Luis E. Sáenz de Miera

Differences of small intestinal bacteria populations in adults and children with/without celiac disease: Effect of age, gluten diet, and disease

Background: Scientific evidence has revealed microecological changes in the intestinal tract of celiac infants. The objective of this work is the study of bacterial differences in the upper small intestine in both adults (healthy, untreated celiac disease [CD], and CD treated with a gluten‐free diet) and children (healthy and untreated CD). Methods: Intestinal bacterial communities were identified by 16S rRNA gene sequencing of DNA extracted from duodenal biopsies. Results: Analysis of the sequences from adults and children showed that this niche was colonized by bacteria affiliated mainly with three phyla: Firmicutes, Proteobacteria, and Bacteroidetes. In total, 89 different genera were identified in adults and 46 in children. Bacterial richness was significantly lower in the children than in the adults. A global principal component analysis of the bacterial communities of both healthy and untreated CD patient groups (including both children and adults) revealed a strong effect of age in principal component 1—clustering all adults and children separately—and a possible effect of the disease in adults with untreated patients clustering separately. Conclusions: There are bacterial differences in the upper small intestine between untreated children CD patients and untreated CD adults due to age. There are bacterial differences in the upper small bacteria microbiota between treated and untreated CD adults due to treatment with a gluten‐free diet. (Inflamm Bowel Dis 2011;)

Characterisation of the soil bacterial community structure and composition of natural and constructed wetlands

In the present study, the pyrosequencing of 16S ribosomal DNA was used to characterise the soil bacterial community composition of a constructed wetland receiving municipal wastewater and a nearby natural wetland. Soil samples were taken from different locations in each wetland (lagoon, zone with T. latifolia, zone with S. atrocinerea). Moreover, the water quality parameters were evaluated (pH, Tª, conductivity, dissolved oxygen, redox potential, nutrients and suspended solids), revealing that the organic matter and nutrient contents were significantly higher in the constructed wetland than in the natural one. In general, the bacterial communities of the natural wetland were more diverse than those of the constructed wetland. The major phylogenic groups of all soils included Proteobacteria, Verrucomicrobia and Chloroflexi, with Proteobacteria being the majority of the community composition. The Verrucomicrobia and Chloroflexi phyla were more abundant in the natural wetland than the constructed wetland; in contrast, the Proteobacteria phylum was more abundant in the constructed wetland than the natural wetland. Beta diversity analyses reveal that the soil bacterial communities in the natural wetland were less dissimilar to each other than to those of the constructed wetland.

Influence of environmental variables on the structure and composition of soil bacterial communities in natural and constructed wetlands

Bacteria are key players in wetland ecosystems, however many essential aspects regarding the ecology of wetland bacterial communities remain unknown. The present study characterizes soil bacterial communities from natural and constructed wetlands through the pyrosequencing of 16S rDNA genes in order to evaluate the influence of wetland variables on bacterial community composition and structure. The results show that the composition of soil bacterial communities was significantly associated with the wetland type (natural or constructed wetland), the type of environment (lagoon, Typha or Salix) and three continuous parameters (SOM, COD and TKN). However, no clear associations were observed with soil pH. Bacterial diversity values were significantly lower in the constructed wetland with the highest inlet nutrient concentrations. The abundances of particular metabolic groups were also related to wetland characteristics.

A Whole Genome DArTseq and SNP Analysis for Genetic Diversity Assessment in Durum Wheat from Central Fertile Crescent

Until now, little attention has been paid to the geographic distribution and evaluation of genetic diversity of durum wheat from the Central Fertile Crescent (modern-day Turkey and Syria). Turkey and Syria are considered as primary centers of wheat diversity, and thousands of locally adapted wheat landraces are still present in the farmers’ small fields. We planned this study to evaluate the genetic diversity of durum wheat landraces from the Central Fertile Crescent by genotyping based on DArTseq and SNP analysis. A total of 39,568 DArTseq and 20,661 SNP markers were used to characterize the genetic characteristic of 91 durum wheat land races. Clustering based on Neighbor joining analysis, principal coordinate as well as Bayesian model implemented in structure, clearly showed that the grouping pattern is not associated with the geographical distribution of the durum wheat due to the mixing of the Turkish and Syrian landraces. Significant correlation between DArTseq and SNP markers was observed in the Mantel test. However, we detected a non-significant relationship between geographical coordinates and DArTseq (r = -0.085) and SNP (r = -0.039) loci. These results showed that unconscious farmer selection and lack of the commercial varieties might have resulted in the exchange of genetic material and this was apparent in the genetic structure of durum wheat in Turkey and Syria. The genomic characterization presented here is an essential step towards a future exploitation of the available durum wheat genetic resources in genomic and breeding programs. The results of this study have also depicted a clear insight about the genetic diversity of wheat accessions from the Central Fertile Crescent.

