BACKGROUND
deep-sea vents are chemosynthetic environments which are considered as extreme as compared to usual life standards due to their physico-chemical characteristics.
for instance, they show high levels of temperature, pressure, heavy metals and sulfide  <cit> .
they are also highly variable, with physico-chemical shifts occurring over very short spatial and temporal scales  <cit> .
although other marine environments  are also variable, hydrothermal environments are particularly unstable due to  the intensity of variations observed in deep-sea vents, e.g., variation of temperature from  <dig> to 2°c over few centimeters  <cit> ,  local random mix of vent fluids with surrounding waters  <cit>  and  the relatively short lifetime of vents  <cit> .
because such unstable environment may be difficult to live in, deep-sea vents are often considered as harsh and stressful.
for example, the vent shrimps rimicaris exoculata, which usually live between 15°c and 30°c, endure sudden changes of thermal conditions due to the convection of fluids and can survive to exposure to very high temperature vent emissions  <cit> .
in terms of biome characteristics, hydrothermal ecosystems are also peculiar, as they show a much higher density of individuals compared with surrounding abyssal plains.
for example, populations of r. exoculata can reach up to  <dig> individuals per square meter  <cit> .
on the other hand, in comparison with coastal environments, hydrothermal vents are associated to a small number of species  <cit>  .
these features, along with the variable nature of the hydrothermal environment, suggest that deep-sea vent organisms may present particular genetic characteristics in relation to their peculiar adaptive abilities.
studying transposable elements  constitute an interesting way to investigate the genomic bases of such adaptive capacities.
several studies have shown that environmental variations can promote genome plasticity through transcriptional activation and mobilization of tes , often in response to specific stimuli such as biotic stress  and abiotic environmental changes   <cit> .
for these reasons, tes have a large impact on genome structure and stability, contribute in particular to variations in genome size  <cit> , and are therefore considered as one of the major sources of genetic variability in eukaryotes  <cit> .
the order decapoda is a great model to investigate the genomic nature of the adaptation of hydrothermal organisms and its possible relationship with tes.
first, decapod crustaceans  are found in various environments, with about  <dig> species described in hydrothermal vents  <cit> .
second, they exhibit great variation in genome size, from  <dig>  pg  in the crab carcinus maenas to  <dig>  pg  in the shrimp sclerocrangon ferox  <cit> , with several species  showing particularly large genomes that are thus liable to harbor high te contents.
given their abundance and diversity, decapods have been greatly underrepresented in studies of tes with only few elements described to date  <cit>  including a ltr retrotransposon, galea <dig>  that we have recently characterized in the hydrothermal galatheid squat lobster munidopsis recta  <cit> .
ltr retrotransposons  and tyrosine recombinase encoding elements  constitute two major groups of retrotransposons  <cit> , a class of tes specific to eukaryotes  <cit> .
several phylogenetic analyses have shown that the pol region of dirs-like elements harbor reverse transcriptase  and rnase h  domains closely related to those of ty3/gypsy-like ltr retrotransposons  <cit> .
however, these elements differ fundamentally in structure from ltr retrotransposons as they are devoid of ltr, do not encode either proteinase and integrase and harbor a methyltransferase domain   <cit> .
moreover, dirs-like retrotransposons replicate through a particular mechanism that involves a tyrosine recombinase  <cit> .
within the dirs-like superfamily  <cit> , two distinct groups have been described: dirs1-like and pat-like elements.
these two groups differ by the nature of their termini  and by phylogenic relationships  <cit> .
in contrast to ltr retrotransposons that have been found in a wide continuous range of species  <cit> , dirs1-like elements have a more patchy distribution being yet reported only in a restricted number of phylogenically diverse eukaryote organisms   <cit> .
although most of the dirs1-like retrotransposon identifications were based on in silico approaches  <cit> , they have not been yet described in several well-studied phylum , and are absent from model organisms such as saccharomyces cerevisae and drosophila melanogaster.
moreover, only few different dirs1-like families have been found per species  <cit>  in contrast to the large variety of ltr retrotransposon families  <cit> .
in the slime mold dictyostelium discoideum, dirs1-like retrotransposons have been shown to harbor an active heat-shock promoter in their inverted terminal repeats  <cit> .
considering the high level of thermal instability in vents, the presence of such promoters could favor te activation in hydrothermal species.
notes: - <dig> accession numbers deposited in ddbj/embl/genbank databases, in repbase database  <cit>  or in retrobase database  <cit> .
hence, studying dirs1-like elements in decapoda could allow both to better understand the role of tes in the adaptation of species in hydrothermal ecosystems and to investigate the dynamics of these peculiar elements.
we focused on ten hydrothermal decapods that represent the three major infraorders observed .
we have detected dirs1-like retrotransposons in all vent shrimps and in m. recta as well as in  <dig> other non-vent decapods.
to our knowledge, this paper constitutes the first study based on dirs1-like element distribution both in a continuous range of species within a single taxon and in species living in various ecosystems.
fifteen new dirs1-like families are at all described among  <dig> out of the  <dig> species tested.
these unusual elements are thus surprisingly more widely distributed among decapods than expected.
RESULTS
fishing out dirs1-like pol sequences in hydrothermal shrimps
in the course of looking for ty3/gypsy-like ltr retrotransposons in three hydrothermal shrimps  using degenerate primers gd <dig> and gd <dig> corresponding to the "rmpfgl" and "lttdas" conserved motifs of the rt domain , we pcr-amplified additional fragments in supplement of those that correspond to ty3/gypsy-like elements .
cloning and sequencing of these additional fragments revealed that they share high sequence identity with the pol region of dirs1-like elements at the protein level.
detailed sequence analysis showed that the fragments observed originated from either gd1/gd <dig> amplifications  or from gd1/gd <dig> amplification with annealing of degenerate primer gd <dig> at a non targeted site .
fishing out of dirs1-like elements using primers designed for ty3/gypsy-like retrotransposons can be explained by the close phylogenic relationships between rt domains of ty3/gypsy-like and dirs1-like elements  <cit> .
although the regions in which the primers were designed are only partly conserved between ty3/gypsy-like and dirs1-like elements, partial conservation may be sufficient for them to anneal on dirs1-like sequences, particularly because primers are degenerate.
to further confirm the presence of dirs1-like elements in these species, we designed a pair of degenerate primers, dd <dig> and dd <dig>  to amplify the region comprised between the "lgflin" and "dlfas" motifs of the dirs1-like elements pol region .
we tested these primers in r. exoculata.
unfortunately, they did not allow for amplifying a single band, but rather led to the amplification of numerous fragments .
after long and tedious optimization of the pcr conditions, we managed to obtain a major amplicon of  <dig> bp, which we then cloned and sequenced.
