Recently evolved combination of unique sulfatase and amidase genes enables bacterial degradation of the wastewater micropollutant acesulfame worldwide

Xenobiotics often challenge the principle of microbial infallibility. One example is acesulfame introduced in the 1980s as zero-calorie sweetener, which was recalcitrant in wastewater treatment plants until the early 2010s. Then, efficient removal has been reported with increasing frequency. By studying acesulfame metabolism in alphaproteobacterial degraders of the genera Bosea and Chelatococcus, we experimentally confirmed the previously postulated route of two subsequent hydrolysis steps via acetoacetamide-N-sulfonate (ANSA) to acetoacetate and sulfamate. Genome comparison of wildtype Bosea sp. 100-5 and an acesulfame degradation-defective mutant revealed the involvement of two plasmid-borne gene clusters. The acesulfame-hydrolyzing sulfatase is strictly manganese-dependent and belongs to the metallo beta-lactamase family. In all degraders analyzed, it is encoded on a highly conserved gene cluster embedded in a composite transposon. The ANSA amidase, on the other hand, is an amidase signature domain enzyme encoded in another gene cluster showing variable length among degrading strains. Transposition of the sulfatase gene cluster between chromosome and plasmid explains how the two catabolic gene clusters recently combined for the degradation of acesulfame. Searching available genomes and metagenomes for the two hydrolases and associated genes indicates that the acesulfame plasmid evolved and spread worldwide in short time. While the sulfatase is unprecedented and unique for acesulfame degraders, the amidase occurs in different genetic environments and likely evolved for the degradation of other substrates. Evolution of the acesulfame degradation pathway might have been supported by the presence of structurally related natural and anthropogenic compounds, such as aminoacyl sulfamate ribonucleotide or sulfonamide antibiotics

New microsatellite loci for water yam (Dioscorea alata, Dioscoreaceae) and cross‐amplification for other Dioscorea species

Dioscorea alata L. is one of the most widely distributed species of the genus in the humid and semihumid tropics and is associated with traditional agriculture. Only a few microsatellite markers have been developed so far for this and other Dioscorea species. We isolated 14 codominant polymorphic microsatellite markers using a microsatellite‐enriched genomic library technique. Ten microsatellite loci were selected, and 80 D. alata accessions from different regions in Brazil were evaluated with nine polymorphic loci. The polymorphism information content (PIC) varied from 0.39 to 0.78 and the power discrimination (PD) ranged from 0.15 to 0.91. Six of the markers showed transferability for the species D. bulbifera, D. cayenensis‐D. rotundata, and D. trifida. The SSR markers obtained are an important tool for further studies aiming to characterize the genetic diversity in D. alata and other Dioscorea spp. accessions986e144e146COORDENAÇÃO DE APERFEIÇOAMENTO DE PESSOAL DE NÍVEL SUPERIOR - CAPESFUNDAÇÃO DE AMPARO À PESQUISA DO ESTADO DE SÃO PAULO - FAPES

NEW MICROSATELLITE LOCI FOR WATER YAM (DIOSCOREA ALATA, DIOSCOREACEAE) AND CROSS-AMPLIFICATION FOR OTHER DIOSCOREA SPECIES

Premise of the study: Dioscorea alata L. is one of the most widely distributed species of the genus in the humid and semihumid tropics and is associated with traditional agriculture. Only a few microsatellite markers have been developed so far for this and other Dioscorea species. Methods and Results: We isolated 14 codominant polymorphic microsatellite markers using a microsatellite-enriched genomic library technique. Ten microsatellite loci were selected, and 80 D. alata accessions from different regions in Brazil were evaluated with nine polymorphic loci. The polymorphism information content (PIC) varied from 0.39 to 0.78 and the power discrimination (PD) ranged from 0.15 to 0.91. Six of the markers showed transferability for the species D. bulbifera, D. cayenensis-D. rotundata, and D. trifida. Conclusions: The SSR markers obtained are an important tool for further studies aiming to characterize the genetic diversity in D. alata and other Dioscorea spp. accessions.FAPESP (Fundacao de Amparo a Pesquisa do Estado de Sao Paulo)CAPES (Coordenacao de Aperfeicoamento de Pessoal de Nivel Superior), Brazi

Antagonistic activity of fungi from anthracnose lesions on Paullinia cupana against Colletotrichum sp.

