Ravi Chitrampalam Research Plant Pathologist, Eurofins BioDiagnostics

It’s arguably a company’s biggest headache, not to mention costly: having seed rejected at the port of entry because officials say it tests positive for a pathogen of concern. This may not be the fault of seed testing officials, but rather their reliance on a singular testing method that can’t differentiate live target cells from the presence of dead ones, often associated with a seed treatment.

Before the advent of molecular technology, seed health testing was solely dependent on traditional methods such as freeze-blotter assay, plating on semi-selective media followed by close observation under the microscope for target-specific characteristics and pathogenicity testing, among others. Any subsequent tests required could take an additional two weeks after plating.

Needless to say, these methods are time consuming and labor intensive, and more importantly require taxonomic expertise to confirm the target of concern at the species level. Furthermore, some pathogens are not capable of being artificially cultivated, which adds to the challenges associated with traditional seed health testing methods.

With the adoption of molecular technologies, most seed testing laboratories have overcome these limitations, as these methods more precisely identify the target of concern, including the unculturable ones. Currently, target specific PCR- (polymerase chain reaction) and qPCR-based methods are used to screen for seed-borne pathogens. Speed and accuracy make molecular diagnostics more attractive, not to mention no longer needing taxonomic expertise.

However, like any other method, molecular technologies also have their challenges. The major deficiency of this method is that live targets cannot be differentiated from the dead ones. Additionally, every target cell is amplified regardless of their metabolic viability, which is a great disadvantage. As a result, seed lots that are treated with disinfectants and shown to be negative for the active target cells by bioassay will test positive using molecular methods. As such, molecular methods are highly recommended as a prescreening method rather than a stand-alone method.

Unfortunately some countries use molecular methods as the stand-alone method to identify the targets in seed, meaning seed lots are sometimes rejected at the port of entry even though they are treated and shown to be negative for the active target by a third-party seed testing lab. This is a costly challenge for those in the seed industry.

I’m excited to say that good news is on the horizon. The recent advances in molecular technology might solve this problem. With the help of DNA-binding dye propidium monoazide (PMA), we can differentiate live targets from dead ones. PMA works by penetrating the dead cells and inhibiting their amplification.

While this technology is very promising, it has not yet been exploited by seed testing laboratories. At Eurofins BioDiagnostics, we’ve initiated a research team to explore the possibility of incorporating PMA components into the standard molecular screening methods to precisely identify the active target in seeds and better serve those in the seed industry.