The scarcity of usable nitrogen frequently limits plant growth. A tight metabolic association with rhizobial bacteria allows legumes to obtain nitrogen compounds by bacterial reduction of dinitrogen (N2) to ammonium (NH4+). We present here the annotated DNA sequence of the alpha-proteobacterium Sinorhizobium meliloti, the symbiont of alfalfa. The tripartite 6.7-megabase (Mb) genome comprises a 3.65-Mb chromosome, and 1.35-Mb pSymA and 1.68-Mb pSymB megaplasmids. Genome sequence analysis indicates that all three elements contribute, in varying degrees, to symbiosis and reveals how this genome may have emerged during evolution. The genome sequence will be useful in understanding the dynamics of interkingdom associations and of life in soil environments.

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Analysis of the 1,683,333-nt sequence of the pSymB megaplasmid from the symbiotic N2-fixing bacterium Sinorhizobium meliloti revealed that the replicon has a high gene density with a total of 1,570 protein-coding regions, with few insertion elements and regions duplicated elsewhere in the genome. The only copies of an essential arg-tRNA gene and the minCDE genes are located on pSymB. Almost 20% of the pSymB sequence carries genes encoding solute uptake systems, most of which were of the ATP-binding cassette family. Many previously unsuspected genes involved in polysaccharide biosynthesis were identified and these, together with the two known distinct exopolysaccharide synthesis gene clusters, show that 14% of the pSymB sequence is dedicated to polysaccharide synthesis. Other recognizable gene clusters include many involved in catabolic activities such as protocatechuate utilization and phosphonate degradation. The functions of these genes are consistent with the notion that pSymB plays a major role in the saprophytic competence of the bacteria in the soil environment.A mong the bacteria, the ␣-proteobacteria appear unusual because of the presence of multiple replicons within the same bacterial strain (1). In the case of Agrobacterium tumefaciens, the causative agent of crown gall disease, the genome contains both a linear and a circular chromosome (2). Many (but not all) of the bacteria that form N 2 -fixing root nodules on leguminous plants are characterized by the presence of multiple plasmids greater than 400 kb in size. In the case of the N 2 -fixing symbiont Sinorhizobium meliloti, there are three replicons, a 3,654-kb circular chromosome (3, 4) and two megaplasmids 1,354 and 1,683 kb in size (5-7). The smaller of the megaplasmids, variously called pSymA, pNod-Nif, or pRmeSU47a, is known to carry many of the genes involved in root nodule formation (nod) and nitrogen fixation (nif ) (8, 9).The 1,683-kb megaplasmid, referred to as pSymB, pExo, or pRmeSU47b, is known to carry various gene clusters involved in exopolysaccharide (EPS) synthesis, C 4 -dicarboxylate transport, and lactose metabolism (10-12). Early studies focused on mutations that abolished synthesis of the succinoglycan EPS, EPS I, because these mutations resulted in a loss of the ability to form normal N 2 -fixing root nodules. This symbiotic defect was rescued by second-site mutations that increased the synthesis of a second galactoglucan EPS (EPS II), whose biosynthetic genes were also located on the pSymB megaplasmid (13,14). Other genes located on pSymB that are required for the formation of N 2 -fixing root nodules include the C 4 -dicarboxylate (dctA) and phosphate transport (phoCDET) genes and the bacA gene (15-18). The presence of large plasmids in bacteria that form associations with plants was described over 20 years ago (19). However, with the exception of the symbiotic genes in relatively small regions of these plasmids, the broader biological role of the plasmids in the biology of the organism has remained obscure. We constructed a ...

Death. These data suggest the existence of a previously uncharacterized historical 35 plague focus that persisted for at least three centuries. We propose that this 36 disease source may have been responsible for the many resurgences of plague in

A phylogenetic portrait of the genus Calicivirus in the family Caliciviridae was developed based upon published sequences and newly characterized calicivirus (CV) strains, including additional Sapporo-like HuCV strains in pediatric diarrhea stool specimens from South Africa, the United Kingdom, and the United States. Distance and parsimony methods were applied to nucleotide and amino acid sequences of human and animal calicivirus 3D RNA-dependent RNA polymerase (approximately 470nt) and capsid hypervariable regions (approximately 1,200nt) to generate phylogenetic trees. Pairwise amino acid identity in the 3D region among the Sapporo-like strains ranged from 61% to 100%. Human and animal caliciviruses (HuCVs and AnCVs) separated into five genogroups: small round-structured viruses (SRSV), Sapporo-like, and hepatitis E virus (HEV)-like HuCVs and rabbit-, and vesicular exanthema of swine virus (VESV)-like AnCVs, each with a distinct genome organization. Each genogroup, including the Sapporo-like HuCVs, subdivided further into subgenogroups. The capsid region trees had higher levels of confidence than the 3D region trees and limited conclusions about genogroups could be drawn from the 3D region analyses. This analysis suggested that CVs include five potential virus subfamilies.

A large amount of information is contained within the phylogenetic relationships between species. In addition to their branching patterns it is also possible to examine other aspects of the biology of the species. The influence that deleterious selection might have is determined here. The likelihood of different phylogenies in the presence of selection is explored to determine the properties of such a likelihood surface. The calculation of likelihoods for a phylogeny in the presence and absence of selection, permits the application of a likelihood ratio test to search for selection. It is shown that even a single selected site can have a strong effect on the likelihood. The method is illustrated with an example from Drosophila melanogaster and suggests that deleterious selection may be acting on transposable elements.

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A whole-genome sequencing technique developed to identify fast neutron-induced deletion mutations revealed that iap1-1 is a new allele of EDS5 (eds5-5). RPS2-AvrRpt2-initiated effector-triggered immunity (ETI) was compromised in iap1-1/eds5-5 with respect to in planta bacterial levels and the hypersensitive response, while intra- and intercellular free salicylic acid (SA) accumulation was greatly reduced, suggesting that SA contributes as both an intracellular signaling molecule and an antimicrobial agent in the intercellular space during ETI. During the compatible interaction between wild-type Col-0 and virulent Pseudomonas syringae pv. tomato (Pst), little intercellular free SA accumulated, which led to the hypothesis that Pst suppresses intercellular SA accumulation. When Col-0 was inoculated with a coronatine-deficient strain of Pst, high levels of intercellular SA accumulation were observed, suggesting that Pst suppresses intercellular SA accumulation using its phytotoxin coronatine. This work suggests that accumulation of SA in the intercellular space is an important component of basal/PAMP-triggered immunity as well as ETI to pathogens that colonize the intercellular space.