Propidium iodide enabled live imaging of Pasteuria sp.- Pratylenchus zeae infection studies under fluorescence microscop
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ORIGINAL ARTICLE
Propidium iodide enabled live imaging of Pasteuria sp.-Pratylenchus zeae infection studies under fluorescence microscopy Francine Perrine-Walker 1,2
&
Khoa Le 1,2
Received: 30 June 2020 / Accepted: 7 October 2020 # Springer-Verlag GmbH Austria, part of Springer Nature 2020
Abstract Live imaging allows observations of cell structures and processes in real time, to monitor dynamic changes within living organisms compared to fixed organisms. Fluorescence microscopy was used to monitor the dynamic infection process of the nematode parasitic bacterium Pasteuria sp. and the sugarcane root-lesion nematode, Pratylenchus zeae. Under fluorescence microscopy, green-autofluorescent globules were observed in live control and Pasteuria sp.-infected nematodes. Only nematodes killed by Pasteuria sp. or heat treated displayed a diffuse pattern of autofluorescence. Propidium iodide (PI), used as a cell membrane integrity indicator, confirmed that the nematode’s cuticle acts as an impermeable barrier. PI stained cells/DNA of heattreated control and Pasteuria sp.-infected P. zeae. PI as a counterstain facilitated the location of Pasteuria endospores on the cuticle surface of P. zeae. No PI staining was observed in sporangia and in endospores within the nematode body. However, PI specifically stained endospores on the cuticle surface and within the cuticle carcass showing, in mature propagules, a ring-like pattern. Live imaging, combined with fluorescence microscopy and fluorescent dyes such as PI, appears useful in live studies on plant nematode interactions with nematophagous bacteria. Keywords Root lesion nematode . Endospore . Infection . Nematode-bacteria interaction . Plant pathogen
Introduction The genus Pasteuria consists of Gram-positive, thallus and endospore-forming bacteria that parasitize invertebrates such as nematodes and small aquatic crustaceans (commonly known as water fleas) in a host-specific manner (Ciancio 2018; Mohan et al. 2011; Overholt et al. 2020;
Handling Editor: Handling Editor: Georg Krohne Electronic supplementary material The online version of this article (https://doi.org/10.1007/s00709-020-01567-0) contains supplementary material, which is available to authorized users. * Francine Perrine-Walker [email protected] 1
School of Life and Environmental Sciences, Life Earth and Environmental Sciences Building (F22), The University of Sydney, Sydney, NSW 2006, Australia
2
Institute of Agriculture, Biomedical Building (C81), The University of Sydney, 1 Central Avenue, Australian Technology Park, Eveleigh, NSW 2015, Australia
Stirling 2014). While Pasteuria can infect more than 300 nematode taxa (Chen and Dickson 1998), most studies have been focused on species capable of parasitizing agriculturally important nematode pests within the Tylenchida family (Ciancio 2018; Mohan et al. 2012; Öztürk et al. 2020; Stirling 1984, 1985, 2008, 2014; Stirling et al. 2017; Sturhan et al. 2005). In addition, the ability of Pasteuria to inhibit host reproduction and to produce endos
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