Lower Statistical Support with Larger Data Sets: Insights from the Ochrophyta Radiation.

It is commonly assumed that increasing the number of characters has the potential to resolve evolutionary radiations. Here, we studied photosynthetic stramenopiles (Ochrophyta) using alignments of heterogeneous origin mitochondrion, plastid, and nucleus. Surprisingly while statistical support for the relationships between the six major Ochrophyta lineages increases when comparing the mitochondrion (6,762 sites) and plastid (21,692 sites) trees, it decreases in the nuclear (209,105 sites) tree. Statistical support is not simply related to the data set size but also to the quantity of phylogenetic signal available at each position and our ability to extract it. Here, we show that this ability for current phylogenetic methods is limited, because conflicting results were obtained when varying taxon sampling. Even though the use of a better fitting model improved signal extraction and reduced the observed conflicts, the plastid data set provided higher statistical support for the ochrophyte radiation than the larger nucleus data set. We propose that the higher support observed in the plastid tree is due to an acceleration of the evolutionary rate in one short deep internal branch, implying that more phylogenetic signal per position is available to resolve the Ochrophyta radiation in the plastid than in the nuclear data set. Our work therefore suggests that, in order to resolve radiations, beyond the obvious use of data sets with more positions, we need to continue developing models of sequence evolution that better extract the phylogenetic signal and design methods to search for genes/characters that contain more signal specifically for short internal branches.

Palantir: a springboard for the analysis of secondary metabolite gene clusters in large-scale genome mining projects.

SUMMARY: To support small and large-scale genome mining projects, we present Post-processing Analysis tooLbox for ANTIsmash Reports (Palantir), a dedicated software suite for handling and refining secondary metabolite biosynthetic gene cluster (BGC) data annotated with the popular antiSMASH pipeline. Palantir provides new functionalities building on NRPS/PKS predictions from antiSMASH, such as improved BGC annotation, module delineation and easy access to sub-sequences at different levels (cluster, gene, module and domain). Moreover, it can parse user-provided antiSMASH reports and reformat them for direct use or storage in a relational database. AVAILABILITY AND IMPLEMENTATION: Palantir is released both as a Perl API available on CPAN (https://metacpan.org/release/Bio-Palantir) and as a web application (http://palantir.uliege.be). As a practical use case, the web interface also features a database built from the mining of 1616 cyanobacterial genomes, of which 1488 were predicted to encode at least one BGC. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Trophic state alters the mechanism whereby energetic coupling between photosynthesis and respiration occurs in Euglena gracilis.

The coupling between mitochondrial respiration and photosynthesis plays an important role in the energetic physiology of green plants and some secondary-red photosynthetic eukaryotes (diatoms), allowing an efficient CO2 assimilation and optimal growth. Using the flagellate Euglena gracilis, we first tested if photosynthesis-respiration coupling occurs in this species harbouring secondary green plastids (i.e. originated from an endosymbiosis between a green alga and a phagotrophic euglenozoan). Second, we tested how the trophic state (mixotrophy and photoautotrophy) of the cell alters the mechanisms involved in the photosynthesis-respiration coupling. Energetic coupling between photosynthesis and respiration was determined by testing the effect of respiratory inhibitors on photosynthesis, and measuring the simultaneous variation of photosynthesis and respiration rates as a function of temperature (i.e. thermal response curves). The mechanism involved in the photosynthesis-respiration coupling was assessed by combining proteomics, biophysical and cytological analyses. Our work shows that there is photosynthesis-respiration coupling and membrane contacts between mitochondria and chloroplasts in E. gracilis. However, whereas in mixotrophy adjustment of the chloroplast ATP/NADPH ratio drives the interaction, in photoautotrophy the coupling is conditioned by CO2 limitation and photorespiration. This indicates that maintenance of photosynthesis-respiration coupling, through plastic metabolic responses, is key to E. gracilis functioning under changing environmental conditions.

Exploring syntenic conservation across genomes for phylogenetic studies of organisms subjected to horizontal gene transfers: A case study with Cyanobacteria and cyanolichens.