sequence analysis revealed that it corresponds actually to a dd2/dd <dig> amplification product, which shares similarity with pol region of dirs1-like elements only in the last  <dig> bp .
due to the technical difficulties to obtain specific amplicons, we did not use this primer pair for other analyses.
to get insights on the genetic diversity of the dirs1-like elements found, we aligned the partial sequences of the pol region that we obtained.
alignment of the  <dig> sequences allowed us to define  <dig> families.
rexalvi1a and cchalvi1a sharing 89% nucleotide sequence identity, they were clustered into a single family that we called alvi <dig> .
on the other hand, rexalvi2a and mfoalvi3a shared too low sequence identity to either alvi <dig>  or to each other  to be clustered and were therefore considered as members of different families that we called alvi <dig> and alvi <dig>
screening for alvi elements in other decapods
to date, dirs1-like retrotransposons have been identified and characterized in only a restricted number of species .
to better understand the evolutionary dynamics of dirs1-like elements, we investigated the diversity and the distribution of the alvi families among several decapoda crustacean genomes.
we first analyzed the distribution of our  <dig> novel dirs1-like families among several decapods, from both hydrothermal and non hydrothermal ecosystems.
for this, we screened for alvi families among the  <dig> decapods species using different combinations of primers  including specific primers of each element and retro1+, a primer shared by all  <dig> elements .
although pairs of specific primers would allow for amplification of closely related elements, we expected that the association of retro1+ with a single specific primer would be particularly useful to extract more divergent sequences from diverse species.
cloning and sequencing of pcr products of expected size led to the identification of either the targeted elements or other dirs1-like elements that belong to another family .
sequence analysis of all the dirs1-like elements found revealed that some were highly degenerate .
these elements were not included for the rest of the study .
among the other sequences, some corresponded to copies of the alvi1- <dig> families, whereas some others could not be clustered to any existing families, and were considered as members of  <dig> new families: alvi <dig>  and alvi <dig> .
we thus decided to include these  <dig> families in our analysis.
however, specific primers could not be designed for the alvi <dig> family, because nucleotidic differences with the other  <dig> families were too spread out on the sequences.
hence, we finally analyzed the distribution of dirs1-like elements in decapods using  <dig> families: alvi1- <dig>
in black bold: amplicon sequences corresponding to the element targeted.
underlined: pcr amplification products in species in which the element targeted was firstly identified.
in italics: amplicon sequences corresponding to another element than the one targeted.
a: pcr amplification product of the expected size observed but not sequenced.
na: no pcr amplification of expected size.
-: not tested.
* see methods for more details.
$ single specific primer associated to the retro1+ primer.
results, presented in table  <dig>  show that alvi elements are only detected in hydrothermal shrimps.
two families  are widely shared among these species while the two others  are restricted to the species where they were firstly identified .
alvi <dig> elements could not be detected in m. fortunata.
using alvi <dig> specific primers on this species we were only able to identify non-targeted elements of the alvi <dig> family , even using a supplementary primer  more restrictive to alvi <dig> family.
non targeted elements were also obtained when we looked for alvi <dig> and alvi <dig> elements , suggesting some specificity troubles especially due to the use of a single specific primer.
for all other decapods, no alvi element could be detected even in other hydrothermal organisms or in the other closely related caridean shrimps  <cit> crangon crangon and palaemon serratus.
in the hydrothermal galatheid crab munidopsis recta, although no pcr amplification fragment of the expected size was obtained, we pcr-amplified an additional fragment  when using retro1+.
sequencing of this fragment revealed that it originated from the retro1+/retro1+ amplification of a new dirs1-like sequence .
it shared low sequence identity to alvi1- <dig> families  and was therefore considered as member of a novel family that we called ymur <dig>
diversity and phylogenetic relationships of alvi dirs1-like retrotransposons
screening for alvi elements among decapods led us to obtain  <dig> partial sequences of the pol region .
to gain power in our investigation of the diversity of these elements, we completed these data with additional sequences.
two complementary approaches were performed: extension of the sequences by pcr walking and pcr amplification of larger internal sequences using specific primers .
seventeen new sequences were thus obtained , enabling to well-cover the pol region of dirs1-like elements at least from the "yldd" rt motif to the end of the rh domain .
all dna sequences obtained were aligned using clustalx program  <cit>  and corresponding alignment was used to analyze the relationships between families.
first, we checked the boundaries of the different families through the construction of a sequence identity matrix using the pairwise gap deletion method .
a group of sequences was considered as a family if its higher intra-group divergence was lower than its inter-groups divergence, without overlap of the two distributions.
the clustering obtained from sequence identity matrix validates the five families that we had previously defined .
among them, the alvi <dig> copies are more diverse  than other families ones.
the alvi <dig> family is also more divergent toward the four other families  whereas alvi2- <dig> families appear to be more closely related as copies from these families differ one by one by only 35% of divergence in average.
second, we analyzed the relationships of the sequences among and between the  <dig> families using a neighbor joining phylogenic analysis with pairwise gap deletion.
the tree obtained  confirmed that each alvi family is well-supported with bootstrap values being greater than 94%.
it also confirmed that the alvi <dig> family highly diverges to the other  <dig> families, with strong bootstrap support  of the separation into an alvi2- <dig> clade, and an alvi <dig> clade.
analysis of the alvi <dig> subtree also revealed a lack of structuration as compared to the other families, this confirming the large sequence diversity observed with the matrix analysis.
on the contrary, although relationships between families within the alvi2- <dig> group can not be resolved by our tree, relationships of sequences within each family are well resolved considering most of bootstrap values being greater than 70%.
it is especially the case for the alvi <dig> family which sequences were obtained from various hydrothermal shrimps.
phylogenic relationships among this family also fit with those of the species  <cit>  in which they were identified, with only few exceptions like the amualvi2a sequence from a. muricola that clusters with alvi <dig> sequences from a. markensis.
screening for dirs1-like elements in decapods
capacity of detection of tes using a pcr approach depends of the choice of the primers, as shown by the identification of some elements using specific/aspecific primer pairs and not using specific/specific pairs.
hence, absence of pcr signal could be due to the divergence of the elements rather than their absence from the species analyzed.
to detect such more distant dirs1-like retrotransposons, we designed degenerate primers using two types of primers: "highly" degenerate and "softly" degenerate .
although both of these approaches have flows , they are complementary and should, when used in parallel, raise the detection sensitivity.
six different combinations of primers were tested , five using "highly" degenerate primers, and one using two "softly" ones .
in black bold: amplicon sequences corresponding to dirs1-like element.
+: pcr amplification products of expected size in species known to harbor dirs1-like elements.
traces: very low amplification of expected size.
na: no amplification of expected size.
