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The employment of biotechnology-based approaches such as somatic embryogenesis has been applied to several plants including Coffea sp. Despite the economic importance of this genus, few information about the role of key regulatory genes in somatic embryogenesis in coffee is available. This work provides information about ABI3 and VAL2 genes performance by RT-qPCR during indirect somatic embryogenesis of Coffea arabica. To achieve this, bioinformatics analysis was performed to identify the genes of the B3 superfamily in the coffee genome. The cell suspensions lines presented similar histological and regeneration patterns, yielding of up to 6.6 embryos per 1 mg of embryogenic aggregates at 7 months. We have identified possible orthologs for VAL2 (Cc06g00410) and ABI3 (Cc01g17380) as well as the other members belonging to superfamily B3. The CaABI3 expression was higher in dedifferentiated competent cells for somatic embryogenesis as compared with non-embryogenic calli. Whereas the expression of VAL2 gene is more active in cotyledonary embryos and plantlets, showing its clear performance in the embryogenesis late stages. The present study suggests that CaABI3 gene could be potentially used as a biomarker for embryogenic process improvement. The good plantlets development obtained from the protocol used may be a reflection of the high expression of CaVAL2 in cotyledonary embryos and plantlets. Key message The activity of CaABI3 is correlated to embryogenic potential with highly expressed in embryogenic masses and expression of the VAL2 gene is increased at the end of the embryogenic process.

Although several ant species are important targets for the development of molecular control strategies, only a few studies focus on identifying and validating reference genes for quantitative reverse transcription polymerase chain reaction (RT-qPCR) data normalization. We provide here an extensive study to identify and validate suitable reference genes for gene expression analysis in the ant Atta sexdens, a threatening agricultural pest in South America. The optimal number of reference genes varies according to each sample and the result generated by RefFinder differed about which is the most suitable reference gene. Results suggest that the RPS16, NADH and SDHB genes were the best reference genes in the sample pool according to stability values. The SNF7 gene expression pattern was stable in all evaluated sample set. In contrast, when using less stable reference genes for normalization a large variability in SNF7 gene expression was recorded. There is no universal reference gene suitable for all conditions under analysis, since these genes can also participate in different cellular functions, thus requiring a systematic validation of possible reference genes for each specific condition. The choice of reference genes on SNF7 gene normalization confirmed that unstable reference genes might drastically change the expression profile analysis of target candidate genes.

Atypical enteropathogenic Escherichia coli (aEPEC) strains are unable to produce the bundle-forming pilus (BFP), which is responsible for the localized adherence pattern, a characteristic of the pathogenicity of typical EPEC strains. The lack of BFP in aEPEC strains suggests that other fimbrial or non-fimbrial adhesins are involved in their adhesion to the host cells. The aim of this study was to investigate the distribution of major subunit fimbrial genes known to be important adherence factors produced by several E. coli pathotypes in a collection of 72 aEPEC strains. Our results demonstrate that a high percentage (94–100%) of aEPEC strains harbored ecpA, fimA, hcpA, and lpfA fimbrial genes. Other fimbrial genes including pilS, pilV, sfpA, daaC, papA, and sfa were detected at lower frequencies (1–8%). Genes encoding fimbrial subunits, which are characteristic of enteroaggregative E. coli or enterotoxigenic E. coli were not found. No correlation was found between fimbrial gene profiles and adherence phenotypes. Since all aEPEC strains contained ecpA, the major pilin gene of the E. coli common pilus (ECP), a subset of ecpA+ strains was analyzed for transcription of ecpRABCDE and production of ECP upon growth in three different culture conditions at 37°C. Transcription of ecpRABCDE occurred in all conditions; however, ECP production was medium dependent. In all, the data suggest that aEPEC strains are highly heterogeneous in terms of their fimbrial gene profiles. Despite lacking BFP production, other mechanisms of cell adherence exist in aEPEC strains to ensure host colonization, e.g., mediated by other prevalent pili such as ECP. Moreover, the production of ECP by aEPEC strains might be influenced by yet unknown post-transcriptional factors.

Garcinia brasiliensis, popularly known as Bacupari, is native to the Amazon and commonly used in folk medicine Fukugetin,

BackgroundCoffee production relies on plantations with varieties from Coffea arabica and Coffea canephora species. The first, the most representative in terms of coffee consumption, is mostly propagated by seeds, which leads to management problems regarding the plantations maintenance, harvest and processing of grains. Therefore, an efficient clonal propagation process is required for this species cultivation, which is possible by reaching a scalable and cost-effective somatic embryogenesis protocol. A key process on somatic embryogenesis induction is the auxin homeostasis performed by Gretchen Hagen 3 (GH3) proteins through amino acid conjugation. In this study, the GH3 family members were identified on C. canephora genome, and by performing analysis related to gene and protein structure and transcriptomic profile on embryogenic tissues, we point a GH3 gene as a potential regulator of auxin homeostasis during early somatic embryogenesis in C. arabica plants.ResultsWe have searched within the published C. canephora genome and found 17 GH3 family members. We checked the conserved domains for GH3 proteins and clustered the members in three main groups according to phylogenetic relationships. We identified amino acids sets in four GH3 proteins that are related to acidic amino acid conjugation to auxin, and using a transcription factor (TF) network approach followed by RT-qPCR we analyzed their possible transcriptional regulators and expression profiles in cells with contrasting embryogenic potential in C. arabica. The CaGH3.15 expression pattern is the most correlated with embryogenic potential and with CaBBM, a C. arabica ortholog of a major somatic embryogenesis regulator.ConclusionTherefore, one out of the GH3 members may be influencing on coffee somatic embryogenesis by auxin conjugation with acidic amino acids, which leads to the phytohormone degradation. It is an indicative that this gene can serve as a molecular marker for coffee cells with embryogenic potential and needs to be further studied on how much determinant it is for this process. This work, together with future studies, can support the improvement of coffee clonal propagation through in vitro derived somatic embryos.

Coffea arabica is the most economically important coffee species worldwide. However, its production is severely limited by diseases such as rust. The mechanisms underlying constitutive defense responses in coffee are still poorly understood, compared with induced defense mechanisms. We aimed to characterize constitutive defense responses of thirteen cultivars of C. arabica. Cultivars were classified under field conditions according to the level of resistance to rust: resistant (R), moderately resistant (MR), and susceptible (S). Based on this classification, the stability of eight reference genes (RGs) was evaluated. The most stable RGs were EF1α, APT1, and 24S. We also evaluated the expression of CaWRKY1, CaPAL1, CaCAD1, and CaPOX1, and activities of PAL, CAD, and POX, which are involved in lignin biosynthesis, and leaf content of total phenolic compounds and lignin. Gene expression and enzymatic activity were not correlated with defense metabolites in the R cultivar group but showed a negative correlation with phenolic compounds in MR cultivars. Cultivar S showed positive correlations of gene expression and enzyme activity with phenolic compounds. These results may assist coffee breeding programs regarding selection of genotypes and in optimization of rust resistance.