8 research outputs found
Genome of the destructive oomycete Phytophthora cinnamomi provides insights into its pathogenicity and adaptive potential
BACKGROUND: Phytophthora cinnamomi is an oomycete pathogen of global relevance. It is considered as one of the
most invasive species, which has caused irreversible damage to natural ecosystems and horticultural crops. There is
currently a lack of a high-quality reference genome for this species despite several attempts that have been made
towards sequencing its genome. The lack of a good quality genome sequence has been a setback for various
genetic and genomic research to be done on this species. As a consequence, little is known regarding its genome
characteristics and how these contribute to its pathogenicity and invasiveness.
RESULTS: In this work we generated a high-quality genome sequence and annotation for P. cinnamomi using a
combination of Oxford Nanopore and Illumina sequencing technologies. The annotation was done using RNA-Seq
data as supporting gene evidence. The final assembly consisted of 133 scaffolds, with an estimated genome size of
109.7 Mb, N50 of 1.18 Mb, and BUSCO completeness score of 97.5%. Genome partitioning analysis revealed that P.
cinnamomi has a two-speed genome characteristic, similar to that of other oomycetes and fungal plant pathogens.
In planta gene expression analysis revealed up-regulation of pathogenicity-related genes, suggesting their
important roles during infection and host degradation.
CONCLUSION: This study has provided a high-quality reference genome and annotation for P. cinnamomi. This is
among the best assembled genomes for any Phytophthora species assembled to date and thus resulted in
improved identification and characterization of pathogenicity-related genes, some of which were undetected in
previous versions of genome assemblies. Phytophthora cinnamomi harbours a large number of effector genes which
are located in the gene-poor regions of the genome. This unique genomic partitioning provides P. cinnamomi with
a high level of adaptability and could contribute to its success as a highly invasive species. Finally, the genome
sequence, its annotation and the pathogenicity effectors identified in this study will serve as an important resource
that will enable future studies to better understand and mitigate the impact of this important pathogen.ADDITIONAL FILE 1 : Figure S1. Genomic profiling using short read data
for the sequenced Phytophthora cinnamomi isolate (GKB4). Figure S2.
GO enrichment analysis of up-regulated genes identified during infection
in Phytophthora cinnamomi.ADDITIONAL FILE 2 : Supplementary Table 1.
Genome assembly statistics of GKB4 and other currently available genome sequences for Phytophthora cinnamomi. Table S2. RxLR effectors identified in the current version of P. cinnamomi genome following the method of McGowin and Fitzpatrick (2017). Table S3. Crinkler effectors identified in the current version of P. cinnamomi genome. Table S4. NLPs identified in the current version of P. cinnamomi genome. Table S5. CAZymes identified from the current version of P. cinnamomi genome. Table S6. Summary of RNA-Seq data generated in this study. Table S7. List of differentially expressed genes between mycelia and in planta infection as identified from DESeq2.The Hans Merensky Foundationhttp://www.biomedcentral.com/bmcgenomicspm2021BiochemistryForestry and Agricultural Biotechnology Institute (FABI)GeneticsMicrobiology and Plant Patholog
Advances in understanding defense mechanisms in Persea americana against Phytophthora cinnamomi
AfricaAvocado (Persea americana) is an economically important fruit crop world-wide, the production of which is challenged by notable root pathogens such as Phytophthora cinnamomi and Rosellinia necatrix. Arguably the most prevalent, P. cinnamomi, is a hemibiotrophic oomycete which causes Phytophthora root rot, leading to reduced yields and eventual tree death. Despite its’ importance, the development of molecular tools and resources have been historically limited, prohibiting significant progress toward understanding this important host-pathogen interaction. The development of a nested qPCR assay capable of quantifying P. cinnamomi during avocado infection has enabled us to distinguish avocado rootstocks as either resistant or tolerant - an important distinction when unraveling the defense response. This review will provide an overview of our current knowledge on the molecular defense pathways utilized in resistant avocado rootstock against P. cinnamomi. Notably, avocado demonstrates a biphasic phytohormone profile in response to P. cinnamomi infection which allows for the timely expression of pathogenesis-related genes via the NPR1 defense response pathway. Cell wall modification via callose deposition and lignification have also been implicated in the resistant response. Recent advances such as composite plant transformation, single nucleotide polymorphism (SNP) analyses as well as genomics and transcriptomics will complement existing molecular, histological, and biochemical assay studies and further elucidate avocado defense mechanisms.The Hans Merensky Foundation as well as the National Research Foundation.http://www.frontiersin.org/Plant_Scienceam2022BiochemistryForestry and Agricultural Biotechnology Institute (FABI)GeneticsMicrobiology and Plant Patholog
Generation of composite Persea americana (Mill.) (avocado) plants : a proof-of-concept study