-: artefacts.
as expected from our previous pcr results, amplification fragments were obtained in hydrothermal shrimps whatever the degenerate primer combination used .
we have shown that m. recta harbors dirs1-like retrotransposons .
using 'highly" degenerate primers, clear amplification fragments of the expected size were only obtained with the gd1/rcdd <dig> combination.
cloning and sequencing allow to obtain three additional related sequences .
two others related sequences  were also obtained using the "softly" degenerate primer combination, one of them  corresponding to an unexpected additional fragment about  <dig> bp .
sequence analysis revealed that the dirs1-like sequences obtained in m. recta can be clustered into  <dig> distinct families  and phylogenic analyses indicates that ymur1- <dig> families positioned themselves as a single well supported clade  .
in other decapods, the two methods gave different results : while using "highly" degenerate primers combinations no amplification of the expected size was obtained, the use of gd1soft/rcdd4soft led to either trace amplification or amplification fragments of the expected size.
cloning and sequencing of all single bands observed allowed us to discover new dirs1-like elements  in  <dig> new species : the two coastal shrimps , the two coastal lobsters ,  <dig> seamount galatheid squat lobsters  and  <dig> coastal crabs .
the sequences obtained in a given species, except x. pilipes, cluster into a single family .
in the crab x. pilipes, the two sequences obtained define two distinct families .
moreover, elements of the galatheid crabs e. annulosa and a. laurentae  can be clustered into a single family we called yannu <dig> .
likewise, elements of the two lobsters n. norvegicus and h. gammarus  clustered into the same ynno <dig> family .
we also detect in the est databases two dirs1-like sequences  of the lobster homarus americanus, that share high similarities with ynno <dig> family .
altogether, this study led us to discover  <dig> new dirs1-like families .
all of these families diverge to alvi1- <dig> and ymur1- <dig> families with < 56% sequence identity .
to study their distribution, two specific primers were designed for each newly discovered dirs1-like family .
they were used to screen all fifteen species in which we had observed the presence of dirs1-like elements  as well as closely related species .
for each family, no pcr signal was obtained except in species in which the families were firstly identified.
this underlines that all elements previously obtained do not result from potential cross-contaminations.
nevertheless, dirs1-like retrotransposons are widely distributed among decapod crustaceans, although this distribution appears to be heterogeneous according to the phylum considered .
dirs1-like elements are yet identified in all caridean shrimps and lobsters tested.
in anomourans the distribution appears very patchy as dirs1-like retrotransposons are characterized in  <dig> out of the  <dig> galatheid squat lobsters but not in porcelain crabs or hermit crab.
finally, the presence of these elements has been only detected in  <dig> out of the  <dig> brachyuran crabs tested with especially no detection in hydrothermal organisms.
discussion
methodological issues
dirs1-like retrotransposons have yet been reported in a restricted number of eukaryote species , in contrast to ltr retrotransposons that show a wide and almost continuous distribution.
however, we found dirs1-like elements in  <dig> out of the  <dig> decapod species studied, with  <dig> novel dirs1-like families identified  and a particularly large diversity observed among hydrothermal shrimps .
to characterize transposable elements among different genomes, two approaches are usually performed: in silico approaches  <cit>  and/or pcr-based procedures  <cit> .
most of previous studies aiming at identifying dirs1-like retrotransposons were based on in silico approaches  <cit> .
however, in the case of decapods, genomic sequences are scarce, with sequencing projects concerning only the creation of est databases .
in particular, to our knowledge no large-scale genome sequencing project is ongoing.
we therefore conducted several pcr-based approaches that enable to screen for partial dirs1-like pol region simultaneously in a large number of species representing numerous decapod taxa .
among the  <dig> sequences that we identified,  <dig> were obtained from pcr-amplified fragments which size differed from the one expected .
analyses of such additional fragments, which can be obtained for different species or using different primer combinations, provide an interesting way to identify new tes.
one issue with dirs-like retrotransposons is that they are sometimes abusively grouped with ty3/gypsy-like elements because their rt/rh domains are closely related.
distinguishing them from each other is however primordial, particularly if dirs-like elements are in fact more frequent that previously supposed.
for that purpose, it is important to check the partial pol sequence of each element for the presence of the mt domain   before assigning them to dirs-like superfamily.
to classify our sequences into families, we used a nucleotide sequence identity matrix built using pairwise deletion method .
this method presents the advantage to compute a distance for each pair of sequences, ignoring only the gaps that are involved in the comparison and therefore including the largest amount of information for the whole dataset.
this method appears as the most appropriate to analyze our sequences, as our dataset is constituted of several pcr-based sequences that only partially overlap, and because many of our sequences harbor small internal indels.
finally, this approach appears efficient as it enables to clearly define the different families that we found .
recently, in an attempt to find criteria for sequence-based te classification for the annotation of tes in large scale genome sequencing projects, the "80-80-80" rule has been proposed  <cit> .
this rule states that two tes belong to the same family if they share 80%  sequence identity over segments longer than  <dig> bp representing at least 80% of the coding domains, internal domains, or terminal repeat regions.
however, as noticed by the authors, this rule is more efficient on terminal repeat regions and non-coding regions than on coding regions like pol.
this property was confirmed with our dataset, thus leading to the conclusion that, although certainly efficient for large-scale genome sequence analysis of tes, this rule is not adapted to our pcr-based analysis.
distribution of alvi families within alvinocarididae genomes
screenings for dirs1-like elements in decapod genomes indicates that the "highly" degenerate primers have a lower detection sensitivity than the "softly" degenerate ones.
oligonucleotide sequences of "softly" degenerate primers correspond to a subset of those present in the "highly" degenerate ones .
nevertheless, the combination of "softly" degenerate primers allowed for identifying dirs1-like elements in at least  <dig> diverse decapods whereas combinations of "highly" degenerate ones detect elements in only the  <dig> alvinocarididae and m. recta .
despite their weaker detection sensitivity, "highly" degenerate primers led to pcr amplifications in the six hydrothermal shrimps whatever the combinations used.
we suggest that these observations may be due to a higher copy number of dirs1-like retrotransposons in these genomes that could counterbalance the lower detection sensitivity of the "highly" degenerate primers.
in complement to degenerate primers, pcr screening using specific primers is a powerful approach to study the distribution of previously characterized elements, as well as for identifying novel members of closely related families.
indeed, screening for alvi1- <dig> families at a small phylogenetic scale  led to the detection of the alvi <dig> and alvi <dig> families.
since all hydrothermal shrimps harbor alvi elements, these ones likely result from the activity of dirs1-like retrotransposons that were already present in the genome of the common ancestor of all alvinocarididae species.
alvi elements may thus belong to relatively ancient lineages of dirs1-like retrotransposons.
they could be as old as several million years, since molecular estimates of divergence show that the vent-endemic caridean shrimp species radiated  <dig>  to  <dig>  million years ago  <cit> .
