Genome Sequence Data Reveal at Least Two Distinct Incursions of the Tropical Race 4 Variant of Fusarium Wilt into South America.

The global banana industry is threatened by one of the most devastating diseases: Fusarium wilt of banana. Fusarium wilt of banana is caused by the soilborne fungus Fusarium oxysporum f. sp. cubense (Foc), which almost annihilated the banana production in the late 1950s. A new strain of Foc, known as tropical race 4 (TR4), attacks a wide range of banana varieties, including Cavendish clones, which are the source of 99% of banana exports. In 2019, Foc TR4 was reported in Colombia, and more recently (2021) in Peru. In this study, we sequenced three fungal isolates identified as Foc TR4 from La Guajira (Colombia) and compared them against 19 whole-genome sequences of Foc TR4 publicly available, including four genome sequences recently released from Peru. To understand the genetic relatedness of the Colombian Foc TR4 isolates and those from Peru, we conducted a phylogenetic analysis based on a genome-wide set of single nucleotide polymorphisms (SNPs). Additionally, we compared the genomes of the 22 available Foc TR4 isolates, looking for the presence-absence of gene polymorphisms and genomic regions. Our results reveal that (i) the Colombian and Peruvian isolates are genetically distant, which could be better explained by independent incursions of the pathogen to the continent, and (ii) there is a high correspondence between the genetic relatedness and geographic origin of Foc TR4. The profile of present/absent genes and the distribution of missing genomic regions showed a high correspondence to the clades recovered in the phylogenetic analysis, supporting the results obtained by SNP-based phylogeny.

Suppression of tomato wilt by cell-free supernatants of Acinetobacter baumannii isolates from wild cacao from the Colombian Amazon.

Tomato vascular wilt caused by Fusarium oxysporum f. sp. lycopersici (Fol) is one of the most limiting diseases of this crop. The use of fungicides and varieties resistant to the pathogen has not provided adequate control of the disease. In this study, siderophore-producing bacteria isolated from wild cocoa trees from the Colombian Amazon were characterized to identify prominent strategies for plant protection. The isolates were taxonomically classified into five different genera. Eight of the fourteen were identified as bacteria of the Acinetobacter baumannii complex. Isolates CBIO024, CBIO086, CBIO117, CBIO123, and CBIO159 belonging to this complex showed the highest efficiency in siderophore synthesis, producing these molecules in a range of 91-129 µmol/L deferoxamine mesylate equivalents. A reduction in disease severity of up to 45% was obtained when plants were pretreated with CBIO117 siderophore-rich cell-free supernatant (SodSid). Regarding the mechanism of action that caused antagonistic activity against Fol, it was found that plants infected only with Fol and plants pretreated with SodSid CBIO117 and infected with Fol showed higher levels of PR1 and ERF1 gene expression than control plants. In contrast, MYC2 gene expression was not induced by the SodSid CBIO117 application. However, it was upregulated in plants infected with Fol and plants pretreated with SodSid CBIO117 and infected with the pathogen. In addition to the disease suppression exerted by SodSid CBIO117, the results suggest that the mechanism underlying this effect is related to an induction of systemic defense through the salicylic acid, ethylene, and priming defense via the jasmonic acid pathway.

Phenotypic and Genotypic Characterization of Phytophthora infestans Isolates Associated with Tomato and Potato Crops in Colombia.

Late blight disease, caused by the plant pathogen Phytophthora infestans, is one of the major threats for tomato and potato crops. Monitoring the populations of P. infestans is important to determine if there are changes in the sensitivity to fungicides and host preference. In this study, microsatellite markers and mitochondrial haplotypes were used to assess the genotype of isolates of P. infestans collected from tomato and potato plants in Colombia. Furthermore, sensitivity to the three fungicides cymoxanil (penetrant fungicide), mefenoxam, and fluopicolide (systemic fungicides), and tomato-potato host preference, were evaluated. Mitochondrial haplotyping showed that isolates collected on tomato were from the genetic groups Ia and Ib, while isolates collected on potatoes belonged to group IIa. Microsatellite analyses showed that isolates from tomato form two groups, including the Ib mitochondrial haplotype (which is genetically close to the US-1 clonal lineage) and the Ia haplotype (related to the EC-3 lineage), whereas Colombian isolates from potato formed a separate group. Furthermore, differences in sensitivity to fungicides were observed. Eighty-one percent of the isolates tested were resistant to mefenoxam with an EC50 >10 µg ml-1. Forty-two percent of the isolates showed an intermediate resistance to cymoxanil. The EC50 values ranged between 1 and 10 µg ml-1. For fluopicolide, 90% of the isolates were sensitive, with EC50 <1 µg ml-1. Host preference assays showed that potato isolates infected both host species. Thus, isolates that infect potatoes may pose a risk for tomato crops nearby.

