We determined the genomic sequence of Nocardia farcinica IFM 10152, a clinical isolate, and revealed the molecular basis of its versatility. The genome consists of a single circular chromosome of 6,021,225 bp with an average G؉C content of 70.8% and two plasmids of 184,027 (pNF1) and 87,093 (pNF2) bp with average G؉C contents of 67.2% and 68.4%, respectively. The chromosome encoded 5,674 putative protein-coding sequences, including many candidate genes for virulence and multidrug resistance as well as secondary metabolism. Analyses of paralogous protein families suggest that gene duplications have resulted in a bacterium that can survive not only in soil environments but also in animal tissues, resulting in disease.N ocardia are filamentous-growing Gram-positive soil saprophytes that belong to the family Actinomycetales, which also includes clinically and industrially important genera such as Mycobacterium, Streptomyces, Corynebacterium, and Rhodococcus. Many species of Nocardia cause the disease nocardiosis in humans and animals on lung, central nervous system, brain, and cutaneous tissues (1), and a species Nocardia asteroides is suspected to be an etiological agent of Parkinson's disease (2). Nocardiosis is on the rise, with an estimated 109-136 new cases occurring annually in Japan (3) and 500-1,000 in the U.S. (www.cdc.gov͞ncidod͞dbmd͞diseaseinfo͞nocardiosis t.htm). However, there are only a few studies on the mechanisms of nocardial virulence. Nocardia species are resistant to many front-line antibiotics. Because treatment for nocardiosis relies heavily on chemotherapy, their intrinsic multiple drug resistance is a serious problem.Another feature of the nocardia is that many strains, even clinical isolates, also have the capability to produce bioactive molecules such as antibiotics (4, 5) and enzymes that are industrially important (6). A monobactam antibiotic, nocardicin, was isolated from Nocardia sp. (7), and a terpenoid brasilicardin with immunosuppressive activity was isolated from a clinical isolate (4).Nocardia farcinica IFM 10152 was isolated from the bronchus of a 68-year-old male Japanese patient. Despite the complicated taxonomy of the nocardia, the species N. farcinica was found to be nearly homogeneous (8). Subsequently, to elucidate the molecular basis of the versatility of the nocardia, we analyzed the genomic sequence of N. farcinica IFM 10152. MethodsGenome Sequencing and Assembly. The nucleotide sequence of the N. farcinica IFM 10152 genome was determined by a wholegenome shotgun strategy. We constructed small-insert (2 kb) and large-insert (10 kb) genomic libraries and generated 127,077 sequences (giving 9-fold coverage) from both ends of the genomic clones. Sequence assembly was carried out by using the PHRED͞PHRAP͞CONSED package (9). Remaining gaps were closed by transcriptional sequencing (Nippon Gene, Toyama, Japan) or by primer walking. There were no ambiguous nucleotides in the genomic sequence that we could determine. Consistency of the final assembly was confirmed in terms of res...

FramePlot is a web-based tool for predicting protein-coding regions in bacterial DNA with a high G + C content, such as Streptomyces. The graphical output provides for easy distinction of protein-coding regions from non-coding regions. The plot is a clickable map. Clicking on an ORF provides not only the nucleotide sequence but also its deduced amino acid sequence. These sequences can then be compared to the NCBI sequence database over the Internet. The program is freely available for academic purposes at http://www.nih.go.jp/jun/cgi-bin/frameplot.pl.

In the period from 1992 to 2001, 303 cases of nocardioses were diagnosed in Japan, with the corresponding etiological agents isolated and characterized. Taxonomic analyses of these 303 strains showed that most nocardial infections were caused by members of the Nocardia asteroides group (72.3%). Speciation showed that 72 strains were N. asteroides, 31 strains were N. cyriacigeorgica, 2 strains were N. beijingensis, 81 strains were N. farcinica, and 33 strains were N. nova. Sixty-six strains of N. brasiliensis were the next most prevalent species of the total Nocardia isolates, followed by 14 strains of N. otitidiscaviarum. Infections by N. transvalensis (3 strains) and N. pseudobrasiliensis (1 strain) were also confirmed. The infections due to N. transvalensis, N. cyriacigeorgica, and N. beijingensis were the first reported in Japan. The most common factor that predisposed individuals to nocardial infection in Japan was therapy by immunosuppressive agents (22.4%), including SLE therapy (3.6%), followed by cancer (6.6%), diabetes (3.6%) and AIDS (2.0%). Nocardial infections occurred more commonly in the elderly, with most of the patients between the ages of 61 and 80 years of age. No significant difference regarding infectivity levels between the sexes was observed.