Molecular evidence of male-to-female sexual transmission of hepatitis C virus after vaginal and anal intercourse.

Hepatitis C virus (HCV) was transmitted from a chronic carrier to his female partner during unprotected anal and vaginal intercourse. Based on HVR1 and phylogenetic tree analysis, the couple had closely related isolates. These findings confirm sexual transmission of HCV without other risk factors.

Two-component systems in Pseudomonas aeruginosa: why so many?

Screening the Pseudomonas aeruginosa genome has led to the identification of the highest number of putative genes encoding two-component regulatory systems of all bacterial genomes sequenced to date (64 and 63 encoding response regulators and histidine kinases, respectively). Sixteen atypical kinases, among them 11 devoid of an Hpt domain, and three independent Hpt modules were retrieved. These data suggest that P. aeruginosa possesses complex control strategies with which to respond to environmental challenges.

ABCdb: an ABC transporter database.

We present the first release of a database devoted to the ATP-binding cassette (ABC) protein domains (ABCdb). The ABC proteins are involved in a wide variety of physiological processes in Archea, Bacteria and Eucaryota where they are encoded by large families of paralogous genes. The majority of ABC domains energize the transport of compounds across the membranes. In bacteria, ABC transporters are involved in the uptake of a wide range of molecules and in mechanisms of virulence and antibiotic resistance. In eukaryotes, most of them are involved in drug resistance and in human cells, many are associated with diseases. Sequence analysis reveals that members of the ABC superfamily can be organized into sub-families and suggests that they have diverged from common ancestral forms. In this release, ABCdb includes the inventory and assembly of the ABC transporter systems of completely sequenced genomes. In addition to the protein entries, the database comprises information on functional domains, sequence motifs, predicted trans-membrane segments, and signal peptides. It also includes a classification in sub-families of the ABC systems as well as a classification of the different partners of the systems. Evolutionary trees and specific sequence patterns are provided for each sub-family. The database is endowed with a powerful query system and it was interfaced with blastP2 program for similarity searches. ABCdb has been developed in the ACeDB format, a database system developed by Jean Thierry-Mieg and Richard Durbin. ABCdb can be accessed via the World Wide Web (http://ir2lcb.cnrs-mrs.fr/ABCdb/).

Covert transmission of hepatitis C virus during bloody fisticuffs.

Hepatitis C virus (HCV) is transmitted primarily through direct percutaneous exposure to infected blood. Sporadic HCV cases exist and may represent more than 10% of HCV transmission. We report the first case of documented transmission of HCV during a fight from a person who unknowingly had chronic HCV infection to a person who subsequently contracted acute hepatitis C. Patient-to-patient transmission was ascertained by sequence analysis of part of the NS5B genome and phylogenetic analysis. This case report suggests that sporadic HCV infection may be a result of blood exposure. This example of transmission could have a major impact in sports such as boxing or rugby. We suggest that in any fight, single use or nondisposable material should be used to dry blood to avoid such contamination.

Bacterial twin-arginine signal peptide-dependent protein translocation pathway: evolution and mechanism.

The recently identified bacterial Tat pathway is capable of exporting proteins with a peculiar twin-arginine signal peptide in folded conformation independently of the Sec machinery. It is structurally and mechanistically similar to the delta pH-dependent pathway used for importing chloroplast proteins into the thylakoid. The tat genes are not ubiquitously present and are absent from half of the completely sequenced bacterial genomes. The presence of the tat genes seems to correlate with genome size and with the presence of important enzymes with a twin-arginine signal peptide. A minimal Tat system requires a copy of tatA and a copy of tatC. The composition and gene order of a tat locus are generally conserved within the same taxonomy group but vary considerably to other groups, which would exclude an acquisition of the Tat system by recent horizontal gene transfer. The tat genes are also found in the genomes of chloroplasts and plant mitochondria but are absent from animal mitochondrial genomes. The topology of evolution trees suggests a bacterial origin of the Tat system. In general, the twin-arginine signal peptide is capable of targeting any passenger protein to the Tat pathway. However, a structural signal carried by the mature part of a passenger protein can override targeting information in a signal peptide under certain circumstances. Tat systems show a substrate-Tat component specificity and a species specificity. The pore size of the Tat channel is estimated as being between 5 and 9 nm. Operational models of the Tat system are proposed.

Protein-coding region discovery in organisms underrepresented in databases.

