O presente trabalho teve como objetivos caracterizar uma coleção de 98 bactérias isoladas de amostras de petróleo e água de formação de reservatórios da Bacia de Campos (RJ) utilizando técnicas de taxonomia molecular e avaliar o potencial de degradação de biomarcadores do petróleo. O sequenciamento e análise filogenética do gene RNAr 16S revelaram Bacillus firmus, megaterium, pumilus, sphaericus, simplex, cereus/B. thuringiensis Marinobacter lutaoensis, Halomonas shengliensis/H. alimentaria/ H.campisalis, Citreicella thiooxidans, Stenotrophomonas maltophilia, Achromobacter xylosoxidans, Micrococcus luteus, Kocuria rosea, Streptomyces alboniger/S. chartreusis /S. moderatus, Staphylococcus hominis e Staphylococcus pasteuri/S. warneri. Os resultados evidenciaram a preferência pela biotransformação do ácido nonadecanóico e esqualano. A caracterização da microbiota presente nos reservatórios e avaliação do potencial de biodegradação pode contribuir para fornecer subsídios para estudos futuros sobre os mecanismos biológicos responsáveis pela biodegradação do petróleo. / This study is aimed to characterize a collection of 98 bacteria isolated from oil and formation water samples derived from reservoirs of the Campos Basin (RJ) using molecular biology-based techniques and to evaluate the degradation potential of petroleum biomarkers. Further sequencing and phylogenetic analysis of 16S rRNA genes revealed species of Bacillus firmus, megaterium, pumilus, sphaericus, simplex, cereus/thuringiensis, Marinobacter lutaoensis, Halomonas shengliensis/H. alimentaria/H. campisalis, Citreicella thiooxidans, Stenotrophomonas maltophilia, Achromobacter xylosoxidans, Micrococcus luteus, Kocuria rosea, Streptomyces alboniger/S. chartreusis/S. moderatus, Staphylococcus hominis and Staphylococcus pasteuri/S. warneri. The results showed the preference of bacteria for the biotransformation of nonadecanoic acid and squalane. The characterization of the microbiota associated to reservoirs and the evaluation of their biodegradation potential may provide subsidies for future studies about the biological mechanisms responsible for petroleum biodegradation.

A afiliação genérica de Fischerella e Hapalosiphon é problemática devido à instabilidade dos caracteres morfológicos. Os gêneros Fischerella e Hapalosiphon são diferenciados pela presença de tricoma multisseriado e uni ou bisseriado, respectivamente. Porém, geneticamente esses caracteres não se mostraram diacríticos para diferenciar gêneros. Estudos moleculares de linhagens isoladas de ecossistemas brasileiros são escassos para Fischerella e inexistentes para Hapalosiphon. Neste estudo, oito linhagens de cianobactérias, pertencentes à família Hapalosiphonaceae, isoladas de água doce e solos brasileiros foram caracterizadas morfologicamente e geneticamente e analisadas para a produção de substâncias bioativas. As análises morfológicas identificaram cinco morfotípos de Fischerella (CENA19, CENA161, CENA212, CENA213, CENA214) e três de Hapalosiphon (CENA63, CENA71, CENA72). As análises filogenéticas do RNAr 16S usando neighbor-joining (NJ) e máxima verossimilhança (MV) colocaram todas as linhagens isoladas em um agrupamento com alto suporte (reamostragens de 99% NJ e MV) contendo membros da ordem Nostocales. Além disso, as linhagens de Fischerella selecionadas para o estudo agruparam-se em um clado interno com alto valor de reamostragem (100% NJ e 86% MV), com exceção da Fischerella CENA19. A posição dessa estirpe na árvore filogenética indica que necessita de revisão taxonômica. As linhagens de solo Hapalosiphon CENA71 e CENA72 também formaram um clado interno separado (99% NJ e 98% MV), mas a linhagem de água doce CENA63 foi colocada em um clado diferente (com valores de reamostragens de 99% NJ e MV), juntamente com linhagens do gênero Hapalosiphon e Westielopsis prolífica SAG 16.93, oriundas de solo. A comparação das análises filogenéticas individuais de regiões dos genes RNAr 16S, rpoC1, rbcL, tufA, e cpcBA-IGS das três linhagens de Hapalosiphon e de duas linhagens de Fischerella, CENA19 e CENA161, mostrou resultados incongruentes devido as diferentes taxas evolutivas desses genes. No entanto, a análise filogenética concatenada desses genes, mostrou que a Fischerella CENA19 agrupou com as duas linhagens de Hapalosiphon CENA71 e CENA72, com alto valor de reamostragem (100%), enquanto que a Fischerella CENA 161 e a Hapalosiphon CENA63 posicionaram-se cada uma em clados separados. Os resultados indicam que a nomenclatura das linhagens de cianobactérias da família Hapalosiphonaceae necessita de revisão. Os extratos intra e extracelulares das linhagens Fischerella sp. CENA161 e CENA19 e Hapalosiphon sp. CENA71 e CENA72 mostraram efeitos inibitórios no crescimento de bactérias patogênicas. As análises