Pathogenic Variability in Bipolaris sorokiniana

Presenter
Major Advisor
Asst. Prof. Pratishtha Adhikari, PhD
Department of Plant Pathology
Agriculture and Forestry University
Rampur, Nepal
Pooja Gurung
PLP-06M-2021
Credit Seminar (PLP699)
3rd Semester, M. Sc. Agriculture
Department of Plant Pathology
Agriculture and Forestry University
Rampur, Nepal
Outline of Presentation

Introduction
The pathogen: Bipolaris sorokiniana
Worldwide Distribution of Pathogen
Host Range
Pathogenicity Factors
Host Response
Pathogenic Variability
Introduction

1809 A.D. Genus: Helminthosporium

Nisikado
(1928) Subgenera

Cylindro-Helminthosporium Eu-Helminthosporium

One or more germtubes from any cell Germination Only from end cells
Straight cylindrical conidia Shape Fusiform often curved

Drechslera Bipolaris
Exserohilum
 Introduction
Differentiating criteria for Bipolaris, Drechslera and Exserohilum

Fig. 1: Hilum structure in Drechslera, Bipolaris Fig. 2: Basal cell germination & progression of
and Exserohilum (left to right) (Alcorn, 1988) septum formation in Drechslera (a,c), Bipolaris
(b,d), and Exserohilum (b,e) (Alcorn, 1988)
The Pathogen: Bipolaris sorokiniana
Fusoid often curved conidia
• Shoemaker (1959) proposed the generic name
Bipolaris for the Helminthosporium species Germinating by one germ tube from
each end (bipolar germination)

• B. sorokiniana is characterized by

Thick-walled, elliptical conidia (60-120µm x

12-20µm) with 5 to 9 cells (Kumar et al.,
2002) Fig. 3: Conidium of B. sorokiniana
Mycelium composed of hyphae interwoven as loose cottony mass,
• In axenic culture appears white or light to dark grey depending on the isolates

Fig. 4: Variation in shades of mycelium of B. sorokiniana cultures

Worldwide Distribution of the Pathogen

Causal agent of foliar spot blotch, root rot, and black points on grains of wheat and
barley (Sivanesan, 1990)

Most common in humid, warm, wheat-growing regions, with South Asia’s Eastern
Gangetic Plains serving as hotspot (Sharma et al., 2022)

Pathogen also occurs in North and Latin America (Duczek and Jones-Flory, 1994),
Brazil (Mehta et al., 1992), in parts of Europe (Kwasna, 1995)

Spot blotch reduces wheat output by 23 to 40 % in Nepal (Sharma and Duveiller, 2006)
Host Range
Table 1: Monocotyledonous hosts of Bipolaris sorokiniana
Cereals Grasses
Triticum aestivum Agropyron pectinatum
Secale cereale Agropyron repens
Hordeum vulgare Alopecurus pratensis
Hordeum murinum Beckmannia eruciformis
Avena sativa Bromus erectus
Bromus inermis
Dactylis glomerata
Festuca heterophylla
Festuca ovina
Lolium perenne
Pennisetum villosum
Poa pratensis
Bakonyi et al. (1997)
Host Response
Hemibiotrophic
Exerts a biotrophic and a subsequent necrotrophic growth phase

Cuticle and cell wall penetration

Development of hyphae within the
invaded, living epidermal host cell
(biotrophic phase)

Fig. 6: Biotrophic growth phase

Hyphae invasion into mesophyll
layer, followed by epidermal &
mesophyll cell death (necrotic
phase)
Host cell collapse: brought about
by toxin secretion
confined to a single epidermal
host cell that is invaded by a
network of infection hyphae
(Kumar et al., 2003)
Fig. 7: Necrotrophic growth phase characterized by invasion of
mesophyll tissue and host cell death. Dead cells appear decolorized
and black arrow heads shows spread of fungal hyphae (Kumar et al.,
2003)
 Pathogenicity Factors

• Bipolaris toxins
Prehelminthosporol: Most abundant and active compound of B. sorokiniana (Carlson et al.,
1991)
Non-host specific Independent of germination Release starts as soon as conidia
comes in contact with water
1.2-2.1µg/mg dry matter of fungal tissue

