Plant Pathology (2019) 68, 1427–1438 Doi: 10.1111/ppa.

13077

REVIEW

A review of the known unknowns in the early stages of

septoria tritici blotch disease of wheat

C. J. Brennana , H. R. Benbowa, E. Mullinsb and F. M. Doohana*

a
UCD School of Biology and Environmental Science, UCD Earth Institute and UCD Institute of Food and Health, College of Science,
University College Dublin, Belfield, Dublin 4; and bDepartment of Crop Science, Teagasc, Oak Park, Carlow, Ireland

Septoria tritici blotch (STB) disease of wheat is caused by the fungal pathogen Zymoseptoria tritici. It is the most
important foliar disease of wheat in western Europe and affects wheat cultivation worldwide. The combination of
intensive fungicide usage, a polycyclic asexual life cycle and an active sexual cycle has led to the emergence of fungal
strains resistant/tolerant to all the major classes of fungicides used in its control. The hallmark of this disease is a long,
symptomless latent phase that precedes the onset of visible symptoms. Understanding the processes that occur during
the symptomless phase of infection is paramount in developing alternative strategies for disease control; however, large
gaps in our knowledge of the disease remain. The known unknowns of the latent stage of infection can be summarized
in three questions. Does the fungus initiate or manipulate host defences to trigger programmed cell death in order to
facilitate nutrient acquisition or is the host acting exclusively? Does the fungus feed during both the latent phase and
the necrotrophic phase like a true hemibiotroph? Does the long latent phase serve a beneficial function for the fungus
or is it simply an artefact of evolution? This review aims to distil observations made during studies that have directly
or indirectly contributed to answering these questions and points towards their most likely answers.

Keywords: host defence, latent phase, septoria tritici blotch, trophic, wheat, Zymoseptoria tritici

2003; Hayes et al., 2015; Dooley et al., 2016) and rapid

Septoria Tritici Blotch
Septoria tritici blotch (STB) is a foliar disease of wheat
caused by the haploid, pathogenic fungus Zymoseptoria
tritici. STB is a polycyclic disease (Fones & Gurr, 2015)
that affects wheat crops worldwide and is the primary
foliar disease of winter wheat in most countries within
western Europe. Yield losses of up to 20% (Fones &
Gurr, 2015) caused by a loss in photosynthetic leaf area
threaten the profitability of wheat; however, yield losses
due to STB can be reduced to 5–10% when integrated
pest management schemes are implemented (Fones &
Gurr, 2015). Heterogeneous host populations in agricul-
tural systems and high fungicide usage (c. 70% of EU
fungicide expenditure is targeted towards controlling
STB; (Torriani et al., 2015)) pose strong selection on
pathogen populations (McDonald & Stukenbrock,
2016). Coupled with frequent sexual recombination in
Z. tritici (Banke et al., 2004; Welch et al., 2018), this
has catered for the rapid evolution of fungal virulence,
resistance to the major classes of fungicides (Lucas,
*E-mail: fiona.doohan@ucd.ie
Published online 18 August 2019
breakdown of host resistance (Cowger et al., 2000;
Stukenbrock & McDonald, 2008; McDonald & Stuken-
brock, 2016; Haueisen et al., 2017). Ongoing research
aimed at controlling STB disease involves a multipronged
approach for understanding host–pathogen gene interac-
tions, resistance breeding and the development of new
fungicide mixtures (Adhikari et al., 2004a,b; Brown
et al., 2015; Heick et al., 2017; Kema et al., 2018; Sain-
tenac et al., 2018).
The Latent Phase of the Disease
The life cycle of Z. tritici is broadly divided into two dis-
tinct stages, namely the symptomless latent phase and
the necrotrophic stage (Fig. 1). The latent phase can be
further subdivided into three stages: transition, ingress
and colonization. The transition stage from the leaf sur-
face to the substomatal cavity is usually completed
between 1 and 3 days post-inoculation (Duncan &
Howard, 2000); however, it has been observed occurring
as early as 12 h after spore germination (Kema et al.,
1996) or much later, following more than 10 days of
epiphytic growth on the leaf surface (Fones et al., 2017;
Table 1).
Ingress takes place through the stomata of the host;
however, attempts at ingress via anticlinal cell walls have

ª 2019 British Society for Plant Pathology 1427

1428 C. J. Brennan et al.

Figure 1 Processes that occur during infection of wheat by septoria tritici blotch (STB). From left to right, the figure depicts the processes known to
occur in the fungus Zymoseptoria tritici (during infection of a susceptible variety), in the susceptible host (during a compatible interaction) and in a
resistant host (during an incompatible interaction). Recognition of the fungal pathogen during the incompatible interaction at 0–1 days post-
inoculation (dpi) allows for the host to initiate a defence response that arrests fungal growth and development; this recognition is absent from the
compatible interaction. Abbreviations: JA, jasmonic acid; SA, salicylic acid; H2O2, hydrogen peroxide; PTI, pattern-triggered immunity; PR genes,
pathogen-responsive genes; PCD, programmed cell death; ETD, effector-triggered defence. [Colour figure can be viewed at wileyonlinelibrary.com].

Table 1 Observed time of stomatal penetration and the associated experimental conditions for six studies on the Zymoseptoria tritici–wheat
interaction.

Spore Resistant Susceptible Relative Stomatal

Z. tritici production Spore wheat wheat Light/dark humidity Temperature penetration
Study isolate medium concentration cultivar cultivar cycle (h) light/dark (%) light/dark (°C) observed
7
Kema et al. IPO87016 PDA 10 Kavkaz/K4500 Shafir 16/8 70/85 22/21 12 HPI
(1996) 1.6.a.4
Duncan & Howard Field isolate V8 107 — Riband 14/10 100 (48 h); 90/90 20/16 1 DPI
(2000)
Rohel et al. T16 PDA 107 — Riband 16/8 100/100 18/18 1 DPI
(2001)
Shetty et al. (2003) IPO323 PDA 106 Stakado Sevin 16/8 100 (72 h); 19/16 5 DPI
50–60/80–90
Shetty et al. (2009) IPO323 PDA 106 Stakado Sevin 16/8 100 (72 h); 19/16 5 DPI
50–60/80–90
Fones et al. (2017) IPO323 YPDA 105–107 — Galaxie 12/12 80/80 20/20 10 DPIab

PDA, potato-dextrose agar; V8, V8 juice agar; YPDA, yeast extract-peptone-dextrose agar; HPI, hours post-inoculation; DPI, days post-inoculation.
a
Can be detected earlier if sought.

also been observed (Duncan & Howard, 2000; Shetty whether the host is resistant or susceptible to Z. tritici
et al., 2003). Although transition and ingression occur at infection and whether or not the Z. tritici isolate is
different rates between resistant and susceptible wheat pathogenic (Cohen & Eyal, 1993; Shetty et al., 2007;
cultivars (Kema et al., 1996), they occur irrespective of Siah et al., 2010). Stomatal ingress avoids the need for