Anthracnose is a cosmopolitan disease caused by Colletotrichum spp. that affects many crops worldwide. Observations have shown that anthracnose leaf lesions may be colonized by several non-pathogenic microorganisms. The relationship of these microorganisms with the pathogen as well as their potential as biocontrol agents is not well known. Guarana (Paullinia cupana Mart. var. sorbilis) is a typical native Amazon crop with unknown microbial diversity. Guarana productivity has been reduced by a fungal disease caused by Colletotrichum sp. In this study, we isolated 15 fungi from guarana anthracnose leaf lesions that belong to five genera: Fusarium, Phomopsis, Leptosphaeria, Microdochium and Pestalotiopsis. Four isolates from the Fusarium sp. (C6 and C10), Pestalotiopsis sp. (C3), and Microdochium sp. (P7) consistently inhibited anthracnose fungal growth in vitro. Except for the Microdochium sp. (P7), these isolates were also able to inhibit the growth of the pathogen in in vivo assays using detached guarana leaves. Some mechanisms related to the growth inhibition of this pathogen were studied. Fusarium sp. (C6) produced chitinases; Fusarium sp. (C6, C10) and Pestalotiopsis sp. (C3) produced antagonistic volatile organic compounds. These three isolates also inhibited the growth of Fusarium spp., a pathogen of several plant species, suggesting their potential broad range of growth inhibition of other phytopathogens

The auxin-producing Bacillus thuringiensis RZ2MS9 promotes the growth and modifies the root architecture of tomato (Solanum lycopersicum cv. Micro-Tom)

Strains of Bacillus thuringiensis (Bt) are commonly commercialized as bioinoculants for insect pest control, but their benefits go beyond their insecticidal property: they can act as plant growth-promoters. Auxins play a major role in the plant growth promotion. However, the mechanism of auxin production by the Bacilli group, and more specifically by Bt strains, is unclear. In previous work, the plant growth-promoting rhizobacterium (PGPR) B. thuringiensis strain RZ2MS9 increased the corn roots. This drew our attention to the strain’s auxin production trait, earlier detected in vitro. Here, we demonstrate that in its genome, RZ2MS9 harbours the complete set of genes required in two pathways that are used for Indole acetic acid (IAA) production. We also detected that the strain produces almost five times more IAA during the stationary phase. The bacterial application increased the shoot dry weight of the Micro-Tom (MT) tomato by 24%. The application also modified MT root architecture, with an increase of 26% in the average lateral root length and inhibition of the axial root. At the cellular level, RZ2MS9-treated MT plants presented elongated root cortical cells with intensified mitotic activity. Altogether, these are the best characterized auxin-associated phenotypes. Besides that, no growth alteration was detected in the auxin-insensitive diageotropic (dgt) plants either with or without the RZ2MS9 inoculation. Our results suggest that auxins play an important role in the ability of B. thuringiensis RZ2MS9 to promote MT growth and provide a better understanding of the auxin production mechanism by a Bt strain

Bacterial communities associated with anthracnose symptomatic and asymptomatic leaves of guarana, an endogenous tropical crop, and their pathogen antagonistic efects

