Evaluation of inoculum potential of pathogens in seeds: relation to physiological quality and DNA quantification by qPCR

Abstract

-Given what is already known in regard to seed health and the availability of molecular methods for detection of the pathogens Stenocarpella maydis and Stenocarpella macrospora in maize seeds, Colletotrichum gossypii var. cephalosporioides in cotton seeds, and Corynespora cassiicola in soybean seeds, the aim of this study was to evaluate seed vigor according to different inoculum potentials. The fungus isolates were inoculated on seeds by the technique of water restriction, through which different inoculum potentials are obtained, corresponding to times of seed exposure of 0, 24, 48, and 96 hours for maize and cotton seeds, and 0, 36, 108, and 144 hours for soybean seeds. The seeds were subjected to germination, electrical conductivity, health, and qPCR tests. Results of the blotter test showed that in most pathosystems, there was a higher incidence of the fungi with an increase in inoculum potential. A decrease in germination percentage was observed in all species as inoculum potential increased, as well as further degradation of seed membranes. The qPCR test confirmed that the most damaged seeds in the tests had higher presence of the pathogens.Index terms: seed pathology, fungi, maize, soybean, cotton.Avaliação do potencial de inóculo de patógenos em sementes: sua relação com a qualidade fisiológica e quantificação do DNA por qPCR RESUMO -Diante do que já se conhece em sanidade de sementes e tendo-se em mãos métodos moleculares para a detecção dos fungos Stenocarpella maydis, Stenocarpella macrospora em sementes de milho, Colletotrichum gossypii var. cephalosporioides em algodão e Corynespora cassiicola em soja, objetivou-se neste estudo avaliar o vigor das sementes em função dos diferentes potenciais de inóculo. Os isolados dos fungos foram inoculados nas sementes por meio da técnica de restrição hídrica pela qual se obtém diferentes potenciais de inóculo, que foram representados por P0, P24, P48 e P96 e P0, P36, P108 e P144, que correspondem à exposição das sementes aos períodos de tempo de 0, 24, 48, 96 horas (milho e algodão) e nos tempos 0, 36, 108 e 144 horas (soja), respectivamente. As sementes foram submetidas aos testes de germinação, condutividade elétrica, sanidade e qPCR. Pelo blotter test, na maioria dos patossistemas, houve maior incidência do fungo com o aumento do potencial de inóculo. Foi observada uma queda na porcentagem de germinação de todas as espécies com o aumento do potencial de inóculo, assim como maior degradação das membranas das sementes. A qPCR confirmou que nas sementes mais prejudicadas havia maior quantidade de inóculo dos patógenos.Termos para indexação: patologia de sementes, fungos, milho, soja, algodão.

Molecular detection of Colletotrichum lindemuthianum in bean seed samples

“…Such result reveals that the longer the exposure time of the seeds to the pathogen the higher the fungal capacity to penetrate and colonize tissues of the seeds. Authors who worked with molecular detection of other fungi in different hosts also observed the same correlation, when using the qPCR technique (Botelho et al, 2015;Guimarães et al, 2017;Vanegas-Berrouet et al, 2014;Sousa et al, 2015).…”

Effect of tear/crack on soybean (Glycine max) seed coat, physiological quality and pathology of the seed

“…Aspergillus is an important storage fungus. Guimarães et al (2017) found that damage caused by pathogens may be directly related to the potential of inoculum and location in the seed (internal or external), with a greater concern if this location is close to the embryo. Even if the fungi do not cause immediate damage, there are latency relationships, which are cumulative during storage.…”

qPCR-based detection of Colletotrichum truncatum in soybean seeds

“…Therefore, accurate, sensitive, and fast diagnostic tools are needed to support seed phytosanitary certification. Species-specific primers in PCR assays can be used in seed certification program not only for presence/ absence result but also to provide a quantitative measure of the pathogens in seeds, other plant parts, and environmental samples (Glynn and Edwards 2010;Botelho et al 2015, Guimarães et al 2017. A PCR assay has been reported for the detection of C. truncatum (Chen et al 2006).…”

“…We developed a qPCR protocol for the specific detection of C. truncatum in soybean seeds; this assay can be a powerful tool to diagnostic laboratories conducting seed phytosanitary certification. Quantitative PCR has been an important tool for pathogen detection in seeds involved in many pathosystems because of its precision, specificity (Guimarães et al 2017), and sensitivity (Barrocas et al 2012). The use of infected soybean seeds may introduce the pathogen into soybeanproducing regions because the seeds are vehicles for the spread of this pathogen and are an important source of primary inoculum (Elmer 2001).…”