Marianne Volleth’s research while affiliated with Otto-von-Guericke University Magdeburg and other places

The vespertilionid bat genus Hesperoptenus, comprising only five species and showing a distribution in the tropical regions from India to Sulawesi, was hitherto underrepresented in recent taxonomic studies. Here, modern cytogenetic techniques were applied to compare the karyotypes of two Hesperoptenus species, H. tomesi and H. blanfordi. Despite having the same chromosome number, 2n = 32, the karyotypes of H. tomesi and H. blanfordi were found to differ extensively. Instead, H. tomesi was found to share all nine derived Robertsonian fusion products with the previously published karyotype of H. doriae (2n = 26), which points to a closer relationship between both species. In line with our chromosomal data, mitochondrial DNA analyses also provided evidence for a closer relationship of H. tomesi with H. doriae and H. tickelli, for which no banded karyotype has been examined. Again, H. blanfordi was clearly separated from the remaining Hesperoptenus species. As a consequence, the subgenus Milithronycteris, comprising all species of Hesperoptenus except the type species H. doriae, is rendered paraphyletic. We therefore suggest to synonymize Milthronycteris with the subgenus Hesperoptenus. A chromosomal character, found in all three Hesperoptenus species examined but absent in other vespertilionid genera studied so far, namely a particular G-banding pattern on the homolog to Myotis chromosome MMY13, may serve as a synapomorphy for the genus Hesperoptenus. Key words: Hesperoptenus doriae, fluorescence in-situ hybridization, COI, cyt b

Phylogenetic relationships and species delimitation in African Vespertilionini have been a long-standing subject of debate and are still controversial, although recent molecular analyses have shed light onto some of these issues. In this study we employed a comparative cytogenetics approach for the delineation of chromosomal homology and for the detection of shared chromosomal characters, which were then used to support proposed phylogenetic relationships. Here, we present karyotype analyses of five African Vespertilionini, Laephotis kirinyaga (2n = 32), Neoromicia guineensis (2n = 26), Pseudoromicia brunnea (2n = 36), Nycticeinops happoldorum (2n = 24), and Nycticeinops schlieffenii (2n = 34), which were complemented by mitochondrial DNA sequence analysis for species classification of all our specimens. Our cytogenetic analyses revealed that three derived Robertsonian fusion chromosomes, 7/11, 8/9, and 10/12, characterize the karyotypes of four African Vespertilionini genera, i.e. Laephotis, Neoromicia, Afronycteris, and Pseudoromicia, for which we propose to constitute a new subtribe, Laephotina. A rare chromosomal rearrangement, an X-autosome translocation, was found in the studied N. guineensis female. The genus Nycticeinops is characterized by a high intrageneric karyotype diversity. In only two of all four analyzed species, i.e. N. happoldorum and N. crassulus, a common chromosomal feature, the fusion product 1/13 was detected. Further, for the recently described East African serotine, L. kirinyaga, we present the second record for West Africa. The cytochrome b sequence of our N. guineensis specimen from Ivory Coast showed 4% divergence to that of its closest relative, N. somalica from Kenya. Key words: Nycticeinops macrocephalus, Nycticeinops eisentrauti, chromosomal homology, COI, RAG2

