Transcriptome analysis of the fungal pathogen Rosellinia necatrix during infection of a susceptible avocado rootstock identifies potential mechanisms of pathogenesis.

BACKGROUND: White root rot disease caused by Rosellinia necatrix is one of the most important threats affecting avocado productivity in tropical and subtropical climates. Control of this disease is complex and nowadays, lies in the use of physical and chemical methods, although none have proven to be fully effective. Detailed understanding of the molecular mechanisms underlying white root rot disease has the potential of aiding future developments in disease resistance and management. In this regard, this study used RNA-Seq technology to compare the transcriptomic profiles of R. necatrix during infection of susceptible avocado 'Dusa' roots with that obtained from the fungus cultured in rich medium. RESULTS: The transcriptomes from three biological replicates of R. necatrix colonizing avocado roots (RGA) and R. necatrix growing on potato dextrose agar media (RGPDA) were analyzed using Illumina sequencing. A total of 12,104 transcripts were obtained, among which 1937 were differentially expressed genes (DEG), 137 exclusively expressed in RGA and 160 in RGPDA. During the root infection process, genes involved in the production of fungal toxins, detoxification and transport of toxic compounds, hormone biosynthesis, gene silencing and plant cell wall degradation were overexpressed. Interestingly, 24 out of the 137 contigs expressed only during R. necatrix growth on avocado roots, were predicted as candidate effector proteins (CEP) with a probability above 60%. The PHI (Pathogen Host Interaction) database revealed that three of the R. necatrix CEP showed homology with previously annotated effectors, already proven experimentally via pathogen-host interaction. CONCLUSIONS: The analysis of the full-length transcriptome of R. necatrix during the infection process is suggesting that the success of this fungus to infect roots of diverse crops might be attributed to the production of different compounds which, singly or in combination, interfere with defense or signaling mechanisms shared among distinct plant families. The transcriptome analysis of R. necatrix during the infection process provides useful information and facilitates further research to a more in -depth understanding of the biology and virulence of this emergent pathogen. In turn, this will make possible to evolve novel strategies for white root rot management in avocado.

Mild water stress-induced priming enhance tolerance to Rosellinia necatrix in susceptible avocado rootstocks.

BACKGROUND: White root rot (WRR) disease caused by Rosellinia necatrix is one of the most important threats affecting avocado orchards in temperate regions. The eradication of WRR is a difficult task and environmentally friendly control methods are needed to lessen its impact. Priming plants with a stressor (biotic or abiotic) can be a strategy to enhance plant defense/tolerance against future stress episodes but, despite the known underlying common mechanisms, few studies use abiotic-priming for improving tolerance to forthcoming biotic-stress and vice versa ('cross-factor priming'). To assess whether cross-factor priming can be a potential method for enhancing avocado tolerance to WRR disease, 'Dusa' avocado rootstocks, susceptible to R. necatrix, were subjected to two levels of water stress (mild-WS and severe-WS) and, after drought-recovery, inoculated with R. necatrix. Physiological response and expression of plant defense related genes after drought-priming as well as the disease progression were evaluated. RESULTS: Water-stressed avocado plants showed lower water potential and stomatal limitations of photosynthesis compared to control plants. In addition, NPQ and qN values increased, indicating the activation of energy dissipating mechanisms closely related to the relief of oxidative stress. This response was proportional to the severity of the water stress and was accompanied by the deregulation of pathogen defense-related genes in the roots. After re-watering, leaf photosynthesis and plant water status recovered rapidly in both treatments, but roots of mild-WS primed plants showed a higher number of overexpressed genes related with plant defense than severe-WS primed plants. Disease progression after inoculating primed plants with R. necatrix was significantly delayed in mild-WS primed plants. CONCLUSIONS: These findings demonstrate that mild-WS can induce a primed state in the WRR susceptible avocado rootstock 'Dusa' and reveal that 'cross-factor priming' with water stress (abiotic stressor) is effective for increasing avocado tolerance against R. necatrix (biotic stressor), underpinning that plant responses against biotic and abiotic stress rely on common mechanisms. Potential applications of these results may involve an enhancement of WRR tolerance of current avocado groves and optimization of water use via low frequency deficit irrigation strategies.

Correlation of Allograft Weight to Recipient Body Weight Index on Renal Function in Kidney Transplantation.