Close-Range Thermography and Reflectance Spectroscopy Support In Vitro and In Vivo Characterization of Colletotrichum spp. Isolates from Mango Fruits.

Colletotrichum causing anthracnose in mango is known for its variable virulence that may have an effect on disease development and efficacy of management strategies. In this study, we characterized Colletotrichum spp. isolated from mango fruits under in vitro and in vivo conditions using close-range thermography and reflectance spectroscopy. Twenty-six isolates were phylogenetically characterized to ascertain species using the internal transcribed spacer sequence. Virulence, spectral (in vivo and in vitro), and thermographic responses (in vivo) of these isolates were analyzed. Isolates were grouped into the Colletotrichum gloeosporioides species complex and classified into eight morphotypes. Mycelial growth, conidia production, sporulation abundance, and area under disease progress curve (AUDPC) varied largely among isolates. Disease symptoms were observed 4 days after inoculation (dai), and, for most morphotypes, changes in tissue temperature were registered at 11 dai, with the greatest decrease at 14 dai with pathogen sporulation. In vitro and in vivo morphotypes shared changes in the spectrum range, and main variations were found in the number of informative spectral bands. In vivo average gross reflectance was higher in disease-inoculated tissue than in healthy uninoculated tissue. Morphotype responses varied depending on AUDPC values and postinoculation time. Discriminant analysis of the spectral response using principal component analysis and partial least squares regression explained 94 to 96.3 and 98 to 99.9% of the variance from in vitro and in vivo tests, respectively. Spectral markers were obtained for four distinct morphotype groups. We found three (550 to 650, 650.1 to 790, and 1,300 to 1,400 nm) and two (520 to 830 and 1,100 to 1,450 nm) regions with highly (P < 0.05) discriminant spectral bands for diseased fruits and morphotype characterization.

A novel Transposable element-derived microRNA participates in plant immunity to rice blast disease.

MicroRNAs (miRNAs) are small non-coding RNAs that direct post-transcriptional gene silencing in plant development and stress responses through cleavage or translational repression of target mRNAs. Here, we report the identification and functional characterization of a new member of the miR812 family in rice (named as miR812w) involved in disease resistance. miR812w is present in cultivated Oryza species, both japonica and indica subspecies, and wild rice species within the Oryza genus, but not in dicotyledonous species. miR812w is a 24nt-long that requires DCL3 for its biogenesis and is loaded into AGO4 proteins. Whereas overexpression of miR812w increased resistance to infection by the rice blast fungus Magnaporthe oryzae, CRISPR/Cas9-mediated MIR812w editing enhances disease susceptibility, supporting that miR812w plays a role in blast resistance. We show that miR812w derives from the Stowaway type of rice MITEs (Miniature Inverted-Repeat Transposable Elements). Moreover, miR812w directs DNA methylation in trans at target genes that have integrated a Stowaway MITE copy into their 3' or 5' untranslated region (ACO3, CIPK10, LRR genes), as well as in cis at the MIR812w locus. The target genes of miR812 were found to be hypo-methylated around the miR812 recognition site, their expression being up-regulated in transgene-free CRISPR/Cas9-edited miR812 plants. These findings further support that, in addition to post-transcriptional regulation of gene expression, miRNAs can exert their regulatory function at the transcriptional level. This relationship between miR812w and Stowaway MITEs integrated into multiple coding genes might eventually create a network for miR812w-mediated regulation of gene expression with implications in rice immunity.

pVHL suppresses Akt/ß-catenin-mediated cell proliferation by inhibiting 14-3-3ζ expression.