Detailed karyotype analysis including fluorescence in-situ hybridization with chromosome-specific and sub-regional probes revealed novel cytogenetic signatures for elucidating phylogenetic relationships in Vespertilionoidea. Two shared derived Robertsonian fusions characterize all members of this superfamily. Further, the majority of Molossidae species show karyotypes with 48 chromosomes, including six of seven species studied here, namely Ozimops petersi, Mormopterus jugularis, Tadarida teniotis, Mops mops, Molossus molossus and M. pretiosus. A shared derived feature of all molossid taxa studied is a variant homolog to Myotis myotis chromosome MMY22. Despite their overall similarity in karyotype composition with one large and three medium- sized bi-armed as well as 19 acrocentric to subtelocentric autosomal pairs, our detailed analysis revealed small differences which shed new light onto molossid phylogenetic relationships. Because of the retention of several ancestral chromosomal characters, O. petersi and M. jugularis represent early offshoots of the molossid tree. A unique derived paracentric inversion in the MMY6 homolog unites all other molossids studied so far. The next species to branch off is T. tadarida, followed by a group composed of M. mops and all studied New World taxa, united by a further derived inversion in the MMY5 homolog. In the light of our findings, we recommend to elevate the subspecies M. t. griseiventer to species rank, because the specimen from Venezuela with a 2n = 42 karyotype differed clearly from the nominate subspecies Molossops t. temminckii with 2n = 48. Further, karyotypes of Miniopterus and Natalus were studied. In the 2n = 46 chromosomal complement of the family Miniopteridae a large number of autapomorphic characters were found. Based on the observation of a common derived condition of the MMY22 homolog, we assume that long- winged bats (Miniopteridae) are closer related to vespertilionids than to molossids. Finally, karyotype comparison in Natalus revealed two autapomorphic characters in the 2n = 36 chromosomal complement. Key words: Cytotaxonomy, chromosomal evolution, bat phylogenetics

Noonan syndrome (NS), the most common among RASopathies, is caused by germline variants in genes encoding components of the RAS-MAPK pathway. Distinct variants, including the recurrent Ser257Leu substitution in RAF1, are associated with severe hypertrophic cardiomyopathy (HCM). Here, we investigated the elusive mechanistic link between NS-associated RAF1S257L and HCM using three-dimensional cardiac bodies and bioartificial cardiac tissues generated from patient-derived induced pluripotent stem cells (iPSCs) harboring the pathogenic RAF1 c.770 C > T missense change. We characterize the molecular, structural, and functional consequences of aberrant RAF1–associated signaling on the cardiac models. Ultrastructural assessment of the sarcomere revealed a shortening of the I-bands along the Z disc area in both iPSC-derived RAF1S257L cardiomyocytes and myocardial tissue biopsies. The aforementioned changes correlated with the isoform shift of titin from a longer (N2BA) to a shorter isoform (N2B) that also affected the active force generation and contractile tensions. The genotype-phenotype correlation was confirmed using cardiomyocyte progeny of an isogenic gene-corrected RAF1S257L-iPSC line and was mainly reversed by MEK inhibition. Collectively, our findings uncovered a direct link between a RASopathy gene variant and the abnormal sarcomere structure resulting in a cardiac dysfunction that remarkably recapitulates the human disease.

The average genome size (GS) of bats, which are the only mammals capable of powered flight, is approximately 18% smaller than that of closely related mammalian orders. The low nuclear DNA content of Chiroptera is comparable to that of birds, which are also characterized by a high metabolic rate. Only a few chiropteran taxa possess notable amounts of constitutive heterochromatin. Here, we studied the karyotypes of two non-related vesper bat species with unusually high amounts of constitutive heterochromatin: Hesperoptenus doriae and Philetor brachypterus. Conventional staining methods and whole-chromosome painting with probes derived from Myotis myotis (2n = 44), showing a karyotype close to that of the presumed ancestor of Vespertilionidae, revealed Robertsonian fusions as the main type of rearrangement leading to the exceptionally reduced diploid chromosome number of 2n = 26 in both species. Moreover, both karyotypes are characterized by large blocks of pericentromeric heterochromatin composed of CMA-positive and DA-DAPI-positive segments. In H. doriae, the heterochromatin accumulation has resulted in a genome size of 3.22 pg (1C), which is 40% greater than the mean genome size for the family. For P. brachypterus, a genome size of 2.94 pg was determined, representing an increase of about 28%. Most notably, in H. doriae, the presence of additional constitutive heterochromatin correlates with an extended mitotic cell cycle duration in vitro. A reduction in diploid chromosome number to 30 or lower is discussed as a possible cause of the accumulation of pericentromeric heterochromatin in Vespertilionidae.

