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- A new cyanobacterial species with a protective effect on lettuce grown under salinity stress: envisaging sustainable agriculture practicesPublication . Brito, Ângela; Rocha, Mariana; Kaštovský, Jan; Vieira, Jorge; Vieira, Cristina P.; Ramos, Vitor; Correia, Manuel; Santos, Marina; Mota, Rita; Roque, Jéssica; Pissarra, João; Melo, Paula; Tamagnini, PaulaIn this work, a new terrestrial cyanobacterial species, Oculatella lusitanica LEGE 161147, was isolated and characterized using a polyphasic approach. Morphologically, O. lusitanica shares characteristics with different Oculatella species (mainly with O. crustae-formantes), lacking distinctive features. However, the phylogeny based on the 16S rRNA gene sequence and the 16S-23S ITS secondary structures support the establishment of this isolate as a new species. O. lusitanica is placed within a clade mainly composed by other Oculatella terrestrial strains; however, it forms a separate lineage. In addition, our species differs from the other Oculatella described so far by lacking the V2 helix within the ITS region. Since cyanobacteria are known to release compounds that promote plant growth and/or increase their tolerance to stresses, the effect of this newly described cyanobacterial species on Lactuca sativa (lettuce) plants development and salinity stress resistance was evaluated. Our results showed that, although the cyanobacterium had no impact on plant growth under the conditions tested, it was able to mitigate the deleterious salinity stress effects on plant size, root and aerial part fresh weight, by eliciting the non-enzymatic antioxidant response system (proline, H2O2 and reduced glutathione). In addition, the microorganism was able to induce a priming effect on lettuce plants by stimulating defensive mechanisms under non-stress conditions, and enhances the activity of nitrogen metabolism-related enzymes glutamate dehydrogenase, glutamine synthetase and nitrate reductase. These results indicate that this native terrestrial cyanobacterial species could be employed as a tool in sustainable agricultural practices.
- Phylogenetic analysis and genetic diversity of the xylariaceous ascomycete Biscogniauxia mediterranea from cork oak forests in different bioclimatesPublication . Costa, Daniela; Ramos, Vitor; Tavares, Rui Manuel; Baptista, Paula; Lino-Neto, TeresaCork oak is a tree species with ecological importance that contributes to economic and social development in the Mediterranean region. Cork oak decline is a major concern for forest sustainability and has negative impacts on cork oak growth and production. This event has been increasingly reported in the last decades and seems to be related with climate changes. Biscogniauxia mediterranea is an endophytic fungus of healthy cork oak trees that turns into a pathogen in trees weaken by environmental stress. Understanding the drivers of B. mediterranea populations diversity and differentiation is expected to allow a better control of cork oak decline and preserve forest sustainability. Endophyte isolates from different cork oak forests were identified as B. mediterranea and their genetic diversity was evaluated using phylogenetic and microsatellite-primed PCR analyses. Genetic diversity and variability of this fungus was correlated with environmental/phytosanitary conditions present in forests/trees from which isolates were collected. High genetic diversity and variability was found in B. mediterranea populations obtained from different forests, suggesting some degree of isolation by distance. Bioclimate was the most significant effect that explained the genetic variability of B. mediterranea, rather than precipitation or temperature intensities alone or disease symptoms. These findings bring new implications for the changing climate to cork oak forests sustainability, cork production and quality.
