Association between bacterial community and cadmium distribution across Colombian cacao crops.

Assessing the bacterial community composition across cacao crops is important to understand its potential role as a modulator of cadmium (Cd) translocation to plant tissues under field conditions; Cd mobility between soil and plants is a complex and multifactorial problem that cannot be captured only by experimentation. Although microbes have been shown to metabolize and drive the speciation of Cd under controlled conditions, regardless of the link between soil bacterial community (SBC) dynamics and Cd mobilization in the rhizosphere, only a few studies have addressed the relationship between soil bacterial community composition (SBCC) and Cd content in cacao seeds (Cdseed). Therefore, this study aimed to explore the association between SBCC and different factors influencing the distribution of Cd across cacao crop systems. This study comprised 225 samples collected across five farms, where we used an amplicon sequencing approach to characterize the bacterial community composition. The soil Cd concentration alone (Cdsoil) was a poor predictor of Cdseed. Still, we found that this relationship was more apparent when the variation within farms was controlled, suggesting a role of heterogeneity within farms in modulating Cd translocation and, thus, seed Cd content. Our results provide evidence of the link between soil bacterial communities and the distribution of Cd across Colombian cacao crops, and highlight the importance of incorporating fine-spatial-scale studies to advance the understanding of factors driving Cd uptake and accumulation in cacao plants. IMPORTANCE: Cadmium (Cd) content in cacao crops is an issue that generates interest due to the commercialization of chocolate for human consumption. Several studies provided evidence about the non-biological factors involved in its translocation into the cacao plant. However, factors related to this process, including soil bacterial community composition (SBCC), still need to be addressed. It is well known that soil microbiome could impact compounds' chemical transformation, including Cd, on the field. Here, we found the first evidence of the link between soil bacterial community composition and Cd concentration in cacao soils and seeds. It highlights the importance of including the variation of bacterial communities to assess the factors driving the Cd translocation into cacao seeds. Moreover, the results highlight the relevance of the spatial heterogeneity within and across cacao farms, influencing the variability of Cd concentrations.

De novo transcriptome sequencing and annotation of the Antarctic polychaete Microspio moorei (Spionidae) with its characterization of the heat stress-related proteins (HSP, SOD & CAT).

We present a de novo transcriptome assembly for the non-model Antarctic polychaete worm Microspio moorei (Spionidae) collected during Antarctic field expedition in Fildes Bay, King George Island, Antarctic Peninsula, in 2017. Here, we report the first transcriptome reference array for Microspio spp. The gene sequences of the spionid worm were annotated from a wide range of functions (i.e., biological, and metabolic processes, catalytic processes, and catalytic activity). HSP70, HSP90 SOD and CAT families were compared to reported annelid transcriptomes and proteomes. The phylogenetic analysis using COI, 16S, and 18S markers effectively clusters the species within the family. However, it also casts uncertainty on the monophyletic nature of the Microspio genera, indicating the necessity for additional data and potentially requiring a reevaluation of its grouping. Within these protein families, 3D model software was used to create one representative of their protein structures. Structural predictions were compared with related reported annelids living at different temperatures and a human X-ray reference. We found structural differences (RMSE >1.8) between the human HSP proteins but no significant differences between the polychaete-predicted proteins (RMSE <1.2). These results encourage further research of heat stress-related proteins, the development of genetic markers for climate change-induced temperature stress, and the study of the underlying mechanisms of the heat response. Moreover, these results motivate the extension of these findings to congeneric species.

Talaromyces santanderensis: A New Cadmium-Tolerant Fungus from Cacao Soils in Colombia.

Inorganic pollutants in Colombian cocoa (Theobroma cacao L.) agrosystems cause problems in the production, quality, and exportation of this raw material worldwide. There has been an increased interest in bioprospecting studies of different fungal species focused on the biosorption of heavy metals. Furthermore, fungi constitute a valuable, profitable, ecological, and efficient natural soil resource that could be considered in the integrated management of cadmium mitigation. This study reports a new species of Talaromyces isolated from a cocoa soil sample collected in San Vicente de Chucurí, Colombia. T. santanderensis is featured by Lemon Yellow (R. Pl. IV) mycelium on CYA, mono-to-biverticillade conidiophores, and acerose phialides. T. santanderensis is distinguished from related species by its growth rate on CYAS and powdery textures on MEA, YES and OA, high acid production on CREA and smaller conidia. It is differentiated from T. lentulus by its growth rate on CYA medium at 37 °C without exudate production, its cream (R. PI. XVI) margin on MEA, and dense sporulation on YES and CYA. Phylogenetic analysis was performed using a polyphasic approach, including different phylogenetic analyses of combined and individual ITS, CaM, BenA, and RPB2 gene sequences that indicate that it is new to science and is named Talaromyces santanderensis sp. nov. This new species belongs to the Talaromyces section and is closely related to T. lentulus, T. soli, T. tumuli, and T. pratensis (inside the T. pinophilus species complex) in the inferred phylogeny. Mycelia growth of the fungal strains was subjected to a range of 0-400 mg/kg Cd and incorporated into malt extract agar (MEA) in triplicates. Fungal radial growth was recorded every three days over a 13-day incubation period and In vitro cadmium tolerance tests showed a high tolerance index (0.81) when the mycelium was exposed to 300 mg/kg of Cd. Results suggest that T. santanderensis showed tolerance to Cd concentrations that exceed the permissible limits for contaminated soils, and it is promising for its use in bioremediation strategies to eliminate Cd from highly contaminated agricultural soils.