The mechanisms controlling degradation of cytosolic ß-catenin are important for regulating ß-catenin co-transcriptional activity. Loss of von Hippel-Lindau protein (pVHL) has been shown to stabilize ß-catenin, increasing ß-catenin transactivation and ß-catenin-mediated cell proliferation. However, the role of phosphoinositide 3-kinase (PI3K)/Akt in the regulation of ß-catenin signaling downstream from pVHL has never been addressed. Here, we report that hyperactivation of PI3K/Akt in cells lacking pVHL contributes to the stabilization and nuclear accumulation of active ß-catenin. PI3K/Akt hyperactivation is facilitated by the up-regulation of 14-3-3ζ and the down-regulation of 14-3-3ε, 14-3-3η and 14-3-3θ. Up-regulation of 14-3-3ζ in response to pVHL is important for the recruitment of PI3K to the cell membrane and for stabilization of soluble ß-catenin. In contrast, 14-3-3ε and 14-3-3η enhanced PI3K/Akt signaling by inhibiting PI3K and PDK1, respectively. Thus, our results demonstrated that 14-3-3 family members enhance PI3K/Akt/ß-catenin signaling in order to increase proliferation. Inhibition of Akt activation and/or 14-3-3 function strongly reduces ß-catenin signaling and decreases cell proliferation. Thus, inhibition of Akt and 14-3-3 function efficiently reduces cell proliferation in 786-0 cells characterized by hyperactivation of ß-catenin signaling due to pVHL loss.

Large-Scale, Ion-Current-Based Proteomic Investigation of the Rat Striatal Proteome in a Model of Short- and Long-Term Cocaine Withdrawal.

Given the tremendous detriments of cocaine dependence, effective diagnosis and patient stratification are critical for successful intervention yet difficult to achieve due to the largely unknown molecular mechanisms involved. To obtain new insights into cocaine dependence and withdrawal, we employed a reproducible, reliable, and large-scale proteomics approach to investigate the striatal proteomes of rats (n = 40, 10 per group) subjected to chronic cocaine exposure, followed by either short- (WD1) or long- (WD22) term withdrawal. By implementing a surfactant-aided precipitation/on-pellet digestion procedure, a reproducible and sensitive nanoLC-Orbitrap MS analysis, and an optimized ion-current-based MS1 quantification pipeline, >2000 nonredundant proteins were quantified confidently without missing data in any replicate. Although cocaine was cleared from the body, 129/37 altered proteins were observed in WD1/WD22 that are implicated in several biological processes related closely to drug-induced neuroplasticity. Although many of these changes recapitulate the findings from independent studies reported over the last two decades, some novel insights were obtained and further validated by immunoassays. For example, significantly elevated striatal protein kinase C activity persisted over the 22 day cocaine withdrawal. Cofilin-1 activity was up-regulated in WD1 and down-regulated in WD22. These discoveries suggest potentially distinct structural plasticity after short- and long-term cocaine withdrawal. In addition, this study provides compelling evidence that blood vessel narrowing, a long-known effect of cocaine use, occurred after long-term but not short-term withdrawal. In summary, this work developed a well-optimized paradigm for ion-current-based quantitative proteomics in brain tissues and obtained novel insights into molecular alterations in the striatum following cocaine exposure and withdrawal.

Transcriptomic and proteomic approach to identify differentially expressed genes and proteins in Arabidopsis thaliana mutants lacking chloroplastic 1 and cytosolic FBPases reveals several levels of metabolic regulation.

