L'objectif a été d'évaluer à l'aide de techniques de phylogénie moléculaire la diversité bactérienne non cultivable dans la neige du Mont Blanc (MtBl) contenant de la poussière saharienne déposée au cours de quatre événements pendant la période 2006-2009. La diversité bactérienne a été évaluée par digestion enzymatique d'ADN et le séquençage partiel de la région V3-V5 du gène et des séquences complètes d'ARNr 16S. Nous avons étudié : (i) de la neige (MtBl) ne contenant aucune poussière saharienne (par ADNr et ARNr) ; (ii) du sable du Sahara (Tunisie) ; (iii) des poussières sahariennes collectées à Grenoble (200 m ASL) et de la neige récupérée au MtBl (4250 m ASL). Le contenu en espèces et en phylotypes dominants a varié dans la neige du MtBl associée aux quatre dépôts de poussière saharienne sur 3 années. Les phylotypes dominants appartiennent aux classes des Actinobacteria, Proteobacteria, Firmicutes, Deinococcus-Thermus, Bacteroidetes et les Cyanobactéria. Cette variabilité semble être davantage causée par les conditions de transport de la poussière. Quinze phylotypes ont été reconnus comme candidats pour colonisation de la neige. Les représentants des genres Massilia, Tumebacillus, Phormidium et Stigonema sont les candidats les plus probables pour la propagation dans la neige. Parmi tous les phylotypes identifiés, 10% ont été classés comme HA-phylotypes basés sur leur similitude (≥98%) avec les représentants du microbiome humain et 11% ont montré moins de 90% de similarité avec les taxons connus. Le séquençage partiel (V3-V5) et complet des gènes codant l'ARNr 16S a permis de décrire la diversité microbienne plus complète et d'en obtenir une image plus détaillée.

L'objectif de cette étude est d'évaluer à l'aide des techniques de phylogénie moléculaire la diversité bactérienne non cultivable dans le manteau neigeux du Mont Blanc (MtBl) contenant de la poussière saharienne déposée au cours de quatre événements pendant la période de 2006 à 2009. Le but final est de découvrir les bactéries qui pourraient être impliquées dans l'établissement du microbiote neigeux.

The objective of this study is to assess the uncultured bacterial diversity in the snowpack of the Mont Blanc (MtBl) glacier containing Saharan dust deposited during four dust events during the period 2006 – 2009 by means of molecular phylogenetics. The final goal is to discover the bacteria that could be involved in the establishment of snow microbiota. Bacterial diversity was evaluated using rybotyping and subsequent sequencing of partial (V3-V5) and full-length 16S rRNA genes. For comparison purpose we also studied following samples: “clean” MtBl snow containing no Saharan dust; Saharan sand collected in Tunisia; Saharan dust collected in Grenoble (200 m a.s.l.) and recovered later on MtBl (4250 m a.s.l.). In order to verify possible microbial activity in situ, both rDNA and rRNA approaches were implemented for the “clean” snow sample. To evaluate the survival/colonization abilities of bacterial phylotypes recovered in snow samples with Saharan dust, we analyzed their closest strain physiology as well as sources of environmental clones using a threshold of ≥98% sequence similarity. For the result interpretation, we also used data on dust elemental composition and dust particles size distribution. As a result 8 clone libraries (including rRNA-based one) were constructed using V3-V5 16S rRNA gene sequences for 5 snow samples (4 with Saharan dust and one “clean”), sample of Saharan dust collected in Grenoble and Saharan sand sample. Furthermore, 4 clone libraries were generated using full-length 16S rRNA gene amplicons obtained from 4 of the above snow samples (three with Saharan dust and one ‘clean'). Species content and dominant phylotypes and their assigning to major divisions varied significantly in alpine snow on a Mont Blanc glacier associated with four depositions of Saharan dust over a 3-year. Dominant phylotypes revealed are belonged to Actinobacteria, Proteobactreia, Firmicutes, Deinococcus-Thermus, Bacteroidetes and Cyanobacteria. Such variability was detected by both partial and full-length 16S rRNA gene sequencing and seems to be caused more by conditions of dust transport than bacterial load from the original dust source. Also the preservation period of dust in snowpack could affect the species composition. Thirteen icy phylotypes as candidates into snow microbiota establishing were recognized in snow containing Saharan dust and only two in “clean” snow sample. Of them, both