Understanding the evolutionary history of symbiotic Cyanobacteria at a fine scale is essential to unveil patterns of associations with their hosts and factors driving their spatiotemporal interactions. As for bacteria in general, Horizontal Gene Transfers (HGT) are expected to be rampant throughout their evolution, which justified the use of single-locus phylogenies in macroevolutionary studies of these photoautotrophic bacteria. Genomic approaches have greatly increased the amount of molecular data available, but the selection of orthologous, congruent genes that are more likely to reflect bacterial macroevolutionary histories remains problematic. In this study, we developed a synteny-based approach and searched for Collinear Orthologous Regions (COR), under the assumption that genes that are present in the same order and orientation across a wide monophyletic clade are less likely to have undergone HGT. We searched sixteen reference Nostocales genomes and identified 99 genes, part of 28 COR comprising three to eight genes each. We then developed a bioinformatic pipeline, designed to minimize inter-genome contamination and processed twelve Nostoc-associated lichen metagenomes. This reduced our original dataset to 90 genes representing 25 COR, which were used to infer phylogenetic relationships within Nostocales and among lichenized Cyanobacteria. This dataset was narrowed down further to 71 genes representing 22 COR by selecting only genes part of one (largest) operon per COR. We found a relatively high level of congruence among trees derived from the 90-gene dataset, but congruence was only slightly higher among genes within a COR compared to genes across COR. However, topological congruence was significantly higher among the 71 genes part of one operon per COR. Nostocales phylogenies resulting from concatenation and species tree approaches based on the 90- and 71-gene datasets were highly congruent, but the most highly supported result was obtained when using synteny, collinearity, and operon information (i.e., 71-gene dataset) as gene selection criteria, which outperformed larger datasets with more genes.

Evaluating the usefulness of alignment filtering methods to reduce the impact of errors on evolutionary inferences.

BACKGROUND: Multiple Sequence Alignments (MSAs) are the starting point of molecular evolutionary analyses. Errors in MSAs generate a non-historical signal that can lead to incorrect inferences. Therefore, numerous efforts have been made to reduce the impact of alignment errors, by improving alignment algorithms and by developing methods to filter out poorly aligned regions. However, MSAs do not only contain alignment errors, but also primary sequence errors. Such errors may originate from sequencing errors, from assembly errors, or from erroneous structural annotations (such as incorrect intron/exon boundaries). Even though their existence is acknowledged, the impact of primary sequence errors on evolutionary inference is poorly characterized. RESULTS: In a first step to fill this gap, we have developed a program called HmmCleaner, which detects and eliminates these errors from MSAs. It uses profile hidden Markov models (pHMM) to identify sequence segments that poorly fit their MSA and selectively removes them. We assessed its performances using > 700 amino-acid MSAs from prokaryotes and eukaryotes, in which we introduced several types of simulated primary sequence errors. The sensitivity of HmmCleaner towards simulated primary sequence errors was > 95%. In a second step, we compared the impact of segment filtering software (HmmCleaner and PREQUAL) relative to commonly used block-filtering software (BMGE and TrimAI) on evolutionary analyses. Using real data from vertebrates, we observed that segment-filtering methods improve the quality of evolutionary inference more than the currently used block-filtering methods. The formers were especially effective at improving branch length inferences, and at reducing false positive rate during detection of positive selection. CONCLUSIONS: Segment filtering methods such as HmmCleaner accurately detect simulated primary sequence errors. Our results suggest that these errors are more detrimental than alignment errors. However, they also show that stochastic (sampling) error is predominant in single-gene evolutionary inferences. Therefore, we argue that MSA filtering should focus on segment instead of block removal and that more studies are required to find the optimal balance between accuracy improvement and stochastic error increase brought by data removal.

Coding and noncoding variants in HFM1, MLH3, MSH4, MSH5, RNF212, and RNF212B affect recombination rate in cattle.

