Diversity and seasonality of ectoparasite burden on two species of Madagascar fruit bat, Eidolon dupreanum and Rousettus madagascariensis

Abstract

Background: Bats are important reservoir hosts for a variety of microparasites, some of which are transmitted by ectoparasite vectors that include mites, fleas, lice, ticks, and bat flies (families Nycteribiidae and Streblidae). All of these ectoparasite taxa are known to parasitize two endemic fruit bats of Madagascar, Eidolon dupreanum and Rousettus madagascariensis. We aimed to describe the diversity of ectoparasite infestation for both bat species through morphological observation and DNA barcoding and elucidate ecological and climatic correlates of seasonal nycteribiid parasitism of these hosts.

Methods: Live E. dupreanum and R. madagascariensis fruit bats were captured monthly in northern and central-eastern Madagascar from 2013-2020. Ectoparasites on all captured bats were counted and identified in the field, then collected into ethanol. Field identification of a subset of samples were confirmed via microscopy and DNA barcoding of the cytochrome C oxidase subunit 1 (COI) and 18S genes. The seasonal abundance of nycteribiid bat flies on both host bats was analyzed using generalized additive models, and the role of climate in driving this seasonality was assessed via cross-correlation analysis combined with generalized linear models. Phylogenetic trees were generated to compare COIand 18S sequences of Madagascar nycteribiid and streblid bat flies with available reference sequences from GenBank.

Results: Ectoparasites corresponding to four broad taxa (mites, ticks, fleas, and bat flies) were recovered from 628 of 873 E. dupreanum and 831 of 862 R. madagascariensis. E. dupreanum were most commonly parasitized by Cyclopodia dubia nycteribiids and R. madagascariensis by Eucampsipoda madagascariensis nycteribiids or Megastrebla wenzeli streblids. We observed significant seasonality in nycteribiid abundance on both bat hosts, which varied by bat sex and was positively correlated with lagged temperature, precipitation, and humidity variables. Barcoding sequences recovered for all three bat fly species grouped with previously reported sequences, confirming morphological species identification. Our study contributes the first DNA barcodes of any kind reported for M. wenzeli and the first 18S barcodes for C. dubia.

Conclusion: This study explores the diversity and abundance of ectoparasite burdens in two Malagasy fruit bat species, highlighting the importance of seasonal ecology and the influence of climate variables on parasitism, which correlates with resource availability.

Keywords: Bat fly; DNA barcoding; Eidolon dupreanum; Madagascar; Nycteribiidae; Pteropodidae; Rousettus madagascariensis; Streblidae; bat ectoparasite.

Fig. 1.

Alluvial plot showing bat host species (center) associations with broad ectoparasite clades (top) and genus-level classifications (bottom). Fleas and bat flies belonging to order Diptera are colored in shades of blue, while mites and ticks belong to class Arachnida (respectively in superorder Acariformes and Parasitiformes) are colored in shades of green. Images taken under the microscope at 40x magnification are shown below the names of the corresponding genera.

Fig. 2.

Seasonal variation in the abundance of Nycteribiidae bat flies counted on (A, B) E. dupreanum and (C,D) R. madagascariensis bats captured at roost sites in central-eastern Madagascar (respectively, Angavobe/Angavokely and Maromizaha caves). Panels (A) and (C) show seasonal ectoparasite count predictions (red line) from best-fit GAMs for male and female bat hosts of each species, with 95% CI by standard error shaded in gray. Translucent background points in black correspond to raw data across all years of the study (2013–2019). Pink background shading corresponds to the gestation period for each species from (61), while blue background shading corresponds to the nutritionally deficient dry season for the region. Panels (B) and (D) show partial effect (y-axis) of bat host mass: forearm residual, respectively for E. dupreanum and R. madagascariensis, on bat fly count. Solid lines (gray for non-significant effects; blue for significant effects) correspond to mean effects, with 95% CIs by standard error shown in translucent shading.

Fig. 3.

Mean monthly nycteribiid count per bat for (A) C. dubia parasitism of E. dupreanum, across the full 2013–2020 time series for Angavokely roost site (gray lines and points; left y-axis), as compared with climate variables of monthly averages of (horizontal panels) diurnal humidity (% relative humidity), total precipitation (mm), and temperature (°C) for the same region (red lines and points; right y-axis). (B) E. madagascariensis parasitism of R. madagascariensis, mirroring the same structure in (A), for the Maromizaha roost site. 95% CIs by standard error are shown for both ectoparasite and climate data.

Fig. 4.

Influence of climate and demographic variables on the abundance of nyteribiid bat fly count for (A,B) C. dubia abundance on E. dupreanum and (C,D) E. madagascariensis abundance on R. madagascariensis. (A,C) Top five GLMs using optimally-lagged climate variables to predict bat fly abundance, ranked by δAICc, for E. dupreanum and R. madagascariensis bat hosts. Rows represent individual models and columns represent predictor variables. (B,D) Incidence rate ratios of each linear predictor from top-fit models shown respectively in (A,C). Significant positive correlates are colored red, significant negative correlates are colored blue, and insignificant correlates are colored grey. 95% CIs by standard error are shown as horizontal error bars.

Fig. 5.

Maximum likelihood phylogeny of COI ectoparasite sequences from untrimmed alignment (RAxML-NG, GTR+I+G4) (81). Bootstrap support values computed using Felsenstein’s method (82) are indicated by shaded circles on each node, corresponding to legend. Sequences are collapsed into single species clades, or where indicated, clades by genera for ease of visualization; see Fig. S6 for full phylogeny with individual sequences labeled. Tip shapes are colored by genera, and tip labels for the three Madagascar clades (E. madagascariensis, C. dubia, M. wenzeli) are highlighted in yellow. Tree is rooted in Drosophila melanogaster, accession number NC_001709. Branch lengths are scaled by nucleotide substitutions per site, corresponding to the scalebar shown.