Phytophthora Root Rot in areas where the pathogen is prevalent. However, advances in
molecular research are hindered by the lack of a high-throughput transient transformation
system in this non-model plant. In this study, a proof-of-concept is demonstrated by the successful
application of Agrobacterium rhizogenes-mediated plant transformation to produce
composite avocado plants. Two ex vitro strategies were assessed on two avocado genotypes
(Itzamna and A0.74): In the first approach, 8-week-old etiolated seedlings were
scarred with a sterile hacksaw blade at the base of the shoot, and in the second, inch-long
incisions were made at the base of the shoot (20-week-old non-etiolated plants) with a sterile
blade to remove the cortical tissue. The scarred/wounded shoot surfaces were treated
with A. rhizogenes strains (K599 or ARqua1) transformed with or without binary plant transformation
vectors pRedRootII (DsRed1 marker), pBYR2e1-GFP (GFP- green fluorescence
protein marker) or pBINUbiGUSint (GUS- beta-glucuronidase marker) with and without rooting
hormone (Dip 'N' Grow) application. The treated shoot regions were air-layered with sterile
moist cocopeat to induce root formation. Results showed that hormone application
significantly increased root induction, while Agrobacterium-only treatments resulted in very
few roots. Combination treatments of hormone+Agrobacterium (-/+ plasmids) showed no
significant difference. Only the ARqua1(+plasmid):A0.74 combination resulted in root transformants,
with hormone+ARqua1(+pBINUbiGUSint) being the most effective treatment with
~17 and 25% composite plants resulting from strategy-1 and strategy-2, respectively. GUS and
GFP-expressing roots accounted for less than 4 and ~11%, respectively, of the total roots/treatment/avocado genotype. The average number of transgenic roots on the composite
plants was less than one per plant in all treatments. PCR and Southern analysis further
confirmed the transgenic nature of the roots expressing the screenable marker genes.
Transgenic roots showed hyper-branching compared to the wild-type roots but this had no
impact on Phytophthora cinnamomi infection. There was no difference in pathogen load 7-
days-post inoculation between transformed and control roots. Strategy-2 involving A0.74:
ARqua1 combination was the best ex vitro approach in producing composite avocado
plants. The approach followed in this proof-of-concept study needs further optimisation involving multiple avocado genotypes and A. rhizogenes strains to achieve enhanced root
transformation efficiencies, which would then serve as an effective high-throughput tool in
the functional screening of host and pathogen genes to improve our understanding of the
avocado-P. cinnamomi interaction.S1 Fig. Representative image showing the composite plant generation attempted in avocado
according to the ex vitro protocol described by [21]. (A) Root induction observed when
in vitro regenerated shoots from avocado zygotic embryos used as explant. (B) Tumor-like
growth with no root induction observed with young apical shoot cuttings as explants.A grant from the Hans Merensky Foundation (HMF), South Africa. SAP was supported financially by University of Pretoria post-doctoral fellowship programme and HMF. BN received student bursaries from National Research Foundation (NRF), South Africa and HMF. JE was fully supported by HMF.http://www.plosone.orgam2017Forestry and Agricultural Biotechnology Institute (FABI)Microbiology and Plant Patholog
Immuno-affinity purification of PglPGIP1, a polygalacturonase-inhibitor protein from pearl millet: studies on its inhibition of fungal polygalacturonases and role in resistance against the downy mildew pathogen.
Molecular cloning of a coiled-coil-nucleotide-binding-site-leucine-rich repeat gene from pearl millet and its expression pattern in response to the downy mildew pathogen
Downy mildew caused by Sclerospora graminicola is a devastating disease of pearl millet. Based on candidate gene approach, a set of 22 resistance gene analogues were identified. The clone RGPM 301 (AY117410) containing a partial sequence shared 83 % similarity to rice R-proteins. A full-length R-gene RGA RGPM 301 of 3552 bp with 2979 bp open reading frame encoding 992 amino acids was isolated by the degenerate primers and rapid amplification of cDNA ends polymerase chain reaction (RACE-PCR) approach. It had a molecular mass of 113.96 kDa and isoelectric point (pI) of 8.71. The sequence alignment and phylogenetic analysis grouped it to a non-TIR NBS LRR group. The quantitative real-time PCR (qRT-PCR) analysis revealed higher accumulation of the transcripts following inoculation with S. graminicola in the resistant cultivar (IP18296) compared to susceptible cultivar (7042S). Further, significant induction in the transcript levels were observed when treated with abiotic elicitor β-aminobutyric acid (BABA) and biotic elicitor Pseudomonas fluorescens. Exogenous application of phytohormones jasmonic acid or salicylic acid also up-regulated the expression levels of RGA RGPM 301. The treatment of cultivar IP18296 with mitogen-activated protein kinase (MPK) inhibitors (PD98059 and U0126) suppressed the levels of RGA RGPM 301. A 3.5 kb RGA RGPM 301 which is a non-TIR NBS-LRR protein was isolated from pearl millet and its up-regulation during downy mildew interaction was demonstrated by qRT-PCR. These studies indicate a role for this RGA in pearl millet downy mildew interaction