this is especially true for the alvi <dig> and alvi <dig> families that are shared among alvinocarididae.
our phylogenetic analysis  also shows that alvi <dig> to alvi <dig> families cluster into a single clade, suggesting that these families diverged from a common ancestor.
the alvi <dig> and alvi <dig> families are each restricted to a single species and may therefore correspond to lineage-specific evolution.
in a. stactophila, the presence of both alvi <dig> and alvi <dig> families suggests that this last could result from the differentiation of a subset of the ancestral family.
in m. fortunata, which is the most distant species among vent shrimps  <cit> , we observe the presence of alvi <dig> instead of alvi <dig> .
two alternative evolutionary scenarios could explain this observation.
if elements of the alvi <dig> family were present in the alvinocarididae ancestor, the distribution observed would result from different lineage-specific burst-like events of different alvi families, as observed for ltr retrotransposons in rice-related species  <cit> , leading to the amplification of alvi <dig> in m. fortunata and this of alvi <dig> in the common ancestor of other related vent shrimps.
on the other hand, if the alvi <dig> family was absent from the alvinocarididae ancestor, it likely results from a lineage-specific evolution of the ancestral alvi <dig> family in the m. fortunata lineage.
in addition, r. exoculata and c. chacei, which are phylogenetically very close  <cit> , harbor the same distribution pattern for dirs1-like families  with close corresponding sequences .
all these observations suggest that the wide distribution of dirs1-like retrotransposons among alvinocarididae is closely related to the phylogeny of species and that the emergence of new elements is linked to the independent evolutionary history of alvi families within each host genomes.
although we cannot privilege one evolutionary scenario, all possible scenarios are in accordance with a simple vertical transmission of the different alvi elements.
aldirs1: a new clade of dirs1-like retrotransposons
previous analyses of phylogenic relationships among the dirs1-like retrotransposons  <cit>  highlighted two clades: the drdirs <dig> clade that groups the dirs1-like elements identified in teleost fishes, and the tcdirs <dig> clade that pools together elements from insects and sea urchins.
to determine the relatedness of our elements with other dirs1-like retrotransposons, we performed a new phylogenic analysis using the maximum likelihood method.
the core dataset included alvi1- <dig> and ymur <dig> families, the only families for which we got sufficiently long sequences, as well as the recently identified elements of another crustacean species, daphnia pulex, and those of the cnidarian nematostella vectensis, which were never reported in phylogenic analyses to date.
the resulting tree  is in accordance to those previously described.
the two dpdirs1- <dig> elements from d. pulex cluster with those of other arthropods  in the tcdirs <dig> clade.
in contrast, dirs1-like elements of decapods are phylogenetically divergent from these, with the alvi1- <dig> and ymur <dig> families forming a distinct clade among the dirs1-like superfamily that we called aldirs1-clade.
these results indicate the presence of amino acid signatures for decapod elements, in particular a tryptophane  instead of a phenylalanine  amino acid in the rt domain at position  <dig> on the alignment .
interestingly, this signature is also shared by the sequences that we obtained in other decapods.
therefore, we propose that the decapod dirs1-like retrotransposons may belong to the aldirs <dig> clade, although further studies on the phylogenic relationships of these elements are needed to confirm this.
for that purpose, two conditions should be fulfilled: to obtain additional sequences to build better amino acids consensus sequences, and to enlarge the sequences obtained for the pol region to improve the number of informative sites.
wide distribution of dirs1-like elements among decapods
our analysis of the distribution of the dirs1-like retrotransposons , the first as such large scale  gives clues to better understand the peculiar scattered distribution of these elements among eukaryote genomes.
three species  harbor at least  <dig> different dirs1-like families.
such results are similar to those observed in other well-studied species such as the sea urchin strongylocentrotus purpuratus  and the zebrafish danio rerio .
in particular, we show that dirs1-like elements can be much more than just a feature of isolate species.
when focusing on a given taxa  the distribution of dirs1-like retrotransposons  appears more or less patchy, depending on the infraorder considered .
three different distribution patterns are observed:  presence of the elements in a large range of species .
presence of the elements in a subpart of the infraorder .
presence of the elements in a restricted number of species .
so, decapoda are the first example revealing that dirs1-like element may be frequent, widely distributed and so probably ancient within an order.
species distribution of dirs1-like retrotransposons  thus appears related to species phylogeny.
some families are widely distributed among closely related species  whereas some others  seem to be only scarcely shared, even among species that belong to the same genus.
considering the environmental conditions of species, dirs1-like retrotransposons are not restricted to hydrothermal species.
however, their higher detection sensitivity in vent organisms  suggests some peculiar features that could be related to a greatest element diversity and/or a higher number of copies.
to better understand the distribution of dirs1-like elements among decapoda, the next step will be to extend our study to other infraorders  or dendrobranchiata ), even to other crustaceans.
such an analysis will also allow for analyzing the vertical transmission of dirs1-like elements suggested by our results.
moreover, we are currently estimating the number of copies of dirs1-like retrotransposons.
this will provide crucial information  to point out possible particularities of vent species in comparison with other decapods and  to understand the dynamics of these elements within decapoda.
we also wonder whether such widespread distribution and diversity of dirs1-like retrotransposons are peculiar features of decapoda, or if similar features can be observed in other clades, for instance one known to harbor dirs1-like elements .
if the widespread distribution of dirs1-like retrotransposons that we observe within a taxon is a usual pattern, then the presence of one element in a given species would indicate that dirs1-like retrotransposons are likely to be found in phylogenetically closely related species.
CONCLUSIONS
because of the small number of dirs1-like elements described to date, little is known about the distribution and the evolutionary dynamics of these retrotransposons, which differ to other tes by their peculiar tyrosine recombinase -based replication model  <cit> .
this paper presents the first study using pcr approaches to specifically search for dirs1-like retrotransposons in several related species.
fifteen new dirs1-like families were identified in  <dig> decapod species which nearly doubles both the number of dirs1-like elements described to date, and the number of species known to harbor such elements.
our study reveals a larger diversity of dirs1-like retrotransposons as we distinguish a third clade  in addition to the tcdirs <dig> and drdirs <dig> clades previously described.
our results provide a new vision of the distribution of the dirs1-like superfamily, and reveals that these elements are not as scattered as previously thought.
it is the first study showing that these retrotransposons could have a widespread distribution within a large order  as well as a great diversity within a family .
these observations open the way to further studies on the evolutionary dynamics of these elements.

