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Since the last systematic review ofChiroderma(big-eyed bats) more than two decades ago, we report on biodiversity surveys that expand the distribution and species diversity of this Neotropical genus. The Caribbean endemic speciesChiroderma improvisumis documented for the first time from Nevis in the northern Lesser Antilles. A broader geographic sampling for a molecular analysis identifies a paraphyletic relationship inChiroderma trinitatumwith respect toChiroderma doriae. Cis-Andean populations ofC. trinitatumare most closely related to the morphologically distinctive and allopatrically distributedC. doriaein the Cerrado and Atlantic Forest of Brazil and Paraguay. The sister taxon to this grouping includes trans-Andean populations ofC. trinitatum, which we recommend to elevate to species status asC. gorgasi. This is an example of a cryptic species becauseC. gorgasiwas previously considered morphologically similar toC. trinitatum, but more detailed examination revealed that it lacks a posterolabial accessory cusp on the lower second premolar and has a narrower breadth of the braincase. We provide an amended description ofChiroderma gorgasi.

Autism Spectrum Disorder (ASD) is genetically complex with ~100 copy number variants and genes involved. To try to establish more definitive genotype and phenotype correlations in ASD, we searched genome sequence data, and the literature, for recurrent predicted damaging sequence-level variants affecting single genes. We identified 18 individuals from 16 unrelated families carrying a heterozygous guanine duplication (c.3679dup; p.Ala1227Glyfs*69) occurring within a string of 8 guanines (genomic location [hg38]g.50,721,512dup) affecting SHANK3, a prototypical ASD gene (0.08% of ASD-affected individuals carried the predicted p.Ala1227Glyfs*69 frameshift variant). Most probands carried de novo mutations, but five individuals in three families inherited it through somatic mosaicism. We scrutinized the phenotype of p.Ala1227Glyfs*69 carriers, and while everyone (17/17) formally tested for ASD carried a diagnosis, there was the variable expression of core ASD features both within and between families. Defining such recurrent mutational mechanisms underlying an ASD outcome is important for genetic counseling and early intervention.

Over the past 30 years (the timespan of a generation), advances in genomics technologies have revealed tremendous and unexpected variation in the human genome and have provided increasingly accurate answers to long-standing questions of how much genetic variation exists in human populations and to what degree the DNA complement changes between parents and offspring. Tracking the characteristics of these inherited and spontaneous (or de novo) variations has been the basis of the study of human genetic disease. From genome-wide microarray and next-generation sequencing scans, we now know that each human genome contains over 3 million single nucleotide variants when compared with the ~ 3 billion base pairs in the human reference genome, along with roughly an order of magnitude more DNA—approximately 30 megabase pairs (Mb)—being ‘structurally variable’, mostly in the form of indels and copy number changes. Additional large-scale variations include balanced inversions (average of 18 Mb) and complex, difficult-to-resolve alterations. Collectively, ~1% of an individual’s genome will differ from the human reference sequence. When comparing across a generation, fewer than 100 new genetic variants are typically detected in the euchromatic portion of a child’s genome. Driven by increasingly higher-resolution and higher-throughput sequencing technologies, newer and more accurate databases of genetic variation (for instance, more comprehensive structural variation data and phasing of combinations of variants along chromosomes) of worldwide populations will emerge to underpin the next era of discovery in human molecular genetics.

We describe a new species of mastiff bat in the genus Molossus (Molossidae), which was previously confused with the common and widely distributed M. molossus, from Guyana and Ecuador based on morphological and molecular differences. It is diagnosed by the following set of morphological characteristics: bicolored dorsal pelage, rounded anterior arch of the atlas, triangular occipital bone, and smaller body and skull size. In a molecular phylogenetic analysis of mitochondrial and nuclear DNA, maximum likelihood and parsimony trees recovered eight clades in the genus and a polyphyletic relationship for the M. molossus species complex. The new species was recovered in a well-supported clade that can be genetically distinguished from other species in the genus by its high level of sequence divergence based on the mitochondrial CO1 gene (8.0–10.1%) and on the nuclear gene beta fibrinogen (1.0–3.1%). It is broadly sympatric with M. molossus sensu stricto in northern South America, but morphologically distinct and genetically divergent.