em espectrômetro de massas Q-TOF MS/MS indicaram a putativa presença de aeruginopeptina, cianopeptolina, fischerelina, aeruginosina, oscilapeptilida, microcistinas e ácido tumonóico nos extratos. No extrato intracelular da Fischerella sp. CENA161 identificou-se três ou quatro variantes de microcistinas, LR, LL, FR e/ou M(O)R. Fragmentos dos genes mcyA, mcyB, mcyC, mcyD, mcyE, mcyG e mcyI dessa linhagem foram seqüenciados. Nas duas análises filogenéticas realizadas com sequências de aminoácidos de McyE e sequências concatenadas de McyD, McyE e McyG, as enzimas da microcistina sintetase ficaram agrupadas de acordo com os gêneros de cianobactérias indicando um padrão de evolução / The generic affiliation of Fischerella and Hapalosiphon is problematic due to instability of morphological characters. The Fischerella and Hapalosiphon genera are differentiated by the presence of trichome multisseriate and uni or bisseriate, respectively. However, genetically these characters were not diacritical to distinguish genera. Molecular studies of strains isolated from Brazilian ecosystems are scarce for Fischerella and absent for Hapalosiphon. In this study, eight cyanobacterial strains, belonging to Hapalosiphonaceae family, isolated from Brazilian freshwater and soil were morphologically and genetically characterized and analyzed for bioactive compound productions. The morphological analyses identified five Fischerella (CENA19, CENA161, CENA212, CENA213, CENA214) and three Hapalosiphon (CENA63, CENA71, CENA72) morphotypes. The neighbor-Joining (NJ) and maximum likelihood (ML) phylogenetic analyses of 16S rRNA placed all isolated strains in high supported (99% NJ and ML of bootstrap) cluster containing members of the order Nostocales. Furthermore, the Fischerella strains studied were grouped in an internal clade with high bootstrap value (100% NJ and 86% ML), with exception of Fischerella CENA19. The position of this strain in the phylogenetic tree indicates that it needs taxonomical revision. The soil Hapalosiphon strains CENA71 and CENA72 also formed a separated tight internal clade (99% NJ and 98% ML), but the freshwater strain CENA63 was placed in a different clade (99% NJ and ML of bootstrap value) together with Hapalosiphon strains genera and Westielopsis prolifica SAG 16.93, originated from soil. The comparison of the phylogenetic analyses of individual regions of the genes 16S rRNA, rpoC1, rbcL, tufA, and cpcBA-IGS from the three Hapalosiphon strains and the two Fischerella strains CENA19 and CENA161 showed incongruent results due to different evolutionary rates of these genes. However, the concatenated phylogenetic analysis of these genes, showed that Fischerella CENA19 grouped with the two Hapalosiphon strains CENA71 and CENA72 with high bootstrap value (100%), while Fischerella CENA 161 and Hapalosiphon CENA63 were positionated each one in separate clades. The results indicate that the nomenclature of cyanobacterial strains from the family Hapalosiphonaceae needs revision. The intra and extracellular extracts of the Fischerella sp. strains CENA161 and CENA19 and Hapalosiphon sp. strains CENA71 and CENA72 showed inhibitory effects on the growth of pathogenic bacteria. The analysis in the mass spectrometer Q-TOF MS/MS indicated the presence of aeruginopeptin, cyanopeptolin, fischerellin, aeruginosin, oscillapeptilide, microcystins and tumonoic acid in the extracts. In the intracellular extracts of Fischerella sp. CENA161, three or four variants of microcystins, LR, LL, FR and/or M(O)R, were identified. Fragments of genes mcyA, mcyB, mcyC, mcyD, mcyE, mcyG and mcyI of this strain were sequenced. In both phylogenetic analyses performed with amino acid sequences of McyE and concatenated sequences of McyD, McyE and McyG, the microcystin synthetase enzymes were grouped according to the cyanobacterial genera, indicating a pattern of evolution

O presente trabalho teve por objetivo avaliar a eficiência de remoção do surfactante aniônico alquilbenzeno linear sulfonado (LAS) em reator anaeróbio de leito granular expandido - EGSB (1,5 litros) com recirculação e alimentação com meio mineral. Além de caracterizar filogeneticamente a diversidade de bactérias na presença do surfactante. O sistema foi operado em condição mesofílica em 4 etapas: (I), (II) e (IV) com TDH de 32 horas, e (III) com TDH de 26 horas. Em todas as etapas a DQO foi em média de 609 \'+ OU -\' 137 mg/L e 14 \'+ OU -\' 1,71 mg/L de LAS afluente. As maiores remoções de LAS foram verificada nas etapas II e IV, com valores de 73,6 \'+ OU -\' 5,6% e 63,6 \'+ OU -\' 6,17%, respectivamente de. Na etapa III essa remoção foi de 47,8 \'+ OU -\' 6,2%. Por meio do balanço de massa constatou-se que 56,6% do total de LAS adicionado foram removidos compreendendo 48,4% por