 deep-sea vents are chemosynthetic environments which are considered as extreme as compared to usual life standards due to their physico-chemical characteristics. for instance, they show high levels of temperature, pressure, heavy metals and sulfide. they are also highly variable, with physico-chemical shifts occurring over very short spatial and temporal scales. although other marine environments are also variable, hydrothermal environments are particularly unstable due to the intensity of variations observed in deep-sea vents, e.g., variation of temperature from to 2°c over few centimeters, local random mix of vent fluids with surrounding waters and the relatively short lifetime of vents. because such unstable environment may be difficult to live in, deep-sea vents are often considered as harsh and stressful. for example, the vent shrimps rimicaris exoculata, which usually live between 15°c and 30°c, endure sudden changes of thermal conditions due to the convection of fluids and can survive to exposure to very high temperature vent emissions. in terms of biome characteristics, hydrothermal ecosystems are also peculiar, as they show a much higher density of individuals compared with surrounding abyssal plains. for example, populations of r. exoculata can reach up to individuals per square meter. on the other hand, in comparison with coastal environments, hydrothermal vents are associated to a small number of species. these features, along with the variable nature of the hydrothermal environment, suggest that deep-sea vent organisms may present particular genetic characteristics in relation to their peculiar adaptive abilities. studying transposable elements constitute an interesting way to investigate the genomic bases of such adaptive capacities. several studies have shown that environmental variations can promote genome plasticity through transcriptional activation and mobilization of tes, often in response to specific stimuli such as biotic stress and abiotic environmental changes. for these reasons, tes have a large impact on genome structure and stability, contribute in particular to variations in genome size, and are therefore considered as one of the major sources of genetic variability in eukaryotes. the order decapoda is a great model to investigate the genomic nature of the adaptation of hydrothermal organisms and its possible relationship with tes. first, decapod crustaceans are found in various environments, with about species described in hydrothermal vents. second, they exhibit great variation in genome size, from pg in the crab carcinus maenas to pg in the shrimp sclerocrangon ferox, with several species showing particularly large genomes that are thus liable to harbor high te contents. given their abundance and diversity, decapods have been greatly underrepresented in studies of tes with only few elements described to date including a ltr retrotransposon, galea that we have recently characterized in the hydrothermal galatheid squat lobster munidopsis recta. ltr retrotransposons and tyrosine recombinase encoding elements constitute two major groups of retrotransposons, a class of tes specific to eukaryotes. several phylogenetic analyses have shown that the pol region of dirs-like elements harbor reverse transcriptase and rnase h domains closely related to those of ty3/gypsy-like ltr retrotransposons. however, these elements differ fundamentally in structure from ltr retrotransposons as they are devoid of ltr, do not encode either proteinase and integrase and harbor a methyltransferase domain. moreover, dirs-like retrotransposons replicate through a particular mechanism that involves a tyrosine recombinase. within the dirs-like superfamily, two distinct groups have been described: dirs1-like and pat-like elements. these two groups differ by the nature of their termini and by phylogenic relationships. in contrast to ltr retrotransposons that have been found in a wide continuous range of species, dirs1-like elements have a more patchy distribution being yet reported only in a restricted number of phylogenically diverse eukaryote organisms. although most of the dirs1-like retrotransposon identifications were based on in silico approaches, they have not been yet described in several well-studied phylum, and are absent from model organisms such as saccharomyces cerevisae and drosophila melanogaster. moreover, only few different dirs1-like families have been found per species in contrast to the large variety of ltr retrotransposon families. in the slime mold dictyostelium discoideum, dirs1-like retrotransposons have been shown to harbor an active heat-shock promoter in their inverted terminal repeats. considering the high level of thermal instability in vents, the presence of such promoters could favor te activation in hydrothermal species. notes: - accession numbers deposited in ddbj/embl/genbank databases, in repbase database or in retrobase database. hence, studying dirs1-like elements in decapoda could allow both to better understand the role of tes in the adaptation of species in hydrothermal ecosystems and to investigate the dynamics of these peculiar elements. we focused on ten hydrothermal decapods that represent the three major infraorders observed. we have detected dirs1-like retrotransposons in all vent shrimps and in m. recta as well as in other non-vent decapods. to our knowledge, this paper constitutes the first study based on dirs1-like element distribution both in a continuous range of species within a single taxon and in species living in various ecosystems. fifteen new dirs1-like families are at all described among out of the species tested. these unusual elements are thus surprisingly more widely distributed among decapods than expected.  fishing out dirs1-like pol sequences in hydrothermal shrimps in the course of looking for ty3/gypsy-like ltr retrotransposons in three hydrothermal shrimps using degenerate primers gd and gd corresponding to the "rmpfgl" and "lttdas" conserved motifs of the rt domain, we pcr-amplified additional fragments in supplement of those that correspond to ty3/gypsy-like elements. cloning and sequencing of these additional fragments revealed that they share high sequence identity with the pol region of dirs1-like elements at the protein level. detailed sequence analysis showed that the fragments observed originated from either gd1/gd amplifications or from gd1/gd amplification with annealing of degenerate primer gd at a non targeted site. fishing out of dirs1-like elements using primers designed for ty3/gypsy-like retrotransposons can be explained by the close phylogenic relationships between rt domains of ty3/gypsy-like and dirs1-like elements. although the regions in which the primers were designed are only partly conserved between ty3/gypsy-like and dirs1-like elements, partial conservation may be sufficient for them to anneal on dirs1-like sequences, particularly because primers are degenerate. to further confirm the presence of dirs1-like elements in these species, we designed a pair of degenerate primers, dd and dd to amplify the region comprised between the "lgflin" and "dlfas" motifs of the dirs1-like elements pol region. we tested these primers in r. exoculata. unfortunately, they did not allow for amplifying a single band, but rather led to the amplification of numerous fragments. after long and tedious optimization of the pcr conditions, we managed to obtain a major amplicon of bp, which we then cloned and sequenced. sequence analysis revealed that it corresponds actually to a dd2/dd amplification product, which shares similarity with pol region of dirs1-like elements only in the last bp. due to the technical difficulties to obtain specific amplicons, we did not use this primer pair for other analyses. to get insights on the genetic diversity of the dirs1-like elements found, we aligned the partial sequences of the pol region that we obtained. alignment of the sequences allowed us to define families. rexalvi1a and cchalvi1a sharing 89% nucleotide sequence identity, they were clustered into a single family that we called alvi. on the other hand, rexalvi2a and mfoalvi3a shared too low sequence identity to either alvi or to each other to be clustered and were therefore considered as members of different families that we called alvi and alvi screening for alvi elements in other decapods to date, dirs1-like retrotransposons have been identified and characterized in only a restricted number of species. to better understand the evolutionary dynamics of dirs1-like elements, we investigated the diversity and the distribution of the alvi families among several decapoda crustacean genomes. we first analyzed the distribution of our novel dirs1-like families among several decapods, from both hydrothermal and non hydrothermal ecosystems. for this, we screened for alvi families among the decapods species using different combinations of primers including specific primers of each element and retro1+, a primer shared by all elements. although pairs