MOESM2 of Bacterial polyextremotolerant bioemulsifiers from arid soils improve water retention capacity and humidity uptake in sandy soil

Additional file 2: Figure S1. Phylogenetic affiliation of partial 16S rRNA gene of the 23 bacterial isolates obtained from chott and desert in the south of Tunisia constructed using MEGA6 package. Neighbor-Joining phylogenetic tree was built using MEGA 6, computing the evolutionary distances using the Kimura 2-parameter model

MOESM1 of Bacterial polyextremotolerant bioemulsifiers from arid soils improve water retention capacity and humidity uptake in sandy soil

Additional file 1:Table S1. List of the 23 bacterial strains used in this work

A Drought Resistance-Promoting Microbiome Is Selected by Root System under Desert Farming

<div><h3>Background</h3><p>Traditional agro-systems in arid areas are a bulwark for preserving soil stability and fertility, in the sight of “reverse desertification”. Nevertheless, the impact of desert farming practices on the diversity and abundance of the plant associated microbiome is poorly characterized, including its functional role in supporting plant development under drought stress.</p> <h3>Methodology/Principal Findings</h3><p>We assessed the structure of the microbiome associated to the drought-sensitive pepper plant (<em>Capsicum annuum</em> L.) cultivated in a traditional Egyptian farm, focusing on microbe contribution to a crucial ecosystem service, i.e. plant growth under water deficit. The root system was dissected by sampling root/soil with a different degree of association to the plant: the endosphere, the rhizosphere and the root surrounding soil that were compared to the uncultivated soil. Bacterial community structure and diversity, determined by using Denaturing Gradient Gel Electrophoresis, differed according to the microhabitat, indicating a selective pressure determined by the plant activity. Similarly, culturable bacteria genera showed different distribution in the three root system fractions. <em>Bacillus</em> spp. (68% of the isolates) were mainly recovered from the endosphere, while rhizosphere and the root surrounding soil fractions were dominated by <em>Klebsiella</em> spp. (61% and 44% respectively). Most of the isolates (95%) presented <em>in vitro</em> multiple plant growth promoting (PGP) activities and stress resistance capabilities, but their distribution was different among the root system fractions analyzed, with enhanced abilities for <em>Bacillus</em> and the rhizobacteria strains. We show that the <em>C. annuum</em> rhizosphere under desert farming enriched populations of PGP bacteria capable of enhancing plant photosynthetic activity and biomass synthesis (up to 40%) under drought stress.</p> <h3>Conclusions/Significance</h3><p>Crop cultivation provides critical ecosystem services in arid lands with the plant root system acting as a “resource island” able to attract and select microbial communities endowed with multiple PGP traits that sustain plant development under water limiting conditions.</p> </div

Diversity indexes of the microbial collection.

<p>The indexes were calculated for the sequences of bacterial strains isolated from the different fractions of the pepper root system and the non-cultivated arid soil. Sequences have been grouped in OTUs based on nucleotide similarity at 99%.</p

Distribution of microbial taxa in the collection of culturable bacterial isolates associated to pepper plants.

<p>Numbers indicate the number of strains assigned to each genus or species, respectively.</p><p>The numbers in parentheses are the total number of isolates for each fraction.</p

Rhizobacteria increased plant resistance to drought stress.

<p>Abbreviations for the figure: CP, (positive) abiotic control, irrigated at the water holding capacity of the soil along all the experiment; NC, (negative) abiotic control, subjected to drought by interrupting water supply for 12 days. (A) Representative images of plants exposed to rhizobacteria compared to untreated plants eight days after the induction of drought. (B) Leaf physiological parameters in treated and untreated plants eight days after the induction of drought.Abbreviations: Pn, net photosynthesis; E, evapo-transpiration; Gs, stomatal conductance; Ci, internal carbon dioxide (CO₂). Student t-test was adopted to statistically analyse the data. *:p≤0,05; **:p≤0,01; ***:p≤0,001. The data reported in the graphs are representative of one replicate experiment. (C) Percentage increase in root fresh weight (FW) and root length (L) of water stressed plants, compared to the abiotic stressed control, set as 0%.</p

Cluster analysis of total microbial communities according to 16S rRNA DGGE profiles.

<p>The cluster analysis of the plot line was obtained from 16S rRNA PCR-DGGE bacterial community profiles, according to Pearson correlation. The analyzed fractions were root tissues (E), rhizosphere (R), root-surrounding soil (S) and bulk soil (B) of three replicate plants of pepper.</p

Abundance of culturable bacteria associated to the different fractions of the pepper root system.

<p>The isolation was performed on different cultivation media. In the table it is reported the amount of bacterial isolates composing the strain collection associated to pepper endosphere and root-associated soil fractions. E, Endosphere; R, rhizosphere; S, root-surrounding soil; B, non-cultivated arid soil.</p

Rhizocompetence of <i>gfp</i>-labelled bacteria on different plant models.

<p>Plant root colonization experiments performed with a <i>Klebsiella pneumoniae</i> strain isolated from the pepper rhizosphere genetically labeled with a <i>gfp</i>. (A) and (B) colonization of <i>Arabidospis thaliana</i> rhizoplane; (C) and (D) colonization of the pepper rhizoplane. Red spots represent root autofluorescence as acquired through the TRICT filter. The scale bars of the different images in the figure correspond to 100 µm.</p

Phylogenetic identification and distribution of bacteria excised and sequenced from DGGE bands.

<p>Identification of the dominant bands in the PCR-DGGE fingerprinting profiles (marked in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0048479#pone.0048479.s001" target="_blank">Fig. S1</a>) and their distribution in the different fractions of the pepper root system. The codes of the different fractions of the pepper root systems are as follow: E, Endosphere; R, rhizosphere; S, root-surrounding soil; B, non-cultivated root-free arid soil. The numbers following the codes indicate the different replicates.</p><p>X: presence of the band in the DGGE profile of the indicated fraction; in bold are indicated the bands that were actually sequenced.</p><p>Sequences of bands with the same mobility in the DGGE gel are reported in the same white/grey boxes. In some cases the different bands showed slightly different sequences whit few nucleotide variations. When the variation resulted within the 3% divergence on the 16S rRNA sequence, the bands where assumed to belong to the same OTU at the 97% identity threshold, as evaluated using DOTUR <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0048479#pone.0048479-Schloss1" target="_blank">[68]</a>.</p