DISCUSSION
In this study we have characterized the genetic and pathotypic
population structure of P. oryzae pathogens infecting traditionalindica varieties in the Yuanyang terraces of rice paddies. This
traditional agrosystem, maintained over centuries
(He et al., 2011), and
where rice disease pressure was reported to be low
(Sheng, 1990),
provides a unique opportunity to decipher the impact of crop diversity
on disease epidemics, especially of rice blast.
Our first important observation based on analysis of microsatellite and
whole genome genetic variation, is the finding of new lineages of the
rice blast pathogen endemic to YYT. Our analysis of population structure
based on microsatellite (513 isolates) and whole genome (46 isolates)
datasets revealed multiple lineages of P. oryzae coexisting in
YYT, with relatively high levels of standing variation compared to
previous results from Gladieux et al., [23] and Saleh et al.(Saleh et al., 2014).
Three genetic lineages endemic to YYT, coexisted with two of the four
worldwide lineages previously described [23]. Although the global
linkage disequilibrium inferred from microsatellite data was
significantly different from 0, analyses of LD-decay, PHI-tests and
reticulations within each of the five lineages provided contrasted
information regarding the existence of recombination (Supplementary
Information 2, Fig. SI2.2). Both mating types were found in sympatry
(same village; data not shown) within the 46 YYT P. oryzae fully
sequenced isolates, but they were found within the same lineage only for
WL1 (Supplementary Information 2, Table SI2.3), which, together with
significant PHI-test and reticulations observed for this lineage, was
consistent with the fact that it has been described as recombinant
(Gladieux et al.,
2018; Latorre et al., 2020; Saleh et al., 2014; Thierry et al., 2021).
Conversely, only Mat-2 isolates were found among the 30 fully sequenced
isolates assigned to lineage YYT1. Therefore, significant PHI-test and
reticulations observed in the minimum spanning network for this lineage
could be due to scarce genetic exchanges among lineages
(Gladieux et al.,
2018), or to footprints of historical recombination. In invasive
pathogens, higher genetic diversity and signatures of recombination are
expected in older, source populations
(Ali et al., 2014;
Thach, Ali, de Vallavieille-Pope, Justesen, & Hovmøller, 2016). Our
observations therefore suggest that YYT area is very close to, if not
included into, the centre of origin of rice-infecting P. oryzaepathogens in continental Southeast Asia hypothesized by previous studies
(Gladieux et al.,
2018; Saleh et al., 2012; Zhong et al., 2018).
Our second important observation is that P. oryzae populations
are not specialized to traditional rice landraces in YYT. For pathogens
mating within or onto their hosts, specialization should drastically
restrict encounters of potential mates and reduce survival of offspring
due to maladaptation of immigrants and hybrid offspring
(Giraud, Gladieux, &
Gavrilets, 2010; Gladieux et al., 2011), which should align the
structure of pathogenic populations on that of the host. Our
genotyping-by-sequencing data show that the rice accessions from YYT are
structured into landraces with relatively high levels of genetic
diversity, both within and among landraces, confirming previous findings
based on 24 microsatellite markers
(Gao et al., 2012).P. oryzae populations are also structured into different
lineages, but our analysis reveals a complete lack of host-pathogen
genetic co-structure. The fact that population subdivision in the
pathogen does not mirror population subdivision in the host strongly
suggests a lack of specialization to the host. However, co-structure
between host and pathogen genealogies might be only detectable at those
loci in the genomes that are specifically involved in coevolutionary
processes (Märkle et al., 2021). The data presented here does not allow
to test this hypothesis since we lack full genomic information on the
plant side, and further genomic analyses are therefore needed to tackle
this issue. GWAs analyses performed on the pathogen side detected loci
that were involved in the interaction with at least two rice accessions,
which is also consistent with a lack of local adaptation. We also
analysed the phenotypic relationships among paired P. oryzae /
rice samples by cross inoculating all isolates on their native and
non-native plants. We showed that nearly all qualitative interactions
were compatible. Liao et al. (2016) sequenced the genomes of two YYT
rice accessions (Acuce and Xiaogu) and showed that they content 7 and 8
known R genes, respectively, without counting all the other unkown R
genes. When 30 representative YYT isolates from indica accessions were
inoculated against the modern rice varieties carrying known resistance
genes, and showed that these P. oryzae isolates had lost may
avirulence functions (Liao et al., 2016). Our result thus confirm that
the great majority of the R genes present in indica YYT accessions were
defeated by the P. oryzae population. Quantitative interactions