of specific primers would allow for amplification of closely related elements, we expected that the association of retro1+ with a single specific primer would be particularly useful to extract more divergent sequences from diverse species. cloning and sequencing of pcr products of expected size led to the identification of either the targeted elements or other dirs1-like elements that belong to another family. sequence analysis of all the dirs1-like elements found revealed that some were highly degenerate. these elements were not included for the rest of the study. among the other sequences, some corresponded to copies of the alvi1- families, whereas some others could not be clustered to any existing families, and were considered as members of new families: alvi and alvi. we thus decided to include these families in our analysis. however, specific primers could not be designed for the alvi family, because nucleotidic differences with the other families were too spread out on the sequences. hence, we finally analyzed the distribution of dirs1-like elements in decapods using families: alvi1- in black bold: amplicon sequences corresponding to the element targeted. underlined: pcr amplification products in species in which the element targeted was firstly identified. in italics: amplicon sequences corresponding to another element than the one targeted. a: pcr amplification product of the expected size observed but not sequenced. na: no pcr amplification of expected size. -: not tested. * see methods for more details. $ single specific primer associated to the retro1+ primer. results, presented in table show that alvi elements are only detected in hydrothermal shrimps. two families are widely shared among these species while the two others are restricted to the species where they were firstly identified. alvi elements could not be detected in m. fortunata. using alvi specific primers on this species we were only able to identify non-targeted elements of the alvi family, even using a supplementary primer more restrictive to alvi family. non targeted elements were also obtained when we looked for alvi and alvi elements, suggesting some specificity troubles especially due to the use of a single specific primer. for all other decapods, no alvi element could be detected even in other hydrothermal organisms or in the other closely related caridean shrimps crangon crangon and palaemon serratus. in the hydrothermal galatheid crab munidopsis recta, although no pcr amplification fragment of the expected size was obtained, we pcr-amplified an additional fragment when using retro1+. sequencing of this fragment revealed that it originated from the retro1+/retro1+ amplification of a new dirs1-like sequence. it shared low sequence identity to alvi1- families and was therefore considered as member of a novel family that we called ymur diversity and phylogenetic relationships of alvi dirs1-like retrotransposons screening for alvi elements among decapods led us to obtain partial sequences of the pol region. to gain power in our investigation of the diversity of these elements, we completed these data with additional sequences. two complementary approaches were performed: extension of the sequences by pcr walking and pcr amplification of larger internal sequences using specific primers. seventeen new sequences were thus obtained, enabling to well-cover the pol region of dirs1-like elements at least from the "yldd" rt motif to the end of the rh domain. all dna sequences obtained were aligned using clustalx program and corresponding alignment was used to analyze the relationships between families. first, we checked the boundaries of the different families through the construction of a sequence identity matrix using the pairwise gap deletion method. a group of sequences was considered as a family if its higher intra-group divergence was lower than its inter-groups divergence, without overlap of the two distributions. the clustering obtained from sequence identity matrix validates the five families that we had previously defined. among them, the alvi copies are more diverse than other families ones. the alvi family is also more divergent toward the four other families whereas alvi2- families appear to be more closely related as copies from these families differ one by one by only 35% of divergence in average. second, we analyzed the relationships of the sequences among and between the families using a neighbor joining phylogenic analysis with pairwise gap deletion. the tree obtained confirmed that each alvi family is well-supported with bootstrap values being greater than 94%. it also confirmed that the alvi family highly diverges to the other families, with strong bootstrap support of the separation into an alvi2- clade, and an alvi clade. analysis of the alvi subtree also revealed a lack of structuration as compared to the other families, this confirming the large sequence diversity observed with the matrix analysis. on the contrary, although relationships between families within the alvi2- group can not be resolved by our tree, relationships of sequences within each family are well resolved considering most of bootstrap values being greater than 70%. it is especially the case for the alvi family which sequences were obtained from various hydrothermal shrimps. phylogenic relationships among this family also fit with those of the species in which they were identified, with only few exceptions like the amualvi2a sequence from a. muricola that clusters with alvi sequences from a. markensis. screening for dirs1-like elements in decapods capacity of detection of tes using a pcr approach depends of the choice of the primers, as shown by the identification of some elements using specific/aspecific primer pairs and not using specific/specific pairs. hence, absence of pcr signal could be due to the divergence of the elements rather than their absence from the species analyzed. to detect such more distant dirs1-like retrotransposons, we designed degenerate primers using two types of primers: "highly" degenerate and "softly" degenerate. although both of these approaches have flows, they are complementary and should, when used in parallel, raise the detection sensitivity. six different combinations of primers were tested, five using "highly" degenerate primers, and one using two "softly" ones. in black bold: amplicon sequences corresponding to dirs1-like element. +: pcr amplification products of expected size in species known to harbor dirs1-like elements. traces: very low amplification of expected size. na: no amplification of expected size. -: artefacts. as expected from our previous pcr results, amplification fragments were obtained in hydrothermal shrimps whatever the degenerate primer combination used. we have shown that m. recta harbors dirs1-like retrotransposons. using'highly" degenerate primers, clear amplification fragments of the expected size were only obtained with the gd1/rcdd combination. cloning and sequencing allow to obtain three additional related sequences. two others related sequences were also obtained using the "softly" degenerate primer combination, one of them corresponding to an unexpected additional fragment about bp. sequence analysis revealed that the dirs1-like sequences obtained in m. recta can be clustered into distinct families and phylogenic analyses indicates that ymur1- families positioned themselves as a single well supported clade. in other decapods, the two methods gave different results: while using "highly" degenerate primers combinations no amplification of the expected size was obtained, the use of gd1soft/rcdd4soft led to either trace amplification or amplification fragments of the expected size. cloning and sequencing of all single bands observed allowed us to discover new dirs1-like elements in new species: the two coastal shrimps, the two coastal lobsters, seamount galatheid squat lobsters and coastal crabs. the sequences obtained in a given species, except x. pilipes, cluster into a single family. in the crab x. pilipes, the two sequences obtained define two distinct families. moreover, elements of the galatheid crabs e. annulosa and a. laurentae can be clustered into a single family we called yannu. likewise, elements of the two lobsters n. norvegicus and h. gammarus clustered into the same ynno family. we also detect in the est databases two dirs1-like sequences of the lobster homarus americanus, that share high similarities with ynno family. altogether, this study led us to discover new dirs1-like families. all of these families diverge to alvi1- and ymur1- families with < 56% sequence identity. to study their distribution, two specific primers were designed for each newly discovered dirs1-like family. they were used to screen all fifteen species in which we had observed the presence of dirs1-like elements as well as closely related species. for each family, no pcr signal was obtained except in species in which the families were firstly identified. this underlines that all elements