Advantages of genome sequencing by long-read sequencer using SMRT technology in medical area

PacBio RS II is the first commercialized third-generation DNA sequencer able to sequence a single molecule DNA in real-time without amplification. PacBio RS II’s sequencing technology is novel and unique, enabling the direct observation of DNA synthesis by DNA polymerase. PacBio RS II confers four major advantages compared to other sequencing technologies: long read lengths, high consensus accuracy, a low degree of bias, and simultaneous capability of epigenetic characterization. These advantages surmount the obstacle of sequencing genomic regions such as high/low G+C, tandem repeat, and interspersed repeat regions. Moreover, PacBio RS II is ideal for whole genome sequencing, targeted sequencing, complex population analysis, RNA sequencing, and epigenetics characterization. With PacBio RS II, we have sequenced and analyzed the genomes of many species, from viruses to humans. Herein, we summarize and review some of our key genome sequencing projects, including full-length viral sequencing, complete bacterial genome and almost-complete plant genome assemblies, and long amplicon sequencing of a disease-associated gene region. We believe that PacBio RS II is not only an effective tool for use in the basic biological sciences but also in the medical/clinical setting.     

Partial purification and characterization of taurodeoxycholate 7α-monooxygenase

Taurodeoxycholate 7α-monooxygenase was partially purified from rat liver microsomes. The enzyme was solubilized with cholate, fractionated with polyethylene glycol and chromatographed on a Sepharose 4B column with cholate as ligand. The enzyme activity was eluted from the column into the fraction eluted with 50 mM phosphate buffer containing cholate and KCl, whereas the benzphetamine demethylase activity was eluted in the non-bound fraction. Thus it was established that both enzymes are different entities. The taurodeoxycholate 7α-monooxygenase activity was reconstituted from the partially purified cytochrome P-450, highly purified NADPH-cytochrome P-450 reductase, dilauroylglyceryl-3-phosphorylcholine and NADPH.     

Cloning, nucleotide sequences, and enzymatic properties of glucose dehydrogenase isozymes from Bacillus megaterium IAM1030.

Bacillus megaterium is known to have several genes that code for isozymes of glucose dehydrogenase. Two of them, gdhI and gdhII, were cloned from B. megaterium IAM1030 in our previous work (T. Mitamura, R. V. Evora, T. Nakai, Y. Makino, S. Negoro, I. Urabe, and H. Okada, J. Ferment. Bioeng. 70:363-369, 1990). In the present study, two new genes, gdhIII and gdhIV, were isolated from the same strain and their nucleotide sequences were identified. Each gene has an open reading frame of 783 bp available to encode a peptide of 261 amino acids. Thus, a total of four glucose dehydrogenase genes have been cloned from B. megaterium IAM1030. In addition, this strain does not seem to have other glucose dehydrogenase genes that can be distinguished from the four cloned genes so far examined by Southern hybridization analysis. The two newly cloned genes were expressed in Escherichia coli cells, and the products, GlcDH-III and GlcDH-IV, were purified and characterized and compared with the other isozymes, GlcDH-I and GlcDH-II, encoded by gdhI and gdhII, respectively. These isozymes showed different mobilities in sodium dodecyl sulfate-polyacrylamide gel electrophoresis (GlcDH-I greater than GlcDH-III = GlcDH-IV greater than GlcDH-II), although they have the same number of amino acid residues. Double-immunodiffusion tests showed that GlcDH-I is immunologically different from the other isozymes and that GlcDH-III and GlcDH-IV are identical to one another but a little different from GlcDH-II. These glucose dehydrogenases were stabilized in the presence of 2 M NaCl. The effect of NaCl was especially large for GlcDH-III, which is most unstable enzyme. Kinetic studies showed that these isozymes are divided into two groups with respect to coenzyme specificity, although they can utilize both NAD and NADP: GlcDH-III and GlcDH-IV prefer NAD, and GlcDH-I and GlcDH-II prefer NADP. The phylogenic relationship of these glucose dehydrogenase genes is also discussed.     

Effectiveness of Trivalent Inactivated Influenza Vaccine in Children Estimated by a Test-Negative Case-Control Design Study Based on Influenza Rapid Diagnostic Test Results

We assessed vaccine effectiveness (VE) against medically attended, laboratory-confirmed influenza in children 6 months to 15 years of age in 22 hospitals in Japan during the 2013–14 season. Our study was conducted according to a test-negative case-control design based on influenza rapid diagnostic test (IRDT) results. Outpatients who came to our clinics with a fever of 38°C or over and had undergone an IRDT were enrolled in this study. Patients with positive IRDT results were recorded as cases, and patients with negative results were recorded as controls. Between November 2013 and March 2014, a total of 4727 pediatric patients (6 months to 15 years of age) were enrolled: 876 were positive for influenza A, 66 for A(H1N1)pdm09 and in the other 810 the subtype was unknown; 1405 were positive for influenza B; and 2445 were negative for influenza. Overall VE was 46% (95% confidence interval [CI], 39–52). Adjusted VE against influenza A, influenza A(H1N1)pdm09, and influenza B was 63% (95% CI, 56–69), 77% (95% CI, 59–87), and 26% (95% CI, 14–36), respectively. Influenza vaccine was not effective against either influenza A or influenza B in infants 6 to 11 months of age. Two doses of influenza vaccine provided better protection against influenza A infection than a single dose did. VE against hospitalization influenza A infection was 76%. Influenza vaccine was effective against influenza A, especially against influenza A(H1N1)pdm09, but was much less effective against influenza B.     

