Elliott et al. BMC Genomics (2018) 19:320 https://doi.org/10.1186/s12864-018-4709-7 RESEARCHARTICLE Open Access Characterization of phenotypic variation and genome aberrations observed among Phytophthora ramorum isolates from diverse hosts Marianne Elliott1, Jennifer Yuzon2, Mathu Malar C3, Sucheta Tripathy3, Mai Bui4, Gary A. Chastagner1, Katie Coats1, David M. Rizzo2, Matteo Garbelotto5 and Takao Kasuga4* Abstract Background: Accumulating evidence suggests that genome plasticity allows filamentous plant pathogens to adapt to changing environments. Recently, the generalist plant pathogen Phytophthora ramorum has been documented to undergo irreversible phenotypic alterations accompanied by chromosomal aberrations when infecting trunks of mature oak trees (genus Quercus). In contrast, genomes and phenotypes of the pathogen derived from the foliage of California bay (Umbellularia californica) are usually stable. We define this phenomenon as host-induced phenotypic diversification (HIPD). P. ramorum also causes a severe foliar blight in some ornamental plants such as Rhododendron spp. and Viburnum spp., and isolates from these hosts occasionally show phenotypes resembling those from oak trunks that carry chromosomal aberrations. The aim of this study was to investigate variations in phenotypes and genomes of P. ramorum isolates from non-oak hosts and substrates to determine whether HIPD changes may be equivalent to those among isolates from oaks. Results: We analyzed genomes of diverse non-oak isolates including those taken from foliage of Rhododendron and other ornamental plants, as well as from natural host species, soil, and water. Isolates recovered from artificially inoculated oak logs were also examined. We identified diverse chromosomal aberrations including copy neutral loss of heterozygosity (cnLOH) and aneuploidy in isolates from non-oak hosts. Most identified aberrations in non-oak hosts were also common among oak isolates; however, trisomy, a frequent type of chromosomal aberration in oak isolates was not observed in isolates from Rhododendron. Conclusion: This work cross-examined phenotypic variation and chromosomal aberrations in P. ramorum isolates from oak and non-oak hosts and substrates. The results suggest that HIPD comparable to that occurring in oak hosts occurs in non- oak environments such as in Rhododendron leaves. Rhododendron leaves are more easily available than mature oak stems and thus can potentially serve as a model host for the investigation of HIPD, the newly described plant-pathogen interaction. Keywords: Invasive pathogens, Aneuploidy, Transposable elements, Loss of heterozygosity * Correspondence: [email protected] 4Crops Pathology and Genetics Research Unit, USDA Agricultural Research Service, Davis, California 95616, USA Full list of author information is available at the end of the article © The Author(s). 2018 Open Access This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated. Elliott et al. BMC Genomics (2018) 19:320 Page 2 of 19 Background EU1, the dominant clonal lineage in Europe. A dead-end The underlying genetic mechanisms of how invasive path- host is one in which the pathogen does not produce the in- ogens adapt to new hosts during co-evolutionary interac- fectious stage [13]. Both oaks and Lawson cypress are con- tions is not well understood. In some systems, initial sidered to be dead-end hosts; therefore, the epidemiological phenotypic plasticity or depauperate interactions occur significance of nwt phenotype is not clear [12]. followed by adaptation to new environments [1–3]. It is worth noting that Sudden Oak Death is actually Phytophthora ramorum is a recent example of an inva- not “sudden” at all. In some cases, individual oak trees sive pathogen attacking more than 130 host plant spe- infected with P. ramorum do not die rapidly. Some live cies across 75 genera [4]. It causes Sudden Oak Death for many years, and in others, the host can recover from (SOD) or Sudden Larch Death (SLD) [5] in forests, and the disease [14]. Such oak survival may be associated Ramorum blight on most of its other hosts including or- with the conversion of P. ramorum to slow growing nwt, namentals in nurseries. As of now, four clonal lineages allowing the host to contain the pathogen, or the patho- of P. ramorum have been described, and all are believed gen may become senescent and therefore may be effect- to be exotic to Europe and North America. Epidemio- ively eradicated [12]. Colony instability, slower growth logical and