A whole-cell-bacterial biosensor for phosphonates allows quantification of glyphosate-based herbicides removal by the highly efficient glyphosate degrader Achromobacter xylosoxidans

MASOTTI, F.; MUÑOZ. A; KRINK, N.; LENCINA, N.; GOTTIG, N.; GARAVAGLIA, B.S.; NIKEL, P.; JORGELINA OTTADO

Congreso; LIX Reunion Anual SAIB; 2023

Glyphosate-based herbicides (GBH) are widely distributed in most of the economically productive lands in which crop production is mainly based on glyphosate-resistant genetically modified plants. Our country possesses one of the most fertile and favorable agricultural land in the world and its crop management is based on these genetically modified crops resistant to glyphosate. Nevertheless, the use of glyphosate has been considered less harmful compared to other pesticides and as consequence of its extensive use, its presence is widespread in different environments. Therefore, it is essential to ensure the detection of GBH in environmental samples to identify polluted hotspots where bioremediation processes might be required. For this purpose, synthetic biology tools were applied in order to obtain a simple, economic and reliable tool as an alternative to the traditional ones based on spectroscopy and chromatography methods and was directly applied to detect remnant glyphosate residues in laboratory samples treated with environmental bacterial isolates. For doing so, previously we have isolated an Agrobacterium tumefaciens CHLDO with ability to degrade glyphosate and that bears a C-P lyase complex codified in a phn cluster, involved in the hydrolysis of the C-P bond present in phosphonates. The transcriptional analysis of phn genes revealed that glyphosate could be performed by means of this C-P lyase pathway. Then, we have constructed a whole-cell-bacterial biosensor using the promoter of the C-P lyase complex from A. tumefaciens CHLDO (PhnG) fused to the fluorescent protein Scarlet in the environmental A. tumefaciens CHLDO chassis. Further, the biosensor was optimized by including an extra copy of phnF, coding for the transcriptional repressor of the cluster. By this Methods, biosensor leakage in the absence of inductor was diminished and the dynamic range, which reveals the fold change between the induced and non-induced state, obtained was acceptable. Here, we used this biosensor to monitor the degradation ability of a previously isolated bacterial strain from soils belonging to fields with repetitious applications of GBH. This strain, Achromobacter xylosoxidans SOS3, belongs to a genus that has been characterized as an excellent glyphosate degrader since it has been described that an Achromobacter sp. remediated the herbicide in soil samples 2–3 folds faster than endogenous microorganisms. By means of the biosensor we could measure glyphosate degradation in cultures of A. xylosoxidans SOS3 grown in minimal medium lacking phosphates and in the presence of 1.5 mM glyphosate. At 3 days of culture, A. xylosoxidans SOS3 was able to degrade all the glyphosate present, highlighting the importance of studying bacterial molecular mechanisms that can be translated to develop useful tools for the analysis of pollutants.