Observations on the Early Establishment of Foliar Endophytic Fungi in Leaf Discs and Living Leaves of a Model Woody Angiosperm, Populus trichocarpa (Salicaceae).

Yu-Ling Huang,A Arnold,Naupaka Zimmerman

doi:10.3390/jof4020058

Yu-Ling Huang, A Arnold + Show 1 more

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https://doi.org/10.3390/jof4020058

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Abstract

Fungal endophytes are diverse and widespread symbionts that occur in the living tissues of all lineages of plants without causing evidence of disease. Culture-based and culture-free studies indicate that they often are abundant in the leaves of woody angiosperms, but only a few studies have visualized endophytic fungi in leaf tissues, and the process through which most endophytes colonize leaves has not been studied thoroughly. We inoculated leaf discs and the living leaves of a model woody angiosperm, Populus trichocarpa, which has endophytes that represent three distantly-related genera (Cladosporium, Penicillium, and Trichoderma). We used scanning electron microscopy and light microscopy to evaluate the timeline and processes by which they colonize leaf tissue. Under laboratory conditions with high humidity, conidia germinated on leaf discs to yield hyphae that grew epiphytically and incidentally entered stomata, but did not grow in a directed fashion toward stomatal openings. No cuticular penetration was observed. The endophytes readily colonized the interiors of leaf discs that were detached from living leaves, and could be visualized within discs with light microscopy. Although they were difficult to visualize within the interior of living leaves following in vivo inoculations, standard methods for isolating foliar endophytes confirmed their presence.

Highlights

Plants in natural and agricultural ecosystems worldwide harbor fungal endophytes in their symptomless tissues, including apparently healthy leaves [1,2]
From observations with Scanning electron microscopy (SEM), we found that leaf discs inoculated with Cladosporium and Penicillium had fungal growth at 14 days after inoculation; as a result, we focused on these two fungi for the sectioning work
Most of the stomata were closed in the SEM samples, but most were open in the light microscopy (LM) samples

Summary

Introduction

Plants in natural and agricultural ecosystems worldwide harbor fungal endophytes in their symptomless tissues, including apparently healthy leaves [1,2]. These endophytes often influence aspects of leaf physiology, gene expression, and function [3,4,5]. Their prevalence and importance prompt inquiries into the processes by which endophytic symbioses establish in living foliage. These endophytes have been found in all of the plant species that have been surveyed far, and are highly diverse in wild and

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