Society published the first guidelines on the standardization of spirometry and diffusing capacity for both technical and clinical staff in 2004. It is therefore essential to distribute the guidelines to both laboratory personnel and general physicians. Furthermore, training workshops are mandatory to improve their understanding of the basics of lung function testing. Recently, there has been increasing interest in noninvasive methods of lung function testing without requiring the patient's cooperation during spontaneous breathing. Three alternative techniques, i.e. the negative expiratory pressure (NEP) method to detect expiratory flow limitation, impulse oscillation system (IOS) to measure respiratory system resistance (Rrs) and reactance (Xrs), and interruption resistance (Rint) to measure respiratory resistance have been introduced. Further study is required to determine the advantage of these methods.

Pulmonary exercise testing predicts prognosis in patients with chronic obstructive pulmonary disease.

OBJECTIVE: In patients with chronic obstructive pulmonary disease (COPD), patient age and initial value of forced expiratory volume in 1 second (FEV1) have been considered the most accurate predictors of mortality among the parameters obtained from pulmonary exercise tests. However, few studies have examined the predictive variables of prognosis among exercise parameters in COPD. We therefore attempted to identify the best index for predicting long-term survival in patients with COPD among the cardiopulmonary variables obtained during exercise testing. PATIENTS AND METHODS: Fifty-eight patients with COPD (50 men and 8 women) without hypoxemia at rest or other serious complications performed resting pulmonary function tests followed by a symptom-limited ramp exercise test on a cycle ergometer with breath-by-breath gas analysis and arterial blood gas sampling. RESULTS: After 3,570+/-1,373 days follow-up (mean+/-SD), 21 died because of deaths by respiratory failure. The overall survival rates calculated by the Kaplan-Meier method were 92.9% and 75.8% at 5 years and 10 years, respectively. In univariate Cox hazards analysis, age, FEV1, VC, RV/TLC, VEmax, VO2max, VCO2max, PaO2max, PacO2max, and PaO2 at rest were found to be significant prognostic indices of survival. However, multivariate analysis revealed only FEV1, PaO2max, and age as independent predictors of mortality. In severe COPD patients (FEV1 <50% predicted, n=35), PaO2max and age also correlated with prognosis, whereas FEV1 did not. CONCLUSION: Pulmonary exercise testing is useful in predicting prognosis in patients with COPD.

[Lung function testing --the Official Guideline of the Japanese Respiratory Society].

This review article introduces the official guideline for the lung function testing firstly established by the Japanese Respiratory Society in November 2004. The members of the Japanese Society of Laboratory Medicine were also included in the working group to make the guideline. A central goal of the guideline is to standardize the skill of lung function testing and thus minimize its variability. The guideline includes the chapters for spirometry, flow-volume curve, and diffusing capacity of the lung, where principles and quality controls are introduced with reference values for each testing. Furthermore, the algorithm for differential diagnosis using the lung function testing is demonstrated in the last chapter. The lung function testing can bring a strong impact on patients' lifestyle and future treatment plan. We sincerely hope that this guideline will contribute to routine laboratory practice.

[The rate of airflow limitation among the elderly undergoing spirometry in a university hospital].

The aim of this study was to examine the rate of airflow limitation among the elderly undergoing spirometry prior to surgical operation, and to evaluate whether or not spirometry is useful for the early detection of COPD. A total of 3,086 patients (1,717 men and 1,369 women) ranging in age from 40 to 93 underwent screening spirometry from January to December 2003 in Tokyo Medical and Dental University Hospital. Among the total population, 2,135 patients (1,188 men and 947 women) were ordered to undergo the spirometry prior to surgical operation, airflow limitation (FEV1/FVC < 70%) was observed in 19.5% (n=417) of the cases and was considerably more frequent in men than in women(27.0%[n=321] vs. 10.1%[n=96]). The rate of FEV1/FVC < 70% in those 2,135 patients increased with age: 6.2%, 9.9%, 19.2%, 32.5% and 34.1% of patients in their 40s, 50s, 60s, 70s, 80s and over, respectively. Among the patients with airflow limitation, 35% of the cases revealed FEV1 > or = 80% predicted; 50%, 50 < or = FEV1 < 80% predicted; 14%, 30 < or = FEV1 < 50% predicted; 1%, FEV1 < 30% predicted. Patients with mild to moderate airflow limitation rarely consulted the pulmonary medicine department. The prevalence of airflow limitation was more frequent among in- and out-patients than in the random sample population. Spirometry prior to surgical operation is useful for the early detection of COPD and these data could be an important source for medical staff seeking a definitive diagnosis of patients revealing airflow limitation.