Plant Pathology (2019) 68, 1427–1438

 The known unknowns
physical penetration of the host tissue via specialized
structures such as an appressorium (Cohen & Eyal,
1993) and may help the pathogen avoid the host plant
defences that would be elicited by direct penetration.
Indeed, several key genes required for appressorium for-
mation in other fungal pathogens are not encoded within
the Z. tritici genome (Goodwin et al., 2011).
After ingress, colonization occurs via intercellular
growth of the fungus from the substomatal cavity into
the apoplastic spaces of the host mesophyll cells (Kema
et al., 1996; Siah et al., 2010). This colonization is the
final stage of the latent phase but the duration varies
depending on the host, isolate and environmental condi-
tions (Hehir et al., 2018).
The Necrotrophic Phase
Zymoseptoria tritici does not form dedicated feeding
structures such as haustoria (Kema et al., 1996) but
instead feeds on the efflux of nutrients released by host
cells upon a loss of their physical integrity. During a
compatible infection, this typically starts around 11–
14 days post-infection, when necrotic symptoms
develop and fungal biomass increases as the fungus
starts its necrotrophic nutrient acquisition; this can be
facilitated by host- or pathogen-driven cell death (Keon
et al., 2007; Reape et al., 2008). In plants, apoptosis-
like programmed cell death (AL-PCD; Danon et al.,
2000) is characterized by cytoplasmic withdrawal from
the cell wall followed by the cell breaking apart into
apoptotic bodies (Dickman et al., 2017). Similar to
AL-PCD, plants can initiate a hypersensitive response
(HR) when under attack by biotrophic pathogens to
restrict the local growth of the pathogen. During infec-
tion by Z. tritici, typical symptoms appear as small
necrotic regions that house the fungus, surrounded by
chlorotic areas undergoing an HR-like response (Fig. 2;
Keon et al., 2007). Within these lesions, reproductive
structures are established in substomatal cavities (Dun-
can & Howard, 2000). Pycnidiospores, the asexual
spores of the fungus, are produced in small, black
structures called pycnidia. Due to their ability to travel
by splash dispersal, pycnidiospores are responsible for
the spread of disease up through the leaf layers of the
host. In contrast, the sexual ascospores of Z. tritici are
responsible for primary infection and are produced in
asci (eight spores per ascus), housed in structures
called pseudothecia. These are ejected forcibly under
high humidity for airborne dispersal (Shaw & Royle,
1989) from residual debris left in the field post-har-
vest.
An Alternative Infection Model
Despite such a seemingly well-categorized model for
infection, recent work carried out by Fones et al. (2017)
has described prolonged fungal epiphytic growth, with
penetration occurring from 10 days post-inoculation.
While germination of fungal spores was observed to
Plant Pathology (2019) 68, 1427–1438
1429
occur just hours after contacting the leaf surface, this
study highlights the possibility that the duration to com-
pletion of the transition stage may take far longer than
previously suggested by Kema et al. (1996). Differences
between studies in terms of laboratory/glasshouse/field
conditions and in the isolates used are likely to influence
the duration of the transition stage. Alongside variability
in disease progression caused by changes in experimental
conditions, Z. tritici isolates themselves exhibit a spec-
trum of host interactions and infection phenotypes. Some
of this variability is genetic; between regionally diverse
isolates, up to 20% of fungal genes may be differentially
expressed (Haueisen et al., 2017). To resolve issues sur-
rounding the timing of stomatal ingression and potential
differences in the field versus glasshouse, comparative
experiments using a range of host varieties (both compat-
ible and incompatible) and a range of strains are
required and are certainly merited as a result of the find-
ings of Fones et al. (2017).
The Known Unknowns
Despite being such a globally important disease of
wheat, there are still large gaps in our understanding of
STB disease. These known unknowns can be summa-
rized in three questions (Fig. 3): Does the long latent
phase serve a beneficial function for the fungus or is it
simply an artefact of evolution? Does the fungus feed
during both the latent phase and the necrotrophic phase
like a true hemibiotroph? Does the fungus initiate or
manipulate host defences to trigger host cell death for
nutrient acquisition, or are defences wholly host-con-
trolled?
Question 1: Does the long latent phase serve a
beneficial function for the fungus or is it simply an
artefact of evolution?
Origination and specialization
While necrotrophic feeding is essential for the comple-
tion of the Z. tritici life cycle, the lack of haustorial
development for nutrient acquisition seems to point away
from the hallmarks of a biotrophic pathogen and
towards that of an endophyte (Goodwin et al., 2011;
S
anchez-Vallet et al., 2015). An absence of dedicated
feeding structures, a non-injurious method of entry, non-
invasive occupation of the apoplast during the latent
phase, a reduced suite of genes encoding putative cell
wall-degrading enzymes (CWDEs) and the partial com-
pletion of the life cycle within a host are all attributes of
Z. tritici. These traits, coupled with the scarcity of diver-
sity observed at the genetic level between fungal endo-
phytes and pathogens (Zuccaro et al., 2011) seem to
suggest possible aberrant endophyte origins for Z. tritici
and that the latent phase of infection serves only as an
artefact of this evolution. If this is truly the case, then
does the latent phase serve any beneficial function for
the pathogen? While a latent phase is by no means
unique to Z. tritici (Shearer & Zadoks, 1972; Parlevliet,
1430 C. J. Brennan et al.

Figure 2 Necrotrophic stage of Zymoseptoria tritici infection of wheat. (a) Collapse of host tissue coinciding with a fungal switch to necrotrophy.
These symptoms typically occur 14 days post-infection. (b) Brown necrotic tissue houses the black asexual fruiting bodies (pycnidia) surrounded by
yellow chlorotic regions undergoing the hypersensitive response, usually occurring 21 days post-infection. [Colour figure can be viewed at
wileyonlinelibrary.com].

Figure 3 Schematic representation of Zymoseptoria tritici infection of susceptible wheat. Hyphal penetration through the stoma to the substomatal
cavity and surrounding mesophyll allows the fungus to manipulate the host from a strictly extracellular position. Questions relating to the function of the
latent phase, early nutrient acquisition and host defence manipulation remain unresolved. [Colour figure can be viewed at wileyonlinelibrary.com].

1975), its utility to the fungus is quite perplexing, partic-
ularly if, as discussed later, the fungus is most probably
not feeding during the symptomless stage.
Based on mutation rate estimates and comparisons to
wild relative populations still capable of infecting multi-
ple grass hosts, it is estimated that Z. tritici’s specializa-
tion for wheat began approximately 10 500 years ago
(Stukenbrock et al., 2006), during the early domestica-
tion of wheat (Stukenbrock et al., 2010). Since its ori-
gins, Z. tritici has retained its ability to grow
independently from its host, a trait absent in obligate
biotrophs. This is no surprise, as obligate biotrophy is
something of an evolutionary one-way street (Kemen
et al., 2011) and lacks the evolutionary flexibility
afforded by a ‘hemibiotrophic’ lifestyle, due to the loss
of a free-living phase (Spanu & K€ amper, 2010). Thus, if
the origin of Z. tritici lies within the ancestry of bio-
trophic endophytes, these would probably be facultative
and not obligate biotrophs, as evidenced by Z. tritici’s
ability to thrive in in vitro conditions.

