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Effects of a fungal invasion on soil bacteria Flavia Pinzari; Anne D. Jungblut; Clark M.D.; Misra R.; Xu X.-M.; Chooneea D. Presentation by F.Pinzari at The World Congress of Soil Science 2022, which took place in Glasgow from 31st July - 5th August 2022 Pinzari F.1,2, Clark M.D.1, Misra R. 3, Chooneea D.3, Xu X.-M.4, Jungblut A.D.1 1Life Sciences Department, Natural History Museum, Cromwell Road, SW7 5BD London, UK 2Institute for Biological Systems, Council of National Research of Italy (CNR), Monterotondo (RM), Italy 3Core Research Laboratories, Molecular Biology, Natural History Museum, London, United Kingdom 4National Institute of Agricultural Botany, East Malling Research Station (EMR), East Malling, UK Fungal bioinoculants have a vast potential in agriculture because they can help increase crop yields and quality and reduce the application of chemicals. Their effectiveness has been widely tested (Malusà et al., 2016). However, little is known about the effect of bioinoculants on microbial assemblages in non-rhizospheric soil. A sudden artificial introduction of a fungal species in soil could theoretically impact the biodiversity of local microbial communities and lead to changes in nutrient availability (van Elsas et al., 2012). We assessed the impact of a competitive fungal inoculum, the globally-used biofertiliser Trichoderma afroharzianum T22, on soil microcosms to understand 1) to what extent the native microbial community richness and relative abundance are influenced by a fungal strain introduced to soil; 2) whether microbial taxa are resilient to the disturbance caused by the fungus; 3) how far the bioinoculant impacts the soil microorganisms functions. We used bacterial 16S rRNA gene amplicon sequencing (Illumina) and a shotgun metagenomic analysis (Oxford Nanopore Sequencing) to analyse the microbial communities in bioreactors after seven weeks of incubation with and without the fungus. The presence of the fungus had a negative impact on the abundance of some groups of bacteria, such as the genus Pseudomonas, and it stimulated the presence of species metabolically linked to the fungus, including chitin degrading Chitinophagaceae. In conclusion, the results suggest that more than an impact on bacteria's overall biodiversity, the fungus has favoured some groups at the expense of others, even creating new food webs and trophic niches. References Malusà E, Pinzari F, Canfora L (2016) Efficacy of Biofertilizers: Challenges to Improve Crop Production. In: D.P. Singh et al. (eds.), Microbial Inoculants in Sustainable Agricultural Productivity: Microbial Inoculants in Sustainable Agricultural Productivity, pp.17-40 Springer India doi.org/10.1007/978-81-322-2644-4_2 van Elsas JD, Chiurazzi M, Mallon CA, Elhottova D, Kristufek V, Salles JF. (2012) Microbial diversity determines the invasion of soil by a bacterial pathogen. Proc Natl Acad Sci U S A. 24;109(4):1159-64. doi: 10.1073/pnas.1109326109. Acknowledgements The NHM-affiliated author Flavia Pinzari was funded by the H2020-MSCA-IF-EF-SE project “AlienInSoil”. This project has received funding from the European Union’s Horizon 2020 research and innovation programme under the Marie Sklodowska-Curie grant agreement No 892048., The NHM-affiliated author Flavia Pinzari was funded by the H2020-MSCA-IF-EF-SE project "AlienInSoil". This project has received funding from the European Union's Horizon 2020 research and innovation programme under the Marie Sklodowska-Curie grant agreement No 892048., {"references":["Malusà E, Pinzari F, Canfora L (2016) Efficacy of Biofertilizers: Challenges to Improve Crop Production. In: D.P. Singh et al. (eds.), Microbial Inoculants in Sustainable Agricultural Productivity: Microbial Inoculants in Sustainable Agricultural Productivity, pp.17-40 Springer India doi.org/10.1007/978-81-322-2644-4_2","van Elsas JD, Chiurazzi M, Mallon CA, Elhottova D, Kristufek V, Salles JF. (2012) Microbial diversity determines the invasion of soil by a bacterial pathogen. Proc Natl Acad Sci U S A. 24;109(4):1159-64. doi: 10.1073/pnas.1109326109."]}

Aims The increasingly widespread use of beneficial microbial inocula in agriculture gives rise to two primary needs: i) the assessment of the environmental risk, i.e. their impact on local soil microbiome and soil properties; ii) being able to track them and monitor their persistence and fate to both optimize their formulation and application method. In previous years, PCR-based methods have detected bacterial or fungal bioinoculant at the species or strain level. However, the selective detection, quantification, and monitoring of target microbial species in a complex ecosystem such as soil require that the tests possess high specificity and sensitivity. Methods and results The work proposes a quantitative real-time PCR detection method using TaqMan chemistry, showing high specificity and sensitivity for thePaenibacillus polymyxaK16 strain. The primer and probe sets were designed using the polymyxin gene cluster targetingpmxCandpmxEsequences. Validation tests showed that these assays allowed a discriminant and specific detection ofP. polymyxa K16in soil. Conclusion The TaqMan-assay developed could thus ensure the necessary level of discrimination required by commercial and regulatory purposes to detect and monitor the bioinoculant in soil.

The interface between biological and geochemical components in surface crust of a saline soil was investigated using X-Ray Diffraction (XRD), and variable pressure scanning electron microscopy (SEM) in combination with Energy Dispersive X-ray Spectrometry (EDS). Mineral compounds such as halite and gypsum were identified crystallized around filaments of cyanobacteria. A total of 92 genera were identified from the bacterial community based on 16S gene pyrosequencing analysis. The occurrence of the gypsum crystals, their shapes and compartmentalization suggested that they separated NaCl from the immediate microenvironment of the cyanobacteria, and that some cyanobacteria and communities of sulfur bacteria may had a physical control over the distinctive halite and gypsum structures produced. This suggests that cyanobacteria might directly or indirectly promote the formation of a protective envelope made of calcium and sulfur-based compounds.