Molossus is one of the most diverse genera of free-tailed bats in the pantropical family Molossidae and occurs though all the Neotropics. Nevertheless, the taxonomy and phylogeny of this group is poorly understood. Here, we present the data on evolutionary relationships of Molossus based on DNA barcodes of COI gene from 346 specimens of Molossus and its sister genus Promops and another New World molossid Eumops. Of these specimens, 50 are new sequences and 296 were obtained from GenBank. In addition, the nuclear gene beta fibrinogen was sequenced from a subset of 35 specimens. These data provide the basis for further exploration and understanding of the phylogenetic relationships of the genus Molossus (Loureiro et al., 2018) [1]. Keywords: Molossidae, Phylogenetics, Neotropics, Evolution

The majority (90%) of native terrestrial mammal species living in the Dominican Republic are bats, and two-thirds of these species are endemic to the Caribbean. However, recent molecular studies using DNA barcoding of the mitochondrial cytochrome c oxidase subunit 1 gene have suggested at least a 25% underestimation of biodiversity in bats throughout the world. A recent survey of bats in the Dominican Republic documented 15 of the 18 known species on the island of Hispaniola. Phylogenetic analysis of 132 individuals resulted in well-supported monophyletic species-level clades (maximal bootstrap values) with intraspecific variation ranging from 0% to 4.7% and interspecific variation ranging from 14.1% to 32.5%. A phylogeographic pattern separating the northern and southern Dominican Republic was recovered in 3 species of bats (Macrotus waterhousii, Pteronotus parnellii, and Pteronotus quadridens). The inclusion of broader geographic sampling across the Neotropics indicated that 3 widely distributed species (Eptesicus fuscus, Molossus molossus, and Monophyllus redmani) have high sequence divergence among insular or between insular and continental populations. Further systematic study is needed to identify morphologically cryptic species and their implications for conservation priorities in the Caribbean.

Although many processes of diversification have been described to explain variation of morphological traits within clades that have obvious differentiation among taxa, not much is known about these patterns in complexes of cryptic species. Molossus is a genus of bats that is mainly Neotropical, occurring from the southeastern United States to southern Argentina, including the Caribbean islands. Molossus comprises some groups of species that are morphologically similar but phylogenetically divergent, and other groups of species that are genetically similar but morphologically distinct. This contrast allows investigation of unequal trait diversification and the evolution of morphological and behavioural characters. In this study, we assessed the role of phylogenetic history in a genus of bat with three cryptic species complexes, and evaluated if morphology and behavior are evolving concertedly. The Genotype by Sequence genomic approach was used to build a species-level phylogenetic tree for Molossus and to estimate the ancestral states of morphological and echolocation call characters. We measured the correlation of phylogenetic distances to morphological and echolocation distances, and tested the relationship between morphology and behavior when the effect of phylogeny is removed. Morphology evolved via a mosaic of convergence and stasis, whereas call design was influenced exclusively through local adaptation and convergent evolution. Furthermore, the frequency of echolocation calls is negatively correlated with the size of the bat, but other characters do not seem to be evolving in concert. We hypothesize that slight variation in both morphology and behaviour among species of the genus might result from niche specialization, and that traits evolve to avoid competition for resources in similar environments.

