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Malate oxidation by plant mitochondria enables the generation of both oxaloacetate and pyruvate for tricarboxylic acid (TCA) cycle function, potentially eliminating the need for pyruvate transport into mitochondria in plants. Here, we show that the absence of the mitochondrial pyruvate carrier 1 (MPC1) causes the co-commitment loss of its putative orthologs, MPC3/MPC4, and eliminates pyruvate transport into Arabidopsis thaliana mitochondria, proving it is essential for MPC complex function. While the loss of either MPC or mitochondrial pyruvate-generating NAD-malic enzyme (NAD-ME) did not cause vegetative phenotypes, the lack of both reduced plant growth and caused an increase in cellular pyruvate levels, indicating a block in respiratory metabolism, and elevated the levels of branched-chain amino acids at night, a sign of alterative substrate provision for respiration. 13C-pyruvate feeding of leaves lacking MPC showed metabolic homeostasis was largely maintained except for alanine and glutamate, indicating that transamination contributes to the restoration of the metabolic network to an operating equilibrium by delivering pyruvate independently of MPC into the matrix. Inhibition of alanine aminotransferases when MPC1 is absent resulted in extremely retarded phenotypes in Arabidopsis, suggesting all pyruvate-supplying enzymes work synergistically to support the TCA cycle for sustained plant growth.

Malate oxidation by plant mitochondria enables the generation of both oxaloacetate (OAA) and pyruvate for tricarboxylic acid (TCA) cycle function, potentially eliminating the need for pyruvate transport into mitochondria in plants. Here we show that the absence of the mitochondrial pyruvate carrier 1 (MPC1) causes the co-commitment loss of its orthologs, MPC3/MPC4, and eliminates pyruvate transport into Arabidopsis mitochondria, proving it is essential for MPC complex function. While the loss of either MPC or mitochondrial pyruvate-generating NAD-malic enzyme (NAD-ME) did not cause vegetative phenotypes, the lack of both reduced plant growth and caused an increase in cellular pyruvate levels, indicating a block in respiratory metabolism, and elevated the levels of branched-chain amino acids at night, a sign of alterative substrate provision for respiration.13C-pyruvate feeding of leaves lacking MPC showed metabolic homeostasis were largely maintained except for alanine and glutamate, indicating that transamination contributes to restoration of the metabolic network to an operating equilibrium by delivering pyruvate independently of MPC into the matrix. Inhibition of alanine aminotransferases (AlaAT) when MPC1 is absent resulted in extremely retarded phenotypes in Arabidopsis, suggesting all pyruvate-supplying enzymes work synergistically to support the TCA cycle for sustained plant growth.

The effects of three endophytic Streptomyces on plant growth and the symbiosis of Lucerne and its rhizobial partner were examined in the presence of three levels of soil nitrogen. Three Streptomyces strains, LuP30 and LuP47B isolated from the roots of Lucerne (Medicago sativa L.) and EN23 isolated from roots of wheat (Triticum aestivum L.) were added as spores to Lucerne seeds (with and without Sinorhizobium meliloti RRI 128) at three levels of applied NH4NO3: 3, 25 and 50 mg/kg of soil. Plant growth increased with the addition of the actinobacteria strains alone from 19 % to 33 %. Co-inoculation of LuP30 with rhizobia strain RRI 128 produced the largest increase in shoot weight (46 %) of Lucerne plants growing in soil with 25 mg/kg NH4NO3. Co-inoculation with each of the actinobacteria with the rhizobia increased the number of nodules by more than 100 % compared with RRI128 alone, 4 weeks after rhizobial inoculation. A labelled 15N experiment showed co-inoculation with rhizobia and LuP30 or LuP47B enhanced N2-fixation 47 % and 72 %, respectively. The actinobacteria significantly improved plant growth and N2-fixation when applied with the rhizobia strain RRI 128 to Lucerne plants growing in soil supplied with 25 mg/kg NH4NO3.

Background and aims Endophytic actinobacteria are known to benefit their hosts by improving plant growth and by reducing the severity of soil borne diseases. In this study, their role in enhancing the growth of lucerne and their interaction with its rhizobial symbiosis is examined. Comparison is made between endophytic actinobacteria isolated from wheat plants and isolates from the roots and nodules of four different legume species: lucerne (Medicago sativa L.), field pea (Pisum sativum L.), subterranean clover (Trifolium subterraneum L.) and burr medic (Medicago polymorpha L.).

High-throughput sequencing libraries are typically limited by the requirement for nanograms to micrograms of input DNA. This bottleneck impedes the microscale analysis of ecosystems and the exploration of low biomass samples. Current methods for amplifying environmental DNA to bypass this bottleneck introduce considerable bias into metagenomic profiles. Here we describe and validate a simple modification of the Illumina Nextera XT DNA library preparation kit which allows creation of shotgun libraries from sub-nanogram amounts of input DNA. Community composition was reproducible down to 100 fg of input DNA based on analysis of a mock community comprising 54 phylogenetically diverse Bacteria and Archaea. The main technical issues with the low input libraries were a greater potential for contamination, limited DNA complexity which has a direct effect on assembly and binning, and an associated higher percentage of read duplicates. We recommend a lower limit of 1 pg (∼100–1,000 microbial cells) to ensure community composition fidelity, and the inclusion of negative controls to identify reagent-specific contaminants. Applying the approach to marine surface water, pronounced differences were observed between bacterial community profiles of microliter volume samples, which we attribute to biological variation. This result is consistent with expected microscale patchiness in marine communities. We thus envision that our benchmarked, slightly modified low input DNA protocol will be beneficial for microscale and low biomass metagenomics., PeerJ, 4, ISSN:2167-8359