Comparison of Database Search Methods for the Detection of Legionella pneumophila in Water Samples Using Metagenomic Analysis

Metagenomic analysis has become a powerful tool to analyze bacterial communities in environmental samples. However, the detection of a specific bacterial species using metagenomic analysis remains difficult due to false positive detections of sequences shared between different bacterial species. In this study, 16S rRNA amplicon and shotgun metagenomic analyses were conducted on samples collected along a stream and ponds in the campus of Hokkaido University. We compared different database search methods for bacterial detection by focusing on Legionella pneumophila. In this study, we used L. pneumophila-specific nested PCR as a gold standard to evaluate the results of the metagenomic analysis. Comparison with the results from L. pneumophila-specific nested PCR indicated that a blastn search of shotgun reads against the NCBI-NT database led to false positive results and had problems with specificity. We also found that a blastn search of shotgun reads against a database of the catalase-peroxidase (katB) gene detected L. pneumophila with the highest area under the receiver operating characteristic curve among the tested search methods; indicating that a blastn search against the katB gene database had better diagnostic ability than searches against other databases. Our results suggest that sequence searches targeting long genes specifically associated with the bacterial species of interest is a prerequisite to detecting the bacterial species in environmental samples using metagenomic analyses

Gnarled-Trunk Evolutionary Model of Influenza A Virus Hemagglutinin

Human influenza A viruses undergo antigenic changes with gradual accumulation of amino acid substitutions on the hemagglutinin (HA) molecule. A strong antigenic mismatch between vaccine and epidemic strains often requires the replacement of influenza vaccines worldwide. To establish a practical model enabling us to predict the future direction of the influenza virus evolution, relative distances of amino acid sequences among past epidemic strains were analyzed by multidimensional scaling (MDS). We found that human influenza viruses have evolved along a gnarled evolutionary pathway with an approximately constant curvature in the MDS-constructed 3D space. The gnarled pathway indicated that evolution on the trunk favored multiple substitutions at the same amino acid positions on HA. The constant curvature was reasonably explained by assuming that the rate of amino acid substitutions varied from one position to another according to a gamma distribution. Furthermore, we utilized the estimated parameters of the gamma distribution to predict the amino acid substitutions on HA in subsequent years. Retrospective prediction tests for 12 years from 1997 to 2009 showed that 70% of actual amino acid substitutions were correctly predicted, and that 45% of predicted amino acid substitutions have been actually observed. Although it remains unsolved how to predict the exact timing of antigenic changes, the present results suggest that our model may have the potential to recognize emerging epidemic strains

Clinicopathological Evaluation of Papillary Thyroid Microcarcinoma

BACKGROUND AND AIMS: Clinicians sometimes encounter papillary thyroid microcarcinoma (PMC) that is less than 10 mm, associated with lymph node metastasis. In this study, we assessed PMC clinicopathologically to clarify risk factors for poor prognosis. PATIENTS AND METHODS: Fifty-one patients who underwent thyroid surgery at Aichi Medical University from September 2009 to October 2016 were included. Patients were divided into two groups, pEX-positive (23 patients) and pEX-negative (28 patients), based on the pathological finding of thyroid capsule invasion. The former indicates that the tumor infiltrated the thyroid capsule and spread to the neighboring tissue, and the latter indicates no capsule invasion. We analyzed factors such as patient characteristics, pathological findings, and serum levels of thyroid hormones in the two groups. RESULTS: No statistical differences were observed between the two groups in gender distribution or age at surgery. Preoperative cancer diagnoses were established for more patients in the pEX-positive group than in the pEX-negative group (n = 21 and 14, respectively; P = 0.004). The mean (±SD) pathological tumor diameter was 5.42 ± 2.77 in the pEX-negative group and 8.32 ± 1.61 in the pEX-positive group (P \u3c 0.001). No significant differences in preoperative serum levels of free T3, free T4, thyroid-stimulating hormone, or thyroglobulin were observed between the two groups. The odds ratio for node positivity in tumors invading thyroid capsules (pEX-positive) compared to those with no capsule invasion (pEX-negative) was 13.20 (95% confidence interval, 3.45-50.42). Immunohistological staining for phosphatase and tensin homolog deleted from chromosome 10 (PTEN) and Akt (protein kinase B) revealed the facilitation of PTEN and suppression of Akt, which might indicate downregulation of the phosphoinositide 3-kinase-Akt (PI3K-Akt) cascade. DISCUSSION: In general, the prognosis of PMC is favorable. However, the prognosis is less favorable in patients with nodal metastasis or extrathyroidal invasion. It is controversial whether resection is required for proven PMCs. For PMCs associated with extrathyroidal invasion, regional lymph node resection with lobectomy should be performed due to the high risk for lymphatic spread. There might be a possibility that the natural progression of PMC seems to be controlled by the facilitation of PTEN. However, a tumor in the lateral peripheral region of the thyroid parenchyma might be associated with capsule invasion followed by lymphatic spread