regulatory consequences of the disease have rates, and lower aggressiveness of nwt phenotypes can caused millions of dollars in losses to ornamental nur- be viewed as an undescribed type of host-pathogen series, larch plantations, and native vegetation [6, 7]. interaction, and generation of the nwt phenotype may The NA1 clonal lineage of P. ramorum is predominant in be a consequence of an active plant defense. Alterna- natural ecosystems and production nurseries in the Pacific tively, genome rearrangements such as formation of Coast region of North America. Despite its primary mode aneuploids and loss of heterozygosity, and the associ- of asexual reproduction and the lack of sexual recombin- ated phenotypic changes can be viewed as a part of ation [8], the pathogen shows diverse colony morphology an evolutionary process in which genome diversifica- and aggressiveness. In culture, for instance, wild type (wt) tion contributes to the adaptation of the invasive P. ramorum grows as a uniform, roughly circular colony pathogen to various new and unfamiliar environmen- mostly appressed to the culture media [9]. However, some tal situations [15]. In addition, de-repression of TEs isolates exhibit a range of phenotypes such as irregular col- observed among nwt isolates [11, 12] has evolutionary ony shape, and unstable or slower growth rate, which we implications as stress-induced changes in TE activity refertoasnon-wildtype(nwt). Additionally, some nwt iso- are claimed to promote structural variation in the lates become senescent, i.e. they cease to grow upon sub- genome and facilitate rapid adaptation in invasive culturing, whereas other show greater aerial growth. species [16]. In either case, the study of HIPD will Inoculation experiments have shown that isolates with nwt provide us insights into host-pathogen interaction in morphology are less aggressive than wt isolates [9–11]. which coevolutionary history is not present, and fur- Among isolates from natural forests, nwt morphology is as- ther study on HIPD may lead us to the development sociated with isolates from oak species (genus Quercus), of novel strategies to manage the pathogen. and nwt isolates are rare among California bay (Umbellu- Inoculation of mature oak trees to unravel the genetic laria californica). California bay is an epidemiologically im- mechanisms underlying host-induced chromosomal alter- portant foliar host, due to the abundant sporulation of the ations is not practical for many reasons, including avail- pathogen it supports on its leaves. ability of specimens, strict regulations of field experiments We have previously demonstrated that the nwt pheno- involving infectious agents as well as heterogeneity of bi- type can experimentally be generated [12]. When wt P. otic and abiotic environments among specimens. How- ramorum isolates from California bay were inoculated into ever, the nwt phenotype has occasionally also been and re-isolated from mature canyon live oak (Q. chrysolep- observed in P. ramorum derived from infectious hosts sis)andShreveoak(Q. parvula var. shrevei), a large per- such as Rhododendron spp. in production nurseries [10]. centage (40–60%) of re-isolates were found to display nwt Due to its size and availability, use of Rhododendron for phenotype and chromosomal alterations such as mono- inoculation studies has a significant advantage over oak. It somy, trisomy, and copy number neutral loss of heterozy- is not yet clear whether nwt phenotypes observed in iso- gosity (cnLOH), which were accompanied by de-repression lates from Rhododendron and other non-oak hosts and of diverse transposable elements (TEs). Because the wt/nwt those from oaks are driven by the same genetic mechan- variation does not associate with population genetic ism. The objective of this research was to characterize markers, and can be induced by inoculation onto Quercus, phenotypic and genomic variations among P. ramorum the concept of host-induced phenotypic diversification isolates from the foliage of ornamental plants and other (HIPD) was formulated [11, 12]. Nwt phenotype has also substrates and compare them to those found in oak iso- been observed among isolates derived from Lawson cy- lates. Taking advantage of a significantly improved version press (Chamaecyparis lawsoniana), which belonged to of P. ramorum genome recently assembled using PacBio Elliott et al. BMC Genomics (2018) 19:320 Page 3 of 19 sequencing, we analyzed genome alterations at high reso- isolates were found in Cluster 1 and Cluster