Poliana Fernanda Giachetto

Large-scale analysis of differential gene expression in coffee genotypes resistant and susceptible to leaf miner–toward the identification of candidate genes for marker assisted-selection

BackgroundA successful development of herbivorous insects into plant tissues depends on coordination of metabolic processes. Plants have evolved complex mechanisms to recognize such attacks, and to trigger a defense response. To understand the transcriptional basis of this response, we compare gene expression profiles of two coffee genotypes, susceptible and resistant to leaf miner (Leucoptera coffella). A total of 22000 EST sequences from the Coffee Genome Database were selected for a microarray analysis. Fluorescence probes were synthesized using mRNA from the infested and non-infested coffee plants. Array hybridization, scanning and data normalization were performed using Nimble Scan® e ArrayStar® platforms. Genes with foldchange values +/-2 were considered differentially expressed. A validation of 18 differentially expressed genes was performed in infected plants using qRT-PCR approach.ResultsThe microarray analysis indicated that resistant plants differ in gene expression profile. We identified relevant transcriptional changes in defense strategies before insect attack. Expression changes (>2.00-fold) were found in resistant plants for 2137 genes (1266 up-regulated and 873 down-regulated). Up-regulated genes include those responsible for defense mechanisms, hypersensitive response and genes involved with cellular function and maintenance. Also, our analyses indicated that differential expression profiles between resistant and susceptible genotypes are observed in the absence of leaf-miner, indicating that defense is already build up in resistant plants, as a priming mechanism. Validation of selected genes pointed to four selected genes as suitable candidates for markers in assisted-selection of novel cultivars.ConclusionsOur results show evidences that coffee defense responses against leaf-miner attack are balanced with other cellular functions. Also analyses suggest a major metabolic reconfiguration that highlights the complexity of this response.

Genome-Wide Detection of CNVs and Their Association with Meat Tenderness in Nelore Cattle

Brazil is one of the largest beef producers and exporters in the world with the Nelore breed representing the vast majority of Brazilian cattle (Bos taurus indicus). Despite the great adaptability of the Nelore breed to tropical climate, meat tenderness (MT) remains to be improved. Several factors including genetic composition can influence MT. In this article, we report a genome-wide analysis of copy number variation (CNV) inferred from Illumina® High Density SNP-chip data for a Nelore population of 723 males. We detected >2,600 CNV regions (CNVRs) representing ≈6.5% of the genome. Comparing our results with previous studies revealed an overlap in ≈1400 CNVRs (>50%). A total of 1,155 CNVRs (43.6%) overlapped 2,750 genes. They were enriched for processes involving guanosine triphosphate (GTP), previously reported to influence skeletal muscle physiology and morphology. Nelore CNVRs also overlapped QTLs for MT reported in other breeds (8.9%, 236 CNVRs) and from a previous study with this population (4.1%, 109 CNVRs). Two CNVRs were also proximal to glutathione metabolism genes that were previously associated with MT. Genome-wide association study of CN state with estimated breeding values derived from meat shear force identified 6 regions, including a region on BTA3 that contains genes of the cAMP and cGMP pathway. Ten CNVRs that overlapped regions associated with MT were successfully validated by qPCR. Our results represent the first comprehensive CNV study in Bos taurus indicus cattle and identify regions in which copy number changes are potentially of importance for the MT phenotype.

New insights from molecular characterization of the tick Rhipicephalus (Boophilus) microplus in Brazil

The Rhipicephalus (Boophilus) microplus complex currently consists of five taxa, namely R. australis, R. annulatus, R. (B.) microplus clade A sensu, R. microplus clade B sensu, and R. (B.) microplus clade C sensu. Mitochondrial DNA-based methods help taxonomists when they are facing the morpho-taxonomic problem of distinguishing members of the R. (B.) microplus complex. The purpose of this study was to perform molecular characterization of ticks in all five regions of Brazil and infer their phylogenetic relationships. Molecular analysis characterized 10 haplotypes of the COX-1 gene. Molecular network analysis revealed that haplotype H-2 was the most dispersed of the studied populations (n = 11). Haplotype H-3 (n = 2) had the greatest genetic differentiation when compared to other Brazilian populations. A Bayesian phylogenetic tree of the COX-1 gene obtained strong support. In addition, it was observed that the population of R. (B.) microplus haplotype H-3 exhibited diverging branches among the other Brazilian populations in the study. The study concludes that the different regions of Brazil have R. (B.) microplus tick populations with distinct haplotypes.

Predominance of syntrophic bacteria, Methanosaeta and Methanoculleus in a two-stage up-flow anaerobic sludge blanket reactor treating coffee processing wastewater at high organic loading rate

The effect of the organic loading rate (OLR) on the performance and microbial composition of a two-stage UASB system treating coffee processing wastewater was assessed. The system was operated with OLR up to 18.2 g COD (L d)-1 and effluent recirculation. Methane production and effluent characteristics were monitored. The microbial composition was examined through next-generation sequencing and qPCR from the anaerobic sludge of the first reactor (R1) operated at low and high OLR. The system showed operational stability, obtaining a maximum methane production of 2.2 L CH4 (L d)-1, with a removal efficiency of COD and phenolic compounds of 84 and 73%, respectively. The performance of R1 at high OLR in steady conditions was associated with an appropriate proportion of nutrients (particularly Fe) and a marked increase of the syntrophic bacteria Syntrophus and Candidatus Cloacimonas, and acetoclastic and hydrogenotrophic methanogens, mainly Methanosaeta, Methanoculleus, Methanobacterium and Methanomassiliicoccus.

Genomic variants revealed by invariably missing genotypes in nelore cattle.

High density genotyping panels have been used in a wide range of applications. From population genetics to genome-wide association studies, this technology still offers the lowest cost and the most consistent solution for generating SNP data. However, in spite of the application, part of the generated data is always discarded from final datasets based on quality control criteria used to remove unreliable markers. Some discarded data consists of markers that failed to generate genotypes, labeled as missing genotypes. A subset of missing genotypes that occur in the whole population under study may be caused by technical issues but can also be explained by the presence of genomic variations that are in the vicinity of the assayed SNP and that prevent genotyping probes from annealing. The latter case may contain relevant information because these missing genotypes might be used to identify population-specific genomic variants. In order to assess which case is more prevalent, we used Illumina HD Bovine chip genotypes from 1,709 Nelore (Bos indicus) samples. We found 3,200 missing genotypes among the whole population. NGS re-sequencing data from 8 sires were used to verify the presence of genomic variations within their flanking regions in 81.56% of these missing genotypes. Furthermore, we discovered 3,300 novel SNPs/Indels, 31% of which are located in genes that may affect traits of importance for the genetic improvement of cattle production.

Influence of Antigenic Mutations in Time Evolution of the Immune Memory --- A Dynamic Modeling

In this paper, we study the behavior of immune memory against antigenic mutation. Using a dynamic model proposed in previous studies, we have performed simulations of several inoculations, where in each virtual sample the viral population undergoes mutations. Our results suggest that the sustainability of the immunizations is dependent on viral variability and that the memory lifetimes are not random, what contradicts what was suggested by recent works. We show that what may cause an apparent random behavior of the immune memory is the antigenic variability.