Plant Pathology (2019) 68, 1427–1438

 The known unknowns
Specialization of microorganisms that live within hosts
can lead to both beneficial and pathogenic relationships.
In the most extreme scenarios, host defence suppression
becomes so effective and the host itself becomes such a
reliable source of nutrients that a free-living phase can
be lost and the relationship becomes obligatory (Kemen
et al., 2011). Facultative microorganisms can also
develop beneficial and pathogenic relationships with their
hosts; beneficial endophytes within one host can cause
pathogenic responses in nonhosts where specialization
has not occurred (Schulz et al., 2002). Interestingly, the
hemibiotrophic fungus Fusarium graminearum also
boasts a shorter, but controversially titled, biotrophic
phase in its infection of wheat (Trail, 2009). When com-
pared to Z. tritici, F. graminearum has far more CWDEs
(88 vs 30, respectively) involved in the degradation of
pectin, cellulose and hemicellulose (do Amaral et al.,
2012). Expression of these CWDEs in Z. tritici is low
during the latent and early necrotrophic phases (Yang
et al., 2013a) and this overall reduction in CWDEs is
common in organisms more closely related to obligate
biotrophs than necrotrophs. Spanu & K€amper (2010)
suggest that these CWDEs may be used more to facilitate
the growth of intercellular hyphae rather than to degrade
cells for nutrients, while simultaneously reducing the risk
of releasing cell wall-derived elicitors that may trigger
host defence responses.
Function of the latent phase
Zymoseptoria tritici has a polycyclic reproductive nature
and a (seemingly) redundant prolonged latent phase. One
could speculate that a shorter latent phase, and therefore
a more rapid switch to necrotrophy, would allow the
fungus to more rapidly complete its life cycle and boast
higher evolutionary success than that of its counterparts
encumbered with a longer latent phase. This would ulti-
mately have culminated in a contemporary necrotroph
instead of the hemibiotroph/latent necrotroph seen today.
Through the use of RNA sequencing, it has been recently
demonstrated that large-scale transcriptional changes in
both the host and the pathogen occur during the switch
to necrotrophy. One influential factor in the mass-alter-
ation of transcription is the remodelling of chromatin
structure via histone modification (Berr et al., 2011). In
the wheat–Z. tritici interaction, chromatin remodelling,
caused by the binding of the TaR1 protein to the
trimethylated lysine 4 of histone H3, may play a crucial
role in evasion of host defence; silencing of TaR1 led to
a truncated latent phase and a reduction in asexual
fecundity of the pathogen (Lee et al., 2015). The authors
propose that by commandeering the host’s transcription
of TaR1, thus suppressing the switch to necrotrophic
growth, the pathogen can reach a critical biomass. This
theory of pathogenic ‘hijacking’ has previously been pre-
sented for the wheat–Z. tritici pathosystem, wherein the
host PCD signalling response used for combating bio-
trophs is manipulated (Rudd et al., 2008). Insights into
why a shortened latent phase would be detrimental to
the fungus can be gleaned from a transcriptomic study
Plant Pathology (2019) 68, 1427–1438
1431
focused on the various stages of the Z. tritici life cycle
(Palma-Guerrero et al., 2015). As Z. tritici’s mating
genes are not currently well characterized, orthologues
from the better characterized Aspergillus spp. (Dyer &
O’Gorman, 2012) were used in the comparison and this
revealed that an abundance of transcripts attributed to
‘sexual sporulation’, ‘meiotic cell cycle processes’ and
‘developmental processes involved in reproduction’ (in
addition to defence transcripts) were expressed during
the latent phase at 3 and 7 days post-inoculation (dpi;
Palma-Guerrero et al., 2015). While the majority of
genes associated with sexual reproduction were up-regu-
lated at 14 dpi, it is interesting to note that several were
only up-regulated during the latent phase. While not
required for sporulation to occur, the latent phase is
notably absent from cultures grown in vitro and so too
are asci.
These recent studies are now revealing an alternative
role for the latent phase, with implications for reproduc-
tion, moving away from nutrient acquisition; this poten-
tially gives previously unexplored importance and
function to the enigmatic latent phase of Z. tritici. With
these insights, it would appear that the latent phase
could serve as a platform that boosts the effectiveness of
the approaching reproductive stage, justifying its contin-
ued prevalence through Darwinian success and reducing
the penalties apparently incurred by a rapid transition to
necrotrophy.
Question 2: Does the fungus feed during both the
latent phase and the necrotrophic phase like a true
hemibiotroph?
Trophic classification
The symptomless latent state is influenced by the
strain 9 host combination, temperature, humidity and
region (Eriksen & Munk, 2003; Haueisen et al., 2017).
It typically lasts for approximately 8–10 dpi; however,
there are numerous reports of shorter and longer latent
periods (Eriksen & Munk, 2003; Fones & Gurr, 2015;
Hehir et al., 2018). The final phase of the latent period
is characterized by apoplastic colonization by the fungus.
Host tissue adjacent to the fungus undergoes cell death
and is fed upon, earning Z. tritici the controversial clas-
sification of ‘hemibiotroph’. Early biotrophic feeding was
reported by Rohel et al. (2001), who used Z. tritici
transformed with a green fluorescent reporter protein
(GFP) regulated by a carbon-source-repressed promoter.
In the presence of carbohydrates, the fungus did not fluo-
resce, indicating repression of the GFP and uptake of
carbohydrates by the fungus (Rohel et al., 2001). How-
ever, unlike conventional biotrophs, there was no signifi-
cant increase in biomass of the fungus during this stage.
There may, of course, have been minute increases in bio-
mass that were not discernible by the qPCR methods
used to detect Z. tritici in planta (Shetty et al., 2007).
Hence, the term ‘latent necrotroph’ as opposed to ‘hemi-
biotroph’ was proposed as a more fitting definition for
Z. tritici (S

