Traditional multilocus phylogeny fails to fully resolve Palearctic ground squirrels (Spermophilus) relationships but reveals a new species endemic to West Siberia.

Previous efforts to reconstruct evolutionary history of Palearctic ground squirrels within the genus Spermophilus have primarily relied on a single mitochondrial marker for phylogenetic data. In this study, we present the first phylogeny with comprehensive taxon sampling of Spermophilus via a conventional multilocus approach utilizing five mitochondrial and five nuclear markers. Through application of the multispecies coalescent model, we constructed a species tree revealing four distinct clades that diverged during the Late Miocene. These clades are 1) S. alaschanicus and S. dauricus from East Asia; 2) S. musicus and S. pygmaeus from East Europe and northwestern Central Asia; 3) the subgenus Colobotis found across Central Asia and its adjacent regions and encompassing S. brevicauda, S. erythrogenys, S. fulvus, S. major, S. pallidicauda, S. ralli, S. relictus, S. selevini, and S. vorontsovi sp. nov.; and 4) a Central/Eastern Europe and Asia Minor clade comprising S. citellus, S. taurensis, S. xanthoprymnus, S. suslicus, and S. odessanus. The latter clade lacked strong support owing to uncertainty of taxonomic placement of S. odessanus and S. suslicus. Resolving relationships within the subgenus Colobotis, which radiated rapidly, remains challenging likely because of incomplete lineage sorting and introgressive hybridization. Most of modern Spermophilus species diversified during the Early-Middle Pleistocene (2.2-1.0 million years ago). We propose a revised taxonomic classification for the genus Spermophilus by recognizing 18 species including a newly identified one (S. vorontsovi sp. nov.), which is found only in a limited area in the southeast of West Siberia. Employing genome-wide single-nucleotide polymorphism genotyping, we substantiated the role of the Ob River as a major barrier ensuring robust isolation of this taxon from S. erythrogenys. Despite its inherent limitations, the traditional multilocus approach remains a valuable tool for resolving relationships and can provide important insights into otherwise poorly understood groups. It is imperative to recognize that additional efforts are needed to definitively determine phylogenetic relationships between certain species of Palearctic ground squirrels.

Molecular systematics of the Sicista tianschanica species complex: a contribution from historical DNA analysis.

The Tianshan birch mouse Sicista tianschanica is an endemic of the Central Asian mountains and has previously been shown to include several karyomorphs ("Terskey", "Talgar", "Dzungar"); however, the taxonomic status of these forms has remained uncertain. We examined the genetic variation in S. tianschanica based on historical DNA samples from museum collections, including the type series. Mitochondrial and nuclear data indicated that the species complex includes two major clades: Northern (N) and Southern (S) (cytb distance 13%). The N clade corresponds to the "Dzungar" karyomorph (Dzungar Alatau, Tarbagatay). The S clade is comprised of four lineages (S1-S4) divergent at 6-8%; the relationships among which are resolved incompletely. The S1 lineage is found in eastern Tianshan and corresponds to the nominal taxon. The S2 is distributed in central and northern Tianshan and corresponds to the "Terskey" karyomorph. The S3 is restricted to Trans-Ili Alatau and belongs to the "Talgar" karyomorph. The S4 is represented by a single specimen from southeastern Dzungar Alatau with "Talgar" karyotype. No interlineage gene flow was revealed. The validity of S. zhetysuica (equivalent to the N clade) is supported. Based on genetic and karyotypic evidence, lineages S2 and S3 are described as distinct species. The status of the S4 requires further investigation.

LINC00973 Induces Proliferation Arrest of Drug-Treated Cancer Cells by Preventing p21 Degradation.

Overcoming drug resistance of cancer cells is the major challenge in molecular oncology. Here, we demonstrate that long non-coding RNA LINC00973 is up-regulated in normal and cancer cells of different origins upon treatment with different chemotherapeutics. Bioinformatics analysis shows that this is a consequence of DNA damage response pathway activation or mitotic arrest. Knockdown of LINC0973 decreases p21 levels, activates cellular proliferation of cancer cells, and suppresses apoptosis of drug-treated cells. We have found that LINC00973 strongly increases p21 protein content, possibly by blocking its degradation. Besides, we have found that ectopic over-expression of LINC00973 inhibits formation of the pro-survival p53-Ser15-P isoform, which preserves chromosome integrity. These results might open a new approach to the development of more efficient anti-cancer drugs.

Phylogenetic relationship and variation of alarm call traits of populations of red-cheeked ground squirrels (Spermophilus erythrogenys sensu lato) suggest taxonomic delineation.

