ABSTRACTFunctional analysis ofBifidobacteriumgenes is essential for understanding host-Bifidobacteriuminteractions with beneficial effects on human health; however, the lack of an effective targeted gene inactivation system in bifidobacteria has prevented the development of functional genomics in this bacterium. Here, we report the development of a markerless gene deletion system involving a double crossover inBifidobacterium longum. Incompatible plasmid vectors were used to facilitate a second crossover step. The conditional replication vector pBS423-ΔrepA, which lacks the plasmid replication generepA, was integrated into the target gene by a first crossover event. Subsequently, the replicative plasmid pTBR101-CM, which harborsrepA, was introduced into this integrant to facilitate the second crossover step and subsequent elimination of the excised conditional replication vector from the cells by plasmid incompatibility. The proposed system was confirmed to work as expected inB. longum105-A using the chromosomal full-length β-galactosidase gene as a target. Markerless gene deletion was tested using theagagene, which encodes α-galactosidase, whose substrates include raffinose. Almost all the pTBR101-CM-transformed strains became double-crossover recombinants after subculture, and 4 out of the 270 double-crossover recombinants had lost the ability to assimilate raffinose. Genotype analysis of these strains revealed markerless gene deletion ofaga. Carbohydrate assimilation analysis and α-galactosidase activity measurement were conducted using both the representative mutant and a plasmid-basedaga-complemented strain. These functional analyses revealed thatagais the only gene encoding a functional α-galactosidase enzyme inB. longum105-A.

It is difficult to determine adverse drug reactions (ADRs) in daily complicated clinical practice in which many kinds of drugs are prescribed. We evaluated how well the Naranjo Algorithm (NA) categorized ADRs among suspected ADRs. The Japan Adverse Drug Events (JADE) study was a prospective cohort study of 3459 inpatients. After all suspected ADRs were reported from research assistants, a single physician reviewer independently assigned an NA score to each. After all NA score of suspected ADRs were scored, two physician reviewers discussed and determined ADRs based on the literature. We investigated the sensitivity and specificity of NA and each component to categorize ADRs among suspected ADRs. A total of 1579 suspected ADRs were reported in 962 patients. Physician reviewers determined 997 ADRs. The percentage of ADRs was 94% if the total NA score reached 5. The modified NA consisted of 5 components that showed high classification abilities; its area under the curve (AUC) was 0.92 for categorizing ADRs, the same as the original. When we set the total NA score cut‐off value to 5, specificity was 0.95 and sensitivity was 0.59. When we reclassified NA components as binary variables, the specificity increased to 0.98 with a cut‐off value of 4 and yielded an AUC of 0.93. In conclusion, we showed that both NA and modified NA could categorize ADRs among suspected ADRs with a high likelihood in daily clinical practice.

BackgroundGuidelines for Type 2 diabetes mellitus (T2DM) management in Japan provide physicians the discretion to select treatment options based on patient pathophysiology of the disease. There exists a wide variation of preference for initial antidiabetes drugs (AD). The current database analysis aimed to understand the real world treatment patterns in drug-naive patients with T2DM in Japan.MethodsWe analyzed data of patients (≥ 18 years) diagnosed with T2DM between October 2012 and September 2016 from the Medical Data Vision, a Diagnosis Procedure Combination database. The primary objective was to determine the proportion of T2DM patients receiving each type of treatment as first-line therapy among the drug-naive cohort.ResultsOf the 436,546 drug-naive patients, 224,761 received their first-line T2DM treatment in the outpatient setting. The mean age of the patient population was 65.6 years at index date. Dipeptidyl peptidase-4 (DPP-4) inhibitor was the most prescribed (56.8%) outpatient AD monotherapy, followed by metformin (15.4%). DPP-4 inhibitors were prescribed over metformin in patients with renal disease (odds ratio [OR]: 4.20; p < 0.0001), coronary heart disease and stroke (OR: 2.22; p < 0.0001). Male (OR: 1.03; p = 0.0026), presence of diabetic complications [retinopathy (OR: 1.33; p < 0.0001), neuropathy (OR: 1.05; p = 0.0037), nephropathy (OR: 1.08; p < 0.0001)] and a high baseline HbA1c (OR: 1.45; p < 0.0001) received treatment intensification during 180 days.ConclusionDPP-4 inhibitors were the most prevalent first-line T2DM treatment followed by metformin in Japan. The findings from this retrospective analysis also support the previously published web survey results and can help understand the real world utilization of T2DM treatment.Trial registration Retrospectively registered

Epicardial leads provide a reliable technique for managing rhythmic disturbance problems in the pediatric population. The only significant predictor of lead failure is the presence of structural heart disease.

The high-resolution soft x-ray photoelectron study of Heusler͑L2 1 ͒-type ͑Fe 2/3 V 1/3 ͒ 100−y Al y ͑y = 23.8− 25.8͒ alloys directly confirms the correlation between their thermoelectric properties and the electronic structures near the Fermi level. It shows the rigid-band-like shift of the main 3d bands and the increase of the photoelectron intensity at the Fermi edge with the small deviation of the Al content from stoichiometry, i.e., y = 25.0. This implies the existence of the pseudogap across the Fermi level in Fe 2 VAl. The Seebeck coefficients evaluated from the present spectroscopic data agree well with the experimental ones as well as the values estimated from a theoretical density of states in a rigid-band model, while the shift is much smaller than what is expected in the rigid-band model. The small modification of the electronic structure in the offstoichiometric ͑Fe 2/3 V 1/3 ͒ 100−y Al y alloys may be attributed to the lower contribution of Al to the electronic states near E F .