anchez-Vallet et al., 2015). Either the fungus
1432 C. J. Brennan et al.
is surviving on subsistence feeding during the initial
phase of infection or quite possibly, as proposed by Keon
et al. (2007), the fungus is using its own energy reserves
during the initial stages of infection and enters a state of
starvation.
Latent phase transcripts
Recent RNA sequencing papers investigating the Z. tritici–
wheat interaction seem to echo the views of Keon et al.
(2007), indicating that there is little if any feeding occur-
ring during early infection (Palma-Guerrero et al., 2015;
Rudd et al., 2015). Lipid reserves can serve as an energy
source for fungal spores and lipids have been detected in
abundance in Z. tritici spores (Rudd et al., 2015). During
the first 3 days of infection, an abundance of transcripts
was recorded, linked to lipid metabolism, alcohol metabo-
lism and chloroperoxidase synthesis. This suggests that the
fungus is catabolizing its own lipid stores (and the subse-
quent alcohols generated during this catabolism) as an
energy source and using this energy to aid in its establish-
ment and possibly in evasion of host defences. Adding cre-
dence to the starvation theory, there was only a single,
significantly expressed, secreted lipase potentially involved
in feeding on plant lipids from the apoplast or even the
waxy cuticle (Palma-Guerrero et al., 2015). The higher
expression of this single transcript associated with intercel-
lular/apoplastic feeding suggests that little, if any, fungal
feeding takes place at the very early stages of infection. At
7 dpi, abundance of transcripts for sterol desaturases, fatty
acid desaturases and alcohol dehydrogenases indicate the
fungus was still using its own lipid reserves. High levels of
fungal protease transcripts were present at this transitional
stage (7–11 dpi) and this large abundance of proteases and
unknown secreted proteins may play a role in fungal feed-
ing and nutrient acquisition. However, it was not until the
necrotrophic stage (11 dpi in Palma-Guerrero et al., 2015)
that high levels of transcripts associated with sugar and
amino acid transport were up-regulated, suggesting feeding
had commenced. Other studies also revealed that while no
significantly detectable biomass accumulation occurred dur-
ing the first 11 days of infection, fungal transcripts associ-
ated with host cell protein catabolism were detected at
10 dpi in the compatible interaction, indicating possible
intercellular fungal feeding (do Amaral et al., 2012; Yang
et al., 2013a,b). Ten days post-inoculation is decidedly late
in the latent phase and compatible interactions have
demonstrated before that the onset of the necrotrophic
phase of infection can begin at 9 dpi (as heralded by
water-soaked lesions appearing on the leaf surface and
complemented by a higher expression of b-1,3-glucan and
chitinase within the host; Keon et al., 2007; Shetty et al.,
2009). Therefore, it is possible that the fungus was transi-
tioning out of the latent phase when these transcripts were
detected.
Latent phase fungal secretions
Studies investigating fungal proteins secreted during
both the compatible (Kettles et al., 2018) and the
incompatible (M’Barek et al., 2015a) interaction have
made progress in answering the question of whether or
not Z. tritici is a hemibiotroph. At present, and based
on previously characterized proteins and current pro-
tein prediction models, it would appear that there is
little evidence that feeding is taking place at a substan-
tial level during the latent phase (Palma-Guerrero
et al., 2015). Though sparse, transcripts that could
possibly be attributed to feeding have been identified
during the latent phase in several studies. Transcripts
associated with fumarase and NAD-dependent malate
dehydrogenase were up-regulated in the studies con-
ducted by Yang et al. (2013a,b) at 4 dpi; however,
from this information alone it cannot be inferred
whether these are secreted or endogenous to the fun-
gus. In the study conducted by Rudd et al. (2015),
transcripts associated with transaldolase were detected
at 1 dpi and multiple glycoside hydrolase transcripts
from a variety of families (62, 10, 5, 43, 3, 76, 30,
35 and clan GH-D) were up-regulated at 1 and 4 dpi.
While these may serve a role in sugar catabolism, their
lack of secretory residues and the fact that they can
also be associated with pathogenesis, antibacterial
defence and normal cellular function means that they
cannot be unambiguously assigned to a dedicated cata-
bolic role in feeding. Furthermore, transcripts associ-
ated with hexose and nitrate transport, which were up-
regulated in Z. tritici during growth in a nutrient-
limited broth, were down-regulated during infection;
this indicates that if feeding is taking place, it would
probably be through alternative nutrient sources (Rudd
et al., 2015). Only a single transcript associated with a
secretory lipase was detected at 1 and 4 dpi in this
study; this result is similar to that of Palma-Guerrero
et al. (2015) at 3 dpi. It is possible that these secreted
lipases may be involved in breaking down lipids as a
source of sustenance or possibly play a role during
spore adhesion, leaf surface colonization or even
apoplastic colonization. This is now increasingly plausi-
ble given the prolonged epiphytic growth observed by
Fones et al. (2017). However, the connection to feed-
ing of a single secreted lipase is tenuous at best when
compared with the abundance of secreted proteins
associated with feeding during the early stages of infec-
tion by other fungal pathogens such as Magnaporthe
oryzae (Kawahara et al., 2012) and F. graminearum
(Lysøe et al., 2011).
While a large amount of the secretions of Z. tritici
appear to be involved in the degradation of host pro-
teins associated with recognition of the pathogen and
initiation of defence during the initial stages of infec-
tion, an unfortunately large abundance of other
secreted proteins remains uncharacterized and
unknown. Because of this, one cannot definitively clas-
sify the feeding habit of the fungus. However, given
the deficit of evidence observed to date (Yang et al.,
2013b, 2015; Rudd et al., 2015), it would
appear that there is minimal fungal feeding taking
place during the latent phase in contrast to the necro-
trophic phase.
Plant Pathology (2019) 68, 1427–1438
 The known unknowns
Question 3: Does the fungus initiate or manipulate host
defences to trigger host cell death for nutrient
acquisition, or are defences wholly host-controlled?
An HR-like response
The HR is one of the most distinguishable plant defence
pathways, involving PCD (Heath, 2000), phytoalexin
production (Keen, 1982; Jabs et al., 1997) and the accu-
mulation of reactive oxygen species (ROS) to restrict fur-
ther pathogen growth and infection (Reape et al., 2008).
This response is usually used by the host to combat bio-
trophic pathogens and can elicit systemic acquired resis-
tance (SAR) through the generation of ROS, such as
hydrogen peroxide (H2O2) superoxide (O2 ) and hydro-
xyl radicals (•OH) (Lamb & Dixon, 1997; Alvarez et al.,
1998). For Z. tritici, H2O2 is inhibitory during the early
latent phase, as it would be to a biotroph, but can be tol-
erated during the necrotrophic stage of the fungus
(Shetty et al., 2003, 2007), primarily due to the produc-
tion of scavenger proteins that reduce the ROS-induced
damage to the fungus (Yang et al., 2013a). The accumu-
lation of H2O2 is accompanied by the collapse of colo-
nized mesophyll cells, causing nutrient leakage into the
apoplast that supports the necrotrophic pathogen (Kema
et al., 1996). While H2O2 accumulation occurs at differ-
ent stages in both compatible and incompatible interac-
tions, the PCD attributed to a classic HR-like response is
confined to the later stages of a compatible interaction
(Rudd et al., 2008). Early accumulation of H2O2 during
an incompatible interaction is strictly regulated by the
host so as not to induce cell death (Stotz et al., 2014).
Despite not colonizing host cells directly, Z. tritici pro-
duces a reduced assortment of CWDEs (in comparison to
other hemibiotrophs; do Amaral et al., 2012) during the
necrotrophic stage (Palma-Guerrero et al., 2015). The
fungal production of ROS-scavenging proteins coupled
with the ability to produce only a limited number of
CWDEs suggests that Z. tritici may have evolved to trig-
ger host cell death, weather the storm of ROS and feed
on the resultant necrotic tissue; this is in contrast to
directly challenging the host defences and independently
attempting to cause necrosis.
Manipulation of host defence
Many necrotrophic (Pandelova et al., 2012) and hemi-
biotrophic (Desmond et al., 2008) pathogens manipulate
wheat host defences to initiate cell death. H2O2 has been
implicated as a signalling molecule for HR but is not
involved in all forms of PCD that occur (Groover et al.,
1997), nor is it universally present in all
plant 9 pathogen combinations investigated for HR
(Heath, 2000). However, the hypothesis that the initia-
tion/manipulation of PCD is a component of the Z. trit-
ici infection process was not supported in a study
conducted by Shetty et al. (2007). Those authors demon-
strated that Z. tritici infection did not increase with
H2O2 infiltration. In fact, the study showed that through
the removal of H2O2 via catalase infiltration, the latent
period of the pathogen was reduced and fungal growth
Plant Pathology (2019) 68, 1427–1438
1433
increased. Furthermore, this study indicated that H2O2 is
damaging to Z. tritici at all stages of its life cycle, but
the fungus is better equipped to tolerate H2O2 in the
later stages of its development (Shetty et al., 2007). Sup-
porting evidence was provided by Rudd et al. (2015) and
corroborated by Palma-Guerrero et al. (2015), who
showed that during early infection of wheat with Z. trit-
ici (as early as 3 dpi), the fungus produces endogenous
catalases in the form of chloroperoxidases, which enable
the fungus to break down H2O2 generated by the host.
As with other necrotrophic pathogens, which can
actively exploit host-produced H2O2 (Govrin & Levine,
2000), Z. tritici has been postulated to not only benefit
from, but also manipulate host H2O2 production (Rudd
et al., 2015; Orton et al., 2016).
Studies of PCD occurring in plants as a result of infec-
tion from Z. tritici have focused primarily on H2O2. For
this reason, the HR-like response of the host cannot be
conclusively and clearly defined without more compre-
hensive studies of the oxidative stress response of wheat
to Z. tritici. For example, other ROS such as hydroxyl
radicals (•OH) and superoxide (O2 ) are produced dur-
ing an HR (Epperlein et al., 1986; Larson, 1988); how-
ever, they are unstable (Able et al., 1998) and
consequently understudied. Hydroxyl radicals have been
implicated in phytoalexin production in soybean (Epper-
lein et al., 1986), and superoxide can be catalysed into
H2O2 during the HR (Larson, 1988). Although they can
act alone, ROS can act in concert with reactive nitrogen
species (RNS), such as nitric oxide (NO) and peroxyni-
trite (ONOO ). In the presence of H2O2, NO will trig-
ger an HR or the expression of plant defence genes
(Fig. 4; Delledonne et al., 1998, 2001; Gr€un et al., 2006)
and can inhibit the detoxification of H2O2 and ONOO
by S-nitrosylation of peroxiredoxin (Dietz et al., 2006;
Romero-Puertas et al., 2007), thus allowing the accumu-
lation of ONOO during the HR (Gaupels et al., 2011).
The cytotoxic ONOO can cross cell membranes and is
potentially involved in mediating NO signalling (Gaupels
et al., 2011) by halting the inhibition of superoxide dis-
mutase (SOD), an enzyme that produces H2O2 from O2
(Holzmeister et al., 2014). Investigations into ROS and
RNS are difficult as these compounds can be short-lived
or indirectly measured during infection. Alternatively, it
may be a better avenue to explore other phenomena that
occur during the HR-like response, such as DNA ladder-
ing cytochrome, (Reape et al., 2008), c release (Keon
et al., 2007), wheat mitogen-activated protein kinase
activation (Rudd et al., 2008) or the roles of fungal
effectors, and subsequently investigate these molecules as
a means of validation that a HR-like response has
occurred.
Studies focusing on the secretome (do Amaral et al.,
2012), transcriptome (Rudd et al., 2015) and proteome
(Yang et al., 2013a, 2015; Yang & Yin, 2015) of Z. trit-
ici during infection of wheat have elucidated the impact
of secreted fungal proteins on the host defence mecha-
nisms. Results of RNA sequencing of a compatible inter-
action between wheat cv. Riband and Z. tritici isolate
1434 C. J. Brennan et al.

