Pathogens within the oomycete genus Phytophthora are among some of the most destructive plant pathogens globally, causing disease and significant losses in important agricultural and forestry crops, damaging the environment, as well as impeding attempts to mitigate climate change [1,2,3,4]. One of the more well-known incidences of Phytophthora disease is the Irish Potato Famine in 1845. This incident was caused by Phytophthora infestans—the causal agent of late blight of potatoes. The disease resulted in the death of half of the potato crop that year and about three-quarters of the crop over the next seven years [3,5]. Other Phytophthora spp., which cause significant impact worldwide, include the causal agents of sudden oak death in California (Phytophthora ramorum), stem rot of soybean (Phytophthora sojae), black shank of tobacco Phytophthora nicotianae), phytophthora root rot of avocado and jarrah dieback of trees in the Jarrah Forest, both caused by Phytophthora cinnamomi [3,6,7]. Despite the economic and ecological relevance of P. cinnamomi, the mechanisms this pathogen utilizes to infect and successfully colonize its hosts are still largely unknown. P. cinnamomi is known to infect plants that are important for agriculture and forestry, with the most significant food losses occurring in avocados. There is little to no knowledge on how P. cinnamomi, a hemi-biotrophic pathogen, maintains a biotrophic lifestyle early in the infection and a necrotrophic lifestyle later in the infection.

Pathogenic lifestyles are centered around feeding on host tissue, where success is dependent on the pathogen’s ability to overcome host defenses. One host defense strategy Phytophthora spp. must evade to sustain their biotrophic phase is the hypersensitive response (HR). The HR is a form of programed cell death (PCD) and is generally the last resort in a host plant’s defense response against a pathogen. The response involves the localized death of cells surrounding the initial site of infection to inhibit the spread of the pathogen. Later in the infection, the HR is favored during the necrotrophic phase. Studies have shown that Phytophthora spp. manipulate the host plant’s cell death machinery to elicit a susceptible outcome [8,9].

Phytophthora spp. harbor a distinct set of genes involved in moderating host–pathogen interactions [10]. These genes encode effectors—small, secreted proteins—that interfere with host defense processes. There are two groups of effectors, cytoplasmic and apoplastic effectors, which are classified by where in the host cell they act. The most well-studied classes of Phytophthora cytoplasmic effectors are Crinklers (CRNs) and RxLRs (Arg-x-Leu-Arg, where x is any amino acid) [4]. Research into Phytophthora effectors has greatly expanded due to the availability of genomic and transcriptomic data, allowing for the prediction of putative effector homologs in Phytophthora spp. [11]. These valuable tools—followed by functional characterization techniques, such as transient transformation in model plants—allow for the identification of effectors that may play crucial roles during infection. However, little genomic research has been conducted on P. cinnamomi, which leaves a gap in knowledge on the mechanisms employed by this pathogen to successfully infect and cause disease in economically and ecologically important plants.

Due to their economic impact, Phytophthora spp. are some of the most studied among oomycetes [4,12]. There is, however, still limited knowledge on the mechanisms utilized to regulate cell death in host plants. It is likely these processes are determined by the delivery of functionally distinct pathogen effectors into the host cell [13].

Plants are immobile organisms and have had to develop morphological, biochemical and physiological adaptations to survive in their environment. PCD is an important mechanism for plant development or defense and can be triggered by both abiotic and biotic stressors [9,14,15]. PCD is described as a genetically controlled process where selected cells are eliminated through a coordinated multi-step fashion [15]. This phenomenon is of considerable importance in agriculture because PCD can significantly affect plant health and subsequent yield [16,17]. Therefore, it is important to understand both the triggers and the pathways through which PCD is elucidated.

There has been some confusion regarding how different forms of PCD should be classified and how terminology should be standardized. Cell death is classified based on the morphological characteristics and, as a result, two major classes of PCD are proposed to occur in plant biology [14]. Class one is vacuolar cell death, which involves the engulfment of the cytoplasm by lytic vacuoles, uptake and degradation of portions of the cytoplasm in the vacuolar lumen and, finally, rupture of the tonoplast followed by a massive release of vacuolar hydrolases. This results in the rapid destruction of the entire protoplast—a cell whose cell wall has been removed by enzymes—or, in some cases, even the entire cell, including the cell wall (Figure 1A). Class two is necrotic cell death, which is distinguished from vacuolar cell death by mitochondrial swelling, absence of the growing lytic vacuoles and early rupture of the plasma membrane, resulting in shrinkage of the protoplast (Figure 1B). Necrosis is regarded as an acute death response, which develops rapidly, taking anywhere from several minutes to a day to complete [14]. The use of morphology to classify PCD has allowed a better understanding of how cell death manifests. Although one limitation in this is that a well-known form of PCD, known as the HR, cannot be ascribed to either class, as its development displays characteristics of both vacuolar and necrotic cell death [14,18,19,20].