We herein study genetic recombination in three cattle populations from France, New Zealand, and the Netherlands. We identify 2,395,177 crossover (CO) events in 94,516 male gametes, and 579,996 CO events in 25,332 female gametes. The average number of COs was found to be larger in males (23.3) than in females (21.4). The heritability of global recombination rate (GRR) was estimated at 0.13 in males and 0.08 in females, with a genetic correlation of 0.66 indicating that shared variants are influencing GRR in both sexes. A genome-wide association study identified seven quantitative trait loci (QTL) for GRR. Fine-mapping following sequence-based imputation in 14,401 animals pinpointed likely causative coding (5) and noncoding (1) variants in genes known to be involved in meiotic recombination (HFM1, MSH4, RNF212, MLH3, MSH5) for 5/7 QTL, and noncoding variants (3) in RNF212B for 1/7 QTL. This suggests that this RNF212 paralog might also be involved in recombination. Most of the identified mutations had significant effects in both sexes, with three of them each accounting for ∼10% of the genetic variance in males.

The African coelacanth genome provides insights into tetrapod evolution.

The discovery of a living coelacanth specimen in 1938 was remarkable, as this lineage of lobe-finned fish was thought to have become extinct 70 million years ago. The modern coelacanth looks remarkably similar to many of its ancient relatives, and its evolutionary proximity to our own fish ancestors provides a glimpse of the fish that first walked on land. Here we report the genome sequence of the African coelacanth, Latimeria chalumnae. Through a phylogenomic analysis, we conclude that the lungfish, and not the coelacanth, is the closest living relative of tetrapods. Coelacanth protein-coding genes are significantly more slowly evolving than those of tetrapods, unlike other genomic features. Analyses of changes in genes and regulatory elements during the vertebrate adaptation to land highlight genes involved in immunity, nitrogen excretion and the development of fins, tail, ear, eye, brain and olfaction. Functional assays of enhancers involved in the fin-to-limb transition and in the emergence of extra-embryonic tissues show the importance of the coelacanth genome as a blueprint for understanding tetrapod evolution.

Number of inadvertent RNA targets for morpholino knockdown in Danio rerio is largely underestimated: evidence from the study of Ser/Arg-rich splicing factors.

Although the involvement of Ser/Arg-rich (SR) proteins in RNA metabolism is well documented, their role in vertebrate development remains elusive. We, therefore, elected to take advantage of the zebrafish model organism to study the SR genes' functions using the splicing morpholino (sMO) microinjection and the programmable site-specific nucleases. Consistent with previous research, we revealed discrepancies between the mutant and morphant phenotypes and we show that these inconsistencies may result from a large number of unsuspected inadvertent morpholino RNA targets. While microinjection of MOs directed against srsf5a (sMOsrsf5a) led to developmental defects, the corresponding homozygous mutants did not display any phenotypic traits. Furthermore, microinjection of sMOsrsf5a into srsf5a-/- led to the previously observed morphant phenotype. Similar findings were observed for other SR genes. sMOsrsf5a alternative target genes were identified using deep mRNA sequencing. We uncovered that only 11 consecutive bases complementary to sMOsrsf5a are sufficient for binding and subsequent blocking of splice sites. In addition, we observed that sMOsrsf5a secondary targets can be reduced by increasing embryos growth temperature after microinjection. Our data contribute to the debate about MO specificity, efficacy and the number of unknown targeted sequences.

Inferring the shallow phylogeny of true salamanders (Salamandra) by multiple phylogenomic approaches.

The rise of high-throughput sequencing techniques provides the unprecedented opportunity to analyse controversial phylogenetic relationships in great depth, but also introduces a risk of being misinterpreted by high node support values influenced by unevenly distributed missing data or unrealistic model assumptions. Here, we use three largely independent phylogenomic data sets to reconstruct the controversial phylogeny of true salamanders of the genus Salamandra, a group of amphibians providing an intriguing model to study the evolution of aposematism and viviparity. For all six species of the genus Salamandra, and two outgroup species from its sister genus Lyciasalamandra, we used RNA sequencing (RNAseq) and restriction site associated DNA sequencing (RADseq) to obtain data for: (1) 3070 nuclear protein-coding genes from RNAseq; (2) 7440 loci obtained by RADseq; and (3) full mitochondrial genomes. The RNAseq and RADseq data sets retrieved fully congruent topologies when each of them was analyzed in a concatenation approach, with high support for: (1) S. infraimmaculata being sister group to all other Salamandra species; (2) S. algira being sister to S. salamandra; (3) these two species being the sister group to a clade containing S. atra, S. corsica and S. lanzai; and (4) the alpine species S. atra and S. lanzai being sister taxa. The phylogeny inferred from the mitochondrial genome sequences differed from these results, most notably by strongly supporting a clade containing S. atra and S. corsica as sister taxa. A different placement of S. corsica was also retrieved when analysing the RNAseq and RADseq data under species tree approaches. Closer examination of gene trees derived from RNAseq revealed that only a low number of them supported each of the alternative placements of S. atra. Furthermore, gene jackknife support for the S. atra - S. lanzai node stabilized only with very large concatenated data sets. The phylogeny of true salamanders thus provides a compelling example of how classical node support metrics such as bootstrap and Bayesian posterior probability can provide high confidence values in a phylogenomic topology even if the phylogenetic signal for some nodes is spurious, highlighting the importance of complementary approaches such as gene jackknifing. Yet, the general congruence among the topologies recovered from the RNAseq and RADseq data sets increases our confidence in the results, and validates the use of phylotranscriptomic approaches for reconstructing shallow relationships among closely related taxa. We hypothesize that the evolution of Salamandra has been characterized by episodes of introgressive hybridization, which would explain the difficulties of fully reconstructing their evolutionary relationships.