Plants are colonized by diverse microorganisms that can substantially impact their health and growth. Understanding bacterial diversity and the relationships between bacteria and phytopathogens may be key to finding effective biocontrol agents. We evaluated the bacterial community associated with anthracnose symptomatic and asymptomatic leaves of guarana, a typical tropical crop. Bacterial communities were assessed through culture-independent techniques based on extensive 16S rRNA sequencing, and cultured bacterial strains were evaluated for their ability to inhibit the growth of Colletotrichum sp. as well as for enzyme and siderophore production. The culture-independent method revealed that Proteobacteria was the most abundant phylum, but many sequences were unclassified. The emergence of anthracnose disease did not significantly affect the bacterial community, but the abundance of the genera Acinetobacter, Pseudomonas and Klebsiella were significantly higher in the symptomatic leaves. In vitro growth of Colletotrichum sp. was inhibited by 11.38% of the cultured bacterial strains, and bacteria with the highest inhibition rates were isolated from symptomatic leaves, while asymptomatic leaves hosted significantly more bacteria that produced amylase and polygalacturonase. The bacterial isolate Bacillus sp. EpD2-5 demonstrated the highest inhibition rate against Colletotrichum sp., whereas the isolates EpD2-12 and FD5-12 from the same genus also had high inhibition rates. These isolates were also able to produce several hydrolytic enzymes and siderophores, indicating that they may be good candidates for the biocontrol of anthracnose. Our work demonstrated the importance of using a polyphasic approach to study microbial communities from plant diseases, and future work should focus on elucidating the roles of culture-independent bacterial communities in guarana anthracnose disease

Screening of tropically derived, multi-trait plant growth- promoting rhizobacteria and evaluation of corn and soybean colonization ability

The present study assessed the plant growth-promoting (PGP) traits and diversity of culturable rhizobacteria associated with guarana (Paullinia cupana), a typical tropical plant. Ninety-six bacteria were isolated, subjected to biochemical tests, and identified by partial or total 16S rDNA sequencing. Proteobacteria and Firmicutes were the dominant rhizospheric phyla found, and Burkholderia and Bacillus were the most abundant genera. Thirteen strains exhibited the four PGP traits evaluated, and most of them belonged to the genus Burkholderia. Two multitrait PGP strains, RZ2MS9 (Bacillus sp.) and RZ2MS16 (Burkholderia ambifaria), expressively promoted corn and soybean growth under greenhouse conditions. Compared to the non-inoculated control, increases in corn root dry weight of 247.8 and 136.9% were obtained with RZ2MS9 and RZ2MS16 inoculation, respectively, at 60 days after seeding. The dry weights of corn and soybean shoots were significantly higher than those of non-inoculated plants, showing increases of more than 47% for both strains and crops. However, soybean root dry weight did not increased after bacterial inoculation with either strain. The colonization behavior of RZ2MS16 was assessed using GFP-labeling combined with fluorescence microscopy and a cultivation-based approach for quantification. RZ2MS16:gfp was able to colonize the roots and shoots of corn and soybean, revealing an endophytic behavior

Bacillus thuringiensis RZ2MS9, a tropical plant growth-promoting rhizobacterium, colonizes maize endophytically and alters the plant's production of volatile organic compounds during co-inoculation with Azospirillum brasilense Ab-V5

The beneficial features of Bacillus thuringiensis (Bt) are not limited to its role as an insecticide; it is also able to promote plant growth interacting with plants and other plant growth-promoting rhizobacterium (PGPR). The PGPR Bt strain RZ2MS9 is a multi-trait maize growth promoter. We obtained a stable mutant of RZ2MS9 labelled with green fluorescent protein (RZ2MS9-GFP). We demonstrated that the Bt RZ2MS9-GFP successfully colonizes maize's roots and leaves endophytically. We evaluated whether RZ2MS9 has an additive effect on plant growth promotion when co-inoculated with Azospirillum brasilense Ab-V5. The two strains combined enhanced maize's roots and shoots dry weight around 50% and 80%, respectively, when compared to the non-inoculated control. However, non-differences were observed comparing RZ2MS9 alone and when co-inoculated with Ab-V5, In addition, we used co-inoculation experiments in glass chambers to analyse the plant's volatile organic compounds (VOCs) production during the maize-RZ2MS9 and maize-RZ2MS9-Ab-V5 interaction. We found that the single and co-inoculation altered maize's VOCs emission profile, with an increase in the production of indoles in the co-inoculation. Collectively, these results increase our knowledge about the interaction between the Bt and maize, and provide a new possibility of combined application with the commercial inoculant A. brasilense Ab-V5