BACKGROUND: The function of renal transplant grafts can be modified by many factors. In one study of graft weight/weight of the recipient, it was concluded to avoid renal transplantation in patients with kidneys with a low ratio between the graft and recipient weight (<2.5 g/kg). OBJECTIVE: The objective of this study was to describe the association between renal allograft weight and renal function 1 month after renal transplantation in the Mexican population. MATERIAL AND METHODS: We studied patients who underwent transplantation from living or cadaveric donors with 1 month of follow-up with a functioning graft. An observational, retrospective, analytic study from January 1, 2014 to November 1, 2014 was conducted. Graft weight, donor weight, recipient weight, age, donor gender, recipient creatinine, and renal function were recorded. RESULTS: A total of 88 patients were included: 35 women (39.8%) and 53 men (60.2%). Sixty (68%) received kidneys from living donors and 28 (31.8%) from deceased donors. Mean recipient body mass index (BMI) was 24.7 (±2.6). Mean graft weight was 152 g (±33.9). Creatinine at 1 month post-transplantation was 1.6 mg/dL (±2.0). Using a linear regression model cold ischemia time was related to serum creatinine at 1 month post-transplantation (P = .020). Using multivariate analysis, significance was observed with respect to these indexes and recipient renal function. Recipient gender also was related and showed statistical significance (P = .015). CONCLUSIONS: Renal graft function depends on many factors including the amount of functional renal mass and nephrons required according to the recipient's weight. The donor kidney weigt (DKW) / receptor body weigt (RBW) index should be considered as selection criteria of donors.

Nondestructive Detection of White Root Rot Disease in Avocado Rootstocks by Leaf Chlorophyll Fluorescence.

White root rot (WRR) disease caused by Rosellinia necatrix is one of the most important threats affecting avocado orchards in temperate regions. In this study, we monitored the progression of WRR disease at the leaf and root levels by the combination of nondestructive chlorophyll fluorescence measurements and confocal laser-scanning microscopy on avocado genotypes susceptible to R. necatrix. Leaf photochemistry was affected at early stages of disease development prior to the appearance of aboveground symptoms, made evident as significant decreases in the trapping efficiency of photosystem-II (Fv'/Fm') and in the steady-state of chlorophyll fluorescence yield (Fs) normalized to the minimal fluorescence yield (F0) (Fs/F0). Decreases in Fv'/Fm' and Fs/F0 were associated with different degrees of fungal penetration, primarily in the lateral roots but not in areas next to the main root collar. Aboveground symptoms were observed only when the fungus reached the root collar. Leaf physiology was also tracked in a tolerant genotype where no changes were observed during disease progression despite the presence of the fungus in the root system. These results highlight the usefulness of this technique for the early detection of fungal infection and the rapid removal of highly susceptible genotypes in rootstock avocado-breeding programs.

Biocontrol bacteria selected by a direct plant protection strategy against avocado white root rot show antagonism as a prevalent trait.

AIM: This study was undertaken to study bacterial strains obtained directly for their efficient direct control of the avocado white root rot, thus avoiding prescreening by any other possible mechanism of biocontrol which could bias the selection. METHODS AND RESULTS: A collection of 330 bacterial isolates was obtained from the roots and soil of healthy avocado trees. One hundred and forty-three representative bacterial isolates were tested in an avocado/Rosellinia test system, resulting in 22 presumptive protective strains, all of them identified mainly as Pseudomonas and Bacillus species. These 22 candidate strains were screened in a more accurate biocontrol trial, confirming protection of some strains (4 out of the 22). Analyses of the potential bacterial traits involved in the biocontrol activity suggest that different traits could act jointly in the final biocontrol response, but any of these traits were neither sufficient nor generalized for all the active bacteria. All the protective strains selected were antagonistic against some fungal root pathogens. CONCLUSIONS: Diverse bacteria with biocontrol activity could be obtained by a direct plant protection strategy of selection. All the biocontrol strains finally selected in this work were antagonistic, showing that antagonism is a prevalent trait in the biocontrol bacteria selected by a direct plant protection strategy. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first report on the isolation of biocontrol bacterial strains using direct plant protection strategy in the system avocado/Rosellinia. Characterization of selected biocontrol bacterial strains obtained by a direct plant protection strategy showed that antagonism is a prevalent trait in the selected strains in this experimental system. This suggests that antagonism could be used as useful strategy to select biocontrol strains.

GFP sheds light on the infection process of avocado roots by Rosellinia necatrix.

In order to monitor Rosellinia necatrix infection of avocado roots, we generated a plasmid vector (pCPXHY1eGFP) constitutively expressing EGFP and developed a protoplast transformation protocol. Using this protocol, four R. necatrix isolates were efficiently transformed and were shown to stably express EGFP homogeneously while not having any observable effect on pathogenicity. Confocal laser scanning microscopy (CLSM) images of avocado roots infected with the highly virulent isolate CH53-GFP demonstrated that fungal penetration of avocado roots occurs simultaneously at several random sites, but it occurs preferentially in the crown region as well as throughout the lenticels and in the junctions between epidermal cells. Not only were R. necatrix hyphae observed invading the epidermal and cortical root cells, but they were also able to penetrate the primary and secondary xylem. Scanning electron microscopy (SEM) images allowed detailed visualisation of the hyphal network generated by invasion of R. necatrix through the epidermal, cortical and vascular cells, including hyphal anastomosis and branching points. To our knowledge, this is the first report describing the construction of GFP-tagged strains belonging to the genus Rosellinia for monitoring white root rot using CLSM and SEM.