Effect of bacterial inoculation, plant genotype and developmental stage on root-associated and endophytic bacterial communities in potato (Solanum tuberosum)

Beneficial bacteria interact with plants by colonizing the rhizosphere and roots followed by further spread through the inner tissues, resulting in endophytic colonization. The major factors contributing to these interactions are not always well understood for most bacterial and plant species. It is believed that specific bacterial functions are required for plant colonization, but also from the plant side specific features are needed, such as plant genotype (cultivar) and developmental stage. Via multivariate analysis we present a quantification of the roles of these components on the composition of root-associated and endophytic bacterial communities in potato plants, by weighing the effects of bacterial inoculation, plant genotype and developmental stage. Spontaneous rifampicin resistant mutants of two bacterial endophytes, Paenibacillus sp. strain E119 and Methylobacterium mesophilicum strain SR1.6/6, were introduced into potato plants of three different cultivars (Eersteling, Robijn and Karnico). Densities of both strains in, or attached to potato plants were measured by selective plating, while the effects of bacterial inoculation, plant genotype and developmental stage on the composition of bacterial, Alphaproteobacterial and Paenibacillus species were determined by PCR-denaturing gradient gel-electrophoresis (DGGE). Multivariate analyses revealed that the composition of bacterial communities was mainly driven by cultivar type and plant developmental stage, while Alphaproteobacterial and Paenibacillus communities were mainly influenced by bacterial inoculation. These results are important for better understanding the effects of bacterial inoculations to plants and their possible effects on the indigenous bacterial communities in relation with other plant factors such as genotype and growth stage

Cultivation of hitherto-uncultured bacteria belonging to the Verrucomicrobia subdivision 1 from the potato (Solanum tuberosum L.) rhizosphere

The role of dominant bacterial groups in the plant rhizosphere, e.g., those belonging to the phyla Acidobacteria and Verrucomicrobia, has, so far, not been elucidated, and this is mainly due to the lack of culturable representatives. This study aimed to isolate hitherto-uncultured bacteria from the potato rhizosphere by a combination of cultivation approaches. An agar medium low in carbon availability (oligotrophic agar medium) and either amended with potato root exudates or catalase or left unamended was used with the aim to improve the culturability of bacteria from the potato rhizosphere. The colony forming unit numbers based on colonies and microcolonies were compared with microscopically determined fluorescence-stained cell numbers. Taxonomical diversity of the colonies was compared with that of library clones made from rhizosphere DNA, on the basis of 16S rRNA gene comparisons. The oligotrophic media amended or not with catalase or rhizosphere extract recovered up to 33.6% of the total bacterial numbers, at least seven times more than the recovery observed on R2A. Four hitherto-uncultured Verrucomicrobia subdivision 1 representatives were recovered on agar, but representatives of this group were not found in the clone library. The use of oligotrophic medium and its modifications enabled the growth of colony numbers, exceeding those on classical agar media. Also, it led to the isolation of hitherto-uncultured bacteria from the potato rhizosphere. Further improvement in cultivation will certainly result in the recovery of other as-yet-unexplored bacteria from the rhizosphere, making these groups accessible for further investigation, e.g., with respect to their possible interactions with plants

Exploring interactions of plant microbiomes

A plethora of microbial cells is present in every gram of soil, and microbes are found extensively in plant and animal tissues. The mechanisms governed by microorganisms in the regulation of physiological processes of their hosts have been extensively studied in the light of recent findings on microbiomes. In plants, the components of these microbiomes may form distinct communities, such as those inhabiting the plant rhizosphere, the endosphere and the phyllosphere. In each of these niches, the "microbial tissue" is established by, and responds to, specific selective pressures. Although there is no clear picture of the overall role of the plant microbiome, there is substantial evidence that these communities are involved in disease control, enhance nutrient acquisition, and affect stress tolerance. In this review, we first summarize features of microbial communities that compose the plant microbiome and further present a series of studies describing the underpinning factors that shape the phylogenetic and functional plant-associated communities. We advocate the idea that understanding the mechanisms by which plants select and interact with their microbiomes may have a direct effect on plant development and health, and further lead to the establishment of novel microbiome-driven strategies, that can cope with the development of a more sustainable agriculture

Caracterização da comunidade bacteriana endofítica de citros por isolamento, PCR específico e DGGE.

