Development of a Simple Hydroponic Assay to Study Vertical and Horizontal Resistance of Soybean and Pathotypes of Phytophthora sojae

Abstract

Phytophthora root rot, caused by Phytophthora sojae, is one of the most damaging diseases of soybean and the introgression of Rps (Resistance to P. sojae) genes into elite soybean lines is arguably the best way to manage this disease. Current bioassays to phenotype the gene-for-gene relationship are hampered with respect to reproducibility and long-term stability of isolates, and do not accurately predict horizontal resistance individually. The aim of our study was to investigate a new way of phenotyping P. sojae isolates and vertical and horizontal resistance in soybean that relies on zoospores inoculated directly into a hydroponic system. Inoculation of P. sojae isolates against a set of eight differentials accurately and reproducibly identified pathotypes over a period of two years. When applied to test vertical resistance of soybean lines with known and unknown Rps genes, the bioassay relied on plant dry weight to correctly identify all genes. In addition, simultaneous inoculations of three P. sojae isolates, collectively carrying eight major virulence factors against 64 soybean lines with known and unknown levels of horizontal resistance, separated the plants into five distinct groups of root rot, allowing the discrimination of lines with various degrees of partial resistance. Based on those results, this bioassay offers several advantages in facilitating efforts in breeding soybean for P. sojae resistance and in identifying virulence factors in P. sojae.

Discriminant haplotypes of avirulence genes of Phytophthora sojae lead to a molecular assay to predict phenotypes

“…Indeed, genetic control of P. sojae through introgression of Rps genes in soybean presupposes a thorough understanding and knowledge of the pathotype diversity of P. sojae isolates in the specific area where the Rps genes are deployed. Up to this point, the only way to determine the pathotype of a given isolate was through cumbersome and long phenotyping procedures, each with its own shortcomings (Dorrance et al ., ; Lebreton et al ., ). Through this unique molecular assay, based on discriminant haplotypes for seven Avr genes of P. sojae , it should now be possible to obtain a rapid and accurate identification of the virulence profile of isolates in order to precisely select soybean material carrying the appropriate Rps genes.…”

“…Indeed, genetic control of P. sojae through introgression of Rps genes in soybean presupposes a thorough understanding and knowledge of the pathotype diversity of P. sojae isolates in the specific area where the Rps genes are deployed. Up to this point, the only way to determine the pathotype of a given isolate was through cumbersome and long phenotyping procedures, each with its own shortcomings (Dorrance et al, 2008;Lebreton et al, 2018). Through this unique molecular assay, based on discriminant Since the intentional deployment of the first Rps gene in soybean, Rps1a, P. sojae has demonstrated a very strong resilience and ability to adapt to selection pressure through rapid mutations of Avr genes (Keeling, 1984;Layton et al, 1986;Drenth et al, 1996;Kaitany et al, 2001).…”

“…Subsequently, 25 isolates collected in the Canadian provinces of Quebec and Ontario between 2010 and 2018 were tested with the multiplex PCR and phenotyped using the hydroponic assay developed by Lebreton et al . (). For the hydroponic assay, zoospores were inoculated into a hydroponic system containing a nutrient solution diluted in water.…”

“…Once the multiplex PCR conditions were optimized, the 31 isolates with known haplotypes, previously sequenced by Arsenault- et al (2018), were analysed to test the efficiency of the molecular assay. Subsequently, 25 isolates collected in the Canadian provinces of Quebec and Ontario between 2010 and 2018 were tested with the multiplex PCR and phenotyped using the hydroponic assay developed by Lebreton et al (2018). For the hydroponic assay, zoospores were inoculated into a hydroponic system containing a nutrient solution diluted in water.…”

“…However, problems with false positives, intermediate responses, and stability of Avr genes/expression over time have hampered its reliability. More recently, Lebreton et al (2018) proposed a new assay to phenotype the isolates of P. sojae that relied on zoospore inoculation in hydroponic solutions to more closely reproduce the natural course of infection. While this bioassay appears to eliminate some of the shortcomings of the hypocotyl assay, it remains labour-intensive and requires several weeks to complete.…”