Phosphorus release and sedimentation in three contiguous shallow brackish lakes, as estimated from changes in phosphorus stock and loading from catchment

In situ phosphorus release rates in three contiguous shallow brackish lakes were calculated by considering the amount of water inflow, changes in salinity and phosphorus stock, and loading from phosphorus inflow based on monthly data. The annual amount of sedimental phosphorus relative to that of phosphorus inflow was different for each of the three water bodies: 16% for Lake Shinji, 3% for the Honjo area, and −8% for Lake Nakaumi, as estimated in a 10-year period from January 1993 to December 2002. During the warm season, the quantity of phosphorus released surpassed sedimentation in these three water bodies. The low annual sedimentation ratio in Lake Nakaumi is related to a large seawater backflow resulting in phosphorus removal, in addition to a stable stratified structure promoting phosphorus release from sediment due to oxygen depletion in the lower layer. In Lake Nakaumi, field data shows that if dissolved oxygen at the sediment surface falls below 2.54 mg L⁻¹, phosphorus release from the sediment begins to be accelerated.     

River to river: First evidence of eel movement between distant rivers via the sea

Environmental Biology of Fishes - Eel movement patterns have been frequently studied to learn about their movements within the fresh- and brackish waters of the same river before their spawning...     

Deficiency of neural recognition molecule NB-2 affects the development of glutamatergic auditory pathways from the ventral cochlear nucleus to the superior olivary complex in mouse

Neural recognition molecule NB-2/contactin 5 is expressed transiently during the first postnatal week in glutamatergic neurons of the central auditory system. Here, we investigated the effect of NB-2 deficiency on the auditory brainstem in mouse. While almost all principal neurons are wrapped with the calyces of Held in the medial nucleus of the trapezoid body (MNTB) in wild type, 8% of principal neurons in NB-2 knockout (KO) mice lack the calyces of Held at postnatal day (P) 6. At P10 and P15, apoptotic principal neurons were detected in NB-2 KO mice, but not in wild type. Apoptotic cells were also increased in the ventral cochlear nucleus (VCN) of NB-2 KO mice, which contains bushy neurons projecting to the MNTB and the lateral superior olive (LSO). At the age of 1 month, the number of principal neurons in the MNTB and of glutamatergic synapses in the LSO was reduced in NB-2 KO mice. Finally, interpeak latencies for auditory brainstem response waves II-III and III-IV were significantly increased in NB-2 KO mice. Together, these findings suggest that NB-2 deficiency causes a deficit in synapse formation and then induces apoptosis in MNTB and VCN neurons, affecting auditory brainstem function.     

韩国华津浦湖的鱼类区系

There are sevenlagoons onthe east coast of Korea,including Kyeongpo Lake,Hyang Lake,Mae Lake,Chung-cho Lake,Yeoungrang Lake,Songji Lake and HwajinpoLake.They are all originally natural and brackish waterlakes.Therefore,these lakes are not only historically veryimportant but also have high values for geological,andecological sciences.Natural lakes like lagoons are different from man-made lakes.That is,water level in natural lakes is almostconstant,cline by waterside is not much,and wet la…     

Trophic position and growth stages of Caddisfly (Stenopsyche marmorata Navas) larvae in the Echi River, Japan

We investigated the trophic position of the larval filter-feeding caddisfly species—Stenopsyche marmorata Navas in the Echi River (Japan) using δ¹³C and δ¹⁵N measurements by instar growth stage. We also analyzed δ¹³C and δ¹⁵N of food sources (periphyton and suspended and benthic POM) along the course of the river. The larvae showed some degree of spatial variation in δ¹³C and δ¹⁵N values with a downstream increase in δ¹³C and δ¹⁵N values. Our results demonstrate that the trophic structure of S. marmorata of different larval stages were spatially varying, depending on food sources. Small-sized instars displayed a higher trophic position than larger instars, confirming that their non-selective feeding behavior leads to consumption of items with high δ¹⁵N values according to the spatial distribution of food sources in the riverine system.     

Monitoring Escape and Feeding Behaviours of Cruiser Fish by Inertial and Magnetic Sensors

A method was developed and applied for monitoring two types of fast-start locomotion (feeding and escape) of a cruiser fish, Japanese amberjacks Seriola quinqueradiata. A data logger, which incorporated a 3-axis gyroscope, a 3-axis accelerometer and a 3-axis magnetometer, was attached to the five fish. The escape, feeding and routine movements of the fish, which were triggered in tank experiments, were then recorded by the data logger and video cameras. The locomotor variables, calculated based on the high resolution measurements by the data logger (500 Hz), were investigated to accurately detect and classify the types of fast-track behaviour. The results show that fast-start locomotion can be detected with a high precision (0.97) and recall rate (0.96) from the routine movements. Two types of fast-start movements were classified with high accuracy (0.84). Accuracy was greater if the data were obtained from the data logger, which combined an accelerometer, a gyroscope and a magnetometer, than if only an accelerometer (0.80) or a gyroscope (0.66) was used.