BACKGROUND: During the photosynthesis, two isoforms of the fructose-1,6-bisphosphatase (FBPase), the chloroplastidial (cFBP1) and the cytosolic (cyFBP), catalyse the first irreversible step during the conversion of triose phosphates (TP) to starch or sucrose, respectively. Deficiency in cyFBP and cFBP1 isoforms provokes an imbalance of the starch/sucrose ratio, causing a dramatic effect on plant development when the plastidial enzyme is lacking. RESULTS: We study the correlation between the transcriptome and proteome profile in rosettes and roots when cFBP1 or cyFBP genes are disrupted in Arabidopsis thaliana knock-out mutants. By using a 70-mer oligonucleotide microarray representing the genome of Arabidopsis we were able to identify 1067 and 1243 genes whose expressions are altered in the rosettes and roots of the cfbp1 mutant respectively; whilst in rosettes and roots of cyfbp mutant 1068 and 1079 genes are being up- or down-regulated respectively. Quantitative real-time PCR validated 100% of a set of 14 selected genes differentially expressed according to our microarray analysis. Two-dimensional (2-D) gel electrophoresis-based proteomic analysis revealed quantitative differences in 36 and 26 proteins regulated in rosettes and roots of cfbp1, respectively, whereas the 18 and 48 others were regulated in rosettes and roots of cyfbp mutant, respectively. The genes differentially expressed and the proteins more or less abundant revealed changes in protein metabolism, RNA regulation, cell signalling and organization, carbon metabolism, redox regulation, and transport together with biotic and abiotic stress. Notably, a significant set (25%) of the proteins identified were also found to be regulated at a transcriptional level. CONCLUSION: This transcriptomic and proteomic analysis is the first comprehensive and comparative study of the gene/protein re-adjustment that occurs in photosynthetic and non-photosynthetic organs of Arabidopsis mutants lacking FBPase isoforms.

Disruption of both chloroplastic and cytosolic FBPase genes results in a dwarf phenotype and important starch and metabolite changes in Arabidopsis thaliana.

In this study, evidence is provided for the role of fructose-1,6-bisphosphatases (FBPases) in plant development and carbohydrate synthesis and distribution by analysing two Arabidopsis thaliana T-DNA knockout mutant lines, cyfbp and cfbp1, and one double mutant cyfbp cfbp1 which affect each FBPase isoform, cytosolic and chloroplastic, respectively. cyFBP is involved in sucrose synthesis, whilst cFBP1 is a key enzyme in the Calvin-Benson cycle. In addition to the smaller rosette size and lower rate of photosynthesis, the lack of cFBP1 in the mutants cfbp1 and cyfbp cfbp1 leads to a lower content of soluble sugars, less starch accumulation, and a greater superoxide dismutase (SOD) activity. The mutants also had some developmental alterations, including stomatal opening defects and increased numbers of root vascular layers. Complementation also confirmed that the mutant phenotypes were caused by disruption of the cFBP1 gene. cyfbp mutant plants without cyFBP showed a higher starch content in the chloroplasts, but this did not greatly affect the phenotype. Notably, the sucrose content in cyfbp was close to that found in the wild type. The cyfbp cfbp1 double mutant displayed features of both parental lines but had the cfbp1 phenotype. All the mutants accumulated fructose-1,6-bisphosphate and triose-phosphate during the light period. These results prove that while the lack of cFBP1 induces important changes in a wide range of metabolites such as amino acids, sugars, and organic acids, the lack of cyFBP activity in Arabidopsis essentially provokes a carbon metabolism imbalance which does not compromise the viability of the double mutant cyfbp cfbp1.

Sex differences and sex-specific regulation of motivated behavior by Melanin-concentrating hormone: a short review.

Recent preclinical research exploring how neuropeptide transmitter systems regulate motivated behavior reveal the increasing importance of sex as a critical biological variable. Neuropeptide systems and their central circuits both contribute to sex differences in a range of motivated behaviors and regulate sex-specific behaviors. In this short review, we explore the current research of how sex as a biological variable influences several distinct motivated behaviors that are modulated by the melanin-concentrating hormone (MCH) neuropeptide system. First, we review how MCH regulates feeding behavior within the context of energy homeostasis differently between male and female rodents. Then, we focus on MCH's role in lactation as a sex-specific process within the context of energy homeostasis. Next, we discuss the sex-specific effects of MCH on maternal behavior. Finally, we summarize the role of MCH in drug-motivated behaviors. While these topics are traditionally investigated from different scientific perspectives, in this short review we discuss how these behaviors share commonalities within the larger context of motivated behaviors, and that sex differences discovered in one area of research may impact our understanding in another. Overall, our review highlights the need for further research into how sex differences in energy regulation associated with reproduction and parental care contribute to regulating motivated behaviors.

"A robust and simple catheter connector assembly for long-term self-administration experiments".

