Dataset Information

Agar plate-based screening methods for the identification of polyester hydrolysis by Pseudomonas species.

ABSTRACT: Hydrolases acting on polyesters like cutin, polycaprolactone or polyethylene terephthalate (PET) are of interest for several biotechnological applications like waste treatment, biocatalysis and sustainable polymer modifications. Recent studies suggest that a large variety of such enzymes are still to be identified and explored in a variety of microorganisms, including bacteria of the genus Pseudomonas. For activity-based screening, methods have been established using agar plates which contain nanoparticles of polycaprolactone or PET prepared by solvent precipitation and evaporation. In this protocol article, we describe a straightforward agar plate-based method using emulsifiable artificial polyesters as substrates, namely Impranil® DLN and liquid polycaprolactone diol (PLD). Thereby, the currently quite narrow set of screening substrates is expanded. We also suggest optional pre-screening with short-chain and middle-chain-length triglycerides as substrates to identify enzymes with lipolytic activity to be further tested for polyesterase activity. We applied these assays to experimentally demonstrate polyesterase activity in bacteria from the P. pertucinogena lineage originating from contaminated soils and diverse marine habitats.

SUBMITTER: Molitor R

PROVIDER: S-EPMC6922526 | biostudies-literature | 2020 Jan

REPOSITORIES: biostudies-literature

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Agar plate-based screening methods for the identification of polyester hydrolysis by Pseudomonas species.

Molitor Rebecka R Bollinger Alexander A Kubicki Sonja S Loeschcke Anita A Jaeger Karl-Erich KE Thies Stephan S

Microbial biotechnology 20190423 1

Hydrolases acting on polyesters like cutin, polycaprolactone or polyethylene terephthalate (PET) are of interest for several biotechnological applications like waste treatment, biocatalysis and sustainable polymer modifications. Recent studies suggest that a large variety of such enzymes are still to be identified and explored in a variety of microorganisms, including bacteria of the genus Pseudomonas. For activity-based screening, methods have been established using agar plates which contain na ...[more]

PMID: 31016871

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Project description:As a bioaerosol sampling standard, Andersen type impactor is widely used since its invention in 1950s, including the investigation of the anthrax attacks in the United States in 2001. However, its related problems such as impaction and desiccation stress as well as particle bounce have not been solved. Here, we improved its biological collection efficiencies by plating a mineral oil layer (100 µL) onto the agar plate. An Andersen six-stage sampler and a BioStage impactor were tested with mineral-oil-spread agar plates in collecting indoor and outdoor bacterial and fungal aerosols. The effects of sampling times (5, 10 and 20 min) were also studied using the BioStage impactor when sampling environmental bioaerosols as well as aerosolized Bacillus subtilis (G+) and Escherichia coli (G-). In addition, particle bounce reduction by mineral-oil-plate was also investigated using an optical particle counter (OPC). Experimental results revealed that use of mineral-oil-spread agar plate can substantially enhance culturable bioaerosol recoveries by Andersen type impactors (p-values<0.05). The recovery enhancement was shown to depend on bioaerosol size, type, sampling time and environment. In general, more enhancements (extra 20%) were observed for last stage of the Andersen six-stage samplers compared to the BioStage impactor for 10 min sampling. When sampling aerosolized B. subtilis, E. coli and environmental aerosols, the enhancement was shown to increase with increasing sampling time, ranging from 50% increase at 5 min to ?100% at 20 min. OPC results indicated that use of mineral oil can effectively reduce the particle bounce with an average of 66% for 10 min sampling. Our work suggests that enhancements for fungal aerosols were primarily attributed to the reduced impaction stress, while for bacterial aerosols reduced impaction, desiccation and particle bounce played major roles. The developed technology can readily enhance the agar-based techniques including those high volume portable samplers for bioaerosol monitoring.

Dataset Information

Agar plate-based screening methods for the identification of polyester hydrolysis by Pseudomonas species.

Publications

Agar plate-based screening methods for the identification of polyester hydrolysis by Pseudomonas species.

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