Amphiphilic nature Aids in softening the waxy cuticle layer of

the leaves prior to infection

Helminthosporol:
Affects membrane permeability thereby, inhibiting mitochondrial
oxidative phosphorylation, the photophosphorylation in chloroplast and
proton pumping across the plasma membrane (Briquet et al., 1998)
Pathogenicity Factors

• Sorokinianin
Condensation product originating from products of
Nakajima (1994)
the sesquiterpene and TCA pathways

Shows inhibitory activity on seed germination

• Hydrolytic enzymes

Cellulose-hydrolysing enzymes:
Glucosidase and Cellobiohydrolase
Pathogenicity Factors

Fig. 8: The structures predominant phytotoxins of Bipolaris sorokiniana

(A) Prehelminthosporol (B) Helminthosporol (C) Sorokinianin (Kumar et al., 2002)
Pathogenic Variability

Bipolaris sorokiniana is a variable fungus due to somatic hybridization, sexual reproduction,

and heterokaryosis (Tinline, 1962)

Major mechanisms involved in the variability of this fungus include heterokaryosis, which
occurs chiefly through anastomosis, enables the parasexual cycle, which is the main source of
genetic diversity

Genetic variability among naturally occurring population is an outcome of evolution and

demographic process (Zhang, 2017)
Paleogeographic analysis suggests that western part of Central Asia transitioned from a
humid to semi-arid climate

This change is thought to have sped up speciation due to habitat alteration

Pathogenic Variability
Phylogeographic Diversity
2.1 million years ago

B. sorokiniana Bipolaris oryzae

within

1.2 million years ago

Asian isolates

0.47 million years ago

Between Indian & Between Indian &
Indian Isolates American Isolates African Isolates

0.38 million years ago 0.35 million years ago 0.05 million years ago
(Sharma et al., 2022)
Pathogenic Variability
Genetic diversity

1. Heterokaryosis 3. Mutation

4. Sexual reproduction
2. Parasexual cycle

Anastomosis Spontaneous
Heterokaryosis occurrence Cochliobolus sativus
Compatibility
Mitotic crossing-over Ascogonia and Spermagonia
het loci
Haplodization Compatible mating type
Pathogenic Variability
Heterokaryosis
• In B. sorokiniana heterokaryosis arising from anastomosis and nuclear migration
between marked strains was reported by Tinline (1962)
condition perpetuated by less than 10% of the conidia

Buller (1993) was the first to outline the process of anastomosis

Mechanistically, hyphal anastomosis has 3 distinct physiological states of participating hyphae

Pre-contact Positive autotropism, peg initiation

Post-contact Growth arrest after physical contact prior to hyphal fusion

Post-fusion Non-self recognition/ Regeneration

Pathogenic Variability

Genetic Control of Anastomosis

het (heterokaryon incompatibility)/

vic (vegetative incompatibility) loci

Individuals that differ in specificity at one or more

het/ vic loci

Heterokaryotic fusion cells are usually

compartmentalized and undergo death by lytic Fig. 9: Hyphal anastomosis shown by
B. sorokiniana
process

Vegetative Incompatibility/ Somatic Incompatibility

Pathogenic Variability

Parasexual cycle

Heterokaryosis Formation of Occasional mitotic Haplodization

Heterozygous diploids crossing over through aneuploidy
(Pontecarvo & Roper, 1952)

Tinline (1962) obtained occasional heterozygous diploid cultures from heterokaryotic ones

Isolation of recombinants constitutes genetic evidence of somatic heterozygous diploids

and parasexuality in the fungus

Supportive evidence of diploidy was chromosome number, conidial size and nuclear
volume (Huang & Tinline, 1974)
Pathogenic Variability

Mutation

Variation caused by mutation reported by Christensen and Davies (1937)

Tinline (1961)
An anisomycin-resistant strain occurred spontaneously in
a spore-populated medium that contained a
concentration of anisomycin that was inhibitory to the
germination and growth of the parent
Pathogenic Variability

Sexual reproduction
Perfect stage: Cochliobolus sativus

Ascogonia & Spermogonia Initiate Asci with 8 ascospores

reported by Shoemaker (1995) Pseudothecial (filiform, helically coiled)
development