High-throughput sequencing analysis reveals the genetic diversity of different regions of the murine norovirus genome during in vitro replication.

In this study, we report the genetic diversity and nucleotide mutation rates of five representative regions of the murine norovirus genome during in vitro passages. The mutation rates were similar in genomic regions encompassing partial coding sequences for non-structural (NS) 1-2, NS5, NS6, NS7 proteins within open reading frame (ORF) 1. In a region encoding a portion of the major capsid protein (VP1) within ORF2 (also including the ORF4 region) and a portion of the minor structural protein (VP2), the mutation rates were estimated to be at least one order of magnitude higher. The VP2 coding region was found to have the highest mutation rate.

On the use of high-throughput sequencing for the study of cyanobacterial diversity in Antarctic aquatic mats.

The study of Antarctic cyanobacterial diversity has been mostly limited to morphological identification and traditional molecular techniques. High-throughput sequencing (HTS) allows a much better understanding of microbial distribution in the environment, but its application is hampered by several methodological and analytical challenges. In this work, we explored the use of HTS as a tool for the study of cyanobacterial diversity in Antarctic aquatic mats. Our results highlight the importance of using artificial communities to validate the parameters of the bioinformatics procedure used to analyze natural communities, since pipeline-dependent biases had a strong effect on the observed community structures. Analysis of microbial mats from five Antarctic lakes and an aquatic biofilm from the Sub-Antarctic showed that HTS is a valuable tool for the assessment of cyanobacterial diversity. The majority of the operational taxonomic units retrieved were related to filamentous taxa such as Leptolyngbya and Phormidium, which are common genera in Antarctic lacustrine microbial mats. However, other phylotypes related to different taxa such as Geitlerinema, Pseudanabaena, Synechococcus, Chamaesiphon, Calothrix, and Coleodesmium were also found. Results revealed a much higher diversity than what had been reported using traditional methods and also highlighted remarkable differences between the cyanobacterial communities of the studied lakes. The aquatic biofilm from the Sub-Antarctic had a distinct cyanobacterial community from the Antarctic lakes, which in turn displayed a salinity-dependent community structure at the phylotype level.

Impact of missing data on phylogenies inferred from empirical phylogenomic data sets.