dominant and minor phylotypes of Cyanobacteria, Proteobacteria, Actinobacteria и Firmicutes were revealed. Data on the closest strain physiology of recognized icy phylotypes suggests that representatives of genera Massilia (Betaproteobacteria), Tumebacillus (Firmicutes), Phormidium and Stigonema (both Cyanobacteria) are most relevant findings in terms of propagation in snow. By analyzing 16S rRNA from the “clean” snow containing no Saharan dust and comparing the data with those obtained for 16S rDNA library, it has been shown that Stigonema-like cyanobacterium identified could be propagating in snow at subzero temperature. Among all identified phylotypes, 10% were categorized as HA-phylotypes based on their con-specificity (≥98% similarity) with normal (non-pathogenic) human microbiome representatives. Furthermore, 11% out of all phylotypes showed less than 90% similarity with known taxa, thus, presenting novel taxa. Sequencing of both partial (V3-V5) and full-length 16S rRNA genes permitted to describe microbial diversity more fully and get more detailed picture. / The objective of this study is to assess the uncultured bacterial diversity in the snowpack of the Mont Blanc (MtBl) glacier containing Saharan dust deposited during four dust events during the period 2006 – 2009 by means of molecular phylogenetics. The final goal is to discover the bacteria that could be involved in the establishment of snow microbiota. Bacterial diversity was evaluated using rybotyping and subsequent sequencing of partial (V3-V5) and full-length 16S rRNA genes. For comparison purpose we also studied following samples: “clean” MtBl snow containing no Saharan dust; Saharan sand collected in Tunisia; Saharan dust collected in Grenoble (200 m a.s.l.) and recovered later on MtBl (4250 m a.s.l.). In order to verify possible microbial activity in situ, both rDNA and rRNA approaches were implemented for the “clean” snow sample. To evaluate the survival/colonization abilities of bacterial phylotypes recovered in snow samples with Saharan dust, we analyzed their closest strain physiology as well as sources of environmental clones using a threshold of ≥98% sequence similarity. For the result interpretation, we also used data on dust elemental composition and dust particles size distribution. As a result 8 clone libraries (including rRNA-based one) were constructed using V3-V5 16S rRNA gene sequences for 5 snow samples (4 with Saharan dust and one “clean”), sample of Saharan dust collected in Grenoble and Saharan sand sample. Furthermore, 4 clone libraries were generated using full-length 16S rRNA gene amplicons obtained from 4 of the above snow samples (three with Saharan dust and one ‘clean'). Species content and dominant phylotypes and their assigning to major divisions varied significantly in alpine snow on a Mont Blanc glacier associated with four depositions of Saharan dust over a 3-year. Dominant phylotypes revealed are belonged to Actinobacteria, Proteobactreia, Firmicutes, Deinococcus-Thermus, Bacteroidetes and Cyanobacteria. Such variability was detected by both partial and full-length 16S rRNA gene sequencing and seems to be caused more by conditions of dust transport than bacterial load from the original dust source. Also the preservation period of dust in snowpack could affect the species composition. Thirteen icy phylotypes as candidates into snow microbiota establishing were recognized in snow containing Saharan dust and only two in “clean” snow sample. Of them, both dominant and minor phylotypes of Cyanobacteria, Proteobacteria, Actinobacteria и Firmicutes were revealed. Data on the closest strain physiology of recognized icy phylotypes suggests that representatives of genera Massilia (Betaproteobacteria), Tumebacillus (Firmicutes), Phormidium and Stigonema (both Cyanobacteria) are most relevant findings in terms of propagation in snow. By analyzing 16S rRNA from the “clean” snow containing no Saharan dust and comparing the data with those obtained for 16S rDNA library, it has been shown that Stigonema-like cyanobacterium identified could be propagating in snow at subzero temperature. Among all identified phylotypes, 10% were categorized as HA-phylotypes based on their con-specificity (≥98% similarity) with normal (non-pathogenic) human microbiome representatives. Furthermore, 11% out of all phylotypes showed less than 90% similarity with known taxa, thus, presenting novel taxa. Sequencing of both partial (V3-V5) and full-length 16S rRNA genes permitted to describe