biodegradação e 8,2% por adsorção. A remoção de matéria orgânica não foi afetada com a adição do LAS e nem pela exposição prolongada a esse surfactante. Entretanto, a estrutura do grânulo foi comprometida quando da adição do surfactante, observado pelo aumento da concentração de sólidos totais efluente de 0,049 g/L na etapa I (sem LAS), 0,128 g/L na etapa II, 0,064 g/L na etapa III e 0,038 g/L na etapa IV, quando da adição de 14 \'+ OU -\' 1,71 mg LAS/L. Além disso, foi notada diminuição do diâmetro médio dos grânulos no decorrer da operação do reator de 0,36 cm nas etapas I e III para 0,34 cm na etapa IV. Por meio da técnica de tubos múltiplos (NMP) foi constatado aumento das bactérias anaeróbias totais e diminuição das arqueias metanogênicas, em função do tempo de operação do reator. As bactérias redutoras de ferro representaram 8% da biomassa anaeróbia na etapa IV. Por meio do seqüenciamento da região 16S do RNAr para o domínio Bacteria da biomassa da extremidade superior do reator e da biomassa do leito, foi verificado semelhança com os seguintes filos Proteobacteria, Firmicutes e Synergistetes. Notou-se diferença significativa entre as bibliotecas de clones para essas duas amostras. / This study aimed to evaluate the efficiency of removal of linear alkylbenzene sulfonate (LAS) in expanded bed reactor (1.5 liters) using granular sludge (EGSB) with recirculation and feed with mineral medium modified. The system was operated at mesophilic condition in four stages: (I) (II) and (IV) with HRT of 32 hours, and (III) with HRT of 26 hours. At all stages the COD averaged 609 \'+ OR -\' 137 mg/L and 14 \'+ OR -\' 1.71 mg/L LAS influent. The higher removals of LAS were found in stages II and IV, respectively, 73.6 \'+ OR -\' 5.6% and 63.6 \'+ OR -\' 6.17%. In stage III this removal was 47.8 \'+ OR -\' 6.2%. Through mass balance was found that 56.6% of total LAS added were removed by biodegradation comprising 48.4% and 8.2% by adsorption. The organic matter removal was not affected by the addition of LAS and not by prolonged exposure to this surfactant. However, the granule structure was compromised after the addition of surfactant, the observed increase in effluent total solids concentration of 0.049 g/L in stage I (no LAS), 0.128 g/L in stage II, 0.064 g/L in stage III and 0.038 g/L in stage IV when adding 14 \'+ OR -\' 1.71 mg/L. Furthermore, it was noticed significant decrease in mean diameter of the granules during the operation of the reactor of 0.36 cm in stages I and III to 0.34 cm in stage IV. Through the multiple tube method (MPN) was found to increase the total anaerobic bacteria and methanogenic archaea decreased depending on the time of reactor operation. Iron-reducing bacteria accounted for 8% of anaerobic bacteria total in step IV. By sequencing the 16S rRNA for the domain Bacteria biomass from the upper end of the reactor and the biomass of the bed, was found similar to the following phyla Proteobacteria, Firmicutes and Synergistetes. Significant difference was noted between the clone libraries for these two samples.

<p>Род <em>Merodon Meigen</em> (Diptera: Syrphidae) се одликује великм бројем врста које имају&nbsp; функцију у опрашивању&nbsp; биљака. У оквиру овог рода својом разноврсношћу издваја се <em>Merodon aureus</em>&nbsp; група&nbsp; врста коју поред фенотипски различитих одликује и присуство већег броја криптичних&nbsp; врста. Због морфолошке сличности криптичне врсте представљају изазов за таксономе, те су молекуларне методе од посебног значаја. У том светлу примарни циљ овог истраживања је био утврђивање молекуларног диверзитета групе и могућности његове примене у таксономији. Истраживање је било засновано на анализи варијабилности секвенци&nbsp; COI&nbsp; и&nbsp; 28S&nbsp; рРНК гена за 718 јединки 41 описане или новоткривене врсте сирфида. Додатно, тестирана је примењивост мултилокусних&nbsp; ISSR маркера у раздвајању врста&nbsp; <em>M.&nbsp; luteomaculatus</em>&nbsp; комплекса. Узорци су тестирани и на присуство<em> Wolbachia</em> с обзиром да она може утицати на еволуцију митохондријалних гена. Добијени резултати су указали на високу варијабилност секвенци&nbsp; COI&nbsp; гена&nbsp; који се показао корисним у&nbsp; утврђивању граница&nbsp; криптичних&nbsp; врста у анализираним комплексима. Секвенце 28Ѕ рРНК гена у већини случајева нису биле од већег значаја за раздвајање врста, али би могле имати потенцијал у&nbsp; раздвајању комплекса&nbsp; или подгрупа&nbsp; врста као допуна анализи&nbsp; секвенци&nbsp; COI&nbsp; гена. Додатно,&nbsp; ISSR&nbsp; маркери су показали потенцијал за примену у молекуларној таксономији. Све анализиране врсте изузев <em>М. balkanicus</em> су биле заражене&nbsp; <em>Wolbachia,</em> али није утврђена јасна веза између варијабилности&nbsp; секвенци&nbsp; митохондријалног&nbsp; COI гена и инфекционог статуса врста. Утврђен образац генетичке варијабилности је вероватно&nbsp; обликован драстичним климатским променама&nbsp; током Плеистоцена и фрагментацијом хабитата. Да би се донели крајњи закључци о таксономском статусу предложених врста потребно је добијене резултате интегрисати са подацима других релевантних&nbsp; научних дисциплина.