O objetivo deste trabalho foi caracterizar a comunidade bacteriana endofítica de plantas assintomáticas (escapes) e afetadas pela clorose variegada dos citros (CVC) por meio de isolamento em meio de cultura, técnica de gradiente desnaturante em gel de eletroforese (DGGE) e detecção de Methylobacterium mesophilicum e Xyllela fastidiosa por meio de PCR específico, para estudar esta comunidade e sua relação com a ocorrência da CVC. A análise da comunidade bacteriana via DGGE permitiu a detecção de X. fastidiosa, bem como Klebsiella sp. e Acinetobacter sp. como endófitos de citros. Foram observados também Curtobacterium sp., Pseudomonas sp., Enterobacter sp. e Bacillus spp. Utilizando primers específicos, Methylobacterium mesophilicum e X. fastidiosa também foram observadas, reforçando hipóteses de que estas bactérias podem estar interagindo no interior da planta hospedeira.Made available in DSpace on 2014-08-29T06:40:32Z (GMT). No. of bitstreams: 1 Caracterizacao.pdf: 239740 bytes, checksum: aee899d2fd3ca589fe2693869037b503 (MD5) Previous issue date: 2006-10-19200

Biological and morphological traits of sugarcane roots in relation to phosphorus uptake

Appropriate management of phosphorus (P) in soil will lead to higher yields and sustainability for sugarcane production. Our study evaluated the importance of differences in root structure and function, and the differential composition of the bacterial community in four sugarcane cultivars, in relation to the efficiency for P uptake and also to assess changes in soil P with distance from the rhizoplane. Experiments were performed in pot trials using a sandy clay loam Ferralsol. In the first experiment, the effect of P application (78.4 mg P kg-1 soil as triple superphosphate) on sugarcane cultivars RB92-579; RB85-5156; RB86-7515 and RB96-6928 was investigated. Secondly, we evaluated P rates of 0; 9.8; 19.6; 39.2 and 78.4 mg kg-1 soil using cultivar RB96-6928 which was shown to be one of the more growth responsive cultivars. The cultivar RB96-6928 exhibited the highest root dry matter and root surface area, while the bacterial communities found in the rhizosphere of these plants were not different from other cultivars, as determined by PCR-DGGE. From the P dose-dependent experiment for this cultivar, optimal plant performance occurred at a P supply up to 38.5 mg P kg-1 soil. Collectively, our results indicate that P efficiency in sugarcane was predominantly associated with the generation of high root biomass and surface area

