Morphological variability or inter-observer bias? A methodological toolkit to improve data quality of multi-researcher datasets for the analysis of morphological variation.

OBJECTIVES: The investigation of morphological variation in animals is widely used in taxonomy, ecology, and evolution. Using large datasets for meta-analyses has dramatically increased, raising concerns about dataset compatibilities and biases introduced by contributions of multiple researchers. MATERIALS AND METHODS: We compiled morphological data on 13 variables for 3073 individual mouse lemurs (Cheirogaleidae, Microcebus spp.) from 25 taxa and 153 different sampling locations, measured by 48 different researchers. We introduced and applied a filtering pipeline and quantified improvements in data quality (Shapiro-Francia statistic, skewness, and excess kurtosis). The filtered dataset was then used to test for genus-wide sexual size dimorphism and the applicability of Rensch's, Allen's, and Bergmann's rules. RESULTS: Our pipeline reduced inter-observer bias (i.e., increased normality of data distributions). Inter-observer reliability of measurements was notably variable, highlighting the need to reduce data collection biases. Although subtle, we found a consistent pattern of sexual size dimorphism across Microcebus, with females being the larger (but not heavier) sex. Sexual size dimorphism was isometric, providing no support for Rensch's rule. Variations in tail length but not in ear size were consistent with the predictions of Allen's rule. Body mass and length followed a pattern contrary to predictions of Bergmann's rule. DISCUSSION: We highlighted the usefulness of large multi-researcher datasets for testing ecological hypotheses after correcting for inter-observer biases. Using genus-wide tests, we outlined generalizable patterns of morphological variability across all mouse lemurs. This new methodological toolkit aims to facilitate future large-scale morphological comparisons for a wide range of taxa and applications.

The genomic diversity of the Eliurus genus in northern Madagascar with a putative new species.

Madagascar exhibits extraordinarily high level of species richness and endemism, while being severely threatened by habitat loss and fragmentation (HL&F). In front of these threats to biodiversity, conservation effort can be directed, for instance, in the documentation of species that are still unknown to science, or in investigating how species respond to HL&F. The tufted-tail rats genus (Eliurus spp.) is the most speciose genus of endemic rodents in Madagascar, with 13 described species, which occupy two major habitat types: dry or humid forests. The large species diversity and association to specific habitat types make the Eliurus genus a suitable model for investigating species adaptation to new environments, as well as response to HL&F (dry vs humid). In the present study, we investigated Eliurus spp. genomic diversity across northern Madagascar, a region covered by both dry and humid fragmented forests. From the mitochondrial DNA (mtDNA) and nuclear genomic (RAD-seq) data of 124 Eliurus individuals sampled in poorly studied forests of northern Madagascar, we identified an undescribed Eliurus taxon (Eliurus sp. nova). We tested the hypothesis of a new Eliurus species using several approaches: i) DNA barcoding; ii) phylogenetic inferences; iii) species delimitation tests based on the Multi-Species Coalescent (MSC) model, iv) genealogical divergence index (gdi); v) an ad-hoc test of isolation-by-distance within versus between sister-taxa, vi) comparisons of %GC content patterns and vii) morphological analyses. All analyses support the recognition of the undescribed lineage as a putative distinct species. In addition, we show that Eliurus myoxinus, a species known from the dry forests of western Madagascar, is, surprisingly, found mostly in humid forests in northern Madagascar. In conclusion, we discuss the implications of such findings in the context of Eliurus species evolution and diversification, and use the distribution of northern Eliurus species as a proxy for reconstructing past changes in forest cover and vegetation type in northern Madagascar.

How ancient forest fragmentation and riparian connectivity generate high levels of genetic diversity in a microendemic Malagasy tree.

Understanding landscape changes is central to predicting evolutionary trajectories and defining conservation practices. While human-driven deforestation is intense throughout Madagascar, exceptions in areas such as the Loky-Manambato region (north) raise questions regarding the causes and age of forest fragmentation. The Loky-Manambato region also harbours a rich and endemic flora, whose evolutionary origin remains poorly understood. We assessed the genetic diversity of an endangered microendemic Malagasy olive species (Noronhia spinifolia Hong-Wa) to better understand the vegetation dynamics in the Loky-Manambato region and its influence on past evolutionary processes. We characterized 72 individuals sampled across eight forests through nuclear and mitochondrial restriction-associated DNA sequencing data and chloroplast microsatellites. Combined population and landscape genetics analyses indicate that N. spinifolia diversity is largely explained by the current forest cover, highlighting a long-standing habitat mosaic in the region. This sustains a major and long-term role of riparian corridors in maintaining connectivity across these antique mosaic habitats, calling for the study of organismal interactions that promote gene flow.

Genetic and morphological diversity of mouse lemurs (Microcebus spp.) in northern Madagascar: The discovery of a putative new species?

Tropical forests harbor extremely high levels of biological diversity and are quickly disappearing. Despite the increasingly recognized high rate of habitat loss, it is expected that new species will be discovered as more effort is put to document tropical biodiversity. Exploring under-studied regions is particularly urgent if we consider the rapid changes in habitat due to anthropogenic activities. Madagascar is known for its extraordinary biological diversity and endemicity. It is also threatened by habitat loss and fragmentation. It holds more than 100 endemic primate species (lemurs). Among these, Microcebus (mouse lemurs) is one of the more diverse genera. We sampled mouse lemurs from several sites across northern Madagascar, including forests never sampled before. We obtained morphological data from 99 Microcebus individuals; we extracted DNA from tissue samples of 42 individuals and amplified two mitochondrial loci (cytb and cox2) commonly used for species identification. Our findings update the distribution of three species (Microcebus tavaratra, Microcebus arnholdi, and Microcebus mamiratra), including a major increase in the distribution area of M. arnholdi. We also report the discovery of a new Microcebus lineage genetically related to M. arnholdi. Several complementary approaches suggest that the newly identified Microcebus lineage might correspond to a new putative species, to be confirmed or rejected with additional data. In addition, morphological analyses showed (a) clear phenotypic differences between M. tavaratra and M. arnholdi, but no clear differences between the new Microcebus lineage and the sister species M. arnholdi; and (b) a significant correlation between climatic variables and morphology, suggesting a possible relationship between species identity, morphology, and environment. By integrating morphological, climatic, genetic, and spatial data of two northern Microcebus species, we show that the spatial distribution of forest-dwelling species may be used as a proxy to reconstruct the past spatial changes in forest cover and vegetation type.