We also found that a blastn search of shotgun reads against a database of the catalase-peroxidase ( katB) gene detected 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.

pneumophila-specific nested PCR as a gold standard to evaluate the results of the metagenomic analysis. We compared different database search methods for bacterial detection by focusing on Legionella pneumophila. 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. 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. Metagenomic analysis has become a powerful tool to analyze bacterial communities in environmental samples. 6Laboratory of Parasitology, Graduate School of Veterinary Medicine, Hokkaido University, Sapporo, Japan.5Faculty of Public Health, Thammasat University, Rangsit Campus, Pathumthani, Thailand.4Global Institute for Collaborative Research and Education, Hokkaido University, Sapporo, Japan.3Division of Collaboration and Education, Research Center for Zoonosis Control, Hokkaido University, Sapporo, Japan.2Precursory Research for Embryonic Science and Technology, Japan Science and Technology Agency, Kawaguchi, Japan.

1Division of Bioinformatics, Research Center for Zoonosis Control, Hokkaido University, Sapporo, Japan.Through an examination of the mechanisms of entry and the factors that regulate it, the investigator hopes to further the understanding of how Legionella causes disease as well as provide insight into novel methods for disease prevention.Jednipit Borthong 1, Ryosuke Omori 1,2, Chihiro Sugimoto 3,4, Orasa Suthienkul 5, Ryo Nakao 6вЂ and Kimihito Ito 1,4,5*вЂ The specific aims of the current proposal are to: 1) isolate and characterize the genes involved in invasion and 2) determine the involvement of these genes in virulence of Legionella. Taken together, these studies indicate that a key factor in the production of virulent Legionella in water supplies is growth within the appropriate environmental host, without which the investigator hypothesizes that human infections will most likely not occur. This mechanism appears to result in enhanced survival of Legionella after entry into activated macrophages. Electron microscopic examination of entry has shown that a novel mechanism, termed ''''''''coiling phagocytosis'''''''', is responsible for bacterial uptake. The investigator's recent data indicate that Legionella grown in an environmental host, the amoebae Acanthamoeba castellani, are approximately two to four logs more invasive for host cells than bacteria grown in the standard laboratory medium. Thus, Legionella must enter a eukaryotic cell in order to survive and the genes involved in entry should be critical for production of disease. It has been found that Legionella replicates almost exclusively intracellularly during infection and in the environment. Approximately 15% of all domestic water supplies are thought to contain Legionella. There have recently been a large number of infections that have arisen aboard cruise ships as well. The first evidence that air-handling devices were the source of infection stems from a confirmed Legionella epidemic in 1976. Legionnaire's disease has been shown to arise in individuals that undergo prolonged exposure to aerosols from domestic water supplies that contain Legionella. Legionella pneumophila causes the severe and sometimes lethal pneumonia termed Legionnaire's disease.

Pbp3 gene and legionella