Distribution area and taxonomic borders within the species complex Spermophilus erythrogenys sensu lato remain questionable. Early evidence suggests that red-cheeked ground squirrels of Southeast Kazakhstan are remarkably different in terms of the acoustic structure of their alarm calls from the red-cheeked ground squirrels of the Kurgan region in Russia. In this study, we analyzed the differences in the acoustic structure of the alarm call and mitochondrial DNA (complete control region, 1005-1006 bp and complete cytochrome b gene, 1140 bp) in 3 populations of red-cheeked ground squirrels (Tara, Altyn-Emel and Balkhash), all located within areas isolated by geographical barriers in Southeast Kazakhstan. We found that the alarm call variables were similar between the 3 study populations and differed by the maximum fundamental frequency (8.46 ± 0.75 kHz) from the values (5.62 ± 0.06 kHz) reported for the red-cheeked ground squirrels from the Kurgan region of Russia. Variation in mtDNA control region was only 3% and variation in cytochrome b gene was only 2.5%. Phylogenetic trees based on cytochrome b gene polymorphism of 44 individuals from the study area and adjacent territories indicated 3 clades with high (98-100%) bootstrap support: "intermedius," "brevicauda" and "iliensis"). We conclude that the 3 study populations in Southeast Kazakhstan belong to the clade intermedius and suggest a taxonomical revision of the species complex Spermophilus erythrogenys sensu lato, including analyses of nuclear DNA and alarm calls for populations of the brevicauda and iliensis clades.

Between-year vocal aging in female red deer (Cervus elaphus).

OBJECTIVES: Studying animal vocal aging has potential implication in the field of animal welfare and for modeling human voice aging. The objective was to examine, using a repeated measures approach, the between-year changes of weight, social discomfort score (bites of other hinds on hind pelt), body condition score (fat reserves) and acoustic variables of the nasal (closed-mouth) and the oral (open-mouth) contact calls produced by farmed red deer hinds (Cervus elaphus) toward their young. RESULTS: Repeated measures ANOVA revealed that with an increase of hind age for 1 year, the acoustic variables of their nasal contact calls (the beginning and maximum fundamental frequencies, the depth of frequency modulation and the peak frequency) decreased, whereas in their oral contact calls only the end fundamental frequency decreased. Duration and power quartiles did not change in any call type. Body weight and body condition score increased between years, whereas discomfort score decreased. Results of this study revealed directly the short-term effects of aging on the acoustics of the nasal contact calls in the same hinds. This study also confirmed that elevated emotional arousal during emission of the oral contact masks the effects of aging on vocalization in female red deer.

Altai pika (Ochotona alpina) alarm calls: individual acoustic variation and the phenomenon of call-synchronous ear folding behavior.

Non-hibernating pikas collect winter food reserves and store them in hay piles. Individualization of alarm calls might allow discrimination between colony members and conspecifics trying to steal food items from a colony pile. We investigated vocal posture, vocal tract length, and individual acoustic variation of alarm calls, emitted by wild-living Altai pikas Ochotona alpina toward a researcher. Recording started when a pika started calling and lasted as long as possible. The alarm call series of 442 individual callers from different colonies consisted of discrete short (0.073-0.157 s), high-frequency (7.31-15.46 kHz), and frequency-modulated calls separated by irregular intervals. Analysis of 442 discrete calls, the second of each series, revealed that 44.34% calls lacked nonlinear phenomena, in 7.02% nonlinear phenomena covered less than half of call duration, and in 48.64% nonlinear phenomena covered more than half of call duration. Peak frequencies varied among individuals but always fitted one of three maxima corresponding to the vocal tract resonance frequencies (formants) calculated for an estimated 45-mm oral vocal tract. Discriminant analysis using variables of 8 calls per series of 36 different callers, each from a different colony, correctly assigned over 90% of the calls to individuals. Consequently, Altai pika alarm calls are individualistic and nonlinear phenomena might further increase this acoustic individualization. Additionally, video analysis revealed a call-synchronous, very fast (0.13-0.23 s) folding, depression, and subsequent re-expansion of the pinna confirming an earlier report of this behavior that apparently contributes to protecting the hearing apparatus from damage by the self-generated high-intensity alarm calls.

One plus one: Binary alarm calls retain individual signature for longer periods than single-note alarms in the European ground squirrel (Spermophilus citellus).

Ground squirrels emit species-specific alarm calls that, among other characteristics, differ by the number of elements. Unlike some species that produce single-element calls, e.g., the Speckled ground squirrel (Spermophilus suslicus), individual European ground squirrels (S. citellus) frequently emit binary-element calls in addition to single-element calls. We tested the hypothesis that the time stability of individuality encoded in alarm calls might be better retained by complicating their acoustic structure by adding extra elements. In a semi-captive colony of individually marked European ground squirrels, we repeatedly recorded alarm calls that were produced towards a human by 12 adult (2 males and 10 females) live-trapped animals. Repeated recordings occurred within time spans of a few hours, 2days and 1year from the first recording. Our results showed that individual calls were highly similar within recordings, but less similar between recordings separated by time spans. Individual differences were best retained when we used nine acoustic variables from both elements. The differences were worse when we used nine variables from only the first element and worst when we used nine variables from only the second element. These results supported the caller reliability hypothesis for species that produce multiple-note alarms, e.g., the Richardson's ground squirrel (S. richardsonii).