</p> / <p>Rod <em>Merodon Meigen</em> (Diptera: Syrphidae) se odlikuje velikm brojem vrsta koje imaju&nbsp; funkciju u oprašivanju&nbsp; biljaka. U okviru ovog roda svojom raznovrsnošću izdvaja se <em>Merodon aureus</em>&nbsp; grupa&nbsp; vrsta koju pored fenotipski različitih odlikuje i prisustvo većeg broja kriptičnih&nbsp; vrsta. Zbog morfološke sličnosti kriptične vrste predstavljaju izazov za taksonome, te su molekularne metode od posebnog značaja. U tom svetlu primarni cilj ovog istraživanja je bio utvrđivanje molekularnog diverziteta grupe i mogućnosti njegove primene u taksonomiji. Istraživanje je bilo zasnovano na analizi varijabilnosti sekvenci&nbsp; COI&nbsp; i&nbsp; 28S&nbsp; rRNK gena za 718 jedinki 41 opisane ili novotkrivene vrste sirfida. Dodatno, testirana je primenjivost multilokusnih&nbsp; ISSR markera u razdvajanju vrsta&nbsp; <em>M.&nbsp; luteomaculatus</em>&nbsp; kompleksa. Uzorci su testirani i na prisustvo<em> Wolbachia</em> s obzirom da ona može uticati na evoluciju mitohondrijalnih gena. Dobijeni rezultati su ukazali na visoku varijabilnost sekvenci&nbsp; COI&nbsp; gena&nbsp; koji se pokazao korisnim u&nbsp; utvrđivanju granica&nbsp; kriptičnih&nbsp; vrsta u analiziranim kompleksima. Sekvence 28Ѕ rRNK gena u većini slučajeva nisu bile od većeg značaja za razdvajanje vrsta, ali bi mogle imati potencijal u&nbsp; razdvajanju kompleksa&nbsp; ili podgrupa&nbsp; vrsta kao dopuna analizi&nbsp; sekvenci&nbsp; COI&nbsp; gena. Dodatno,&nbsp; ISSR&nbsp; markeri su pokazali potencijal za primenu u molekularnoj taksonomiji. Sve analizirane vrste izuzev <em>M. balkanicus</em> su bile zaražene&nbsp; <em>Wolbachia,</em> ali nije utvrđena jasna veza između varijabilnosti&nbsp; sekvenci&nbsp; mitohondrijalnog&nbsp; COI gena i infekcionog statusa vrsta. Utvrđen obrazac genetičke varijabilnosti je verovatno&nbsp; oblikovan drastičnim klimatskim promenama&nbsp; tokom Pleistocena i fragmentacijom habitata. Da bi se doneli krajnji zaključci o taksonomskom statusu predloženih vrsta potrebno je dobijene rezultate integrisati sa podacima drugih relevantnih&nbsp; naučnih disciplina.</p> / <p>Genus<em>&nbsp; Merodon&nbsp; Meigen&nbsp;</em> (Diptera: Syrphidae)&nbsp; is characterized by a large number of species which have a function in plants pollination. Within this genus, <em>Merodon </em>aureus species group is distinguished by its diversity and it is characterized not only by phenotypically divergent species but also with a large number of cryptic species. Because of the high morphological similarity, cryptic species are a challenge for taxonomist, thus molecular methods are of special importance. The primary goal of this research was to determine the molecular diversity of the group and the possibility of its application in taxonomy. The study was based on the sequences variability analysis of COI and 28S rRNA genes for 718 individuals belonging to 41 described or newly discovered hoverfly species. Additionally, the applicability in species delimitation of multilocus marker ISSR was tested on&nbsp; <em>M. luteomaculatus</em>&nbsp; species complex. Specimens have also been tested for<em> Wolbachia</em> since&nbsp; it may affect the evolution of mitochondrial genes. The obtained results indicated a high variability of the COI gene&nbsp; sequences that proved useful for determining the&nbsp; cryptic&nbsp; species boundaries in the analyzed complexes. Sequences of the 28S rRNA gene in&nbsp;&nbsp; most cases were not of much significance for the species delimitation, but they could have the potential to separate species complexes&nbsp; оr subgroups&nbsp; as a complement to the analysis of the COI gene&nbsp; sequences. Additionally, the ISSR markers showed potential for application in molecular taxonomy. All analyzed species&nbsp; except&nbsp; <em>M. balkanicus</em>&nbsp; were infected with&nbsp; <em>Wolbachia</em>, but no clear&nbsp; relation&nbsp; was established between the sequence variability of the mitochondrial COI gene and&nbsp; the infectious status of species. The established pattern of genetic variability is probably shaped&nbsp; by drastic climatic changes during Pleistocene and habitats fragmentation. In order to&nbsp; achieve&nbsp; the final conclusions on the taxonomic status of the proposed species, it is necessary to integrate the obtained results with the data of other&nbsp; relevant scientific disciplines.</p>

No description available.