indicated that P. oryzae isolates did not perform significantly
better on their native than on their non-native plants, and thatP. oryzae isolates originating from plants of the same landrace
did not perform significantly better on this landrace than on any other
landraces, thus leading to rejection of the “home versus away”
criteria of local adaptation
(Blanquart, Klatz,
Nuismer, & Gandon, 2013). Together, the results of our analysis of
population genetic and pathotypic structure therefore reveals a complete
lack of P. oryzae specialization to rice landraces and thus, a
lack of adaptation to specific hosts, at least within the indica host
compartment. Some studies have shown adaptation to specific host lines
(Goyeau, Halkett,
Zapater, Carlier, & Lannou, 2007; Goyeau, Park, Schaeffer, & Lannou,
2006), though this was not the case here. Although our results are
consistent with a generalist life style in P. oryzae population,
maladaptation cannot be definitely ruled out. Maladaptation in pathogens
describes the case where isolates performance is significantly better on
non-native hosts than on native hosts
(Kniskern, Barrett, &
Bergelsong, 2011). Given that we have assessed performance using an
overall infection trait (the percentage of diseased leaf surface), we
cannot exclude that other traits involved in fungal fitness that are not
captured by our index (e.g. number of lesions, proportion of HR lesions
among infective lesions, growth speed of lesion, sporulation capacity)
would reveal such a maladaptation pattern.
Rice landraces and their P. oryzae pathogens from the Yuanyang
terraces therefore represent a model system in which the pathogen is
specialized to indica and japonica rice subspecies (Liao
et al. 2016), but not specialized to the various landraces ofindica rice. This is consistent with predictions that the nature
of the mechanism underlying immunity in the host or avirulence in the
pathogen and the magnitude of divergence in immune systems between hosts
has an impact on the likelihood of pathogen specialization
(Giraud et al., 2010;
Schulze-Lefert & Panstruga, 2011). Liao et al.
(2016) showed that in
the YYT area where japonica and indica rice subspecies are cultivated in
sympatry, differences in immune systems between indica andjaponica subspecies would prevent the emergence of populations
with a generalist lifestyle on both hosts, because a large effector
complement is required to infect japonica rice, whileindica rice has a larger repertoire of immune receptors and
therefore greater capacity to detect effectors that will trigger
immunity. Differences in repertoires of immune receptors amongindica landraces might be sufficient to lead to the emergence of
specialized P. oryzae populations in stable and homogeneous
conditions. The maintenance of generalist P. oryzaegenotypes highlighted by our results could be explained by the elevated
heterogeneity of the “host landscape” in the YYT area, with
spatio-temporal variation in rice genotypes distribution (elevated
genetic diversity within and among rice landraces, spatial mosaics of
paddy fields sown with different landraces, and temporal turnover of the
mosaics) impeding the emergence of specialized pathogens on specificindica plant genotypes or landraces. This could also be possible
in case of lack of selection pressure due to R genes in the host, where
the host lack many R genes, the absence of R genes in YYT landraces is
highly unlikely. Unlike populations of P. oryzae from modern
agrosystems, which tend to be largely clonal and infecting relatively
stable and homogenous host populations
(Gladieux et al.,
2018; Zhong et al., 2018), populations infecting YYT landraces displayed
higher genotypic and genetic diversities with occurrence of
recombination. This last feature may contribute to the maintenance of
the generalist lifestyle by re-shuffling virulence alleles amongP. oryzae pathogens. Finally, the Yuanyang Terraces bring to
light an interesting observation, in which the populations of the
pathogen are diverse and recombinant - characteristics of populations
with high adaptive potential - while they generate less yield losses
than the clonal populations observed in other parts of the globe. This
suggests that the epidemiological models that should help the
re-engineering of agrosystems should not ignore the scenarios leading to
the emergence of recombinant and diverse pathogenic populations, as
observed in our case study. Strategies should be adopted to avoid the
emergence of specialist pathogen lineages and encourage the evolution of
generalist pathogen which causes lesser disease burden. Varietal mixture
at the spatial scale (even if not within the field) and temporal scale
along with the encouragement of diverse sources of germplasm could be
useful in this regard.
This study could inspire future work to foster the adoption of dynamic
diversity (McDonald,
2014) to decrease the disease burden at the landscape level. This could
be translated to other areas and other pathosystems to ensure a low
disease burden without rapid emergence of virulences and low fungicide
application.