Progress in sequencing technology allows researchers to assemble ever-larger supermatrices for phylogenomic inference. However, current phylogenomic studies often rest on patchy data sets, with some having 80% missing (or ambiguous) data or more. Though early simulations had suggested that missing data per se do not harm phylogenetic inference when using sufficiently large data sets, Lemmon et al. (Lemmon AR, Brown JM, Stanger-Hall K, Lemmon EM. 2009. The effect of ambiguous data on phylogenetic estimates obtained by maximum likelihood and Bayesian inference. Syst Biol. 58:130-145.) have recently cast doubt on this consensus in a study based on the introduction of parsimony-uninformative incomplete characters. In this work, we empirically reassess the issue of missing data in phylogenomics while exploring possible interactions with the model of sequence evolution. First, we note that parsimony-uninformative incomplete characters are actually informative in a probabilistic framework. A reanalysis of Lemmon's data set with this in mind gives a very different interpretation of their results and shows that some of their conclusions may be unfounded. Second, we investigate the effect of the progressive introduction of missing data in a complete supermatrix (126 genes × 39 species) capable of resolving animal relationships. These analyses demonstrate that missing data perturb phylogenetic inference slightly beyond the expected decrease in resolving power. In particular, they exacerbate systematic errors by reducing the number of species effectively available for the detection of multiple substitutions. Consequently, large sparse supermatrices are more sensitive to phylogenetic artifacts than smaller but less incomplete data sets, which argue for experimental designs aimed at collecting a modest number (~50) of highly covered genes. Our results further confirm that including incomplete yet short-branch taxa (i.e., slowly evolving species or close outgroups) can help to eschew artifacts, as predicted by simulations. Finally, it appears that selecting an adequate model of sequence evolution (e.g., the site-heterogeneous CAT model instead of the site-homogeneous WAG model) is more beneficial to phylogenetic accuracy than reducing the level of missing data.

Assessing the effect of the CLPG mutation on the microRNA catalog of skeletal muscle using high-throughput sequencing.

The callipyge phenotype is a monogenic muscular hypertrophy that is only expressed in heterozygous sheep receiving the CLPG mutation from their sire. The wild-type phenotype of CLPG/CLPG animals is thought to result from translational inhibition of paternally expressed DLK1 transcripts by maternally expressed miRNAs. To identify the miRNA responsible for this trans effect, we used high-throughput sequencing to exhaustively catalog miRNAs expressed in skeletal muscle of sheep of the four CLPG genotypes. We have identified 747 miRNA species of which 110 map to the DLK1-GTL2 or callipyge domain. We demonstrate that the latter are imprinted and preferentially expressed from the maternal allele. We show that the CLPG mutation affects their level of expression in cis (∼3.2-fold increase) as well as in trans (∼1.8-fold increase). In CLPG/CLPG animals, miRNAs from the DLK1-GTL2 domain account for ∼20% of miRNAs in skeletal muscle. We show that the CLPG genotype affects the levels of A-to-I editing of at least five pri-miRNAs of the DLK1-GTL2 domain, but that levels of editing of mature miRNAs are always minor. We present suggestive evidence that the miRNAs from the domain target the ORF of DLK1, thereby causing the trans inhibition underlying polar overdominance. We highlight the limitations of high-throughput sequencing for digital gene expression profiling as a result of biased and inconsistent amplification of specific miRNAs.

A single ancient origin for prototypical serine/arginine-rich splicing factors.

Eukaryotic precursor mRNA splicing is a process involving a very complex RNA-protein edifice. Serine/arginine-rich (SR) proteins play essential roles in precursor mRNA constitutive and alternative splicing and have been suggested to be crucial in plant-specific forms of developmental regulation and environmental adaptation. Despite their functional importance, little is known about their origin and evolutionary history. SR splicing factors have a modular organization featuring at least one RNA recognition motif (RRM) domain and a carboxyl-terminal region enriched in serine/arginine dipeptides. To investigate the evolution of SR proteins, we infer phylogenies for more than 12,000 RRM domains representing more than 200 broadly sampled organisms. Our analyses reveal that the RRM domain is not restricted to eukaryotes and that all prototypical SR proteins share a single ancient origin, including the plant-specific SR45 protein. Based on these findings, we propose a scenario for their diversification into four natural families, each corresponding to a main SR architecture, and a dozen subfamilies, of which we profile both sequence conservation and composition. Finally, using operational criteria for computational discovery and classification, we catalog SR proteins in 20 model organisms, with a focus on green algae and land plants. Altogether, our study confirms the homogeneity and antiquity of SR splicing factors while establishing robust phylogenetic relationships between animal and plant proteins, which should enable functional analyses of lesser characterized SR family members, especially in green plants.

Microbiota Profiling on Veterinary Faculty Restroom Surfaces and Source Tracking.