Antitumoral, Antioxidant And Antimicrobial Molecules From Combretum Rupicola

This investigation describes the anticancer, antioxidant and antimicrobial properties of the extracts of Combretum rupicola, a native plant from Northeast of Brazil. Methanolic, ethyl acetate and chloroform extracts from leaves of C. rupicola were evaluated in relation to their potential in inhibiting the cell growth and cytotoxic properties against nine human cancer cell lines (MCF-7, NCI-ADR/RES, OVCAR-3, PC-3, HT-29, NCIH460, 786-O, UACC-62, K-562). In addition, antioxidant activity of these extracts was measured using DPPH as radical scavenging assay and the antimicrobial activities against nine pathogens were tested by agar diffusion method. Preliminary results showed that the this plant demonstrates to have moderate activity against bacteria but, on the other side, the extracts showed significant anticancer activitiesagainst four cell lines and the most significant activity was observed against MCF-7 (65.9 μg mL-1), highest inhibitory concentration IC50 0.22 μg mL-1 for antioxidant activity. These founds are the first scientific reports on secondary metabolites with biological activities of C. rupicola, suggesting the potential of this botanic species for pharmaceutical industry.41B422B428Butler, M.S., The role of natural product chemistry in drug Discovery (2004) J of Nat Prod, 67 (12), pp. 2141-2153Ripa, F.A., Nahar, L., Haque, M., Islam, M.M., Antibacterial, Cytotoxic and Antioxidant Activity of Crude Extract of Marsilea Quadrifolia (2009) Eur J of Scien Res, 33 (1), pp. 123-129Santos, S.N., Oliveira, L.K.X., Melo, I.S., Velozo, E.S., Roque, M.R.A., Antifungal activity of bacterial strains from the rhizosphere of Stachytarpheta crassifólia (2011) Afri J of Biot, 10 (25), pp. 4996-5000Pettit, G.R., Singh, S.B., Niven, M.L., Hamel, E., Schmidt, J.M., Isolation, Structure, and Synthesis of Combretastatins A-1 and B- 1, Potent New Inhibitors of Microtubule Assembly, Derived from Combretum caffrum (1987) J of Nat Prod, 50 (1), pp. 119-131Tan, F.X., Shi, S.H., Zhong, Y., Gong, X., Wang, Y.G., Phylogenetic relationships of Combretoideae (Combretaceae) inferred from plastid, nuclear gene and spacer sequences (2002) J of Plant Res, 115, pp. 475-481Fyhrquist, P., Mwasumbi, L., Haeggstrom, C.A., Vuorela, H., Hiltunen, R., Vuorela, P., Microscopic diagnosis of tumors of the central nervous system by phase contrast microscopy (2002) J of Ethnoph, 79, pp. 169-177McGaw, L.J., Rabe, T., Sparg, S.G., Jager, A.K., Eloff, J.N., Van Staden, J., An investigation on the biological activity of Combretum species (2001) J of Ethnoph, 75, pp. 45-50Ridley, H.N., Notes on the Botany of Fernando Noronha (1890) Botany, 27 (181), pp. 1-95Eloff, J.N., Katerere, D.R., McGaw, L.J., The biological activity and chemistry of the southern African Combretaceae (2008) J of Ethnoph, 119, pp. 686-699Grammer, A., Antibiotic sensitivity and Assay test (1976) Collins CH and Lyne PM, Eds. Microbiological methods, p. 235. , Butterworths, LondonPerformance standards for antimicrobial susceptibility testing, 14th information supplement (2004) Document M100-S14. The National Committee for Clinical Laboratory Standards, , NCCLS, 4th NCCLS documents M100- S14, Wayne, PAMilardovic, S., Ivekovic, D., Grabaric, B.S., A novel amperometric method for antioxidant activity determination using DPPH free radical (2006) Bioelectro, 68, pp. 175-180Santos, S.N., Castanha, R.F., Haber, L.L., Scramin, S., Marques, M.O.M., Melo, I.S., Determinação quantitativa da atividade antioxidante de extratos brutos de microrganismos pelo método de captura de radical livre DPPH (2011) Téc. Embrapa Meio Ambiente, 1, pp. 1-5Skehan, P., Storeng, R., Scudiero, D., Monks, A., McMahon, J., Vistica, D., New Colorimetric Cytotoxicity Assay for Anticancer-Drug Screening (1990) J Nat Canc Inst, 82, pp. 1107-1112Holbeck, S.L., Update on NCI in vitro drug screen utilities (2004) Eur J Canc, 40, pp. 785-793Eloff, J.N., The antibacterial activity of 27 Southern African members of the Combretaceae (1999) J of Ethnoph, 67, pp. 355-360Silva, O., Duarte, A., Cabrita, J., Pimentel, M., Diniz, A., Gomes, E., Antimicrobial activity of Guinea-Bissau traditional remedies (1996) J of Ethnop, 50, pp. 55-59Baba-Moussa, F., Akpagana, K., Bouchet, P., Antifungal activities of seven West African Combretaceae used in traditional medicine (1998) J of Ethnoph, 66, pp. 335-338Eloff, J.N., Picard, J., Masoko, P., Resistance of animal fungal pathogens to solvents used in bioassays (2007) S Afri J of Bot, 73, pp. 667-669Mourão, F., Umeo, S.H., Takemura, O.S., Linde, G.A., Colauto, N.B., Antioxidant activity of Agaricus brasiliensis basidiocarps on different maturation phases (2011) Braz J of Micr, 42, pp. 197-202Zheng, W., Wang, S.Y., Antioxidant activity and phenolic compounds in selected herbs (2001) J Agric Food Che, 49 (11), pp. 5165-5170Masoko, P., Eloff, J.N., Screening of twenty-four South African combretum and six terminalia species (Combretaceae) for antioxidant activities (2007) Afri J of Trad med, 4 (2), pp. 231-23