The prediction of coding sequences has received a lot of attention during the last decade. We can distinguish two kinds of methods, those that rely on training with sets of example and counter-example sequences, and those that exploit the intrinsic properties of the DNA sequences to be analyzed. The former are generally more powerful but their domains of application are limited by the availability of a training set. The latter avoid this drawback but can only be applied to sequences that are long enough to allow computation of the statistics. Here, we present a method that fills the gap between the two approaches. A learning step is applied using a set of sequences that are assumed to contain coding and non-coding regions, but with the boundaries of these regions unknown. A test step then uses the discriminant function obtained during the learning to predict coding regions in sequences from the same organism. The learning relies upon a correspondence analysis and prediction is presented on a graphical display. The method has been evaluated on a sample of yeast sequences, and the analysis of a set of expressed sequence tags from the Eucalyptus globulus-Pisolithus tinctorius ectomycorrhiza illustrates the relevance of the approach in its biological context.

Mother-to-infant transmission of hepatitis C virus: molecular evidence of superinfection by homologous virus in children.

BACKGROUND/AIM: Vertical transmission of hepatitis C virus (HCV) is well established but its incidence is low. To assess the molecular evidence of mother-to-infant transmission or intrafamilial transmission of HCV, the NS5 B region and the hypervariable region 1 (HVR1) of the E2/NS1 region of the HCV genome from each member of a family were investigated. METHODS: A 35-year-old mother with chronic hepatitis C virus infection and her four infected boys were studied. The same HCV 1a genotype was found in all five. Phylogenetic analysis was done by the neighbor-joining, the maximum likelihood, and the maximum parsimony methods. RESULTS: Comparison of the phylogenetic trees in the NS5B and HVR1 regions showed that the sequences in the children were more closely related to the population of variants of their own mother than to any genotype la sequence available in the databases. However, four HVR1 clones from two brothers (E2 and E3) had a strong homology, but were significantly divergent from the variants of the mother. CONCLUSIONS: These results suggest that a cluster of HCV strains exists in the family and that E3 could have been superinfected by E2 HCV strains and reciprocally. In conclusion, phylogenetic analysis through variable regions of the genome suggests that at least two modes of transmission are involved in this family: perinatal and horizontal.

Inventory, assembly and analysis of Bacillus subtilis ABC transport systems.

We have undertaken the inventory and assembly of the ATP binding cassette (ABC) transporter systems in the complete genome of Bacillus subtilis. We combined the identification of the three protein partners that compose an ABC transporter (nucleotide-binding domain, NBD; membrane spanning domain, MSD; and solute-binding protein, SBP) with constraints on the genetic organization. This strategy allowed the identification of 86 NBDs in 78 proteins, 103 MSD proteins and 37 SBPs. The analysis of transcriptional units allows the reconstruction of 59 ABC transporters, which include at least one NBD and one MSD. A particular class of five dimeric ATPases was not associated to MSD partners and is assumed to be involved either in macrolide resistance or regulation of translation elongation. In addition, we have detected five genes encoding ATPases without any gene coding for MSD protein in their neighborhood and 11 operons that encode only the membrane and solute-binding proteins. On the bases of similarities, three ATP-binding proteins are proposed to energize ten incomplete systems, suggesting that one ATPase may be recruited by more than one transporter. Finally, we estimate that the B. subtilis genome encodes for at least 78 ABC transporters that have been split in 38 importers and 40 extruders. The ABC systems have been further classified into 11 sub-families according to the tree obtained from the NBDs and the clustering of the MSDs and the SBPs. Comparisons with Escherichia coli show that the extruders are over-represented in B. subtilis, corresponding to an expansion of the sub-families of antibiotic and drug resistance systems.

Integrated mapping and sequencing of a 115 kb DNA fragment from Bacillus subtilis: sequence analysis of a 21 kb segment containing the sigL locus.

A sequence strategy which combines a low redundancy shotgun approach and directed sequencing has been elaborated. Essentially, the sequences, as well as the size of the fragments utilized for a low coverage shotgun approach, were exploited for the construction of a physical map of the region to be sequenced. The latter considerably simplified the subsequent directed sequencing steps. We report the physical mapping of a 115 kb segment which covers nearly 100 kb of the hisA-cysB region of the Bacillus subtilis chromosome and contains previously sequenced genes sigL and sacB. Sequencing and analysis of a 21305 bp segment, which includes the sigL locus, revealed 21 ORFs, apparently belonging to at least seven transcription units. This segment has a G + C content greater than 47%, compared to 43% characteristic of the flanking regions, and mainly consists of genes whose products seem to be involved in the synthesis of an exopolysaccharide. These observations leave open the possibility that the analysed fragment has been acquired through horizontal transfer.