previously obtained do not result from potential cross-contaminations. nevertheless, dirs1-like retrotransposons are widely distributed among decapod crustaceans, although this distribution appears to be heterogeneous according to the phylum considered. dirs1-like elements are yet identified in all caridean shrimps and lobsters tested. in anomourans the distribution appears very patchy as dirs1-like retrotransposons are characterized in out of the galatheid squat lobsters but not in porcelain crabs or hermit crab. finally, the presence of these elements has been only detected in out of the brachyuran crabs tested with especially no detection in hydrothermal organisms. discussion methodological issues dirs1-like retrotransposons have yet been reported in a restricted number of eukaryote species, in contrast to ltr retrotransposons that show a wide and almost continuous distribution. however, we found dirs1-like elements in out of the decapod species studied, with novel dirs1-like families identified and a particularly large diversity observed among hydrothermal shrimps. to characterize transposable elements among different genomes, two approaches are usually performed: in silico approaches and/or pcr-based procedures. most of previous studies aiming at identifying dirs1-like retrotransposons were based on in silico approaches. however, in the case of decapods, genomic sequences are scarce, with sequencing projects concerning only the creation of est databases. in particular, to our knowledge no large-scale genome sequencing project is ongoing. we therefore conducted several pcr-based approaches that enable to screen for partial dirs1-like pol region simultaneously in a large number of species representing numerous decapod taxa. among the sequences that we identified, were obtained from pcr-amplified fragments which size differed from the one expected. analyses of such additional fragments, which can be obtained for different species or using different primer combinations, provide an interesting way to identify new tes. one issue with dirs-like retrotransposons is that they are sometimes abusively grouped with ty3/gypsy-like elements because their rt/rh domains are closely related. distinguishing them from each other is however primordial, particularly if dirs-like elements are in fact more frequent that previously supposed. for that purpose, it is important to check the partial pol sequence of each element for the presence of the mt domain before assigning them to dirs-like superfamily. to classify our sequences into families, we used a nucleotide sequence identity matrix built using pairwise deletion method. this method presents the advantage to compute a distance for each pair of sequences, ignoring only the gaps that are involved in the comparison and therefore including the largest amount of information for the whole dataset. this method appears as the most appropriate to analyze our sequences, as our dataset is constituted of several pcr-based sequences that only partially overlap, and because many of our sequences harbor small internal indels. finally, this approach appears efficient as it enables to clearly define the different families that we found. recently, in an attempt to find criteria for sequence-based te classification for the annotation of tes in large scale genome sequencing projects, the "80-80-80" rule has been proposed. this rule states that two tes belong to the same family if they share 80% sequence identity over segments longer than bp representing at least 80% of the coding domains, internal domains, or terminal repeat regions. however, as noticed by the authors, this rule is more efficient on terminal repeat regions and non-coding regions than on coding regions like pol. this property was confirmed with our dataset, thus leading to the conclusion that, although certainly efficient for large-scale genome sequence analysis of tes, this rule is not adapted to our pcr-based analysis. distribution of alvi families within alvinocarididae genomes screenings for dirs1-like elements in decapod genomes indicates that the "highly" degenerate primers have a lower detection sensitivity than the "softly" degenerate ones. oligonucleotide sequences of "softly" degenerate primers correspond to a subset of those present in the "highly" degenerate ones. nevertheless, the combination of "softly" degenerate primers allowed for identifying dirs1-like elements in at least diverse decapods whereas combinations of "highly" degenerate ones detect elements in only the alvinocarididae and m. recta. despite their weaker detection sensitivity, "highly" degenerate primers led to pcr amplifications in the six hydrothermal shrimps whatever the combinations used. we suggest that these observations may be due to a higher copy number of dirs1-like retrotransposons in these genomes that could counterbalance the lower detection sensitivity of the "highly" degenerate primers. in complement to degenerate primers, pcr screening using specific primers is a powerful approach to study the distribution of previously characterized elements, as well as for identifying novel members of closely related families. indeed, screening for alvi1- families at a small phylogenetic scale led to the detection of the alvi and alvi families. since all hydrothermal shrimps harbor alvi elements, these ones likely result from the activity of dirs1-like retrotransposons that were already present in the genome of the common ancestor of all alvinocarididae species. alvi elements may thus belong to relatively ancient lineages of dirs1-like retrotransposons. they could be as old as several million years, since molecular estimates of divergence show that the vent-endemic caridean shrimp species radiated to million years ago. this is especially true for the alvi and alvi families that are shared among alvinocarididae. our phylogenetic analysis also shows that alvi to alvi families cluster into a single clade, suggesting that these families diverged from a common ancestor. the alvi and alvi families are each restricted to a single species and may therefore correspond to lineage-specific evolution. in a. stactophila, the presence of both alvi and alvi families suggests that this last could result from the differentiation of a subset of the ancestral family. in m. fortunata, which is the most distant species among vent shrimps, we observe the presence of alvi instead of alvi. two alternative evolutionary scenarios could explain this observation. if elements of the alvi family were present in the alvinocarididae ancestor, the distribution observed would result from different lineage-specific burst-like events of different alvi families, as observed for ltr retrotransposons in rice-related species, leading to the amplification of alvi in m. fortunata and this of alvi in the common ancestor of other related vent shrimps. on the other hand, if the alvi family was absent from the alvinocarididae ancestor, it likely results from a lineage-specific evolution of the ancestral alvi family in the m. fortunata lineage. in addition, r. exoculata and c. chacei, which are phylogenetically very close, harbor the same distribution pattern for dirs1-like families with close corresponding sequences. all these observations suggest that the wide distribution of dirs1-like retrotransposons among alvinocarididae is closely related to the phylogeny of species and that the emergence of new elements is linked to the independent evolutionary history of alvi families within each host genomes. although we cannot privilege one evolutionary scenario, all possible scenarios are in accordance with a simple vertical transmission of the different alvi elements. aldirs1: a new clade of dirs1-like retrotransposons previous analyses of phylogenic relationships among the dirs1-like retrotransposons highlighted two clades: the drdirs clade that groups the dirs1-like elements identified in teleost fishes, and the tcdirs clade that pools together elements from insects and sea urchins. to determine the relatedness of our elements with other dirs1-like retrotransposons, we performed a new phylogenic analysis using the maximum likelihood method. the core dataset included alvi1- and ymur families, the only families for which we got sufficiently long sequences, as well as the recently identified elements of another crustacean species, daphnia pulex, and those of the cnidarian nematostella vectensis, which were never reported in phylogenic analyses to date. the resulting tree is in accordance to those previously described. the two dpdirs1- elements from d. pulex cluster with those of other arthropods in the tcdirs clade. in contrast, dirs1-like elements of decapods are phylogenetically divergent from these, with the alvi1- and ymur families forming a distinct clade among the dirs1-like superfamily that we called aldirs1-clade. these results indicate the presence of amino acid signatures for decapod elements, in particular a tryptophane instead of a phenylalanine amino acid in the rt domain at