Figure 4 Adapted figure of the balanced model created by Delledonne et al. (2001). (a), (b), (c) and (d) refer to four potential outcomes of the
interactions between reactive oxygen species and nitrogen reactive species. O2 , superoxide; SOD, superoxide dismutase; H2O2, hydrogen
peroxide; NO, nitric oxide; ONOO , peroxynitrite. Large font denotes an abundance of a given molecule, strikethrough denotes absence, inhibition
or failure to occur. (a) The balanced model, wherein NO and H2O2 occur in a balanced amount in the presence of SOD, inducing cell death. (b)
Where SOD is absent, O2 is not converted to H2O2 and cell death does not occur. (d) An abundance of O2 causes a lack of interaction between
NO and H2O2 meaning no cell death occurs. (d) An abundance of NO that interacts with O2 leads to a lack of O2 for conversion to H2O2 through
SOD-mediated dismutation. Cell death does not occur.

IPO323 strongly suggested that the commandeering of
host defence coincides with the switch to necrotrophy
and that complex carbohydrates (key constituents of cell
walls) become a significant energy source for the fungus
during late infection (Rudd et al., 2015). Analysis of the
secretome of the fungus itself revealed that H2O2 is used
as a substrate for the generation of beneficial fungal
compounds, such as antibiotics, to reduce resource com-
petition (Orton & Brown, 2016) while simultaneously
reducing ROS-induced damage. The link between cell
death and necrotrophic growth is supported by a
microarray analysis of leaves from a susceptible wheat
cultivar inoculated with Z. tritici (Keon et al., 2007).
Here, the authors showed an increase in host cell death
and ROS accumulation by using qPCR and ROS staining
as visible symptoms appeared. Furthermore, during the
later stages of infection (c. 14 dpi), a suite of fungal
genes involved in oxidative stress and ROS detoxifica-
tion, such as catalases and peroxidases, were up-regu-
lated, demonstrating fungal adaptation to oxidative
stress (Keon et al., 2007). Although the role of these
transcripts in pathogenicity has not been proven, evi-
dence is amassing that an HR-like response is most prob-
ably in the interest of the fungus and an inevitable part
of a compatible interaction.
The role of fungal effectors
In 2011, the first Z. tritici effectors were functionally
categorized and identified as lysin domain-containing
effectors (LysM) associated with the suppression of
pattern-triggered immunity (PTI) through interference in
the chitin-induced plant defence (Marshall et al., 2011).
These LysM effectors were capable of binding to chitin
and one of them (namely Mg3LysM) impeded chitin-trig-
gered defences in wheat. Furthermore, mutant strains of
Z. tritici lacking Mg3LysM were generated that were
impaired in several aspects of infection, from coloniza-
tion to asexual reproduction; in addition, the mutant
was more easily detected by the host, as evidenced by a
heightened host defence response during the latent phase
of infection. Complementing this work was the identifi-
cation of the wheat homologues of a plasma membrane
receptor kinase CERK1 and companion receptor-like
kinase CEBiP required for chitin recognition in rice (Lee
et al., 2014). In wheat plants where the homologues of
these genes were silenced (via virus-induced gene silenc-
ing), the Mg3LysM mutant strain of Z. tritici was patho-
genic, highlighting the active role Z. tritici takes in
defence manipulation of the host (Marshall et al., 2011).
Recent analysis illustrated the ability of 14 Z. tritici
effectors to induce host cell death in the nonhost