Historical events, habitat preferences, and geographic barriers might result in distinct genetic patterns in insular versus mainland populations. Comparison between these two biogeographic systems provides an opportunity to investigate the relative role of isolation in phylogeographic patterns and to elucidate the importance of evolution and demographic history in population structure. Herein, we use a genotype‐by‐sequencing approach (GBS) to explore population structure within three species of mastiff bats (Molossus molossus, M. coibensis, and M. milleri), which represent different ecological histories and geographical distributions in the genus. We tested the hypotheses that oceanic straits serve as barriers to dispersal in Caribbean bats and that isolated island populations are more likely to experience genetic drift and bottlenecks in comparison with highly connected ones, thus leading to different phylogeographic patterns. We show that population structures vary according to general habitat preferences, levels of population isolation, and historical fluctuations in climate. In our dataset, mainland geographic barriers played only a small role in isolation of lineages. However, oceanic straits posed a partial barrier to the dispersal for some populations within some species (M. milleri), but do not seem to disrupt gene flow in others (M. molossus). Lineages on distant islands undergo genetic bottlenecks more frequently than island lineages closer to the mainland, which have a greater exchange of haplotypes., In this manuscript, we use the genotype‐by‐sequencing approach on three species of Mastiff bats with different ecological histories and geographical distributions to explore population genetics parameters and better understand the role of geographic barriers in the dispersal and gene flow in bats. Our analyses show that the population structure of each species varies according to environmental factors, levels of population isolation, and demographic histories. We show that oceanic straights seem to pose a partial barrier for some species and that more isolated lineages on islands tend to undergo genetic bottlenecks more than connected lineages closer to the mainland.

Mammals are one of the better known groups of animals, and in the Neotropics bats typically comprise about half of the mammalian species diversity. But, well resolved species-level phylogenies are still lacking for most taxa of bats. One broadly distributed genus is the mastiff bats, Molossus. Species within this genus are morphologically very similar, which results in a confusing and unstable taxonomy. In addition, low levels of genetic divergence among some clades make resolution of phylogenetic relationships difficult. Most authors recognize Molossus as being monophyletic, however, phylogenetic relationships within the genus remain poorly understood based on traditional Sanger sequencing of individual genes. We propose a more comprehensive framework based on large-scale genomic data derived from Next Generation Sequencing techniques to better understand evolutionary relationships within a group of closely related species with a rich taxonomic history. In this study, we utilized the NGS method of Genotype by Sequencing (GBS) to test the monophyly of the genus, understand evolutionary relationships within Molossus and investigate the genetic integrity of currently recognized species. Given that both de novo and reference genome pipelines are often used in the assembly of Single Nucleotide Polymorphism data from GBS, and that several tree inference methodologies have been proposed for SNP data, we test whether different alignments and phylogenetic approaches produce similar results. We also examined how the process of SNP identification and mapping can affect the consistency of the analyses. Our data provide the first high resolution phylogeny for the genus Molossus, bringing new insights into recognition of species boundaries and relationships among taxa. This study clarifies the taxonomy of Molossus and elevates the number of species in the genus from 11 to 14. We suggest the revalidation of the names M. nigricans, and M. fluminensis, which were synonymized under the name M. rufus; and M. bondae, previously synonymized under the name M. currentium. Different alignments and phylogenetic inferences produce consistent results, supporting use of SNP approach in addressing evolutionary questions on a macroevolutionary scale where the genetic distance among clades is low.

Bats are excellent models for studying the molecular basis of sensory adaptation. In Chiroptera, a sensory trade-off has been proposed between the visual and auditory systems, though the extent of this association has yet to be fully examined. To investigate whether variation in visual performance is associated with echolocation, we experimentally assayed the dim-light visual pigment rhodopsin from bat species with differing echolocation abilities. While spectral tuning properties were similar among bats, we found that the rate of decay of their light-activated state was significantly slower in a nonecholocating bat relative to species that use distinct echolocation strategies, consistent with a sensory trade-off hypothesis. We also found that these rates of decay were remarkably slower compared with those of other mammals, likely indicating an adaptation to dim light. To examine whether functional changes in rhodopsin are associated with shifts in selection intensity upon bat Rh1 sequences, we implemented selection analyses using codon-based likelihood clade models. While no shifts in selection were identified in response to diverse echolocation abilities of bats, we detected a significant increase in the intensity of evolutionary constraint accompanying the diversification of Chiroptera. Taken together, this suggests that substitutions that modulate the stability of the light-activated rhodopsin state were likely maintained through intensified constraint after bats diversified, being finely tuned in response to novel sensory specializations. Our study demonstrates the power of combining experimental and computational approaches for investigating functional mechanisms underlying the evolution of complex sensory adaptations.