Comparative analysis of the composition of bacterial communities from two constructed wetlands for municipal and swine wastewater treatment

This work provides information about bacterial community structure in natural wastewater treatment systems treating different types of wastewater. The diversity and composition of bacterial communities associated with the rhizosphere of Typha latifolia and Salix atrocinerea were studied and compared among two different natural wastewater treatment systems, using the direct sequencing of the 16S ribosomal RNA codifying genes. Phylogenetic affiliations of the bacteria detected allowed us to define the main groups present in these particular ecosystems. Moreover, bacterial community structure was studied through two diversity indices. Ten identified and five non-identified phyla were found in the samples; the phylum Proteobacteria was the predominant group in the four ecosystems. The results showed a bacterial community dominated by beta-proteobacteria and a lower diversity value in the swine wastewater treatment system. The municipal wastewater treatment system presented a high diverse community in both macrophytes (Typha latifolia and Salix atrocinerea), with gamma-proteobacteria and alpha-proteobacteria, respectively, as the most abundant groups.

High genetic diversity in a world-wide collection of Lathyrus sativus L. revealed by isozymatic analysis

Grasspea ( Lathyrus sativus L.) is an annual, herbaceous, drought-resistant legume and staple crop in Asian and African countries. Little is known about the nature and the amount of genetic diversity present in existing grasspea seed collections, yet this information is pivotal for future breeding programmes, such as those striving to reduce high neurotoxin levels present in seeds. Here we report on the level of genetic diversity within a world-wide collection of L. sativus , determined by isozymatic analysis. Although grasspea is generally considered a predominantly self-pollinating species, we found that the population genetic structure of these accessions showed a considerable outcrossing rate of 36%. The identification of a mixed mating system in L. sativus has significant implications for collecting and multiplying genetic resources for conservation and for future breeding purposes. In addition, we determined the genetic closeness of grasspea accessions from different geographical regions around the world. While we noticed an allelic richness in this species that was conserved across the regions, we did not find any evidence of high genetic identity between accessions, even when originating from the same geographical location. Instead, we found that greater genetic variability existed at the intra-regional level than at the inter-regional level.

New Insights into Capsicum spp Relatedness and the Diversification Process of Capsicum annuum in Spain

The successful exploitation of germplasm banks, harbouring plant genetic resources indispensable for plant breeding, will depend on our ability to characterize their genetic diversity. The Vegetable Germplasm Bank of Zaragoza (BGHZ) (Spain) holds an important Capsicum annuum collection, where most of the Spanish pepper variability is represented, as well as several accessions of other domesticated and non-domesticated Capsicum spp from all over the five continents. In the present work, a total of 51 C. annuum landraces (mainly from Spain) and 51 accessions from nine Capsicum species maintained at the BGHZ were evaluated using 39 microsatellite (SSR) markers spanning the whole genome. The 39 polymorphic markers allowed the detection of 381 alleles, with an average of 9.8 alleles per locus. A sizeable proportion of alleles (41.2%) were recorded as specific alleles and the majority of these were present at very low frequencies (rare alleles). Multivariate and model-based analyses partitioned the collection in seven clusters comprising the ten different Capsicum spp analysed: C. annuum, C. chinense, C. frutescens, C. pubescens, C. bacatum, C. chacoense and C. eximium. The data clearly showed the close relationships between C. chinense and C. frutescens. C. cardenasii and C. eximium were indistinguishable as a single, morphologically variable species. Moreover, C. chacoense was placed between C. baccatum and C. pubescens complexes. The C. annuum group was structured into three main clusters, mostly according to the pepper fruit shape, size and potential pungency. Results suggest that the diversification of C. annuum in Spain may occur from a rather limited gene pool, still represented by few landraces with ancestral traits. This ancient population would suffer from local selection at the distinct geographical regions of Spain, giving way to pungent and elongated fruited peppers in the South and Center, while sweet blocky and triangular types in Northern Spain.