A computer filtering method to drive out tiny genes from the yeast genome.

The authors of the first yeast chromosome sequence defined a minimum threshold requirement of 100 codons, above which an open reading frame (ORF) is retained as a putative coding sequence. However, at least 58 yeast genes shorter than 100 codons have an assigned protein function. Therefore, the yeast genome may contain other tiny but functionally important genes that are discarded from analyses by this simple filtering rule. We have established discriminant functions from the in-phase hexamer frequencies of functional genes and of simulated ORFs derived from a stationary Markov chain model. Fifty-two out of the 58 genes were recognized as coding ORFs by our discriminating method. The test was also applied to all the small ORFs (36 to 100 codons) found in the intergenic regions of published chromosomes. It retained 140 new potential tiny coding sequences, among which we identified seven new genes by similarity searches. Our method, used conjointly with similarity searches, can also highlight sequencing errors resulting from the disruption of the coding frame of longer ORFs. This method, by its ability to detect potential coding ORFs, can be a very useful tool for functional analysis.

A frameshift error detection algorithm for DNA sequencing projects.

During the determination of DNA sequences, frameshift errors are not the most frequent but they are the most bothersome as they corrupt the amino acid sequence over several residues. Detection of such errors by sequence alignment is only possible when related sequences are found in the databases. To avoid this limitation, we have developed a new tool based on the distribution of non-overlapping 3-tuples or 6-tuples in the three frames of an ORF. The method relies upon the result of a correspondence analysis. It has been extensively tested on Bacillus subtilis and Saccharomyces cerevisiae sequences and has also been examined with human sequences. The results indicate that it can detect frameshift errors affecting as few as 20 bp with a low rate of false positives (no more than 1.0/1000 bp scanned). The proposed algorithm can be used to scan a large collection of data, but it is mainly intended for laboratory practice as a tool for checking the quality of the sequences produced during a sequencing project.

Analysis of errors in finished DNA sequences: the surfactin operon of Bacillus subtilis as an example.

Increased productivity in DNA sequencing would not be valid without a straightforward detection and estimation of errors in finished sequences. The sequence of the surfactin operon from Bacillus subtilis was obtained by two different groups and by chance we were also working on the same chromosome region. Taking advantage of this situation we report in this paper, the number and nature of errors found in the overlapping part of the DNA sequences obtained by the three laboratories. The coincidence of some of the errors with compression in sequence ladders and with secondary DNA structures as well as the detection of frameshift errors using computer programs, are demonstrated. Finally we discuss the definition of a new sequencing strategy that might minimize both the error rate and the cost of sequencing.

A master sequence related to a free left Alu monomer (FLAM) at the origin of the B1 family in rodent genomes.

The question of the origin of the B1 family of rodents is addressed. The modern B1 elements are similar to the left Alu monomer, but with a 9 bp deletion and a 29 bp duplication. Search of databases for B1 elements that do not exhibit those modern features revealed sequence fragments that are very similar to the free left Alu monomers (FLAMs) described in the primate genomes. In addition, the analysis reveals elements that have 10 bp or 7 bp deletion in place of the 9 bp deletion but without the 29 bp tandem duplication. The elements described define families of proto B1 elements (referred as PB1, PB1D10 and PB1D7) that appeared before the first modern B1 element. A phylogenetic reconstruction suggest that the origin of Alu and B1 families took place before the divergence between the primate and the rodent lineages and that each family has followed different evolutionary routes since this radiation.

Emergence of master sequences in families of retroposons derived from 7sl RNA.

The past few years have brought new insight into the evolution of families of retroposons. These are composed of a very small number of master sequences able to duplicate, and a large majority of copies that are inactive for retroposition. During the course of time, successive replacements of master sequences have produced waves of amplification that are recognizable as subfamilies. In the Alu and the B1 families, one can distinguish two evolutionary periods. The first involves only monomeric elements that are now extinguished (fossil elements) and is characterized by deep remodeling of the sequences. This period ends, in primates, with the fusion of a free left and a free right Alu monomer, producing the first modern Alu dimeric element; in rodents it ends with a tandem duplication of 29 bp to create the first modern B1 element. The second period is characterized by a great stability of the master sequences. The observed turn-over of master sequences is still an enigma. However, analysis of the contemporary master sequences and of the oldest master sequences provide some clues. Here, we review the very first stages of the appearance of the Alu and the B1 families in mammalian genomes.

Fast identification of repetitive elements in biological sequences.