Le jujubier (ou Ziziphus mauritiana Lam.) est une espèce à usages multiples (fruits, fourrage, bois de service) prioritaire pour le reboisement et l’arboriculture fruitière dans le Sahel. Dans ce contexte où les sols sont souvent dégradés et pauvres en minéraux (P en particulier), la mycorhization et la fertilisation phosphatée pourraient jouer un rôle important dans l’amélioration de la croissance et de la productivité des jujubiers.L’objectif principal de ce travail était d’améliorer la croissance et la production fruitière de Z. mauritiana par l’inoculation mycorhizienne dans deux vergers au Sénégal. Il avait pour objectifs spécifiques (i) d’évaluer les réponses à l’inoculation avec des CMAs de différentes espèces de Ziziphus et de provenances de Z. mauritiana en serre, (ii) d’évaluer l’impact de l’inoculation avec R. irregularis IR27 sur la croissance, la survie et la production fruitière de Z. mauritiana, (iii) d’évaluer l’impact de l’inoculation sur la diversité des communautés de CMAs associés à Z. mauritiana en plantation et (iv) de déterminer la persistance de R. irregularis IR27 dans les racines de Z. mauritiana en plantation. Le champignon R. irregularis IR27 s’est avéré le plus efficace parmi les CMAs testés dans ce travail. Le couple Z. mauritiana /R. irregularis IR27 a donc été choisi comme modèle pour étudier l’impact de l’inoculation sur la production fruitière de deux provenances, Gola (variété indienne sélectionnée pour ses fruits de grosse taille) et Tasset (provenance locale à fruits de petite taille) dans deux sites contrastés (Amally et Keur Mangari). Nos résultats ont montré un effet positif de l’inoculation sur la croissance, la survie et le taux de mycorhization de Z. mauritiana à 13 et 24 mois respectivement à Amally et à Keur Mangari. L’inoculation a également augmenté la production fruitière des jujubiers à 18 et 30 mois de plantation à Keur Mangari. Ces résultats montrent la grande capacité de R. irregularis IR27 à compétir face aux CMAs indigènes. Le séquençage Illumina MiSeq du gène 18S a permis de révéler un impact négatif de l’inoculation sur la diversité et la richesse des communautés de CMAs natifs à Amally contrairement à Keur Mangari où l’inoculation n’a pas eu d’impact ni sur la diversité ni sur la richesse des CMAs. Le gène RPB1 s’est révélé pertinent comme marqueur pour détecter R. irregularis IR27 dans les racines de Z. mauritiana inoculés et évaluer par qPCR l’intensité de la colonisation racinaire des jujubiers par R. irregularis IR27 qui a représenté 11 à 13% à 13 mois de plantation à Amally et 12 à 15% à 24 mois de plantation à Keur Mangari. Cependant, il s’avère important d’évaluer à plus long terme l’impact de R. irregularis IR27 et son devenir dans les racines de Z. mauritiana en plantation dans une large gamme de conditions environnementales. / The jujube (or Ziziphus mauritiana Lam.) is an important multipurpose species (e.g. fruits, fodder, wood) for reforestation and fruit farming in the Sahel. In this context where soils are often degraded and deficient in P, mycorrhization and phosphorus fertilization could play a major role on improvement of jujube growth and productivity. The main objective of this work was to improve growth and fruit production of Z. mauritiana by mycorrhizal inoculation in two orchards at Senegal. This work aims (i) to evaluate the responses of different species of Ziziphus and provenances of Z. mauritiana to inoculation with AMF in greenhouse conditions, (ii) to assess the impact of inoculation with R. irregularis IR27 on growth, survival and fruit production of Z. mauritiana, (iii) to assess the impact of inoculation on diversity of native AMF communities associated to Z. mauritiana after planting and (iv) to determinate the persistence of R. irregularis IR27 in roots of Z. mauritiana after planting.The fungus R. irregularis IR27 proved to be the most effective AMF tested in this work. The pair Z. mauritiana /R. irregularis IR27 has been chosen as model to study the impact of inoculation on fruit production of two provenances, Gola (Indian variety selected for its large size fruits) and Tasset (local cultivar with small-sized fruits) in two sites with contrasting rainfall (Amally and Keur Mangari). Our results showed a positive effect of inoculation on growth, survival and mycorrhizal colonization of Z. mauritiana plants at 13 and 24 months after planting at Amally and Keur Mangari respectively. Inoculation increased also fruit production of jujubes at 18 and 30 months after planting at Keur Mangari. These results indicated the high ability of R. irregularis to compete with indigenous AMF. The MiSeq Illumina sequencing of 18S rRNA gene revealed a negative impact of inoculation on AMF richness and diversity at Amally, unlike at Keur Mangari where inoculation had no impact on AMF richness and diversity. The RPB1 gene proved to be an appropriate marker to detect of R. irregularis IR27 in inoculated Z. mauritiana roots and to evaluate by qPCR the root colonization of R. irregularis IR27 which accounted for 11 to 13% at 13 months after planting at Amally and 12 to 15% at 24 months after planting at Keur Mangari. Therefore, it is important to assess at long-term the impact of R. irregularis IR27 and its persistence in inoculated Z. mauritiana roots in large environmental conditions.