As many as 17 subspecies are currently recognized for the lesser false vampire, Megaderma spasma along its wide distributional range, from India to Southeast Asia. Cytogenetic studies have been so far restricted to specimens from only four geographical regions, namely Malaysia, Thailand, China and the Philippines. We examined the karyotypes of M. spasma medium from Peninsular Malaysia and M. s. trifolium from Malaysian Borneo with conventional and molecular cytogenetic techniques, using whole chromosome painting probes from the vespertilionid Myotis myotis and the hipposiderid Aselliscus stoliczkanus. Both Megaderma subspecies showed the same diploid chromosome number, 2n = 38, but differed concerning interstitial heterochromatic segments and number and shape of heterochromatic B chromosomes. Comparison with published data revealed that the Bornean specimens ́ karyotype best resembled the chromosomal set of the Chinese specimen. In contrast, a single male specimen from Malaysian Borneo showed a different karyotype (2n = 40) without B chromosomes and interstitial heterochromatin, indicating that it may represent a cryptic, yet unknown Megaderma species. Key words: karyotype, fluorescence in-situ hybridization, cytotaxonomy, chromosomal evolution

Classical and molecular cytogenetic methods were applied to study the karyotypes of one species of Hipposideridae and three taxa of the Rhinolophidae subgenus Aquias from Malaysian Borneo. Except for four chromosomal pairs with autapomorphic arm combinations, the karyotype of Coelops robinsoni was found to be similar to the closely related Aselliscus stoliczkanus. From the three Rhinolophus taxa studied, only R. trifoliatus was found to share the karyotype with conspecifics from Peninsular Malaysia. In contrast, the karyotype of R. luctus foetidus from Sarawak, Borneo differed in the composition of the Y-autosomal translocation products from the closely related R. morio from Peninsular Malaysia, formerly also a subspecies of R. luctus. Therefore, elevation to specific rank is suggested for R. l. foetidus. Examination of the chromosomal set of male R. sedulus specimens from Borneo with 2n = 45 and a Neo-X1X2Y sex chromosome system revealed extreme differences to the karyotype of specimens from Peninsular Malaysia with 2n = 28, to date also classified as R. sedulus. Therefore, with Sarawak, Borneo, as the type locality for R. sedulus, the taxon from Peninsular Malaysia is here described as a new species. Key words: sex-autosome translocations, cytotaxonomy, fluorescence in-situ hybridization

The maternal mode of mitochondrial DNA (mtDNA) inheritance is central to human genetics. Recently, evidence for bi-parental inheritance of mtDNA was claimed for individuals of three pedigrees that suffered mitochondrial disorders. We sequenced mtDNA using both direct Sanger and Massively Parallel Sequencing in several tissues of eleven maternally related and other affiliated healthy individuals of a family pedigree and observed mixed mitotypes in eight individuals. Cells without nuclear DNA, i.e. thrombocytes and hair shafts, only showed the mitotype of haplogroup (hg) V. Skin biopsies were prepared to generate ρ° cells void of mtDNA, sequencing of which resulted in a hg U4c1 mitotype. The position of the Mega-NUMT sequence was determined by fluorescence in situ hybridization and two different quantitative PCR assays were used to determine the number of contributing mtDNA copies. Thus, evidence for the presence of repetitive, full mitogenome Mega-NUMTs matching haplogroup U4c1 in various tissues of eight maternally related individuals was provided. Multi-copy Mega-NUMTs mimic mixtures of mtDNA that cannot be experimentally avoided and thus may appear in diverse fields of mtDNA research and diagnostics. We demonstrate that hair shaft mtDNA sequencing provides a simple but reliable approach to exclude NUMTs as source of misleading results.

To find a mate, males and females of phaneropterine bush-crickets engage in duets. The male starts a duet and a female responds if she is prepared to mate. This pattern is exactly followed in the acoustic communication of a species of the genus Anacaulomera from the State of Bahia in Brazil which we newly describe in this manuscript. However, the male also acoustically defends the duet. As soon as it hears a female response, it adds an additional component to its own signal to make the localisation of the female difficult for rivals. At the same time the male reduces the interval between its songs for a faster exchange of information about the location of the partner, and reduces the intensity of the song that it can be heard only from nearby. Besides the nearly perfect anti-eavesdropping modifications of the duet the new species is remarkable concerning its karyotype. The meiotic behaviour of chromosomes in the male may indicate the origin of a new type of sex determination mechanism.