We have developed a fast filtering method for searching repetitive sequences in databases that allows the simultaneous identification of different families of repetitive elements during the same scanning. It discriminates between repetitive elements and non-related sequences by comparing the frequencies of k-words found in both groups of sequences. The distance used to sort out the sequences is based on a weighting of the k-words, which is obtained by performing a correspondence analysis on learning sets of correctly chosen sequences. The identification of Alu elements in human sequences is given as an illustration of the method. The Alu sequences are divided in four distinct groups of elements: the left and right monomers located on the direct and on the complementary strands. The results obtained on the test sets show that a very good discrimination is achieved with a word length of 6 b.p. Indeed, only 0.5% of the non-Alu sequences were incorrectly predicted as Alu elements for a threshold value allowing the identification of all Alu monomers. The misclassification of the different Alu monomers (1.4%) in the four groups of examples occurs only when the left and the right monomers are in the same orientation. Moreover, during the scanning of 63 GenBank sequences longer than 10 Kb, all the Alu elements were correctly identified (616 elements) and only a few non-Alu sequences were wrongly predicted as Alu elements (22 fragments). There is a real need for this kind of method since most of the repetitive elements are not annotated in the database entries. This method can then be used for a systematic screening of new sequences before their insertion in databases. It can also allow the creation of specific databases devoted to repetitive elements, which is a required step for any further analysis of those elements.

Origin of the Alu family: a family of Alu-like monomers gave birth to the left and the right arms of the Alu elements.

The Alu dimeric elements are a common feature of the primate genomes, where they constitute a family of related sequences (1). The identification of a free left Alu monomer (FLAM) family plus a free right Alu monomer (FRAM) family suggests that the dimeric structure results from the fusion of a FLAM sequence with a FRAM sequence (2). Here, we describe a very old Alu-like monomeric family, referred to as FAM for fossil Alu monomer. This family arose from a 7SL RNA sequence and gave birth to the FLAM and FRAM families. From the results obtained, the evolution of the Alu family can be subdivided into two phases. The first phase, which involves only monomeric elements, is characterized by deep remodelling of the progenitor sequences and ends with the appearance of the first Alu dimeric element through the fusion of a FLAM and a FRAM element. The second phase, still in progress, starts with the first Alu dimeric element. This phase is characterized by the stabilization of the progenitor sequences.

Fusion of a free left Alu monomer and a free right Alu monomer at the origin of the Alu family in the primate genomes.

In the primate genome, a typical Alu element corresponds to a dimeric structure composed of two different but related monomeric sequences arranged in tandem. However, the analysis of primate sequences found in GenBank reveals the presence of free left and free right Alu elements. Here, we report the statistical study of those monomeric elements. We found that only a small fraction of them results from a deletion of a dimeric Alu sequence. The majority derives from the amplification of monomeric progenitor sequences and constitutes two families of monomeric elements: a family of free left Alu monomers that is composed of two subfamilies and a small family of free right Alu monomers. Both families predated the dimeric Alu elements, and a phylogenetic analysis strongly suggests that the first progenitor of the dimeric Alu family arose through the fusion of a free left monomer with a free right monomer.

Successive waves of fixation of B1 variants in rodent lineage history.

A new method of analyzing phylogenetic relations among members of sequence family (Quentin 1988) discriminates at least six possible B1 subfamilies in the mouse genome. Several additional and independent observations suggest that these groupings have evolutionary significance, and that successive waves of fixation of new variants occur during rodent lineage history. We have reason to believe that, in a genome, the founder sequences of different families of retroposons are in competition with regard to the amplification/fixation process.

[Role of arteriography and x-ray computed tomography in the current evaluation of pigmented villonodular synovitis]. / Place de l'artériographie et de la tomodensitométrie dans le bilan actuel des synovites villo-nodulaires pigmentées.

The angiographic characteristics of five surgically proven cases of pigmented villonodular synovitis are reported. A CT scan was performed in one case. CT scan arthrography is very useful when it demonstrates regions of high attenuation. The angiography never demonstrated in the five cases capillary blush nor arteriovenous shunting, but a regular hypervascular mass. These findings are helpful for the surgeon suggesting the best surgical approach and the best site of biopsy.

The Alu family developed through successive waves of fixation closely connected with primate lineage history.

A new method of analyzing phylogenetic relations among members of the sequence family is presented and applied to human Alu sequences upon which work has been published. This method, based upon a correspondence analysis, works with large samples and yields easily interpretable graphical representations. Results obtained argue in favor of a new evolutionary scheme for Alu sequences, implying successive waves of amplification/fixation closely connected to primate lineage history.