C'est essentiellement sur les bactéries libres que portent les études récentes en écologie microbienne d'eau douce, et seulement quelques études ont concerné les communautés bactériennes attachées. Dans cette étude basée sur l'analyse des séquences du gène 16SARNr, la diversité des communautés bactériennes attachées ainsi que de la fraction libre a été étudiée sur deux systèmes d'eau douce ; un lac mésotrophe (le lac du Bourget) et un lac hypereutrophe (le lac Villerest). La diversité des Actinobacteria, Betaproteobacteria et Verrucomicrobia libres et attachées a été étudié en relation avec les variables environnementales, dans le lac du Bourget pendant deux périodes à dominances contrastées de phytoplancton. L'analyse des résultats a montré une différence au niveau phylogénétique entre les communautés bactériennes attachées et libres des trois groupes de bactéries étudiées. Le clade betaI, dominait les Betaproteobacteria des fractions libres et attachées, avec 57,8% de la totalité des unités taxinomiques opérationnelles (OTUs). Pour les Actinobacteria, le groupe d'acIV a été détecté comme le plus abondant, suivi par acI avec respectivement 45% et 25% du total des OTUs. De même, les groupes Verrucomicrobia d'eau douce, à savoir CREPA29, FukuN18, CL120-10 sont apparus comme les plus importants, avec 22,3%, 16,15% et 14,61% des OTUs respectivement. Cette étude a permis de définir 15 nouveaux clades putatifs représentant la diversité bactérienne d'eau douce des Betaproteobacteria (lbI-lbVIII), Actinobacteria (acLBI) et Verrucomicrobia (CRE-PA29, FukuS27, BourFI-BourFIV). Par ailleurs, 12 groupes représentant la diversité de phylogénétique des Betaproteobacteria, Actinobacteria et Verrucomicrobia contiennent exclusivement des OTUs de la fraction attachée. La dynamique saisonnière souligne les changements des phylotypes bactériens distincts pour les deux communautés attaché et libre. Les Actinobacteria dans la fraction attachée était associée avec la biomasse des Chrysophyceae et N-NO3, et les Betaproteobacteria avec la biomasse de Chlorophyceae et de la richesse du phytoplancton tandis que les Verrucomicrobia de cette même fraction ont semblé être principalement influencés par la richesse du phytoplancton, l′abondance des rotifères et les nutriments inorganiques (N-NO3, SiO2). D'autre part, dans les communautés libres, peu de clades d'actinobacterie dépendent des nutriments ou du phytoplancton, alors que les Betaproteobacteria et Verrucomicrobia ont été principalement associés avec les paramètres biologiques (i.e. phytoplancton et copépodes). Pendant le bloom des cyanobacteries (Microcystis sp.) dans le lac Villerest, les Betaproteobacteria, Gammaproteobacteria, Alphaproteobacteria, Bacteroidetes et Actinobacteria ont été détectés comme taxa dominants dans les banques de clones du gène 16S ARNr. Toutefois Verrucomicrobia et Deinococcus-Thermus sont apparus relativement moins abondants dans les deux fractions, tandis que,Gemmatimonadetes, Acidobacteria, Chloroflexi, Planctomycetes, Deltaproteobacteria, Firmicutes et Op11 sont apparus comme des phyla mineurs dans la banque de clones des communautés bactériennes attaché et libre. Les Betaproteobacteria (n=118) attachées sont apparus comme le groupe dominant, suivi par Gammaproteobacteria (n=74) et Bacteroidetes (n=52). L'analyse phylogénétique des séquences obtenues pour la banque de clone de la fraction libre a montré que la plupart des OTUs appartiennent à Betaproteobacteria (n=192), suivi par Bacteroidetes (n=132) et Actinobacteria (n=61). Tandis que les Gammaproteobacteria (n=42) et Alphaproteobacteria (n=42) sont présents dans des proportions égales dans la banque de clones du 16S ARNr libre. (...) / The free-living bacteria point of view dominates in recent research of freshwater microbial ecology, only a few studies have focused on attached bacterial communities. In present study, based on 16S rRNA gene sequences, diversity of attached and free-living bacterial community was investigated from two freshwater aquatic systems ; a mesotrophic lake Bourget and a hypereutrophic lake Villerest. The diversity of attached and free-living Actinobacteria, Betaproteobacteria and Verrucomicrobia, in relation to environmental variables was investigated from lake Bourget during two contrasting periods of phytoplankton dominance. Comparison analyses showed a phylogenetic difference between attached and free-living bacterial communities of all three studied bacterial groups. The betaI, appeared as most dominant among all clades representing phylogenetic diversity of freshwater Betaproteobacteria, for both attached and free-living fractions, contributing to 57.8% of of the total retrieved opertational taxonomic units (OTUs). For Actinobacteria, the acIV cluster was detected as dominant, followed by acI accounting for 45% and 25% of the total retrieved OTUs respectively. Similarly, freshwater Verrucomicrobia cluster namely, CRE-PA29, FukuN18, CL120-10 appeared as dominant, comprising 22.3%, 16.15% and 14.61% of the total retrieved OTUs respectively. This study allowed defining 15 new putative clades representing the freshwater bacterial divesity of Betaproteobacteria (lbI-lbVIII), Actinobacteria (acLBI) and Verrucomicrobia (CRE-PA29, FukuS27, BourFI-BourFIV). In addition, 12 clusters representing the phylogenetic diversity of Betaproteobacteria, Actinobacteria and Verrucomicrobia were exclusively comprised of OTUs from the attached