Mating type alleles for sexual

compatibility (Tinline, 1951)
A and a

Despite a seemingly simplicity in the mating type system, mature pseudothecia

do not develop in all compatible pairings

Under natural conditions, the perfect stage was only found in Zambia (Raemaekers, 1988)
Involvement of multiple genes
(Harding & Tinline, 1983) Impaired gene function arrests development
Present Status

Genetic variability reported in Virulence diversity reported in

Brasil (Oliveira et al., 2002) Uruguay (Gamba & Estramill, 2002)

Australia (Meldrum et al., 2004)
India (Saharan et al., 2008)
Syria (Arabi & Jawhar, 2004)
Poland (Saharan et al., 2008)
Canada (Ghazvini & Tekauz, 2007)
Iran ((Zilfaghary et al., 2021)
Pakistan (Asad et al., 2007)
India (Singh et al., 2021)
References

 Arabi, M., & Jawhar, M. (2004). Identification of Cochliobolus sativus (spot blotch) isolates expressing differential
virulence on barley genotypes in Syria. Journal of Phytopathology, 152(8–9), 461–464.
 Asad, S., Iftikhar, S., Munir, A., Ahmad, I., & Ayub, N. (2007). Pathogenic diversity in Bipolaris sorokiniana isolates
collected from different wheat growing areas of the Punjab and NWFP of Pakistan. Pakistan Journal of Botany,
39(6), 2225–2231.
 Bakonyi, J., Aponyi, I., & Fischl, G. (1997). Diseases caused by Bipolaris sorokiniana and Drechslera tritici repentis
in Hungary. Helminthosporium Blights of Wheat: Spot Blotch and Tan Spot, 80–85.
 Baturo-Ciesniewska, A. (2011). Genetic variability and pathogenicity among polish isolates of Bipolaris
sorokiniana from spring barley. Journal of Plant Pathology, 93(2), 291–302.
 Briquet, M., Vilret, D., Goblet, P., Mesa, M., & Eloy, M.-C. (1998). Plant cell membranes as biochemical targets of
the phytotoxin helminthosporol. Journal of Bioenergetics and Biomembranes, 30, 285–295.
 Buller, A.-H. R. (1933). Researches on fungi, Volume V.
 Carlson, H., Nilsson, P., Jansson, H. B., & Odham, G. (1991). Characterization and determination of
prehelminthosporol, a toxin from the plant pathogenic fungus Bipolaris sorokiniana, using liquid
chromatography/mass spectrometry. Journal of Microbiological Methods, 13(4), 259–269.
 Christensen, J. J., & Davies, F. R. (1937). Nature of variation in Helminthosporium sativum. Mycologia, 29(1), 85–
99.
 References
 Duczek, L. J., & Jones-Flory, L. L. (1993). Relationships between common root rot, tillering, and yield loss in spring
wheat and barley. Canadian Journal of Plant Pathology, 15(3), 153–158.
 Gamba, F., & Estramill, E. (2002). Variation in virulence within Uruguayan population of Cochliobolus sativus. 2nd
International Workshop Barley Leaf Blights., 59–62.
 Ghazvini, H., & Tekauz, A. (2007). Virulence diversity in the population of Bipolaris sorokiniana. Plant Disease, 91(7),
814–821.
 Harding, H., & Tinline, R. D. (1983). The existence of differentially fertile strains in two populations of Cochliobolus
sativus. Canadian Journal of Plant Pathology, 5(1), 17–20.
 Huang, H. C., & Tinline, R. D. (1974). Somatic mitosis in haploid and diploid strains of Cochliobolus sativus. Canadian
Journal of Botany, 52(7), 1561–1568.
 Kumar, J., Schäfer, P., Hückelhoven, R., Langen, G., Baltruschat, H., Stein, E., Nagarajan, S., & Kogel, K. H. (2002).
Bipolaris sorokiniana, a cereal pathogen of global concern: Cytological and molecular approaches towards better
control. In Molecular Plant Pathology (Vol. 3, Issue 4, pp. 185–195). John Wiley & Sons, Ltd.
https://doi.org/10.1046/j.1364-3703.2002.00120.x