Predicting the Antigenic Structure of the Pandemic (H1N1) 2009 Influenza Virus Hemagglutinin

The pandemic influenza virus (2009 H1N1) was recently introduced into the human population. The hemagglutinin (HA) gene of 2009 H1N1 is derived from “classical swine H1N1” virus, which likely shares a common ancestor with the human H1N1 virus that caused the pandemic in 1918, whose descendant viruses are still circulating in the human population with highly altered antigenicity of HA. However, information on the structural basis to compare the HA antigenicity among 2009 H1N1, the 1918 pandemic, and seasonal human H1N1 viruses has been lacking. By homology modeling of the HA structure, here we show that HAs of 2009 H1N1 and the 1918 pandemic virus share a significant number of amino acid residues in known antigenic sites, suggesting the existence of common epitopes for neutralizing antibodies cross-reactive to both HAs. It was noted that the early human H1N1 viruses isolated in the 1930s–1940s still harbored some of the original epitopes that are also found in 2009 H1N1. Interestingly, while 2009 H1N1 HA lacks the multiple N-glycosylations that have been found to be associated with an antigenic change of the human H1N1 virus during the early epidemic of this virus, 2009 H1N1 HA still retains unique three-codon motifs, some of which became N-glycosylation sites via a single nucleotide mutation in the human H1N1 virus. We thus hypothesize that the 2009 H1N1 HA antigenic sites involving the conserved amino acids will soon be targeted by antibody-mediated selection pressure in humans. Indeed, amino acid substitutions predicted here are occurring in the recent 2009 H1N1 variants. The present study suggests that antibodies elicited by natural infection with the 1918 pandemic or its early descendant viruses play a role in specific immunity against 2009 H1N1, and provides an insight into future likely antigenic changes in the evolutionary process of 2009 H1N1 in the human population

Anthropogenic interferences lead to gut microbiome dysbiosis in Asian elephants and may alter adaptation processes to surrounding environments

Human activities interfere with wild animals and lead to the loss of many animal populations. Therefore, efforts have been made to understand how wildlife can rebound from anthropogenic disturbances. An essential mechanism to adapt to environmental and social changes is the fluctuations in the host gut microbiome. Here we give a comprehensive description of anthropogenically induced microbiome alterations in Asian elephants (n = 30). We detected gut microbial changes due to overseas translocation, captivity and deworming. We found that microbes belonging to Planococcaceae had the highest contribution in the microbiome alterations after translocation, while Clostridiaceae, Spirochaetaceae and Bacteroidia were the most affected after captivity. However, deworming significantly changed the abundance of Flavobacteriaceae, Sphingobacteriaceae, Xanthomonadaceae, Weeksellaceae and Burkholderiaceae. These findings may provide fundamental ideas to help guide the preservation tactics and probiotic replacement therapies of a dysbiosed gut microbiome in Asian elephants. More generally, these results show the severity of anthropogenic activities at the level of gut microbiome, altering the adaptation processes to new environments and the subsequent capability to maintain normal physiological processes in animals

Reference values for the locomotive syndrome risk test quantifying mobility of 8681 adults aged 20–89 years: A cross-sectional nationwide study in Japan

Background The locomotive syndrome risk test was developed to quantify the decrease in mobility among adults, which could eventually lead to disability. The purpose of this study was to establish reference values for the locomotive syndrome risk test for adults and investigate the influence of age and sex. Methods We analyzed 8681 independent community dwellers (3607 men, 5074 women). Data pertaining to locomotive syndrome risk test (the two-step test, the stand-up test, and the 25-question geriatric locomotive function scale [GLFS-25]) scores were collected from seven administrative areas of Japan. Results The reference values of the three test scores were generated and all three test scores gradually decreased among young-to-middle-aged individuals and rapidly decreased in individuals aged over 60 years. The stand-up test score began decreasing significantly from the age of 30 years. The trajectories of decrease in the two-step test score with age was slightly different between men and women especially among the middle-aged individuals. The two physical test scores were more sensitive to aging than the self-reported test score. Conclusion The reference values generated in this study could be employed to determine whether an individual has mobility comparable to independent community dwellers of the same age and sex

Cross-Protective Potential of a Novel Monoclonal Antibody Directed against Antigenic Site B of the Hemagglutinin of Influenza A Viruses