In this study, we aimed to develop a comprehensive microbial source amplicon database tailored for source tracking in veterinary settings. We rigorously tested our locally curated source tracking database by selecting a frequently accessed environment by veterinary students and veterinarians. By exploring the composition of resident microbiota and identifying potential sources of contamination, including animals, the environment, and human beings, we aimed to provide valuable insights into the dynamics of microbial transmission within veterinary facilities. The 16S rDNA amplicon sequencing was used to determine the bacterial taxonomic profiles of restroom surfaces. Bacterial sources were identified by linking our metadata-enriched local database to the microbiota profiling analysis using high-quality sequences. Microbiota profiling shows the dominance of four phyla: Actinobacteria, Bacteroidetes, Proteobacteria, and Firmicutes. If the restroom cleaning process did not appear to impact microbiota composition, significant differences regarding bacterial distribution were observed between male and female users in different sampling campaigns. Combining 16S rDNA profiling to our specific sources labeling pipeline, we found aquatic and human sources were the primary environment keywords in our campaigns. The probable presence of known animal sources (bovids, insects, equids, suids…) associated with bacterial genera such as Chryseobacterium, Bergeyella, Fibrobacter, and Syntrophococcus was also involved in restroom surfaces, emphasizing the proximity between these restrooms and the exchange of bacteria between people involved in animals handling. To summarize, we have demonstrated that DNA sequence-based source tracking may be integrated with high-throughput bacterial community analysis to enrich microbial investigation of potential bacterial contamination sources, especially for little known or poorly identified taxa. However, more research is needed to determine the tool's utility in other applications.

Patrocles: a database of polymorphic miRNA-mediated gene regulation in vertebrates.

The Patrocles database (http://www.patrocles.org/) compiles DNA sequence polymorphisms (DSPs) that are predicted to perturb miRNA-mediated gene regulation. Distinctive features include: (i) the coverage of seven vertebrate species in its present release, aiming for more when information becomes available, (ii) the coverage of the three compartments involved in the silencing process (i.e. targets, miRNA precursors and silencing machinery), (iii) contextual information that enables users to prioritize candidate 'Patrocles DSPs', including graphical information on miRNA-target coexpression and eQTL effect of genotype on target expression levels, (iv) the inclusion of Copy Number Variants and eQTL information that affect miRNA precursors as well as genes encoding components of the silencing machinery and (v) a tool (Patrocles finder) that allows the user to determine whether her favorite DSP may perturb miRNA-mediated gene regulation of custom target sequences. To support the biological relevance of Patrocles' content, we searched for signatures of selection acting on 'Patrocles single nucleotide polymorphisms (pSNPs)' in human and mice. As expected, we found a strong signature of purifying selection against not only SNPs that destroy conserved target sites but also against SNPs that create novel, illegitimate target sites, which is reminiscent of the Texel mutation in sheep.

Contamination detection in genomic data: more is not enough.

The decreasing cost of sequencing and concomitant augmentation of publicly available genomes have created an acute need for automated software to assess genomic contamination. During the last 6 years, 18 programs have been published, each with its own strengths and weaknesses. Deciding which tools to use becomes more and more difficult without an understanding of the underlying algorithms. We review these programs, benchmarking six of them, and present their main operating principles. This article is intended to guide researchers in the selection of appropriate tools for specific applications. Finally, we present future challenges in the developing field of contamination detection.

An Extended Reservoir of Class-D Beta-Lactamases in Non-Clinical Bacterial Strains.

Bacterial genes coding for antibiotic resistance represent a major issue in the fight against bacterial pathogens. Among those, genes encoding beta-lactamases target penicillin and related compounds such as carbapenems, which are critical for human health. Beta-lactamases are classified into classes A, B, C, and D, based on their amino acid sequence. Class D enzymes are also known as OXA beta-lactamases, due to the ability of the first enzymes described in this class to hydrolyze oxacillin. While hundreds of class D beta-lactamases with different activity profiles have been isolated from clinical strains, their nomenclature remains very uninformative. In this work, we have carried out a comprehensive survey of a reference database of 80,490 genomes and identified 24,916 OXA-domain containing proteins. These were deduplicated and their representative sequences clustered into 45 non-singleton groups derived from a phylogenetic tree of 1,413 OXA-domain sequences, including five clusters that include the C-terminal domain of the BlaR membrane receptors. Interestingly, 801 known class D beta-lactamases fell into only 18 clusters. To probe the unknown diversity of the class, we selected 10 protein sequences in 10 uncharacterized clusters and studied the activity profile of the corresponding enzymes. A beta-lactamase activity could be detected for seven of them. Three enzymes (OXA-1089, OXA-1090 and OXA-1091) were active against oxacillin and two against imipenem. These results indicate that, as already reported, environmental bacteria constitute a large reservoir of resistance genes that can be transferred to clinical strains, whether through plasmid exchange or hitchhiking with the help of transposase genes. IMPORTANCE The transmission of genes coding for resistance factors from environmental to nosocomial strains is a major component in the development of bacterial resistance toward antibiotics. Our survey of class D beta-lactamase genes in genomic databases highlighted the high sequence diversity of the enzymes that are able to recognize and/or hydrolyze beta-lactam antibiotics. Among those, we could also identify new beta-lactamases that are able to hydrolyze carbapenems, one of the last resort antibiotic families used in human antimicrobial chemotherapy. Therefore, it can be expected that the use of this antibiotic family will fuel the emergence of new beta-lactamases into clinically relevant strains.