The power of oral and nasal calls to discriminate individual mothers and offspring in red deer, Cervus elaphus.

BACKGROUND: In most species, acoustical cues are crucial for mother-offspring recognition. Studies of a few species of ungulates showed that potential for individual recognition may differ between nasal and oral contact calls. RESULTS: Vocalizations of 28 hinds and 31 calves of farmed Iberian red deer (Cervus elaphus hispanicus) were examined with discriminant function analyses (DFA) to determine whether acoustic structure of their oral and nasal contact calls encodes information about the caller's identity. Contact calls were elicited by brief separation of individually identified animals by a distance over 10 m or by a bar fence. Both oral and nasal calls of both hinds and calves showed high potential to discriminate individuals. In hinds, individuality was significantly higher in the oral than in the nasal calls, whereas in calves, individuality was equally well expressed in both oral and nasal calls. For calves, the maximum fundamental frequency was higher and the duration was longer in oral calls than in nasal calls. For hinds, the maximum fundamental frequency and the duration were indistinguishable between oral and nasal calls. Compared to the pooled sample of oral and nasal calls, separate oral or nasal call samples provided better classifying accuracy to individual in either hinds or calves. Nevertheless, in both hinds and calves, even in the pooled sample of oral and nasal calls, the degree of individual identity was 2-3 times greater than expected by chance. For hinds that provided calls in both years, cross-validation of calls collected in 2012 with discriminant functions created with calls from 2011 showed a strong decrease of classifying accuracy to individual. CONCLUSIONS: These results suggest different potentials of nasal and oral calls to allow the discrimination of individuals among hinds, but not among red deer calves. The high potential of individual recognition even with the pooled sample of oral and nasal calls allows mother and young to remember only one set of acoustic variables for mutual vocal recognition. Poor between-year stability of individual characteristics of hind oral and nasal calls would require updating keys to individual recognition each calving season.

The potential to encode sex, age, and individual identity in the alarm calls of three species of Marmotinae.

In addition to encoding referential information and information about the sender's motivation, mammalian alarm calls may encode information about other attributes of the sender, providing the potential for recognition among kin, mates, and neighbors. Here, we examined 96 speckled ground squirrels (Spermophilus suslicus), 100 yellow ground squirrels (Spermophilus fulvus) and 85 yellow-bellied marmots (Marmota flaviventris) to determine whether their alarm calls differed between species in their ability to encode information about the caller's sex, age, and identity. Alarm calls were elicited by approaching individually identified animals in live-traps. We assume this experimental design modeled a naturally occurring predatory event, when receivers should acquire information about attributes of a caller from a single bout of alarm calls. In each species, variation that allows identification of the caller's identity was greater than variation allowing identification of age or sex. We discuss these results in relation to each species' biology and sociality.

Stability of acoustic individuality in the alarm calls of wild yellow ground squirrels Spermophilus fulvus and contrasting calls from trapped and free-ranging callers.

The questions of individuality and stability of cues to identity in vocal signals are of considerable importance from theoretical and conservation perspectives. While individuality in alarm calls has been reported for many sciurids, it is not well-documented that the vocal identity encoded in the alarm calls is stable between different encounters with predators. Previous studies of two obligate hibernating rodents, speckled ground squirrels Spermophilus suslicus, and yellow ground squirrels Spermophilus fulvus demonstrated that, after hibernation, most individuals could not be identified reliably by their alarm calls. Moreover, in most speckled ground squirrels, individual patterns of alarm calls changed progressively over as little as 2 weeks. However, these previous data have been obtained using the collection of alarm calls from trapped animals. Here, we examined ten free-ranging dye-marked yellow ground squirrels to determine whether their alarm calls retain the cues to individuality between two encounters of surrogate predators (humans), separated on average by 3 days. Discriminant function analysis showed that the alarm calls of individual yellow ground squirrels were very similar within a recording session, providing very high individual distinctiveness. However, in six of the ten animals, the alarm calls were unstable between recording sessions. Also, we examined ten dye-marked individuals for consistency of acoustic characteristics of their alarm calls between the encounters of humans, differing in techniques of call collection, from free-ranging vs trapped animals. We found differences only in two variables, both related to sound degradation in the environment. Data are discussed in relation to hypotheses explaining the adaptive utility of acoustic individuality in alarm calls.