fraction. The seasonal dynamics of environmental variables have been reflected as changes in distinct bacterial phylotypes for both attached and free-living communities. The attached bacterial communities of Actinobacteria showed affiliation with Chrysophyceae biomass and N-NO3, while attached Betaproteobacteria were affiliated with biomass of Chlorophyceae and phytoplankton richness. Similarly attached verrucomicrobial communities appeared to be mainly influenced by phytoplankton richness, rotifers abundances and inorganic nutrients (NNO3,SiO2). On the other hand, within free-living communities, few actinobacterial clades were found to be dependent on either nutrients or phytoplankton communities, whereas Betaproteobacteria and Verrucomicrobia were mainly associated with biological parameters (i.e. phytoplankton and copepods communities). In another study during a cyanobacterial (Microcystis sp.) bloom from lake Villerest, Betaproteobacteria, Gammaproteobacteria, Alphaproteobacteria, Bacteroidetes and Actinobacteria were detected as prevalent taxa among the 16S rRNA gene clone libraries, however, Verrucomicrobia and Deinococcus-Thermus appeared as comparatively less abundant bacterial groups in both fractions. Whereas, Gemmatimonadetes, Acidobacteria, Chloroflexi, Planctomycetes, Deltaproteobacteria, Firmicutes and Op11 were appeared as minor phyla in clone libraries of attached and free-living bacterial communities. For attached bacterial communities Betaproteobacteria (n=118) appeared as most dominant group, followed by Gammaproteobacteria (n=74) and Bacteroidetes (n=52). The phylogenetic analysis of the sequences obtained for the clone library from free-living fraction showed that most of the OTUs belonged to Betaproteobacteria (n=192) followed in decreasing order by Bacteroidetes (n=132) and Actinobacteria (n=61) whereas Gammaproteobacteria (n=42) and Alphaproteobacteria (n=42) appeared in equal proportion in free-living 16S rRNA clone libraries. (...)

Interactions between plant roots and soil microorganisms in the rhizosphere are critical for plant growth. However, understanding of precisely how root exudates influence the diversity and activity of rhizosphere microorganisms is limited. The main objective of this study was to investigate the effect of radiata pine (Pinus radiata) root exudates on rhizosphere soil microbial communities, with an emphasis on the role of low molecular weight organic anions. The study involved the development and validation of new methods for investigating rhizosphere processes in a purpose-built facility. This included development of an in situ sampling technique using an anion exchange membrane strip to collect a range of organic anions exuded from radiata pine roots grown in large-scale rhizotrons. These included tartarate, quinate, formate, malate, malonate, shikimate, lactate, acetate, maleate, citrate, succinate and fumarate. Soil microbial activity and diversity were determined using dehydrogenase activity and denaturing gradient gel electrophoresis. Links between organic anions in root exudates and rhizosphere soil microbial community structures were investigated by comparing wild type and genetically modified radiata pine trees which were grown in rhizotrons for 10 months. As expected, there was considerable temporal and spatial variability in the amounts and composition of organic anions collected, and there were no consistent or significant differences determined between the two tree lines. Significant differences in rhizosphere microbial communities were detected between wild type and genetically modified pine trees; however, they were inconsistent throughout the experiment. The shifts in microbial communities could have been related to changes in exudate production and composition. Based on results from the main rhizotron experiment, a microcosm study was carried out to investigate the influence of selected pine root exudate sugars (glucose, sucrose and fructose) and organic anions (quinate, lactate and maleate) on soil microbial activity and diversity. Soil microbial activity increased up to 3-fold in all of the sugar and organic anion treatments compared to the control, except for a mixture of sugars and maleate where it decreased. The corresponding impacts on soil microbial diversity were assessed using denaturing gradient gel electrophoresis and 16S rRNA phylochips. Addition of the exudate compounds had a dramatic impact on the composition and diversity of the soil microbial community. A large number of bacterial taxa (88 to 1043) responded positively to the presence of exudate compounds, although some taxa (12 to 24) responded negatively. Organic anions had a greater impact on microbial communities than sugars, which indicated that they may have important roles in rhizosphere ecology of radiata pine. In addition, a diverse range of potentially beneficial bacterial taxa were detected in soil amended with organic anions, indicating specific regulation of rhizosphere microbial communities by root exudates. This project highlighted the considerable challenges and difficulties involved in detailed investigation of in situ rhizosphere processes. Nonetheless, the findings of this study represent a significant contribution to advancing understanding of relationships between root exudates and soil microbial diversity, which will be further enhanced by refinement and application of the specific methodologies and techniques developed.