Intravenous self-administration in rats is used widely to study the reinforcing effects of drugs and serves as the gold standard for assessing their use and misuse potential. One challenge that researchers often encounter when scaling up experiments is balancing the cost, time investment to construct, and robustness of each implanted catheter. These catheters include multiple components such as surgical meshing and a variety of entry ports designed to facilitate the connection of the rat to a catheter port tethering system. Other considerations include maintaining the catheters free of blockage during the extent of the drug self-administration experiment. These large-scale studies provide ample opportunity for the catheter system to fail. The failure and replacement of commercially purchased catheters leads to ballooning expenses, and the failure of in-lab manufactured catheters requires the manufacture of reserves, also increasing costs, as these handmade products are inherently more variable. We have developed a catheter system that combines a commercially available implantable back-mounted entry connector system with inexpensive medical items such as surgical mesh, sutures, and an air-tight back flow prevention system to bolster the overall success of self-administration experiments.•Method to bolster commercially available jugular catheter components for long-lasting self-administration experiments.•Reduces the overall cost per unit of self-administration experiments.•Easily assembled by laboratory students and staff.

Synthetic exendin-4 disrupts responding to reward predictive incentive cues in male rats.

Synthetic exendin-4 (EX4, exenatide), is a GLP-1 receptor agonist used clinically to treat glycemia in Type-2 diabetes mellitus. EX4 also promotes weight loss and alters food reward-seeking behaviors in part due to activation of GLP-1 receptors in the mesolimbic dopamine system. Evidence suggests that GLP-1 receptor activity can directly attenuate cue-induced reward seeking. Here, we tested the effects of EX4 (0.6, 1.2, and 2.4 µg/kg, i.p.) on incentive cue (IC) responding, using a task where rats emit a nosepoke response during an intermittent reward-predictive IC to obtain a sucrose reward. EX4 dose-dependently attenuated responding to ICs and increased the latencies to respond to the IC and enter the sucrose reward cup. Moreover, EX4 dose-dependently decreased the total number of active port nosepokes for every cue presented. There was no effect of EX4 on the number of reward cup entries per reward earned, a related reward-seeking metric with similar locomotor demand. There was a dose-dependent interaction between the EX4 dose and session time on the responding to ICs and nosepoke response latency. The interaction indicated that effects of EX4 at the beginning and end of the session differed by the dose of EX4, suggesting dose-dependent pharmacokinetic effects. EX4 had no effect on free sucrose consumption behavior (i.e., total volume consumed, bout size, number of bouts) within the range of total sucrose volumes obtainable during the IC task (~3.5 ml). However, when rats were given unrestricted access for 1 h, where rats obtained much larger total volumes of sucrose (~30 ml), we observed some dose-dependent EX4 effects on drinking behavior, including decreases in total volume consumed. Together, these findings suggest that activation of the GLP-1 receptor modulates the incentive properties of cues attributed with motivational significance.

Assembly of an inexpensive rat jugular catheter button based on a split-septum needleless intravenous system.

Rat intravenous self-administration is a widely-used animal model in the study of substance use disorders. Rats are tethered to a drug delivery system usually through a port or button that interfaces the drug delivery system with a chronic indwelling jugular vein catheter. These buttons can be purchased commercially but are costly, presenting a significant economic barrier for many researchers. Many researchers manufacture buttons in-house from a combination of individual custom made and commercially available components, resulting in large variation in terms of how the animals are handled and the longevity of catheter patency. We have developed a jugular catheter button that uses a split septum port to provide snap-on entry of a blunt cannula allowing for quick and easy attachment of the i.v. tubing. The port is constructed from commercially available split septum ports, surgical mesh and small metal cannula. The system is "needleless" which decreases the risk of infection and improves safety. The split-septum buttons are easily sterilized in-house adding to the reliability and decreases in the risk of infection. We have used this easily constructed, and inexpensive button for i.v. self-administration experiments in which 80 % of the rats maintained patency for a minimum of 35 days.•Inexpensive method to construct a self-administration backport button.•Utilizes inexpensive components already found in a research laboratory or commercially available.•Can be sterilized in-house without degrading glue or components.

Complete genome sequence of Pediococcus acidilactici A40, a bacterium with biocontrol and plant growth-promoting properties.

We report the complete genome assembly of Pediococcus acidilactici A40, a bacterium with biocontrol and plant growth-promoting properties, obtained from Colombia.