- Potential of the endophyte Penicillium commune in the control of olive anthracnose via induction of antifungal volatiles in host plantPublication . Silva, Sofia; Costa, Helgeneusa Neto da; Lopes, Teresa; Ramos, Vitor; Rodrigues, Nuno; Pereira, J.A.; Lino-Neto, Teresa; Baptista, PaulaOlive anthracnose, caused by several Colletotrichum species, is the most economically harmful fruit disease of the olive crop. This work aimed to evaluate the ability of the endophyte Penicillium commune CIMO 14FM009 to protect the olive tree against Colletotrichum nymphaeae via induction of plant volatile organic compounds (VOCs). Accordingly, olive tree branches were inoculated with the endophyte and one month later with the pathogen. After 0, 3, and 24 h of pathogen inoculation, the volatile composition of leaves and fruits was analyzed by HSSPME- GC/MS, and compared with controls (branches inoculated with buffer, endophyte, or pathogen). The effect of plant-derived volatiles on C. nymphaeae was also evaluated. Penicillium commune induced the release of VOCs on the olive trees, with the capacity to reduce significantly the growth (up to 1.4-fold) and sporulation (up to 1.2-fold) of C. nymphaeae. This effect was most notorious on olives than on leaves, and occurred 3 h after pathogen-challenge, suggesting the need for a stressful stimulus for the production of antifungal VOCs. The observed inhibition was associated to a specific set of VOCs released from olives (mostly belonging to the alcohols and esters chemical classes) and leaves (mostly belonging to the alkenes). Curiously, a set of VOCs belonging to alkene, alkane and ester classes, were emitted exclusively in olive branches inoculated with C. nymphaeae. These findings provide new possibilities for controlling olive anthracnose using P. commune and/or volatiles, which efficacy should be tested in future works.
- The Extremophile Endolithella mcmurdoensis gen. et sp. nov. (Trebouxiophyceae, Chlorellaceae), a new Chlorella-like Endolithic alga from AntarcticaPublication . Martins, Teresa P.; Ramos, Vitor; Hentschke, Guilherme S.; Castelo-Branco, Raquel; Rego, Adriana; Monteiro, Maria; Brito, Ângela; Tamagnini, Paula; Cary, S. Craig; Vasconcelos, Vítor; Krienitz, Lothar; Magalhães, Catarina; Leão, Pedro N.The McMurdo Dry Valleys constitute the largest ice-free region of Antarctica and one of the most extreme deserts on Earth. Despite the low temperatures, dry and poor soils and katabatic winds, some microbes are able to take advantage of endolithic microenvironments, inhabiting the pore spaces of soil and constituting photosynthesis-based communities. We isolated a green microalga, Endolithella mcmurdoensis gen. et sp. nov, from an endolithic sandstone sample collected in the McMurdo Dry Valleys (Victoria Land, East Antarctica) during the K020 expedition, in January 2013. The single non-axenic isolate (E. mcmurdoensis LEGE Z-009) exhibits cup-shaped chloroplasts, electron-dense bodies, and polyphosphate granules but our analysis did not reveal any diagnostic morphological characters. On the basis of phylogenetic analysis of the 18S rRNA (SSU) gene, the isolate was found to represent a new genus within the family Chlorellaceae.
- Actinobacteria and cyanobacteria diversity in terrestrial Antarctic microenvironments evaluated by culture-dependent and independent methodsPublication . Rego, Adriana; Raio, Francisco; Martins, Teresa P.; Ribeiro, Hugo; Sousa, António G.G.; Séneca, Joana; Baptista, Mafalda S.; Lee, Charles K.; Craig Cary, S.; Ramos, Vitor; Carvalho, Maria F.; Leão, Pedro N.; Magalhães, CatarinaBacterial diversity from McMurdo Dry Valleys in Antarctica, the coldest desert on earth, has become more easily assessed with the development of High Throughput Sequencing (HTS) techniques. However, some of the diversity remains inaccessible by the power of sequencing. In this study, we combine cultivation and HTS techniques to survey actinobacteria and cyanobacteria diversity along different soil and endolithic micro-environments of Victoria Valley in McMurdo Dry Valleys. Our results demonstrate that the Dry Valleys actinobacteria and cyanobacteria distribution is driven by environmental forces, in particular the effect of water availability and endolithic environments clearly conditioned the distribution of those communities. Data derived from HTS show that the percentage of cyanobacteria decreases from about 20% in the sample closest to the water source to negligible values on the last three samples of the transect with less water availability. Inversely, actinobacteria relative abundance increases from about 20% in wet soils to over 50% in the driest samples. Over 30% of the total HTS data set was composed of actinobacterial strains, mainly distributed by 5 families: Sporichthyaceae, Euzebyaceae, Patulibacteraceae, Nocardioidaceae, and Rubrobacteraceae. However, the 11 actinobacterial strains isolated in this study, belonged to Micrococcaceae and Dermacoccaceae families that were underrepresented in the HTS data set. A total of 10 cyanobacterial strains from the order Synechococcales were also isolated, distributed by 4 different genera (Nodosilinea, Leptolyngbya, Pectolyngbya, and Acaryochloris-like). In agreement with the cultivation results, Leptolyngbya was identified as dominant genus in the HTS data set. Acaryochloris-like cyanobacteria were found exclusively in the endolithic sample and represented 44% of the total 16S rRNA sequences, although despite our efforts we were not able to properly isolate any strain from this Acaryochloris-related group. The importance of combining cultivation and sequencing techniques is highlighted, as we have shown that culture-dependent methods employed in this study were able to retrieve actinobacteria and cyanobacteria taxa that were not detected in HTS data set, suggesting that the combination of both strategies can be usefull to recover both abundant and rare members of the communities.
