Direct and indirect effects of fungi and oomycetes on leaf litter degradation by freshwater macroinvertebrates

Abstract

The decomposition processes in lakes are documented in few reports only, and it is therefore important to improve our understanding of the process of leaf conditioning and its effects on the benthic community in lakes. In the present thesis I investigated how the identity of microbial colonisers affects the consumption by macroinvertebrates organisms were examined. For the studies presented here the shredder Gammarus roeselii and small-particle-feeder Limnomysis benedeni, a recent invader of Lake Constance were chosen because of their numerical importance in the littoral benthic community of Lake Constance. Here two experiments were performed, in which alder leaves were exposed in the littoral of Lake Constance. Regular leaf subsamples were analysed for chemical and physical leaf parameters, and the consumption rates of G. roeselii were determined in laboratory food choice assays with autoclaved and leached alder leaves as additional food items. In addition to leaf toughness the bulk leaf parameters N-, C-, P-, protein- and polyphenol content were measured and the ergosterol content was determined by HPLC. Consumption rates of littoral exposed leaves were statistically analysed for effects of leaf parameters using permutation based tests and a linear model approach. In both experiments leaf parameters changed and consumption by G. roeselii increased significantly with conditioning in the littoral. The negative correlation of polyphenols with shredder feeding corroborated the known repellence by polyphenols. Notwithstanding studies by others the N- and the protein content decreased over the first time of exposure in the littoral, which suggested that leaf colonising micro-organisms (fungi and oomycetes) could not compensate for leaching of N-containing constituents. In both exposition experiments increasing ergosterol content over exposition time pointed at increasing metabolically active eumycotic fungal biomass on the littoral-exposed leaves, and therefore aquatic fungi and oomycetes were isolated from leaves.The fungal and oomycete isolates were cultured and identified based on the sequence of internal transcribed spacer (ITS) regions of rDNA, and the sequences obtained during these studies were deposits into GenBank®. In freshwaters, fungi are regarded as the most important microbial component on decaying leaves, and here 9 ascomycete and 4 oomycete strains were isolated from conditioned leaf litter. Here, for the first time, oomycetes were demonstrated to affect leaf parameters to a similar extent as other fungal strains and to positively affect consumption by Gammarus, which suggest that oomycetes have a greater impact on leaf litter decomposition in freshwaters than hitherto assumed. In order to experimentally separate effects of leaching and colonisation by fungi or oomycetes, experiments with single isolates growing on autoclaved leaves were performed, in which effects of single strains on leaf parameters and on consumption by G. roeselii were assessed. The majority of the different fungi and oomycete isolates on leaves were significantly preferred over controls, and consumption rates by G. roeselii proved to be strain-specific. The leaf parameters N, C, P, protein and polyphenol were affected by colonisation with single isolates, and the magnitude of the effects was strongly strain-specific. Statistical analysis with a linear model revealed that polyphenol and protein levels were major determinants of the consumption rate of Gammarus, suggesting that fungi and oomycetes might indirectly steer consumption by altering the leaf litter content of protein and polyphenols, in particular during later stages of conditioning in the field. Shredders discriminate between leaves colonised by different fungal and oomycete species, but mechanistically, the mediation of preference by fungi on leaves is not well understood. In order to test the hypothesis, that the strain-specific preference of G. roeselii is mediated by attractants or repellents that are constituents of fungi or oomycetes, selected fungal and oomycete strains were grown either in synthetic or leaf extract medium. Mycelia were extracted with solvents methanol or methylene chloride:methanol (2:1, v:v). Leaves covered with these extracts were subjected to choice feeding assays with G. roeselii. Methanol extracts proved to be repellent, and lipid extracts had no effect on the preference of G. roeselii. These results were contrary to the effects of the single isolates on leaves and suggested that compounds others than lipids or those extracted by methanol mediated the preference of G. roeselii. The repellent effect of the extracts of fungi or oomycetes was strongly affected by the C-source in the growth medium. The benthic mysid L. benedeni has recently invaded Lake Constance. Controlled laboratory experiments revealed that this mysid fed as well on shortly as well as on extensively leached leaf litter of several tree species. The interaction of the measured leaf parameters C- and polyphenol content explained 74% of the attractiveness of the leaf litter for the mysid, which suggested that feeding of L. benedeni is hindered by the waxes and cutin of the cuticula and by the lignocellulose structure of the leaf. 3 fungi growing on leaves elicited an intermediate feeding activity by L. benedeni, compared to that of littoral-exposed and autoclaved leaf litter. This suggests that L. benedeni feeds unselectively on the different microbial colonisers on decaying leaves. For the first time it was demonstrated that L. benedeni is a benthic leaf consumer that might potentially facilitate leaf degradation in Lake Constance.