Biology subjects

Murillo-Roos, M.

Publications and source records attributed to Murillo-Roos, M..

2 recordsLinked to original sources

bioRxiv2021

Niche separation in cross-feeding sustains bacterial strain diversity across nutrient environments and may increase chances for survival in nutrient-limited leaf apoplasts

The leaf microbiome plays a crucial role in plants health and resilience to stress. Like in other hosts, successful colonization is dependent on multiple factors, among them, resource accessibility. The apoplast is an important site of plant-microbe interactions where nutrients are tightly regulated. While leaf pathogens have evolved elaborate strategies to obtain nutrients there, it is not yet clear how commensals survive without most of these adaptations. Resource limitation can promote metabolic interactions, which in turn shape and stabilize microbiomes but this has not been addressed in detail in leaves. Here, we investigated whether and how the nutrient environment might influence metabolic exchange and assembly of bacterial communities in Flaveria trinervia and F. robusta leaves. We enriched bacteria from both plant species in-vitro in minimal media with sucrose as a carbon source, and with or without amino acids. After enrichment, we studied the genetic and metabolic diversity within the communities. Enriched Pseudomonas koreensis strains could cross-feed from diverse leaf bacteria. Although P. koreensis could not utilize sucrose, cross-feeding diverse metabolites from Pantoea sp ensured their survival in the sucrose-only enrichments. The Pseudomonas strains had high genetic similarity ([~]99.8% ANI) but still displayed clear niche partitioning, enabling them to simultaneously cross-feed from Pantoea. Interestingly, cross-feeders were only enriched from F. robusta and not from F. trinervia. Untargeted metabolomics analysis of the leaf apoplasts revealed contrasting nutrient environments, with greater concentrations of high-cost amino acids in F. trinervia. Additionally, P. koreensis strains were better able to survive without a cross-feeding partner in these richer apoplasts. Thus, cross feeding might arise as an adaptation to cope with nutrient limitations in the apoplast. Understanding how apoplast resources influence metabolic interactions could therefore provide plant breeders targets to manipulate leaf microbiome shape and stability.

microbiology↗

bioRxiv2020

Obtaining deeper insights into microbiome diversity using a simple method to block host and non-targets in amplicon sequencing

Microbiome profiling is revolutionizing our understanding of biological mechanisms such as metaorganismal (host+microbiome) assembly, functions and adaptation. Amplicon sequencing of multiple conserved, phylogenetically informative loci is an instrumental tool for characterization of the highly diverse microbiomes of natural systems. Investigations in many study systems are hindered by loss of essential sequencing depth due to amplification of non-target DNA from hosts or overabundant microorganisms. This issue requires urgent attention to address ecologically relevant problems using high throughput, high resolution microbial profiling. Here, we introduce a simple, low cost and highly flexible method using standard oligonucleotides ("blocking oligos") to block amplification of non-targets and an R package to aid in their design. They can be dropped into practically any two-step amplicon sequencing library preparation pipeline. We apply them in leaves, a system presenting exceptional challenges with host and non-target microbial amplification. Blocking oligos designed for use in eight target loci reduce undesirable amplification of host and non-target microbial DNA by up to 90%. In addition, 16S and 18S "universal" plant blocking oligos efficiently block most plant hosts, leading to increased microbial alpha diversity discovery without biasing beta diversity measurements. By blocking only chloroplast 16S amplification, we show that blocking oligos do not compromise quantitative microbial load information inherent to plant-associated amplicon sequencing data. Using these tools, we generated a near-complete survey of the Arabidopsis thaliana leaf microbiome based on diversity data from eight loci and discuss complementarity of commonly used amplicon sequencing regions for describing leaf microbiota. The blocking oligo approach has potential to make new questions in a variety of study systems more tractable by making amplicon sequencing more targeted, leading to deeper, systems-based insights into microbial discovery.

microbiology↗