Plant Pathology (2019) 68, 1427–1438

 The known unknowns
Nicotiana benthamiana (Kettles et al., 2016). Five of the
effectors were also considerably up-regulated in N. ben-
thamiana compared to in vitro-grown Z. tritici (Kettles
et al., 2016). Interestingly, the cell death phenotype fol-
lowing infiltration of at least three of the effectors was
dependent on expression of two host resistance genes
(NbBAK1 and NbSOBIR1), suggesting that apoplastic
recognition of the effectors is crucial for a downstream
cell death response (Kettles et al., 2016), in keeping with
the effector-triggered branch of the zig-zag model of
plant immunity (Jones & Dangl, 2006). Although there
is no evidence that these effectors directly induce cell
death when infiltrated into a wheat host, their expression
levels peak during the switch from symptomless to necro-
trophic growth in wheat (Rudd et al., 2015), further
implicating them in the host–pathogen relationship.
More recently, in 2017, the Z. tritici ribonuclease
effector Zt6, toxic to bacteria, yeasts and tobacco but
not Z. tritici itself, was shown to elicit cell death in
wheat as well as serving a secondary role in antimicro-
bial competition (Kettles et al., 2018). Not only does
Zt6 have dual functionality, it also displays a double
peak in expression, first at 1 dpi, followed by a second
peak at 14 dpi, coinciding, as do the effectors discovered
by Rudd et al. (2015), with the switch to necrotrophy.
However, unlike the NbBAK1 and NbSOBIR1 effectors,
the ability of Zt6 to induce cell death is independent of
host receptor-like kinase resistance genes (Kettles et al.,
2018). Although this evidence points to a role in
pathogenicity or virulence of the fungus, DZt6 mutants
had no loss of virulence when inoculated onto wheat
leaves (Kettles et al., 2018). This suggests, therefore, that
perhaps the primary role of Zt6 is not toxicity towards
the plant, but rather to competing microorganisms.
The Z. tritici avirulence factor AvrStb6 is a small cys-
teine-rich effector protein that interacts with STB6 (a
wall-associated receptor kinase-like protein and the first
major STB resistance gene to be cloned), which confers
gene-for-gene resistance in wheat to Z. tritici (Zhong
et al., 2017; Kema et al., 2018; Saintenac et al., 2018).
A direct interaction between STB6 and AvrStb6 has not
been demonstrated (Saintenac et al., 2018) and the func-
tion of the AvrStb6 effector in the absence of stb6
remains unknown (Brunner & McDonald, 2018). Con-
ventional effector-triggered immune responses such as
these are usually followed by an HR (Dickman & Fluhr,
2013). However, this is not the case in the stb6/AvrStb6
interaction, which more closely follows effector-triggered
defence rather than immunity; in this model, an apoplas-
tic fungus detected by receptor-like proteins can be
arrested without the elicitation of an HR-like response
(Jones & Dangl, 2006; Stotz et al., 2014). This, in turn,
suggests that an HR-like response is advantageous to
Z. tritici and its lack or inhibition is detrimental to colo-
nization by the fungus. Effects of interactions between
host resistance genes and pathogen effector genes are not
restricted to qualitative resistance as quantitative resis-
tance was also observed in the recently discovered stb7/
Avr3D1 interaction (Meile et al., 2018).
Plant Pathology (2019) 68, 1427–1438
1435
Discovery of similar resistance gene interactions or
expansion of the current stb6/AvrStb6 model with effec-
tors known to cause cell death, together with a better
understanding of their respective interactors, will add to
the evidence for manipulation of the host’s defences by
Z. tritici; such research will undoubtedly help to deci-
pher the nature of dedicated pathogen attack and host
defence mechanisms against STB disease. Therefore, in
answer to the question ‘Does the fungus initiate or
manipulate host defences to trigger cell death for nutrient
acquisition, or are defences wholly host-controlled?’, it
would appear that the secretion of fungal effectors are
actively responsible for manipulation of host defences
and may even contribute to the cell death seen during
later stages of fungal infection.
Concluding Remarks
The collective efforts of those involved in the develop-
ment and application of both contemporary molecular
and sequencing technologies to the Z. tritici–wheat inter-
action have gradually begun to shift the paradigm of
infection from the unknown to known. Steady accumula-
tion of information on the transcriptomes, proteomes,
secretome, defence/evasion pathways and key genes of
both the host and the pathogen has finally begun to
paint a clearer (albeit still low resolution) picture of the
overall infection cycle, including that of the once wholly
elusive latent phase of infection (Fig. 1). Based on pre-
sent evidence, it is suggested that the latent phase is not
an inactive state nor a state involved in feeding, but one
dedicated to host defence suppression and reproductive
preparation. Explorations into the roles and functions of
the unknown fungal effectors secreted by Z. tritici have
all but conclusively demonstrated that HR-like manipula-
tion is taking place and future work will undoubtedly
highlight more candidates involved in triggering the
switch to necrotrophy. Furthermore, elucidation of the
roles of individual or families of host susceptibility genes
or fungal virulence genes responsible for compatible
interactions could lead to them becoming targets for
manipulation by breeding or mutagenesis to elicit resis-
tance or control the fungus; such methods could be used
alongside the continued use of contemporary chemical
control and the development of novel chemistries.
Exploratory and comparative ‘omics’ research, particu-
larly RNA-Seq and proteomics, can have high costs. This
unfortunately leads to the selection of the
pathogen 9 host combinations most easy to work with,
narrowing the scope of understanding of incompatible
interactions. For understandable reasons, not least the
cost of analysis such as RNA-Seq, there is a lack of
information on the biology underlying host and nonhost
resistance to Z. tritici. Several studies have been con-
ducted using virulent Z. tritici 9 susceptible host combi-
nations (Keon et al., 2005, 2007; Yang et al., 2013a;
Palma-Guerrero et al., 2015; Rudd et al., 2015) but
fewer using avirulent Z. tritici 9 resistant host combina-
tions where infection is arrested (M’Barek et al., 2015b;
1436 C. J. Brennan et al.
Yang et al., 2015). Understanding the latent phase in
both the compatible and incompatible interactions (and
the differences that are represented in both) will allow
for a more all-encompassing approach to understanding
and ultimately combating Z. tritici.
Acknowledgements
Ciaran Brennan was funded by the Irish Department of
Agriculture, Food and the Marine Research Stimulus
grant CIVYL (RSF 11/S/121). Harriet Benbow was
funded by Science Foundation Ireland PI Research
Award 14/1A/2508.
References
Able AJ, Guest DI, Sutherland MW, 1998. Use of a new tetrazolium-
based assay to study the production of superoxide radicals by tobacco
cell cultures challenged with avirulent zoospores of Phytophthora
parasitica varnicotianae. Plant Physiology 117, 491–9.
Adhikari T, Yang X, Cavaletto J et al., 2004a. Molecular mapping of
Stb1, a potentially durable gene for resistance to Septoria tritici blotch
in wheat. Theoretical and Applied Genetics 109, 944–53.
Adhikari TB, Cavaletto JR, Dubcovsky J, Gieco JO, Schlatter AR,
Goodwin SB, 2004b. Molecular mapping of the Stb4 gene for
resistance to Septoria tritici blotch in wheat. Phytopathology 94,
1198–206.
Adhikari TB, Balaji B, Breeden J, Goodwin SB, 2007. Resistance of
wheat to Mycosphaerella graminicola involves early and late peaks of
gene expression. Physiological and Molecular Plant Pathology 71, 55–
68.
Alvarez ME, Pennell RI, Meijer P-J, Ishikawa A, Dixon RA, Lamb C,
1998. Reactive oxygen intermediates mediate a systemic signal
network in the establishment of plant immunity. Cell 92, 773–84.
do Amaral AM, Antoniw J, Rudd JJ, Hammond-Kosack KE, 2012.
Defining the predicted protein secretome of the fungal wheat leaf
pathogen Mycosphaerella graminicola. PLoS One 7, e49904.
Banke S, Peschon A, McDonald BA, 2004. Phylogenetic analysis of