position on the alignment. interestingly, this signature is also shared by the sequences that we obtained in other decapods. therefore, we propose that the decapod dirs1-like retrotransposons may belong to the aldirs clade, although further studies on the phylogenic relationships of these elements are needed to confirm this. for that purpose, two conditions should be fulfilled: to obtain additional sequences to build better amino acids consensus sequences, and to enlarge the sequences obtained for the pol region to improve the number of informative sites. wide distribution of dirs1-like elements among decapods our analysis of the distribution of the dirs1-like retrotransposons, the first as such large scale gives clues to better understand the peculiar scattered distribution of these elements among eukaryote genomes. three species harbor at least different dirs1-like families. such results are similar to those observed in other well-studied species such as the sea urchin strongylocentrotus purpuratus and the zebrafish danio rerio. in particular, we show that dirs1-like elements can be much more than just a feature of isolate species. when focusing on a given taxa the distribution of dirs1-like retrotransposons appears more or less patchy, depending on the infraorder considered. three different distribution patterns are observed: presence of the elements in a large range of species. presence of the elements in a subpart of the infraorder. presence of the elements in a restricted number of species. so, decapoda are the first example revealing that dirs1-like element may be frequent, widely distributed and so probably ancient within an order. species distribution of dirs1-like retrotransposons thus appears related to species phylogeny. some families are widely distributed among closely related species whereas some others seem to be only scarcely shared, even among species that belong to the same genus. considering the environmental conditions of species, dirs1-like retrotransposons are not restricted to hydrothermal species. however, their higher detection sensitivity in vent organisms suggests some peculiar features that could be related to a greatest element diversity and/or a higher number of copies. to better understand the distribution of dirs1-like elements among decapoda, the next step will be to extend our study to other infraorders or dendrobranchiata ), even to other crustaceans. such an analysis will also allow for analyzing the vertical transmission of dirs1-like elements suggested by our results. moreover, we are currently estimating the number of copies of dirs1-like retrotransposons. this will provide crucial information to point out possible particularities of vent species in comparison with other decapods and to understand the dynamics of these elements within decapoda. we also wonder whether such widespread distribution and diversity of dirs1-like retrotransposons are peculiar features of decapoda, or if similar features can be observed in other clades, for instance one known to harbor dirs1-like elements. if the widespread distribution of dirs1-like retrotransposons that we observe within a taxon is a usual pattern, then the presence of one element in a given species would indicate that dirs1-like retrotransposons are likely to be found in phylogenetically closely related species.  because of the small number of dirs1-like elements described to date, little is known about the distribution and the evolutionary dynamics of these retrotransposons, which differ to other tes by their peculiar tyrosine recombinase -based replication model. this paper presents the first study using pcr approaches to specifically search for dirs1-like retrotransposons in several related species. fifteen new dirs1-like families were identified in decapod species which nearly doubles both the number of dirs1-like elements described to date, and the number of species known to harbor such elements. our study reveals a larger diversity of dirs1-like retrotransposons as we distinguish a third clade in addition to the tcdirs and drdirs clades previously described. our results provide a new vision of the distribution of the dirs1-like superfamily, and reveals that these elements are not as scattered as previously thought. it is the first study showing that these retrotransposons could have a widespread distribution within a large order as well as a great diversity within a family. these observations open the way to further studies on the evolutionary dynamics of these elements. 
hydrothermal organisms show a particularly great diversity of dirs1-like elements with families characterized among alvinocarididae shrimps and in the galatheid crab munidopsis recta. they also suggest some peculiar features of these retrotransposons in hydrothermal organisms where a great diversity of elements is already observed. phylogenic analyses show that these elements are divergent toward the dirs1-like families previously described in other crustaceans and arthropods and form a new clade called aldirs at larger scale, the distribution of dirs1-like retrotransposons appears more or less patchy depending on the taxa considered. using pcr approaches, we have identified new dirs1-like families in diverse decapod species. considering their mutational abilities, tes can contribute to the genetic diversity and evolution of organisms. our results lead to nearly double both the number of dirs1-like elements described to date, and the number of species known to harbor these ones. transposable elements are major constituents of eukaryote genomes and have a great impact on genome structure and stability. dirs1-like retrotransposons are a particular group of retrotransposons according to their mode of transposition that implies a tyrosine recombinase. in this study, we provide the first degenerate primers designed to look specifically for dirs1-like retrotransposons. finally, this paper constitutes the first essential step which allows for considering further studies based on the dynamics of the dirs1-like retrotransposons among several genomes. in this paper, we determine the distribution of dirs1-like elements among decapod species, of them living in hydrothermal vents that correspond to particularly unstable environments. they allowed for revealing for the first time a widespread distribution of these elements among a large phylum, here the order decapoda. indeed, a scattered distribution can be observed in the infraorder brachyura whereas all the species tested in infraorders caridea and astacidea harbor some dirs1-like elements. to date, they have been described in a restricted number of species in comparison with the ltr retrotransposons.
 transposable elements are major constituents of eukaryote genomes and have a great impact on genome structure and stability. considering their mutational abilities, tes can contribute to the genetic diversity and evolution of organisms. knowledge of their distribution among several genomes is an essential condition to study their dynamics and to better understand their role in species evolution. dirs1-like retrotransposons are a particular group of retrotransposons according to their mode of transposition that implies a tyrosine recombinase. to date, they have been described in a restricted number of species in comparison with the ltr retrotransposons. in this paper, we determine the distribution of dirs1-like elements among decapod species, of them living in hydrothermal vents that correspond to particularly unstable environments.  using pcr approaches, we have identified new dirs1-like families in diverse decapod species. hydrothermal organisms show a particularly great diversity of dirs1-like elements with families characterized among alvinocarididae shrimps and in the galatheid crab munidopsis recta. phylogenic analyses show that these elements are divergent toward the dirs1-like families previously described in other crustaceans and arthropods and form a new clade called aldirs at larger scale, the distribution of dirs1-like retrotransposons appears more or less patchy depending on the taxa considered. indeed, a scattered distribution can be observed in the infraorder brachyura whereas all the species tested in infraorders caridea and astacidea harbor some dirs1-like elements.  our results lead to nearly double both the number of dirs1-like elements described to date, and the number of species known to harbor these ones. in this study, we provide the first degenerate primers designed to look specifically for dirs1-like retrotransposons. they allowed for revealing for the first time a widespread distribution of these elements among a large phylum, here the order decapoda. they also suggest some peculiar features of these retrotransposons in hydrothermal organisms where a great diversity of elements is already observed. finally, this paper constitutes the first essential step which allows for considering further studies based on the dynamics of the dirs1-like retrotransposons among several genomes. 