The HR is a special form of PCD, involving rapid localized cell death at the point of pathogen penetration [16,23]. The host plant utilizes HR to limit biotrophic pathogen growth and generates long-range signals for systemic acquired resistance (SAR) [24]. Thus, another PCD classification system was developed to accommodate the placement of the HR. This system classifies forms of PCD based on what functions they play in the host plant, rather than by their morphology or pathways. Two classes were described: developmentally controlled PCD (dPCD) and pathogen-triggered PCD (pPCD). During vegetative and reproductive development, dPCD occurs and is often a final differentiation step for specific cell types [25]. Conversely, pPCD is elicited in the host plant by invading agents and can benefit either the plant or pathogen, depending on the studied plant–pathogen interaction [26]. An additional class has also been proposed to describe PCD resulting from environmental stress, termed ePCD [25]. The ePCD classification includes stresses, such as temperature or irradiation, or biotic aggressors, such as pathogens [15]. pPCD is specific to pathogen-triggered cell death, whereas ePCD includes all external stressors as PCD triggers. The use of ePCD as a classification may, however, be problematic, since different PCD pathways may be in play during both abiotic and biotic-triggered PCD.

Plant PCD pathways are not as well understood as animal cell death. Animals have a core PCD machinery that is mainly regulated post-translationally [17,27], whereas it is not known whether the different forms of plant PCD share the same core machinery or whether the similarities they share were independently adopted to fulfill analogous roles for different pathways [9]. When looking at the two main plant PCD forms—dPCD and pPCD—there are marked differences as well as commonalities in their proposed pathways. For one, a vacuolar type of cell death is associated with dPCD, and features of both necrosis and vacuolar PCD are seen in pPCD [9]. There is also evidence of transcriptional regulation and signaling in both forms of plant PCD, but in different contexts. Unfortunately, there are still gaps in our knowledge regarding pPCD. This is largely due to predominance of previous dPCD-centered investigations. In addition, variability has been seen in pPCD responses to a multitude of different abiotic and biotic factors, whereas dPCD is a relatively conserved process across all plant species. This section serves to summarize our current knowledge on the transcriptional regulation, hormonal signaling and triggers involved in pPCD.

The stimulation and repression of cell death pathways by transcription regulators has been seen in animal PCD [28,29], and recent evidence indicates that some level of transcriptional control of PCD is also likely in plants [30,31,32,33,34]. Different classes of transcription factors (TFs), including members of NAC, ethylene-responsive element-binding factors (ERFs) and WRKY families, have been shown to play roles in cell fate regulation in response to different stresses. NAC TFs have been linked to the regulation of PCD triggered by both abiotic and biotic stresses [34]. One example is that of OsNAC4, which has been shown to be a key positive regulator of the HR by modulating the expression of almost 150 genes in rice, such as Copper Zinc Superoxide Dismutase 1 (CSD1) gene and BAX Inhibitor 1(BI-1) gene [35]. ERF TFs also play a role in the regulation of the HR, where the conditional expression of NbCD1—from Nicotiana benthamiana—in response to multiple HR elicitors is sufficient to induce the HR [30]. Numerous WRKY TFs are involved in the regulation of cell death, and they may play a role in the suppression of the HR during initial infection of the necrotrophic fungus, Botrytis cinerea, in Arabidopsis [36], through the activation or suppression of antagonistic signaling pathways, such as salicylic acid (SA), ethylene (ET) and jasmonic acid (JA) mediated pathways. Although there is a large body of research on TFs and their role in pPCD, there is still a lack of knowledge on Phytophthora pathogens and the involvement of TFs in eliciting or suppressing PCD during Phytophthora infection.