- Parakomarekiella sesnandensis gen. et sp. nov. (Nostocales, Cyanobacteria) isolated from the Old Cathedral of Coimbra, Portugal (UNESCO World Heritage Site)Publication . Soares, Fabiana; Ramos, Vitor; Trovão, João; Cardoso, Susana M.; Tiago, Igor; Portugal, AntónioIn a study that aimed to fingerprint the phototrophic community of the biodeteriorated walls of the Old Cathedral of Coimbra (UNESCO World Heritage Site), an unknown Nostoc/Komarekiella-like cyanobacterium was isolated. We employed a polyphasic approach based on morphological, ecological and phylogenetic analyses of the partial 16S and the whole 16S-23S ITS rRNA regions to characterize this organism. Our strains shared 16S rRNA gene sequence similarity of 99–100% with GenBank-assigned ‘Nostoc sp.’ sequences, and 98% similarity with the genera Goleter, Roholtiella, Aulosira and Komarekiella. Despite being morphologically similar to Komarekiella, phylogenetic analyses placed our strains in a separated genus-level clade, distant from Nostoc sensu stricto and with Komarekiella as the closest taxon. Considering all molecular, phylogenetic and ecological data, we here propose Parakomarekiella sesnandensis F.Soares, V.Ramos and A.Portugal, gen. et sp. nov.
- A novel molecular diagnostic method for the gut content analysis of Philaenus DNAPublication . Rodrigues, Isabel; Ramos, Vitor; Benhadi-Marín, Jacinto; Moreno, Aránzazu; Fereres, Alberto; Pereira, J.A.; Baptista, PaulaPhilaenus spumarius is a vector of Xylella fastidiosa, one of the most dangerous plants pathogenic bacteria worldwide. There is currently no control measure against this pathogen. Thus, the development of vector control strategies, like generalist predators, such as spiders, could be essential to limit the spread of this vector-borne pathogen. In this study, a polymerase chain reaction (PCR)-based approach was developed to principally detect DNA of P. spumarius in the spider’s gut. Accordingly, 20 primer pairs, targeting the mitochondrial cytochrome oxidase I (COI) and cytochrome b (cytB) genes, were tested for specificity, sensitivity, and efficiency in detecting P. spumarius DNA. Overall, two primer sets, targeting COI gene (COI_Ph71F/COI_Ph941R) and the cytB gene (cytB_ Ph85F/cytB_Ph635R), showed the highest specificity and sensitivity, being able to amplify 870 pb and 550 bp fragments, respectively, with P. spumarius DNA concentrations 100-fold lower than that of the DNA of non-target species. Among these two primer sets, the cytB_Ph85F/cytB_Ph635R was able to detect P. spumarius in the spider Xysticus acerbus, reaching 50% detection success 82 h after feeding. The feasibility of this primer set to detect predation of P. spumarius by spiders was confirmed in the field, where 20% of the collected spiders presented positive amplifications.