Sulphur-oxidising and Sulphate-reducing Communities in Brazilian Mangrove Sediments

Mangrove soils are anaerobic environments rich in sulphate and organic matter. Although the sulphur cycle is one of the major actors in this ecosystem, little is known regarding the sulphur bacteria communities in mangrove soils. We investigated the abundance, composition and diversity of sulphur-oxidizing (SOB) and sulphate-reducing (SRB) bacteria in sediments from three Brazilian mangrove communities: two contaminated, one with oil (OilMgv) and one with urban waste and sludge (AntMgv), and one pristine (PrsMgv). The community structures were assessed using quantitative real-time polymerase chain reaction (qPCR), polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) and clone libraries, using genes for the enzymes adenosine-5′-phosphosulphate reductase (aprA) and sulphite reductase (Dsr) (dsrB). The abundance for qPCR showed the ratio dsrB/aprA to be variable among mangroves and higher according to the gradient observed for oil contamination in the OilMgv. The PCR-DGGE patterns analysed by Nonmetric Multidimensional Scaling revealed differences among the structures of the three mangrove communities. The clone libraries showed that Betaproteobacteria, Gammaproteobacteria and Deltaproteobacteria were the most abundant groups associated with sulphur cycling in mangrove sediments. We conclude that the microbial SOB and SRB communities in mangrove soils are different in each mangrove forest and that such microbial communities could possibly be used as a proxy for contamination in mangrove forests.

Cultivation of hitherto-uncultured bacteria belonging to the Verrucomicrobia subdivision 1 from the potato (Solanum tuberosum L.) rhizosphere

The role of dominant bacterial groups in the plant rhizosphere, e.g., those belonging to the phyla Acidobacteria and Verrucomicrobia, has, so far, not been elucidated, and this is mainly due to the lack of culturable representatives. This study aimed to isolate hitherto-uncultured bacteria from the potato rhizosphere by a combination of cultivation approaches. An agar medium low in carbon availability (oligotrophic agar medium) and either amended with potato root exudates or catalase or left unamended was used with the aim to improve the culturability of bacteria from the potato rhizosphere. The colony forming unit numbers based on colonies and microcolonies were compared with microscopically determined fluorescence-stained cell numbers. Taxonomical diversity of the colonies was compared with that of library clones made from rhizosphere DNA, on the basis of 16S rRNA gene comparisons. The oligotrophic media amended or not with catalase or rhizosphere extract recovered up to 33.6% of the total bacterial numbers, at least seven times more than the recovery observed on R2A. Four hitherto-uncultured Verrucomicrobia subdivision 1 representatives were recovered on agar, but representatives of this group were not found in the clone library. The use of oligotrophic medium and its modifications enabled the growth of colony numbers, exceeding those on classical agar media. Also, it led to the isolation of hitherto-uncultured bacteria from the potato rhizosphere. Further improvement in cultivation will certainly result in the recovery of other as-yet-unexplored bacteria from the rhizosphere, making these groups accessible for further investigation, e.g., with respect to their possible interactions with plants.Netherlands Genomics InitiativeDutch Ministry of Agriculture, Nature, and Food quality[KB4