• Kwasna, H. (1995). Ecology, taxonomy and nomenclature of Helminthosporia-history and actual situation. In J.
Chelkowsi (Ed.), Helminthosporia-Metabolites, Biology, Plant Diseases: Bipolaris, Drechslera, Exserohilum (pp. 27--
60). Institute of Plant Genetics, Polish Academy of Science.
References
 Meldrum, S. I., Ogle, H. J., & Platz, G. J. (2004). Pathotypes of Cochliobolus sativus on barley in Australia.
Australasian Plant Pathology, 33, 190–114.
 Nakajima, H., Isomi, K., Hamasaki, T., & Ichinoe, M. (1994). Sorokinianin: a novel phytotoxin produced by the
phytopathogenic fungus Bipolaris sorokiniana. Tetrahedron Letters, 35(51), 9597–9600.
 Oliveira, A. M. R. de, Matsumura, T. S., Prestes, A. M., & Van Der Sand, S. T. (2002). Intraspecific variability of
Bipolaris sorokiniana isolates determined by random-amplified polymorphic DNA (RAPD). Genet. Mol. Res, 1(4),
350–358.
 Pontecorvo, G., Roper, J. A., Chemmons, L. M., Macdonald, K. D., & Bufton, A. W. J. (1953). The genetics of
Aspergillus nidulans. Advances in Genetics, 5, 141–238.
 Raemaekers, R. H. (1988). Helminthosporium sativum: disease complex on wheat and sources of resistance in
Zambia. Wheat Production Constraints in Tropical Environments. Chiang Mai, Thailand. 19-23 Jan 1987.
 Saharan, M. S., Singh, D. P., Chauhan, P. K., & Karwasara, S. S. (2008). Genetic variability in Bipolaris sorokiniana
isolates causing spot blotch of wheat in India. Indian Phytopathology, 61(2), 268–272.
 Sharma, P., Mishra, S., Singroha, G., Kumar, R. S., Singh, S. K., & Singh, G. P. (2022). Phylogeographic diversity
analysis of Bipolaris sorokiniana (Sacc.) shoemaker causing spot blotch disease in wheat and barley. Genes, 13(12),
2206.
 Sharma, R. C., & Duveiller, E. (2006). Spot blotch continues to cause substantial grain yield reductions under
resource-limited farming conditions. Journal of Phytopathology, 154(7–8), 482–488.
https://doi.org/10.1111/j.1439-0434.2006.01134.x
 References
 Shoemaker, R. A. (1959). Nomenclature of Drechslera and Bipolaris, grass parasites segregated from
‘Helminthosporium.’ Canadian Journal of Botany, 37(5), 879–887.
 Singh, R., Singh, R. V., Mishra, L. K., & Chauhan, A. (2021). Analysis of cultural and pathogenic diversity in Bipolaris
sorokiniana causing spot blotch of bread wheat in North India. International Research Journa of Pure and Applied
Chemistry, 22(4), 12–22.
 Sivanesan, A. (1990). List of sets, index of species, and list of accepted names for some obsolete species names in CMI
Descriptions of Pathogenic Fungi and Bacteria Sets 1--100, Nos. 1--1000, issued January 1964--March 1990.
Mycopathologia, 111(2), 91–108.
 Tinline, R. D. (1951). Studies on the perfect stage of Helminthosporium sativum. Canadian Journal of Botany, 29(1087),
467–480.
 Tinline, R. D. (1961). Cochliobolus sativus: IV. Drug-resistant, color, and nutritionally exacting mutants. Canadian Journal
of Botany, 39(7), 1695–1704.
 Tinline, R. D. (1962). Cochliobolus sativus: V. heterokaryosis and parasexuality. Canadian Journal of Botany, 40(3), 425–
437.
 Tinline, R. D., & Dickson, J. G. (1958). Cochliobolus sativus. I. Perithecial development and the inheritance of spore color
and mating type. Mycologia, 50(5), 697–706.
 Zhang, W., Manawasinghe, I. S., Zhao, W., Xu, J., Brooks, S., Zhao, X., Hyde, K. D., Chethana, K. W. T., Liu, J., Li, X., &
others. (2017). Multiple gene genealogy reveals high genetic diversity and evidence for multiple origins of Chinese
Plasmopara viticola population. Scientific Reports, 7(1), 17304.
Any Queries

Thank you