Different phytohormones play a role in dPCD, such as JA, auxin, strigolactones and ET—ET being the most characterized dPCD hormone [37,38,39]. Phytohormones control the dPCD processes via transcriptional regulation of genes, such as proteases and nucleases, to gradually build up dPCD competence during cellular differentiation. This contrasts with pPCD, where no preparation is required, and the cells are always ready to initiate an immune response upon pathogen attack [9]. The infection strategy of a plant pathogen—whether the pathogen adopts a biotrophic, necrotrophic or hemi-biotrophic lifestyle—determines the underlying mechanism for phytohormone-regulated pPCD in the host during plant–pathogen interactions [40,41]. It has been shown that SA plays an essential role in host defense response against biotrophic and the early stages of hemi-biotrophic pathogens, whereas JA and ET play an important role in the host defense response against necrotrophic and the later stages of hemi-biotrophic pathogens [41]. SA is the only phytohormone shown to play an essential role in the establishment of pPCD, allowing immunity toward biotrophic pathogens and susceptibility to necrotrophic pathogens. [42,43]. It has been found that some pathogens interfere with cellular SA biosynthesis or signaling through the delivery of effector proteins. [26]. For example, penetration-specific effector 1 (PSE1) from Phytophthora parasitica inhibits SA-mediated cell death and increased pathogen growth by promoting auxin accumulation at infection sites [44]. Due to the importance of phytohormones in the different trophic interactions, it would be of value to investigate their roles in the maintenance and switch from the biotrophic to necrotrophic stage in hemi-biotrophic pathogens, such as Phytophthora. This will shed light on how Phytophthora is able to successfully infect a host plant and avoid the hosts’ defense responses.

dPCD requires preparation before PCD can be triggered/executed. Several cytoplasmic signals are implicated in dPCD triggering, such as calcium fluxes, accumulation of reactive oxygen species (ROS) and cytoplasmic acidification [45]. During the self-incompatibility (SI) response—the inability of a plant with functional pollen to set seeds when self-pollinated—in Papaver rhoeas, calcium influx triggers a signaling cascade, which results in rapid PCD of the incompatible pollen tubes [46]. In contrast, pPCD requires no preparation and is only triggered upon pathogen attack. The main pPCD trigger is cytoplasmic immune receptor-mediated recognition at the site of attack [47]. Calcium influxes, as well as accumulation of SA, ROS and nitric oxides (NO), are triggered upon pathogen perception during pPCD. SA signaling subsequently amplifies the ROS burst in a positive feedback loop, creating a toxic environment [48]. Some necrotrophic pathogens have been known to ‘hijack’ PCD machinery, where pathogens, such as Cochliobolus victoriae, secrete PCD triggering toxins [49,50]. Common triggers that are recognized by host receptors are effectors. Different Phytophthora effectors and their role in host PCD will be discussed in a later section.

Different forms of pPCD will benefit either the plant or pathogen, depending on the type of plant–pathogen interaction and the trophic lifestyle of the pathogen [4,9,25]. Most Phytophthora spp. are hemi-biotrophic pathogens, meaning they feature a biotrophic life stage during early infection followed by a switch to necrotrophy during the later stages of host tissue colonization [4,51]. As the HR is generally considered most effective against biotrophic pathogens, while potentially benefiting necrotrophic pathogens, hemi-biotrophic pathogens—such as Phytophthora—are at a distinct advantage [52,53]. This response is a race between the host and pathogen, where the pathogen attempts to tip the balance toward suppression of host defense, and the host tries to launch an effective defense response to prevent infection [4].

Phytophthora spp. may have developed a strategy to ‘hijack’ a plant’s HR machinery, suppressing the HR during the biotrophic stage and inducing it during the necrotrophic stage [24,54,55]. This ‘hijack’ strategy is further supported by the production of haustoria that deliver defense-controlling pathogenicity factors and effectors, which function in keeping the host cell alive [56,57]. Conversely, the switch to necrotrophy, which involves the upregulation of specialized effector genes—such as Nep1-like proteins (NLPs)—aims to deliberately kill the host cell [58]. This is further supported by the similarities in metabolic enzyme expression between P. infestans during the necrotrophic stage and the necrotrophic pathogen Pythium ultimum [59]. This strategy increases the virulence of Phytophthora spp. through the differential expression and delivery of effectors at different stages of host plant infection and colonization [13,54,55,60,61,62].