Draft Genome Sequence and De Novo Assembly of a Fusarium oxysporum f. sp. lycopersici Isolate Collected from the Andean Region in Colombia.

We report a draft genome assembly of the causal agent of tomato vascular wilt, Fusarium oxysporum f. sp. lycopersici isolate 59, obtained from the Andean region in Colombia.

Activation of VTA GABA neurons disrupts reward seeking by altering temporal processing.

The role of ventral tegmental area (VTA) dopamine in reward, cue processing, and interval timing is well characterized. Using a combinatorial viral approach to target activating DREADDs (Designer Receptors Exclusively Activated by Designer Drugs, hM3D) to GABAergic neurons in the VTA of male rats, we previously showed that activation disrupts responding to reward-predictive cues. Here we explored how VTA GABA neurons influence the perception of time in two fixed interval (FI) tasks, one where the reward or interval is not paired with predictive cues (Non-Cued FI), and another where the start of the FI is signaled by a constant tone that continues until the rewarded response is emitted (Cued FI). Under vehicle conditions in both tasks, responding was characterized by "scalloping" over the 30 s FI, in which responding increased towards the end of the FI. However, when VTA GABA neurons were activated in the Non-Cued FI, the time between the end of the 30 s interval and when the rats made a reinforced response increased. Additionally, post-reinforcement pauses and overall session length increased. In the Cued FI task, VTA GABA activation produced erratic responding, with a decrease in earned rewards. Thus, while both tasks were disrupted by VTA GABA activation, responding that is constrained by a cue was more sensitive to this manipulation, possibly due to convergent effects on timing and cue processing. Together these results demonstrate that VTA GABA activity disrupts the perception of interval timing, particularly when the timing is set by cues.

Chemogenetic Activation of Mesoaccumbal Gamma-Aminobutyric Acid Projections Selectively Tunes Responses to Predictive Cues When Reward Value Is Abruptly Decreased.

BACKGROUND: Mesolimbic circuits regulate the attribution of motivational significance to incentive cues that predict reward, yet this network also plays a key role in adapting reward-seeking behavior when the contingencies linked to a cue unexpectedly change. Here, we asked whether mesoaccumbal GABA (gamma-aminobutyric acid) projections enhance adaptive responding to incentive cues of abruptly altered reward value, and whether these effects were distinct from global activation of all ventral tegmental area GABA circuits. METHODS: We used a viral targeting system to chemogenetically activate mesoaccumbal GABA projections in male rats during a novel cue-dependent operant value-shifting task, in which the volume of a sucrose reward associated with a predictive cue is suddenly altered, from the beginning and throughout the session. We compared the results with global activation of ventral tegmental area GABA neurons, which will activate local inhibitory circuits and long loop projections. RESULTS: We found that activation of mesoaccumbal GABA projections decreases responding to incentive cues associated with smaller-than-expected rewards. This tuning of behavioral responses was specific to cues associated with smaller-than-expected rewards but did not impact measures related to consuming the reward. In marked contrast, activating all ventral tegmental area GABA neurons resulted in a uniform decrease in responding to incentive cues irrespective of changes in the size of the reward. CONCLUSIONS: Targeted activation of mesoaccumbal GABA neurons facilitates adaptation in reward-seeking behaviors. This suggests that these projections may play a very specific role in associative learning processes.

Efficacy of Disinfectants against Fusarium oxysporum f. sp. cubense Tropical Race 4 Isolated from La Guajira, Colombia.

Banana, the main export fruit for Colombia, is threatened by Fusarium wilt (FWB), caused by Fusarium oxysporum f. sp. cubense (Foc), tropical race 4 (TR4). Pathogen containment through disinfecting tools, machinery, shoes, and any means that may carry contaminated soil particles with proper disinfectants is at the forefront of disease management. In this study, the biocide efficacy of 10 commercial quaternary ammonium compounds (QACs) products and one based on glutaraldehyde (GA) were evaluated on both reproductive structures (microconidia and macroconidia) and survival spores (chlamydospores) of Foc TR4 (strain 140038) isolated from La Guajira, Colombia. QACs were evaluated at 1200 ppm and two exposure times: <1 and 15 min in the absence or presence of soil. For GA disinfectant, four different concentrations (500, 800, 1200, and 2000 ppm) were evaluated at both contact times in the presence of soil. In the absence of soil, all QACs showed 100% biocidal efficiency against microconidia, macroconidia, and chlamydospores at both <1 and 15 min. The presence of soil decreased the efficacy of disinfectants, but some of them, such as QAC3_1st, QAC7_4th, and QAC5_4th, showed 98%, 98%, and 100% efficacy against Foc TR4 chlamydospores, respectively, after <1 min of contact time. For instance, the GA-based disinfectant was able to eliminate all Foc TR4 propagules after 15 min for all concentrations tested.

