Photoacoustic Sensing As A Non-Destructive Optical Tool To Detect Seed-Borne Phytopathogens

In a context where global food demand is continuously rising, the need for innovative, highly specific, sensitive, and low cost methods for the early detection of seed-borne phytopathogens is a key step for growing healthy and safe crops. In addition to the economic losses resulted from a reduced production, seed-borne pathogens may easily spread worldwide by means of infected seeds1, that are often asymptomatic. Here we focus on the development of a new optical detection method for the bacterial plant pathogen C. flaccumfaciens pv. flaccumfaciens (Cff) (Hedges) Collins&Jones. Cff is a Gram positive, aerobic, nonspore-forming bacterium, and one of the most important biotic constraints affecting the production of edible legumes all over the world. As no effective chemicals against this bacterium are known, the development of specific and sensitive diagnostic tests for the rapid detection of Cff on plant materials is fundamental to prevent disease outbreaks. Currently, the available methods for Cff detection are the visual inspection of suspected seed lots or molecular PCR-based methods applied also on-site, such as "loop-mediated isothermal amplification" (LAMP) method. The last are unable to discriminate a viable from a not-viable inoculum and are based on destructive procedures, besides to be time consuming, laborious, expensive, and difficult to automate. For the above requirements, non-destructive methods such as optical techniques are of great interest, but nowadays the few examples of optical technologies employed for plant diseases diagnosis are mainly based on Vis-NIR reflectance spectroscopy which is limited to superficial features of the sample, especially in the visible region. Sensing techniques based on photoacoustic effect, which combine spectroscopic specificity with the deep penetration of ultrasound in biological tissues, hold the potential to overcome the main limits of pure optical techniques. The photoacoustic principle rests on the use of short optical pulses to excite specific chromophores to trigger a cascade of consecutive events that include the steps of optical absorbance, photothermal conversion and heat propagation, and thermoelastic conversion and acoustic propagation. As a consequence, photoacoustic sensing is an hybrid technique which combines the best advantages of optical contrast and acoustic detection. Photoacoustic imaging and sensing have been mainly exploited in biomedical applications. Conversely, the huge potential of such technologies in the agri-food sector as sensitive, precise and low cost diagnostic techniques is still unexplored. Here, we propose the use of photoacoustic sensing for the non-destructive detection of the quarantine bacterial phytopathogen Cff, by exploiting the optical absorbance of inner specific chromophores (likely carotenoids) it produces. We demonstrated the feasibility to optically detect in vitro grown Cff colonies, by selecting the most suitable excitation wavelengths. These results may open the way to innovative applications of photoacoustic sensing to non-destructively assess health and quality of plants and seeds.