Des recherches en sciences judiciaires ont montré récemment une possible corrélation entre le temps d’entreposage d’échantillons de fluides corporels et la dégradation de l’ARN dans ceux-ci. Le moment où une tache a été déposée sur une scène de crime peut être important pour déterminer la pertinence d’un échantillon dans une enquête. Dans ce mémoire, nous rapportons les profils de dégradation de quatre ARN différents mesurés par RT-qPCR, soit l’ARN ribosomique 18S et les ARNm de la β-actine, de la glyceraldehyde-3-phosphate déhydrogénase et de la cyclophiline A, obtenus de taches de sang, de salive et de sperme, entreposés à la température de la pièce ou au congélateur à -80°C sur une période de 6 mois. Nos résultats montrent une faible variation interindividuelle pour le sang et le sperme, mais une différence importante entre les donneurs pour la salive. De plus, le profil de dégradation est semblable pour tous les transcrits, mais diffère entre les fluides. La congélation des échantillons stabilise les ARN avant leur analyse. Finalement, la quantité d’ARN détecté est en relation avec le temps d’entreposage et pourrait être utilisée afin d’estimer l’âge des échantillons lorsque l’impact des conditions d’entreposage sur la dégradation de l’ARN sera mieux connu. / Recent studies in forensic science have shown a possible correlation between the degradation rate of some RNA transcripts and the age of bloodstains. The time of deposition of a stain can be of major importance to determine the relevance of a sample in a forensic investigation. In this thesis, we describe the degradation profiles of the 18S ribosomal RNA and the β-actin, glyceraldehyde-3-phosphate dehydrogenase and cyclophilin A mRNAs, measured by RT-qPCR and obtained from dried blood, semen and saliva stains stored at room temperature or frozen at -80°C up to 6 months. Our results showed low inter-individual variation for blood and semen stains, but a high variation was observed between donors for saliva. Moreover, degradation profile of each transcripts was similar, but differed between fluids. Freezing samples prevented RNA degradation over time. Finally, RNA quantity was in relation with the time of storage and could be used to estimate the time since deposition of a stain when the effects of various storage conditions on RNA degradation profiles will be better documented. / Projet de recherche réalisé en collaboration avec la section Biologie/ADN du Laboratoire de sciences judiciaires et de médecine légale (LSJML) de Montréal.

Le mutualisme entre les plantes et les champignons arbusculaires mycorhiziens est extrêmement répandu (~ 80% des plantes sont colonisées par ces organismes) et ancien (il ya plus de 450 millions d'années). Cette relation symbiotique est une composante essentielle du fonctionnement des écosystèmes et de leur productivité, et est fortement impliqué dans le cycle de deux éléments clés: le phosphore et le carbone. Le maintien de ce mutualisme est devenu particulièrement important dans le contexte actuel de perte de biodiversité. Un des objectifs de cette thèse était de comprendre la stabilité de ce mutualisme. L'accent a tout d'abord été mis sur les échanges de nutriments impliqués dans cette symbiose, en testant si la plante hôte et les symbiotes fongiques sont capables de discriminer leurs différents partenaires, et d'allouer davantage de ressources aux partenaires fournissant plus de nutriments. J'ai ensuite étudié la possibilité de l'implication de la plante hôte dans la protection des symbiotes mycorhiziens via un transfert de métabolites secondaires dans les hyphes. Nous avons alors pu emettre une nouvelle hypothèse suggérant que la protection en métabolites secondaires venant de la plante serait positivement corrélée avec le niveau de coopération (à savoir le transfert des nutriments) du champignon symbiotique. L'echelle d'étude est ensuite passée de l'individu à la communauté en étudiant les effets de la diminution de la diversité végétale sur la diversité des symbiotes racinaires. Pour ce faire, des analyses moléculaires et des outils novateurs ont été utilisés, tels que le séquençage à haut débit. Pour faciliter encore l'étude des séquences obtenues et d'autres séquences fongiques, j'ai collaboré avec des collègues afin de créer une base de données 'Phymyco-DB' rendue publique en 2012. Enfin, je discute de l'implication du mutualisme mycorhizien dans le contexte des systèmes agricoles actuels et propose de nouvelles trajectoires pour gérer ces systèmes. Ce projet de thèse apporte un nouvel éclairage sur la façon dont fonctionnent ces interactions entre les plantes et champignons MA et sur la manière dont ils façonnent les processus écologiques et les trajectoires évolutives dans les écosystèmes naturels et agricoles. Ces points sont d'une importance majeure pour développer une agriculture plus écologiquement intensive et durable. Le projet a fourni de nouvelles connaissances et perspectives sur la perte de la diversité végétale, et ses conséquences pour la stabilité de la symbiose AM. Comme les champignons mycorhiziens sont essentiels dans les processus des écosystèmes et l'entretien de la fertilité des sols, ce travail devrait avoir un large impact dans (i) la politique de protection des sols, (ii) la recherche sur l'amélioration des plantes et (iii) la conception de systèmes agricoles durables.