Development of a Method for Detecting and Estimating Moniliophthora roreri Spore Loads Based on Spore Traps and qPCR.

Frosty pod rot, caused by Moniliophthora roreri, is the most damaging disease of cacao in Latin America and, to better comprehend its epidemiology, we must understand its dissemination and proliferation. However, we do not know how M. roreri spores loads fluctuate in time and space due to the lack of a reliable technique to quantify M. roreri spores in the fields. Therefore, we developed a method that relies on spore traps and qPCR to detect and quantify M. roreri spore loads. This study demonstrated that the qPCR protocol can detect down to 0.025 ng of M. roreri DNA and quantify between 0.006 ng and 60 ng. Moreover, it demonstrated that qPCR protocol can detect and quantify DNA extracted from spore suspension and spore traps containing at least 2.9 × 104 M. roreri spores. However, the variability of the estimates for spore samples was high. Finally, we described a spore-trap device designed to carry spore traps in the field. The qPCR protocol and spore-trap device here developed will help in the understanding of the M. roreri dissemination patterns since they can be used to assess the environmental loads of M. roreri spore in cacao fields.

The Complete Chloroplast Genome of Plukenetia volubilis Provides Insights Into the Organelle Inheritance.

Plukenetia volubilis L. (Malpighiales: Euphorbiaceae), also known as Sacha inchi, is considered a promising crop due to its high seed content of unsaturated fatty acids (UFAs), all of them highly valuable for food and cosmetic industries, but the genetic basis of oil biosynthesis of this non-model plant is still insufficient. Here, we sequenced the total DNA of Sacha inchi by using Illumina and Nanopore technologies and approached a de novo reconstruction of the whole nucleotide sequence and the organization of its 164,111 bp length of the chloroplast genome, displaying two copies of an inverted repeat sequence [inverted repeat A (IRA) and inverted repeat B (IRB)] of 28,209 bp, each one separating a small single copy (SSC) region of 17,860 bp and a large single copy (LSC) region of 89,833 bp. We detected two large inversions on the chloroplast genome that were not presented in the previously reported sequence and studied a promising cpDNA marker, useful in phylogenetic approaches. This chloroplast DNA (cpDNA) marker was used on a set of five distinct Colombian cultivars of P. volubilis from different geographical locations to reveal their phylogenetic relationships. Thus, we evaluated if it has enough resolution to genotype cultivars, intending to crossbreed parents and following marker's trace down to the F1 generation. We finally elucidated, by using molecular and cytological methods on cut flower buds, that the inheritance mode of P. volubilis cpDNA is maternally transmitted and proposed that it occurs as long as it is physically excluded during pollen development. This de novo chloroplast genome will provide a valuable resource for studying this promising crop, allowing the determination of the organellar inheritance mechanism of some critical phenotypic traits and enabling the use of genetic engineering in breeding programs to develop new varieties.

Fungal Endophytes of Tahiti Lime (Citrus citrus × latifolia) and Their Potential for Control of Colletotrichum acutatum J. H. Simmonds Causing Anthracnose.

Colletotrichum acutatum is one of the causal agents of anthracnose in several crops, and of post-flowering fruit drop (PFD) in citrus and key lime anthracnose (KLA). The pathogen normally attacks flowers, causing lesions only in open flowers. Under very favorable conditions, however, it can also affect flower buds and small fruits, causing complete rotting of the fruit and a premature fall, resulting in major economic crop losses. We isolated endophytic fungi from Tahiti lime to evaluate its diversity, verify its antagonistic capacity against the phytopathogen Colletotrichum acutatum C-100 in dual tests, and evaluate the ability of various endophytic agents to control flowers with induced anthracnose. 138 fungal isolates were obtained from 486 fragments of branches, leaves, and fruit; from which 15 species were identified morphologically. A higher isolation frequency was found in branches and leaves, with a normal level of diversity compared to other citrus species. Of the 15 morphospecies, 5 were trialed against C. acutatum in antagonism tests, resulting in a finding of positive inhibition. 2 endophytic fungi from the antagonism tests demonstrated high inhibition of the phytopathogen, and were thus used in in vivo tests with Tahiti lime flowers, applied in a spore solution. Spore solutions of two molecularly identified species, Xylaria adscendens, and Trichoderma atroviride, reduced the lesions caused by the phytopathogen in these in vivo tests. The finding that these endophytes react antagonistically against C. acutatum may make them good candidates for further biological control research in an agroindustry that requires environmental sustainability.