globally distributed Mycosphaerella graminicola populations based on
three DNA sequence loci. Fungal Genetics and Biology 41, 226–38.
Berr A, Shafiq S, Shen W-H, 2011. Histone modifications in
transcriptional activation during plant development. Biochimica et
Biophysica Acta – Gene Regulatory Mechanisms 1809,
567–76.
Brown JK, Chartrain L, Lasserre-Zuber P, Saintenac C, 2015. Genetics of
resistance to Zymoseptoria tritici and applications to wheat breeding.
Fungal Genetics and Biology 79, 33–41.
Brunner PC, McDonald BA, 2018. Evolutionary analyses of the
avirulence effector AvrStb6 in global populations of Zymoseptoria
tritici identify candidate amino acids involved in recognition.
Molecular Plant Pathology 19, 1836–46.
Cohen L, Eyal Z, 1993. The histology of processes associated with the
infection of resistant and susceptible wheat cultivars with Septoria
tritici. Plant Pathology 42, 737–43.
Cowger C, Hoffer M, Mundt C, 2000. Specific adaptation by
Mycosphaerella graminicola to a resistant wheat cultivar. Plant
Pathology 49, 445–51.
Danon A, Delorme V, Mailhac N, Gallois P, 2000. Plant programmed
cell death: a common way to die. Plant Physiology and Biochemistry
38, 647–55.
Delledonne M, Xia Y, Dixon RA, Lamb C, 1998. Nitric oxide functions
as a signal in plant disease resistance. Nature 394, 585–8.
Delledonne M, Zeier J, Marocco A, Lamb C, 2001. Signal interactions
between nitric oxide and reactive oxygen intermediates in the plant
hypersensitive disease resistance response. Proceedings of the National
Academy of Sciences of the United States of America 98, 13454–9.
Desmond OJ, Manners JM, Stephens AE et al., 2008. The Fusarium
mycotoxin deoxynivalenol elicits hydrogen peroxide production,
programmed cell death and defence responses in wheat. Molecular
Plant Pathology 9, 435–45.
Dickman MB, Fluhr R, 2013. Centrality of host cell death in plant–
microbe interactions. Annual Review of Phytopathology 51, 543–70.
Dickman M, Williams B, Li Y, De Figueiredo P, Wolpert T, 2017.
Reassessing apoptosis in plants. Nature Plants 3, 773–9.
Dietz K-J, Jacob S, Oelze M-L et al., 2006. The function of
peroxiredoxins in plant organelle redox metabolism. Journal of
Experimental Botany 57, 1697–709.
Dooley H, Shaw MW, Mehenni-Ciz J, Spink J, Kildea S, 2016. Detection
of Zymoseptoria tritici SDHI-insensitive field isolates carrying the
SdhC-H152R and SdhD-R47W substitutions. Pest Management
Science 72, 2203–7.
Duncan KE, Howard RJ, 2000. Cytological analysis of wheat infection
by the leaf blotch pathogen Mycosphaerella graminicola. Mycological
Research 104, 1074–82.
Dyer PS, O’Gorman CM, 2012. Sexual development and cryptic
sexuality in fungi: insights from Aspergillus species. FEMS
Microbiology Reviews 36, 165–92.
Epperlein M, Noronha-Dutra A, Strange R, 1986. Involvement of the
hydroxyl radical in the abiotic elicitation of phytoalexins in legumes.
Physiological and Molecular Plant Pathology 28, 67–77.
Eriksen L, Munk L, 2003. The occurrence of Mycosphaerella
graminicola and its anamorph Septoria tritici in winter wheat during
the growing season. European Journal of Plant Pathology 109, 253–9.
Fones H, Gurr S, 2015. The impact of septoria tritici blotch disease on
wheat: an EU perspective. Fungal Genetics and Biology 79, 3–7.
Fones HN, Eyles CJ, Kay W, Cowper J, Gurr SJ, 2017. A role for
random, humidity-dependent epiphytic growth prior to invasion of
wheat by Zymoseptoria tritici. Fungal Genetics and Biology 106, 51–
60.
Gaupels F, Spiazzi-Vandelle E, Yang D, Delledonne M, 2011. Detection
of peroxynitrite accumulation in Arabidopsis thaliana during the
hypersensitive defense response. Nitric Oxide 25, 222–8.
Goodwin SB, M’Barek SB, Dhillon B et al., 2011. Finished genome of
the fungal wheat pathogen Mycosphaerella graminicola reveals
dispensome structure, chromosome plasticity, and stealth pathogenesis.
PLoS Genetics 7, e1002070.
Govrin EM, Levine A, 2000. The hypersensitive response facilitates plant
infection by the necrotrophic pathogen Botrytis cinerea. Current
Biology 10, 751–7.
Groover A, Dewitt N, Heidel A, Jones A, 1997. Programmed cell death
of plant tracheary elements differentiating in vitro. Protoplasma 196,
197–211.
Gr€un S, Lindermayr C, Sell S, Durner J, 2006. Nitric oxide and gene
regulation in plants. Journal of Experimental Botany 57, 507–16.
Haueisen J, Moeller M, Eschenbrenner CJ et al., 2017. Extremely flexible
infection programs in a fungal plant pathogen. bioRxiv, 229997.
Hayes LE, Sackett KA, Anderson NP, Flowers MD, Mundt CC, 2015.
Evidence of selection for fungicide resistance in Zymoseptoria tritici
populations on wheat in western Oregon. Plant Disease 100, 483–9.
Heath MC, 2000. Hypersensitive response-related death in programmed
cell death in higher plants. Plant Molecular Biology 44, 321–34.
Hehir JG, Connolly C, O’Driscoll A et al., 2018. Temporal and spatial
field evaluations highlight the importance of the pre-symptomatic
phase in supporting strong partial resistance in Triticum aestivum
against Zymoseptoria tritici. Plant Pathology 67, 573–83.
Heick TM, Justesen AF, Jørgensen LN, 2017. Anti-resistance strategies
for fungicides against wheat pathogen Zymoseptoria tritici with focus
on DMI fungicides. Crop Protection 99, 108–17.
Holzmeister C, Gaupels F, Geerlof A et al, 2014. Differential inhibition
of Arabidopsis superoxide dismutases by peroxynitrite-mediated
tyrosine nitration. Journal of Experimental Botany 66, 989–99.
Jabs T, Tsch€ope M, Colling C, Hahlbrock K, Scheel D, 1997. Elicitor-
stimulated ion fluxes and O2 from the oxidative burst are essential
components in triggering defense gene activation and phytoalexin
Plant Pathology (2019) 68, 1427–1438
 The known unknowns
synthesis in parsley. Proceedings of the National Academy of Sciences
of the United States of America 94, 4800–5.
Jones JD, Dangl JL, 2006. The plant immune system. Nature 444, 323–9.
Kawahara Y, Oono Y, Kanamori H, Matsumoto T, Itoh T, Minami E,
2012. Simultaneous RNA-seq analysis of a mixed transcriptome of rice
and blast fungus interaction. PLoS One 7, e49423.
Keen N, 1982. Specific recognition in gene-for-gene host–parasite
systems. Advances in Plant Pathology. 1, 35–82.
Kema GH, Yu D, Rijkenberg FH, Shaw MW, Baayen RP, 1996.
Histology of the pathogenesis of Mycosphaerella graminicola in wheat.
Phytopathology 86, 777–86.
Kema GH, Gohari AM, Aouini L et al., 2018. Stress and sexual
reproduction affect the dynamics of the wheat pathogen effector
AvrStb6 and strobilurin resistance. Nature Genetics 50, 375–80.
Kemen E, Gardiner A, Schultz-Larsen T et al., 2011. Gene gain and loss
during evolution of obligate parasitism in the white rust pathogen of
Arabidopsis thaliana. PLoS Biology 9, e1001094.
Keon J, Rudd JJ, Antoniw J, Skinner W, Hargreaves J, Hammond-
Kosack K, 2005. Metabolic and stress adaptation by Mycosphaerella
graminicola during sporulation in its host revealed through microarray
transcription profiling. Molecular Plant Pathology 6, 527–40.
Keon J, Antoniw J, Carzaniga R et al., 2007. Transcriptional adaptation
of Mycosphaerella graminicola to programmed cell death (PCD) of its
susceptible wheat host. Molecular Plant–Microbe Interactions 20,
178–93.
Kettles GJ, Bayon C, Canning G, Rudd JJ, Kanyuka K, 2016. Apoplastic
recognition of multiple candidate effectors from the wheat pathogen
Zymoseptoria tritici in the nonhost plant Nicotiana benthamiana. New
Phytologist 213, 338–50.
Kettles GJ, Bayon C, Sparks CA, Canning G, Kanyuka K, Rudd JJ, 2018.
Characterization of an antimicrobial and phytotoxic ribonuclease
secreted by the fungal wheat pathogen Zymoseptoria tritici. New
Phytologist 217, 320–31.
Lamb C, Dixon RA, 1997. The oxidative burst in plant disease
resistance. Annual Review of Plant Biology 48, 251–75.
Larson RA, 1988. The antioxidants of higher plants. Phytochemistry 27,
969–78.
Lee W-S, Rudd JJ, Hammond-Kosack KE, Kanyuka K, 2014.
Mycosphaerella graminicola LysM effector-mediated stealth
pathogenesis subverts recognition through both CERK1 and CEBiP
homologues in wheat. Molecular Plant–Microbe Interactions 27, 236–
43.
Lee J, Orosa B, Millyard L et al., 2015. Functional analysis of a wheat
homeodomain protein, TaR1, reveals that host chromatin remodelling
influences the dynamics of the switch to necrotrophic growth in the
