Tracing the Origins of Agricultural Products with Barcoded Microbial Spores

The globalization of agricultural trade has greatly improved our access to cereals, vegetables, and fruits. However, complicated trade routes and supply chains make it challenging to trace the origin of agricultural products. This is particularly problematic when the source of food contamination cannot be determined with the currently available methods. For example, a recent recall of contaminated romaine lettuce from North American shelves caused significant economic losses and raised public concerns on food safety before a lengthy investigation pinpointed the origin of the outbreak. A recent study by Springer's team from the Harvard Medical School has described the so-called Barcoded Microbial Spores (BMS) method, which enables safe, rapid, and high-resolution tracing of the origins of agricultural products (Qian et al., 2020).

Dormant microbial spores are renowned for their extreme resistance to harsh conditions and their ability to survive in natural environments for many decades. Those tiny and durable structures have been utilized by Springer's team to engineer BMS for the purpose of tracing object provenance. To develop BMS, tandem DNA barcodes composed of custom-designed, non-redundant DNA sequences were transformed into the genomes of a common spore-forming bacterium (Bacillus subtilis) and fungus (Saccharomyces cerevisiae). To minimize risk to the environment, the engineered BMS organisms were impaired in their germination and growth under natural conditions either by removing genes essential for germination (B. subtilis) or by boiling (S. cerevisiae).

After these barcoded spores were inoculated onto various surfaces of interest, origin and contact tracing was conducted using a Cas13a-based RNA detection technique known as SHERLOCK (Gootenberg et al., 2017) (Figure 1 ). Notably, the SHERLOCK assay has been used in testing for Zika and SARS-CoV-2 (causing COVID-19) viruses. By combining the SHERLOCK assay with a highly efficient spore lysis procedure, the authors could detect BMS with single-molecule sensitivity, and by pairing with SANGER sequencing, could correctly pinpoint the origins of inoculated leafy plant samples from a mixture without cross contamination (Figure 1).

Figure 1.

Schematic Illustration of the Entire Procedure of BMS-based Tracing Technology, Including BMS Labeling of Leafy Vegetables and the SHERLOCK Assay.

RPA, recombinase polymerase amplification. Note that Cas13a non-specifically cuts near RNA (reporter RNA here) upon activation by its RNA target.

BMS technology holds great potential in the precise tracing of agricultural products and thus the sources of foodborne illnesses. The use of the custom-designed 28-bp Cas13a target as the barcode enables the specificity of BMS sequence to have near-unlimited variation. Moreover, BMS can persist on sand, soil, carpet, and wood for months, independent of weather conditions, and were detectable on food products, even after washing, boiling, microwaving, or frying. Critically, the isothermal nature of the SHERLOCK assay enables paper spot in-field testing, and primary presence/absence screening can be conducted using a mobile phone, thereby enabling high-throughput field screening followed by the laboratory identification of the specific BMS. Further studies are needed to trace BMS on diverse food and agricultural fields to determine wide reliability of BMS technology.