Obtaining retrotransposon sequences, analysis of their genomic distribution and use of retrotransposon-derived genetic markers in lentil (Lens culinaris Medik.)

Retrotransposons with long terminal repeats (LTR-RTs) are widespread mobile elements in eukaryotic genomes. We obtained a total of 81 partial LTR-RT sequences from lentil corresponding to internal retrotransposon components and LTRs. Sequences were obtained by PCR from genomic DNA. Approximately 37% of the LTR-RT internal sequences presented premature stop codons, pointing out that these elements must be non-autonomous. LTR sequences were obtained using the iPBS technique which amplifies sequences between LTR-RTs. A total of 193 retrotransposon-derived genetic markers, mainly iPBS, were used to obtain a genetic linkage map from 94 F7 inbred recombinant lines derived from the cross between the cultivar Lupa and the wild ancestor L. culinaris subsp. orientalis. The genetic map included 136 markers located in eight linkage groups. Clusters of tightly linked retrotransposon-derived markers were detected in linkage groups LG1, LG2, and LG6, hence denoting a non-random genomic distribution. Phylogenetic analyses identified the LTR-RT families in which internal and LTR sequences are included. Ty3-gypsy elements were more frequent than Ty1-copia, mainly due to the high Ogre element frequency in lentil, as also occurs in other species of the tribe Vicieae. LTR and internal sequences were used to analyze in silico their distribution among the contigs of the lentil draft genome. Up to 8.8% of the lentil contigs evidenced the presence of at least one LTR-RT similar sequence. A statistical analysis suggested a non-random distribution of these elements within of the lentil genome. In most cases (between 97% and 72%, depending on the LTR-RT type) none of the internal sequences flanked by the LTR sequence pair was detected, suggesting that defective and non-autonomous LTR-RTs are very frequent in lentil. Results support that LTR-RTs are abundant and widespread throughout of the lentil genome and that they are a suitable source of genetic markers useful to carry out further genetic analyses.

A genome-wide identification and comparative analysis of the lentil MLO genes

Powdery mildew is a widespread fungal plant disease that can cause significant losses in many crops. Some MLO genes (Mildew resistance locus O) have proved to confer a durable resistance to powdery mildew in several species. Resistance granted by the MLO gene family members has prompted an increasing interest in characterizing these genes and implementing their use in plant breeding. Lentil (Lens culinaris Medik.) is a widely grown food legume almost exclusively consumed as dry seed with an average world production of 4.5 million tons. Powdery mildew causes severe losses on certain lentil cultivars under particular environmental conditions. Data mining of the lentil CDC Redberry draft genome allowed to identify up to 15 gene sequences with homology to known MLO genes, designated as LcMLOs. Further characterization of these gene sequences and their deduced protein sequences demonstrated conformity with key MLO protein characteristics such as the presence of transmembrane and calmodulin binding domains, as well as that of other conserved motifs. Phylogenetic and other comparative analyses revealed that LcMLO1 and LcMLO3 are the most likely gene orthologs related to powdery mildew response in other species, sharing a high similarity with other known resistance genes of dicot species, such as pea PsMLO1 and Medicago truncatula MtMLO1 and MtMLO3. Sets of primers were designed as tools to PCR amplify the genomic sequences of LcMLO1 and LcMLO3, also to screen lentil germplasm in search of resistance mutants. Primers were used to obtain the complete sequences of these two genes in all of the six wild lentil relatives. Respective to each gene, all Lens sequences shared a high similarity. Likewise, we used these primers to screen a working collection of 58 cultivated and 23 wild lentil accessions in search of length polymorphisms present in these two genes. All these data widen the insights on this gene family and can be useful for breeding programs in lentil and close related species.