Bats represent one of the largest and most striking nocturnal mammalian radiations, exhibiting many visual system specializations for performance in light-limited environments. Despite representing the greatest ecological diversity and species richness in Chiroptera, Neotropical lineages have been undersampled in molecular studies, limiting the potential for identifying signatures of selection on visual genes associated with differences in bat ecology. Here, we investigated how diverse ecological pressures mediate long-term shifts in selection upon long-wavelength (Lws) and short-wavelength (Sws1) opsins, photosensitive cone pigments that form the basis of colour vision in most mammals, including bats. We used codon-based likelihood clade models to test whether ecological variables associated with reliance on visual information (e.g. echolocation ability and diet) or exposure to varying light environments (e.g. roosting behaviour and foraging habitat) mediated shifts in evolutionary rates in bat cone opsin genes. Using additional cone opsin sequences from newly sequenced eye transcriptomes of six Neotropical bat species, we found significant evidence for different ecological pressures influencing the evolution of the cone opsins. WhileLwsis evolving under significantly lower constraint in highly specialized high-duty cycle echolocating lineages, which have enhanced sonar ability to detect and track targets, variation inSws1constraint was significantly associated with foraging habitat, exhibiting elevated rates of evolution in species that forage among vegetation. This suggests that increased reliance on echolocation as well as the spectral environment experienced by foraging bats may differentially influence the evolution of different cone opsins. Our study demonstrates that different ecological variables may underlie contrasting evolutionary patterns in bat visual opsins, and highlights the suitability of clade models for testing ecological hypotheses of visual evolution.

• Herein we propose a framework for assembling and analyzing Genotype by Sequencing (GBS) data to better understand evolutionary relationships within a group of closely related species using the mastiff bats (Molossus) as our model system. Many species within this genus have low-levels of genetic variation within and between morphologically distinct species, and the relationships among them remain unresolved using traditional Sanger sequencing methods. Given that both de novo and reference genome pipelines can be used to assemble next generation sequences, and that several tree inference methodologies have been proposed for single nucleotide polymorphism (SNP) data, we test whether different alignments and phylogenetic approaches produce similar results. We also examined how the process of SNP identification and mapping can affect the consistency of the analyses. Different alignments and phylogenetic inferences produced consistent results, supporting the GBS approach for answering evolutionary questions on a macroevolutionary scale when the genetic distance among phenotypically identifiable clades is low. We highlight the importance of exploring the relationships among groups using different assembly assumptions and also distinct phylogenetic inference methods, particularly when addressing phylogenetic questions in genetic and morphologically conservative taxa. • The method uses the comparison of several filter settings, alignments, and tree inference approaches on Genotype by Sequencing data. • Consistent results were found among several approaches. • The methodology successfully recovered well supported species boundaries and phylogenetic relationships among species of mastiff bats not hypothesized by previous methods., Graphical abstract Image, graphical abstract

The genus Molossus of the free-tailed bat family Molossidae is morphologically conservative and the level of genetic divergence is also low among many species, which has masked the actual species diversity in the genus. We analyzed the evolutionary relationships among species within Molossus using three mitochondrial (COI, Cytb, and 16S RNA), and two nuclear genes (RAG2 and β-fib) from more than 400 specimens across the Neotropics. We also investigated the genetic integrity of currently recognized species, especially those with broad geographic distributions and controversial taxonomic arrangements. Maximum likelihood and Bayesian phylogenetic reconstructions show that some recognized species are not monophyletic. The use of combined mitochondrial and nuclear genes as well as the use of only mitochondrial markers revealed more robust phylogenies than those for individual data sets and those restricted to the two nuclear genes. We recovered the recently described M. fentoni and M. alvarezi, validated M. milleri and M. verrilli as distinct species relative to M. molossus, and confirmed that the name M. barnesi should be considered as a junior synonym of M. coibensis. Additionally, M. rufus appears to be a complex of cryptic species, showing that the broadly distributed genus Molossus is more diverse than previously reported.