A Fast-Growing Oleaginous Strain of Coelastrella Capable of Astaxanthin and Canthaxanthin Accumulation in Phototrophy and Heterotrophy.

Considering the importance of microalgae as a promising feedstock for the production of both low- and high-value products, such as lipids and pigments, it is desirable to isolate strains which simultaneously accumulate these two types of products and grow in various conditions in order to widen their biotechnological applicability. A novel freshwater strain from the genus Coelastrella was isolated in Belgium. Compared to other Coelastrella species, the isolate presented rapid growth in phototrophy, dividing 3.5 times per day at a light intensity of 400 µmol·m-2·s-1 and 5% CO2. In addition, nitrogen depletion was associated with the accumulation of astaxanthin, canthaxanthin, and fatty acids, which reached ~30% of dry weight, and a majority of SFAs and MUFAs, which are good precursors for biodiesel. This strain also accumulated astaxanthin and canthaxanthin in heterotrophy. Although the content was very low in this latter condition, it is an interesting feature considering the biotechnological potential of the microalgal heterotrophic growth. Thus, due to its rapid growth in the light, its carotenogenesis, and its fatty acids characteristics, the newly identified Coelastrella strain could be considered as a potential candidate for biorefinery purposes of both low- and high-values products.

Phylogenomic Analyses of Snodgrassella Isolates from Honeybees and Bumblebees Reveal Taxonomic and Functional Diversity.

Snodgrassella is a genus of Betaproteobacteria that lives in the gut of honeybees (Apis spp.) and bumblebees (Bombus spp). It is part of a conserved microbiome that is composed of a few core phylotypes and is essential for bee health and metabolism. Phylogenomic analyses using whole-genome sequences of 75 Snodgrassella strains from 4 species of honeybees and 14 species of bumblebees showed that these strains formed a monophyletic lineage within the Neisseriaceae family, that Snodgrassella isolates from Asian honeybees diverged early from the other species in their evolution, that isolates from honeybees and bumblebees were well separated, and that this genus consists of at least seven species. We propose to formally name two new Snodgrassella species that were isolated from bumblebees: i.e., Snodgrassella gandavensis sp. nov. and Snodgrassella communis sp. nov. Possible evolutionary scenarios for 107 species- or group-specific genes revealed very limited evidence for horizontal gene transfer. Functional analyses revealed the importance of small proteins, defense mechanisms, amino acid transport and metabolism, inorganic ion transport and metabolism and carbohydrate transport and metabolism among these 107 specific genes. IMPORTANCE The microbiome of honeybees (Apis spp.) and bumblebees (Bombus spp.) is highly conserved and represented by few phylotypes. This simplicity in taxon composition makes the bee's microbiome an emergent model organism for the study of gut microbial communities. Since the description of the Snodgrassella genus, which was isolated from the gut of honeybees and bumblebees in 2013, a single species (i.e., Snodgrassella alvi), has been named. Here, we demonstrate that this genus is actually composed of at least seven species, two of which (Snodgrassella gandavensis sp. nov. and Snodgrassella communis sp. nov.) are formally described and named in the present publication. We also report the presence of 107 genes specific to Snodgrassella species, showing notably the importance of small proteins and defense mechanisms in this genus.