Efficient direct shoot organogenesis and genetic stability in micropropagated sacha inchi (Plukenetia volubilis L.).

OBJECTIVE: It is necessary to improve biotech platforms based on in vitro cell tissue culture to support sacha inchi (Plukenetia volubilis L.) research programs and draw on the nutritional value of the high polyunsaturated fatty acid content of its oilseed. Here, we developed a rapid and efficient method for induction and direct in vitro shoot development for this species. RESULTS: Shoots were generated from hypocotyl explants. The highest organogenic response was obtained in woody plant medium supplemented with 1 mg/L thidiazuron and 0.5 mg/L zeatin supplemented with L-glutamine, adenine hemisulfate, and L-arginine. Shoots obtained using this medium were transferred and subcultivated with different concentrations of indole-3-butyric acid and 1-naphthylacetic acid for rooting. For the first time, a histological analysis was performed supporting direct organogenic development in this species. The plantlets obtained were transferred ex vitro with a survival percentage of 80%. The genetic stability of the plants recovered was confirmed by randomly amplified polymorphic DNA analysis. All results indicate that it would be possible to stimulate direct shoot formation from hypocotyls to support the sustainable use of this species.

Bacillus subtilis EA-CB0575 genome reveals clues for plant growth promotion and potential for sustainable agriculture.

Bacillus subtilis is a remarkably diverse bacterial species that displays many ecological functions. Given its genomic diversity, the strain Bacillus subtilis EA-CB0575, isolated from the rhizosphere of a banana plant, was sequenced and assembled to determine the genomic potential associated with its plant growth promotion potential. The genome was sequenced by Illumina technology and assembled using Velvet 1.2.10, resulting in a whole genome of 4.09 Mb with 4332 genes. Genes involved in the production of indoles, siderophores, lipopeptides, volatile compounds, phytase, bacilibactin, and nitrogenase were predicted by gene annotation or by metabolic pathway prediction by RAST. These potential traits were determined using in vitro biochemical tests, finding that B. subtilis EA-CB0575 produces two families of lipopeptides (surfactin and fengycin), solubilizes phosphate, fixes nitrogen, and produces indole and siderophores compounds. Finally, strain EA-CB0575 increased 34.60% the total dry weight (TDW) of tomato plants with respect to non-inoculated plants at greenhouse level. These results suggest that the identification of strain-specific genes and predicted metabolic pathways might explain the strain potential to promote plant growth by several mechanisms of action, accelerating the development of plant biostimulants for sustainable agricultural.

Inducible Antibacterial Activity in the Bacillales by Triphenyl Tetrazolium Chloride.

The world is in the midst of an antimicrobial resistance crisis, driving a need to discover novel antibiotic substances. Using chemical cues as inducers to unveil a microorganism's full metabolic potential is considered a successful strategy. To this end, we investigated an inducible antagonistic behavior in multiple isolates of the order Bacillales, where large inhibition zones were produced against Ralstonia solanacearum only when grown in the presence of the indicator triphenyl tetrazolium chloride (TTC). This bioactivity was produced in a TTC-dose dependent manner. Escherichia coli and Staphylococcus sp. isolates were also inhibited by Bacillus sp. strains in TTC presence, to a lesser extent. Knockout mutants and transcriptomic analysis of B. subtilis NCIB 3610 cells revealed that genes from the L-histidine biosynthetic pathway, the purine, pyrimidine de novo synthesis and salvage and interconversion routes, were significantly upregulated. Chemical space studied through metabolomic analysis, showed increased presence of nitrogenous compounds in extracts from induced bacteria. The metabolites orotic acid and L-phenylalaninamide were tested against R. solanacearum, E. coli, Staphylococcus sp. and B. subtilis, and exhibited activity against pathogens only in the presence of TTC, suggesting a biotransformation of nitrogenous compounds in Bacillus sp. cells as the plausible cause of the inducible antagonistic behavior.

Construction of probe of the plant growth-promoting bacteria Bacillus subtilis useful for fluorescence in situ hybridization.

Strains of Bacillus subtilis are plant growth-promoting bacteria (PGPB) of many crops and are used as inoculants. PGPB colonization is an important trait for success of a PGPB on plants. A specific probe, based on the 16 s rRNA of Bacillus subtilis, was designed and evaluated to distinguishing, by fluorescence in situ hybridization (FISH), between this species and the closely related Bacillus amyloliquefaciens. The selected target for the probe was between nucleotides 465 and 483 of the gene, where three different nucleotides can be identified. The designed probe successfully hybridized with several strains of Bacillus subtilis, but failed to hybridize not only with B. amyloliquefaciens, but also with other strains such as Bacillus altitudinis, Bacillus cereus, Bacillus gibsonii, Bacillus megaterium, Bacillus pumilus; and with the external phylogenetic strains Azospirillum brasilense Cd, Micrococcus sp. and Paenibacillus sp. The results showed the specificity of this molecular probe for B. subtilis.