Combining transcriptome analysis and GWAS for identification and validation of marker genes in the Physalis peruviana-Fusarium oxysporum pathosystem.

Vascular wilt, caused by the pathogen Fusarium oxysporum f. sp. physali (Foph), is a major disease of cape gooseberry (Physalis peruviana L.) in Andean countries. Despite the economic losses caused by this disease, there are few studies related to molecular mechanisms in the P. peruviana-Foph pathosystem as a useful tool for crop improvement. This study evaluates eight candidate genes associated with this pathosystem, using real-time quantitative PCR (RT-qPCR). The genes were identified and selected from 1,653 differentially expressed genes (DEGs) derived from RNA-Seq analysis and from a previous genome-wide association study (GWAS) of this plant-pathogen interaction. Based on the RT-qPCR analysis, the tubuline (TUB) reference gene was selected for its highly stable expression in cape gooseberry. The RT-qPCR validation of the candidate genes revealed the biological variation in their expression according to their known biological function. Three genes related to the first line of resistance/defense responses were highly expressed earlier during infection in a susceptible genotype, while three others were overexpressed later, mostly in the tolerant genotype. These genes are mainly involved in signaling pathways after pathogen recognition, mediated by hormones such as ethylene and salicylic acid. This study provided the first insight to uncover the molecular mechanism from the P. peruviana-Foph pathosystem. The genes validated here have important implications in the disease progress and allow a better understanding of the defense response in cape gooseberry at the molecular level. Derived molecular markers from these genes could facilitate the identification of tolerant/susceptible genotypes for use in breeding schemes.

In planta gene expression analysis of Xanthomonas oryzae pathovar oryzae, African strain MAI1.

BACKGROUND: Bacterial leaf blight causes significant yield losses in rice crops throughout Asia and Africa. Although both the Asian and African strains of the pathogen, Xanthomonas oryzae pv. oryzae (Xoo), induce similar symptoms, they are nevertheless genetically different, with the African strains being more closely related to the Asian X. oryzae pv. oryzicola (Xoc). RESULTS: Changes in gene expression of the African Xoo strain MAI1 in the susceptible rice cultivar Nipponbare were profiled, using an SSH Xoo DNA microarray. Microarray hybridization was performed comparing bacteria recovered from plant tissues at 1, 3, and 6 days after inoculation (dai) with bacteria grown in vitro. A total of 710 bacterial genes were found to be differentially expressed, with 407 up-regulated and 303 down-regulated. Expression profiling indicated that less than 20% of the 710 bacterial transcripts were induced in the first 24 h after inoculation, whereas 63% were differentially expressed at 6 dai. The 710 differentially expressed genes were one-end sequenced. 535 sequences were obtained from which 147 non-redundant sequences were identified. Differentially expressed genes were related to metabolism, secretion and transport, pathogen adherence to plant tissues, plant cell-wall degradation, IS elements, and virulence. In addition, various other genes encoding proteins with unknown function or showing no similarity to other proteins were also induced. The Xoo MAI1 non-redundant set of sequences was compared against several X. oryzae genomes, revealing a specific group of genes that was present only in MAI1. Numerous IS elements were also found to be differentially expressed. Quantitative real-time PCR confirmed 86% of the identified profile on a set of 14 genes selected according to the microarray analysis. CONCLUSIONS: This is the first report to compare the expression of Xoo genes in planta across different time points during infection. This work shows that as-yet-unidentified and potentially new virulence factors are appearing in an emerging African pathogen. It also confirms that African Xoo strains do differ from their Asian counterparts, even at the transcriptional level.