phytopathogenic fungus Zymoseptoria tritici. New Phytologist 206,
598–605.
Lucas J, 2003. Resistance to Qol fungicides: implications for cereal
disease management in Europe. Pesticide Outlook 14, 268–70.
Lysøe E, Seong K-Y, Kistler HC, 2011. The transcriptome of Fusarium
graminearum during the infection of wheat. Molecular Plant–Microbe
Interactions 24, 995–1000.
M’Barek SB, Cordewener JH, Ghaffary SMT et al., 2015a. FPLC and
liquid-chromatography mass spectrometry identify candidate necrosis-
inducing proteins from culture filtrates of the fungal wheat pathogen
Zymoseptoria tritici. Fungal Genetics and Biology 79, 54–62.
M’Barek SB, Cordewener JH, van der Lee TA et al., 2015b. Proteome
catalog of Zymoseptoria tritici captured during pathogenesis in wheat.
Fungal Genetics and Biology 79, 42–53.
Marshall R, Kombrink A, Motteram J et al., 2011. Analysis of two in
planta expressed LysM effector homologs from the fungus
Mycosphaerella graminicola reveals novel functional properties and
varying contributions to virulence on wheat. Plant Physiology 156,
756–69.
McDonald BA, Stukenbrock EH, 2016. Rapid emergence of pathogens in
agro-ecosystems: global threats to agricultural sustainability and food
security. Philosophical Transactions of the Royal Society B: Biological
Sciences 371, 20160026.
Plant Pathology (2019) 68, 1427–1438
1437
Meile L, Croll D, Brunner PC et al., 2018. A fungal avirulence factor
encoded in a highly plastic genomic region triggers partial resistance to
septoria tritici blotch. New Phytologist 219, 1048–61.
Orton ES, Brown JK, 2016. Reduction of growth and reproduction of
the biotrophic fungus Blumeria graminis in the presence of a
necrotrophic pathogen. Frontiers in Plant Science 7, 742.
Orton E, Rudd J, Brown J, 2016. Early molecular signatures of responses
of wheat to Zymoseptoria tritici in compatible and incompatible
interactions. Plant Pathology 66, 450–9.
Palma-Guerrero J, Torriani SF, Zala M et al., 2015. Comparative
transcriptome analyses of Zymoseptoria tritici strains show complex
lifestyle transitions and intra-specific variability in transcription
profiles. Molecular Plant Pathology 17, 845–59.
Pandelova I, Figueroa M, Wilhelm LJ et al., 2012. Host-selective toxins
of Pyrenophora tritici-repentis induce common responses associated
with host susceptibility. PLoS One 7, e40240.
Parlevliet J, 1975. Partial resistance of barley to leaf rust, Puccinia
hordei. I. Effect of cultivar and development stage on latent period.
Euphytica 24, 21–7.
Reape TJ, Molony EM, McCabe PF, 2008. Programmed cell death in
plants: distinguishing between different modes. Journal of
Experimental Botany 59, 435–44.
Rohel EA, Payne AC, Fraaije BA, Hollomon DW, 2001. Exploring
infection of wheat and carbohydrate metabolism in Mycosphaerella
graminicola transformants with differentially regulated green
fluorescent protein expression. Molecular Plant–Microbe Interactions
14, 156–63.
Romero-Puertas MC, Laxa M, Matte A et al., 2007. S-nitrosylation of
peroxiredoxin II E promotes peroxynitrite-mediated tyrosine nitration.
The Plant Cell 19, 4120–30.
Rudd JJ, Keon J, Hammond-Kosack KE, 2008. The wheat mitogen-
activated protein kinases TaMPK3 and TaMPK6 are differentially
regulated at multiple levels during compatible disease interactions with
Mycosphaerella graminicola. Plant Physiology 147, 802–15.
Rudd J, Kanyuka K, Hassani-Pak K et al., 2015. Transcriptome and
metabolite profiling the infection cycle of Zymoseptoria tritici on
wheat (Triticum aestivum) reveals a biphasic interaction with plant
immunity involving differential pathogen chromosomal contributions,
and a variation on the hemibiotrophic lifestyle definition. Plant
Physiology 167, 1158–85.
Saintenac C, Lee W-S, Cambon F et al., 2018. Wheat receptor-kinase-
like protein Stb6 controls gene-for-gene resistance to fungal pathogen
Zymoseptoria tritici. Nature Genetics 50, 368–74.
S
anchez-Vallet A, McDonald MC, Solomon PS, McDonald BA, 2015. Is
Zymoseptoria tritici a hemibiotroph? Fungal Genetics and Biology 79,
29–32.
Schulz B, Boyle C, Draeger S, R€ ommert A-K, Krohn K, 2002.
Endophytic fungi: a source of novel biologically active secondary
metabolites. Mycological Research 106, 996–1004.
Shaw M, Royle D, 1989. Airborne inoculum as a major source of
Septoria tritici (Mycosphaerella graminicola) infections in winter
wheat crops in the UK. Plant Pathology 38, 35–43.
Shearer B, Zadoks J, 1972. The latent period of Septoria nodorum in
wheat. 1. The effect of temperature and moisture treatments under
controlled conditions. Netherlands Journal of Plant Pathology 78,
231–41.
Shetty NP, Kristensen B, Newman M-A, Møller K, Gregersen PL,
Jørgensen HL, 2003. Association of hydrogen peroxide with restriction
of Septoria tritici in resistant wheat. Physiological and Molecular Plant
Pathology 62, 333–46.
Shetty NP, Mehrabi R, L€ utken H et al., 2007. Role of hydrogen peroxide
during the interaction between the hemibiotrophic fungal pathogen
Septoria tritici and wheat. New Phytologist 174, 637–47.
Shetty NP, Jensen JD, Knudsen A et al., 2009. Effects of b-1,3-glucan
from Septoria tritici on structural defence responses in wheat. Journal
of Experimental Botany 60, 4287–300.
Siah A, Deweer C, Duyme F et al., 2010. Correlation of in planta endo-
b-1,4-xylanase activity with the necrotrophic phase of the
1438 C. J. Brennan et al.
hemibiotrophic fungus Mycosphaerella graminicola. Plant Pathology
59, 661–70.
Spanu P, K€ amper J, 2010. Genomics of biotrophy in fungi and
oomycetes—emerging patterns. Current Opinion in Plant Biology 13,
409–14.
Stergiopoulos I, Zwiers L-H, De Waard MA, 2003. The ABC transporter
MgAtr4 is a virulence factor of Mycosphaerella graminicola that
affects colonization of substomatal cavities in wheat leaves. Molecular
Plant–Microbe Interactions 16, 689–98.
Stotz HU, Mitrousia GK, de Wit PJ, Fitt BD, 2014. Effector-triggered
defence against apoplastic fungal pathogens. Trends in Plant Science
19, 491–500.
Stukenbrock EH, McDonald BA, 2008. The origins of plant pathogens in
agro-ecosystems. Annual Review of Phytopathology 46, 75–100.
Stukenbrock EH, Banke S, Javan-Nikkhah M, McDonald BA, 2006.
Origin and domestication of the fungal wheat pathogen
Mycosphaerella graminicola via sympatric speciation. Molecular
Biology and Evolution 24, 398–411.
Stukenbrock EH, Jørgensen FG, Zala M, Hansen TT, McDonald BA,
Schierup MH, 2010. Whole-genome and chromosome evolution
associated with host adaptation and speciation of the wheat pathogen
Mycosphaerella graminicola. PLoS Genetics 6, e1001189.
Torriani SF, Melichar JP, Mills C, Pain N, Sierotzki H, Courbot M,
2015. Zymoseptoria tritici: a major threat to wheat production,
integrated approaches to control. Fungal Genetics and Biology 79,
8–12.
Trail F, 2009. For blighted waves of grain: Fusarium graminearum in the
postgenomics era. Plant Physiology 149, 103–10.
Welch T, Feechan A, Kildea S, 2018. Effect of host resistance on genetic
structure of core and accessory chromosomes in Irish Zymoseptoria
tritici populations. European Journal of Plant Pathology 150, 139–48.
Yang F, Yin Q, 2015. Comprehensive proteomic analysis of the wheat
pathogenic fungus Zymoseptoria tritici. Proteomics 16, 98–101.
Yang F, Li W, Jørgensen HJ, 2013a. Transcriptional reprogramming of
wheat and the hemibiotrophic pathogen Septoria tritici during two
phases of the compatible interaction. PLoS One 8, e81606.
Yang F, Melo-Braga MN, Larsen MR, Jørgensen HJ, Palmisano G,
2013b. Battle through signaling between wheat and the fungal
pathogen Septoria tritici revealed by proteomics and
phosphoproteomics. Molecular & Cellular Proteomics 12, 2497–508.
Yang F, Li W, Derbyshire M, Larsen MR, Rudd JJ, Palmisano G, 2015.
Unraveling incompatibility between wheat and the fungal pathogen
Zymoseptoria tritici through apoplastic proteomics. BMC Genomics
16, 362.
Zhong Z, Marcel TC, Hartmann FE et al., 2017. A small secreted
protein in Zymoseptoria tritici is responsible for avirulence on wheat
cultivars carrying the Stb6 resistance gene. New Phytologist 214, 619–
31.
Zuccaro A, Lahrmann U, G€ uldener U et al., 2011. Endophytic life
strategies decoded by genome and transcriptome analyses of the
mutualistic root symbiont Piriformospora indica. PLoS Pathogens 7,
e1002290.
Plant Pathology (2019) 68, 1427–1438