The hemagglutinin (HA) of influenza A viruses has been classified into sixteen distinct subtypes (H1–H16) to date. The HA subtypes of influenza A viruses are principally defined as serotypes determined by neutralization or hemagglutination inhibition tests using polyclonal antisera to the respective HA subtypes, which have little cross-reactivity to the other HA subtypes. Thus, it is generally believed that the neutralizing antibodies are not broadly cross-reactive among HA subtypes. In this study, we generated a novel monoclonal antibody (MAb) specific to HA, designated MAb S139/1, which showed heterosubtypic cross-reactive neutralization and hemagglutination inhibition of influenza A viruses. This MAb was found to have broad reactivity to many other viruses (H1, H2, H3, H5, H9, and H13 subtypes) in enzyme-linked immunosorbent assays. We further found that MAb S139/1 showed neutralization and hemagglutination-inhibition activities against particular strains of H1, H2, H3, and H13 subtypes of influenza A viruses. Mutant viruses that escaped neutralization by MAb S139/1 were selected from the A/Aichi/2/68 (H3N2), A/Adachi/2/57 (H2N2), and A/WSN/33 (H1N1) strains, and sequence analysis of the HA genes of these escape mutants revealed amino acid substitutions at positions 156, 158, and 193 (H3 numbering). A molecular modeling study showed that these amino acids were located on the globular head of the HA and formed a novel conformational epitope adjacent to the receptor-binding domain of HA. Furthermore, passive immunization of mice with MAb S139/1 provided heterosubtypic protection. These results demonstrate that MAb S139/1 binds to a common antigenic site shared among a variety of HA subtypes and neutralizes viral infectivity in vitro and in vivo by affecting viral attachment to cells. The present study supports the notion that cross-reactive antibodies play some roles in heterosubtypic immunity against influenza A virus infection, and underscores the potential therapeutic utility of cross-reactive antibodies against influenza

Symmetric Item Set Mining Method Using Zero-suppressed BDDs and Application to Biological Data

In this paper, we present a method of finding symmetric items in a combinatorial item set database. The techniques for finding symmetric variables in Boolean functions have been studied for long time in the area of VLSI logic design, and the BDD (Binary Decision Diagram) -based methods are presented to solve such a problem. Recently, we have developed an efficient method for handling databases using ZBDDs (Zero-suppressed BDDs), a particular type of BDDs. In our ZBDD-based data structure, the symmetric item sets can be found efficiently as well as for Boolean functions. We implemented the program of symmetric item set mining, and applied it to actual biological data on the amino acid sequences of influenza viruses. We found a number of symmetric items from the database, some of which indicate interesting relationships in the amino acid mutation patterns. The result shows that our method is helpful for extracting hidden interesting information in real-life databases

A visual environment for dynamic web application composition

HTML-based interface technologies enable end-users to easily use various remote Web applications. However, it is difficult for end-users to compose new integrated tools of both existing Web applications and legacy local applications such as spreadsheets, chart tools and database. In this paper, the authors propose a new framework where end-users can wrap remote Web applications into visual components called pads, and functionally combine them together through drag & drop-paste operations. The authors use, as the basis, a meme media architecture IntelligentPad that was proposed by the second author. In the IntelligentPad architecture, each visual component called a pad has slots as data I/O ports. By pasting a pad onto another pad users can integrate their functionalities. The framework presented in this paper allows users to visually create a wrapper pad for any Web application by defining HTML nodes within the Web application to work as slots. Examples of such a node include input-forms and text strings on Web pages. Users can directly manipulate both wrapped Web applications and wrapped local legacy tools on their desktop screen to define application linkages among them. Since no programming expertise is required to wrap Web applications or to functionally combine them together, end-users can build new integrated tools of both wrapped Web applications and local legacy applications

Data Mining in Amino Acid Sequences of H3N2 Influenza Viruses Isolated during 1968 to 2006

The hemagglutinin (HA) of influenza viruses undergoes antigenic drift to escape from antibody-mediated immune pressure. In order to predict possible structural changes of the HA molecules in future, it is important to understand the patterns of amino acid mutations and structural changes in the past. We performed a retrospective and comprehensive analysis of structural changes in H3 hemagglutinins of human influenza viruses isolated during 1968 to 2006. Amino acid sequence data of more than 2000 strains have been collected from NCBI Influenza virus resources. Information theoretic analysis of the collected sequences revealed a number of simultaneous mutations of amino acids at two or more different positions (correlated mutations). We also calculated the net charge of the HA1 subunit, based on the number of charged amino acid residues, and found that the net charge increased linearly from 1968 to 1984 and, after then, has been saturated. This level of the net charge may be an upper limit for H3 HA to be functional. It is noted that ”correlated mutations” with the conversion of acidic and basic amino acid residues between two different positions were frequently found after 1984, suggesting that these mutations contributed to counterbalancing effect to keep the net charge of the HA . These approaches may open the way to find the direction of future antigenic drift of influenza viruses