Dynamics of Scaphoideus titanus population in southern South Tyrol (Italy) and detection of Grapevine Flavescence Dorée phytoplasma in the insect with a multiplex real-time PCR

In South Tyrol the population of Scaphoideus titanus, the vector of Grapevine Flavescence dorée phytoplasma (FD), has been steadily increasing since 2010. The present work provides an overview of the monitoring activity of Scaphoideus titanus in South Tyrolean vineyards coupled with the description of a sensitive and reliable detection method of FD in the insect vector. We have developed an endogenous control of Scaphoideus titanus to be used in a multiplex real-time PCR, amplifying the map gene of FD phytoplasma. We present evidence on the performance of this new vector reference and its compatibility with FD detection

Metaproteogenomic analysis of microbial communities in the phyllosphere and rhizosphere of rice

The above- and below-ground parts of rice plants create specific habitats for various microorganisms. In this study, we characterized the phyllosphere and rhizosphere microbiota of rice cultivars using a metaproteogenomic approach to get insight into the physiology of the bacteria and archaea that live in association with rice. The metaproteomic datasets gave rise to a total of about 4600 identified proteins and indicated the presence of one-carbon conversion processes in the rhizosphere as well as in the phyllosphere. Proteins involved in methanogenesis and methanotrophy were found in the rhizosphere, whereas methanol-based methylotrophy linked to the genus Methylobacterium dominated within the protein repertoire of the phyllosphere microbiota. Further, physiological traits of differential importance in phyllosphere versus rhizosphere bacteria included transport processes and stress responses, which were more conspicuous in the phyllosphere samples. In contrast, dinitrogenase reductase was exclusively identified in the rhizosphere, despite the presence of nifH genes also in diverse phyllosphere bacteria

Synthetic vanillate-regulated promoter for graded gene expression in Sphingomonas

Regulated promoters are an important basic genetic tool allowing, for example, gene-dosage and gene depletion studies. We have previously described a cumate-inducible promoter (P(Q5)) that is functional in diverse Alphaproteobacteria. This promoter has been engineered by combining a synthetic minimal promoter, P(syn2), and operator sites and the repressor of the Pseudomonas putida F1 cym/cmt system. In the present study, we engineered a vanillate-regulated promoter using P(syn2) and the regulatory elements of the Caulobacter crescentus vanR-vanAB system. We show that the resulting promoter, which we called P(V10), responds rapidly to the inducer vanillate with an induction ratio of about two orders of magnitude in Sphingomonas melonis Fr1. In contrast to the switch-like behavior of P(Q5), P(V10) shows a linear dose-response curve at intermediate vanillate concentrations, allowing graded gene expression. P(V10) is functionally compatible with and independent of P(Q5) and cumate, and vice versa, suggesting that both systems can be used simultaneously

Community proteogenomics reveals insights into the physiology of phyllosphere bacteria

Aerial plant surfaces represent the largest biological interface on Earth and provide essential services as sites of carbon dioxide fixation, molecular oxygen release, and primary biomass production. Rather than existing as axenic organisms, plants are colonized by microorganisms that affect both their health and growth. To gain insight into the physiology of phyllosphere bacteria under in situ conditions, we performed a culture-independent analysis of the microbiota associated with leaves of soybean, clover, and Arabidopsis thaliana plants using a metaproteogenomic approach. We found a high consistency of the communities on the 3 different plant species, both with respect to the predominant community members (including the alphaproteobacterial genera Sphingomonas and Methylo bacterium) and with respect to their proteomes. Observed known proteins of Methylobacterium were to a large extent related to the ability of these bacteria to use methanol as a source of carbon and energy. A remarkably high expression of various TonB-dependent receptors was observed for Sphingomonas. Because these outer membrane proteins are involved in transport processes of various carbohydrates, a particularly large substrate utilization pattern for Sphingomonads can be assumed to occur in the phyllosphere. These adaptations at the genus level can be expected to contribute to the success and coexistence of these 2 taxa on plant leaves. We anticipate that our results will form the basis for the identification of unique traits of phyllosphere bacteria, and for uncovering previously unrecorded mechanisms of bacteria-plant and bacteria-bacteria relationships