Zoological Research

389−400

Authors: Quan Li;Feng Cheng;Stephen  M. Jackson;Kristofer  M. Helgen;Wen-Yu Song;Shao-Ying Liu;Daosavanh Sanamxay;Song Li;Fei Li;Yun Xiong;Jun Sun;Hong-Jiao Wang;Xue-Long Jiang

The flying squirrels (Pteromyini, Rodentia) are the most diverse and widely distributed group of gliding mammals. Taxonomic boundaries and relationships within flying squirrels remain an area of active research in mammalogy. The discovery of new specimens of Pteromys (Hylopetes) leonardi Thomas, 1921, previously considered a synonym of Hylopetes alboniger, in Yunnan Province, China allowed a morphological and genetic reassessment of the status of this taxon. Phylogenetic reconstruction was implemented using sequences of two mitochondrial (12S ribosomal RNA and 16S ribosomal RNA) and one nuclear (interphotoreceptor retinoid-binding protein) gene fragments. Morphological assessments involved examinations of features preserved on skins, skulls, and penises of museum specimens, supplemented with principal component analysis of craniometric data. Together these assessments revealed that this taxon should be recognized not only as a distinct species, but should also be placed within a new genus, described here as Priapomys gen. nov.

406−411

Authors: Ke-Chu Zhang;Qi-Hai Zhou;Huai-Liang Xu;Zhong-Hao Huang

Changes in abiotic and biotic factors can affect the efficiency of biological systems in animals, forcing them to adjust their behaviors in response to daily and seasonal variations. From September  2016  to  August  2017,  we  collected  ranging behavior  data  on  four  groups  of  white-headed  langurs (Trachypithecus leucocephalus)  in  the  Guangxi  Chongzuo White-Headed  Langur  National  Nature  Reserve,  Guangxi, southwest  China.  We  simultaneously  analyzed  how  multiple ecological factors affect langur ranging behavior, which should facilitate  our  understanding  of  the  potential  mechanisms underlying  their  adaptation  to  limestone  habitats.  Results showed that langur ranging behavior was significantly affected by  diet  composition,  food  availability,  and  climatic  factors. Specifically, moving time and daily path length increased with the increase in dietary diversity. Furthermore, moving time and daily  path  length  were  positively  associated  with  the availability  of  fruit  and  relative  humidity  of  the  forest,  and moderately associated with temperature and relative humidity of bare rock. Our study demonstrated that langurs maintain stable  moving  and  feeding  times  and  exhibit  a  short  daily travel  distance,  likely  adopting  an  energy-conserving behavioral  strategy  in  response  to  food  shortages  and  high temperatures  in  the  fragmented  karst  forest.  These  results highlight the importance of food availability and temperature in shaping the ranging behavior of these karst-dwelling primates.

417−422

Authors: Wen-Ping He;Jian Zhou;Zhe Li;Ting-Sen Jing;Chun-Hua Li;Yue-Jing Yang;Meng-Bin Xiang;Chao-Wei Zhou;Guang-Jun Lv;Hong-Yan Xu;Hui Luo;Hua Ye

The  Chinese  longsnout  catfish  (Leiocassis longirostris Günther) is one of the most economically important freshwater fish  in  China.  As  wild  populations  have  declined  sharply  in recent years, it is also a valuable model for research on sexual dimorphism, comparative biology, and conservation. However, the  current  lack  of  high-quality  chromosome-level  genome information  for  the  species  hinders  the  advancement  of comparative  genomic  analysis  and  evolutionary  studies. Therefore, we constructed the first high-quality chromosome- level reference genome for L. longirostris. The total genome was  703.19  Mb,  with  389  contigs  and  contig  N50  length  of 4.29  Mb.  Using  high-throughput  chromosome  conformation capture (Hi-C) data, the genome sequences (685.53 Mb) were scaffolded into 26 chromosomes ranging from 17.36 to 43.97 Mb, resulting in a chromosomal anchoring rate for the genome of  97.44%.  In  total, 23 708 protein-coding  genes  were identified in the genome. Phylogenetic analysis indicated that L. longirostris and  its  closest  related  species P. fulvidraco diverged  approximately  26.6  million  years  ago.  This  high- quality  reference  genome  of L. longirostris should  pave  the way  for  future  genomic  comparisons  and  evolutionary research.

428−432

Authors: Christian Roos;Roberto Portela Miguez;Richard Sabin;Edward E. Louis Jr.;Michael Hofreiter;Dietmar Zinner

Accurate information on name-bearing types, including corresponding type localities, is essential for proper taxonomy. However, such geographic information is often missing or unreliable. The localities of type specimens collected 100–200 years ago can be difficult to trace due to changes in local names or simple inaccuracies. Such a case can be found for the gray-backed sportive lemur (Lepilemur dorsalis), with its type locality imprecisely fixed as Northwest Madagascar. In recent years, eight species have been newly described for the Inter-River-Systems (IRSs) of this region, however the designation of L. dorsalis remains controversial due to a lack of a precise type locality. Here, we sequenced the complete mitochondrial genomes (mitogenomes) of type specimens of L. dorsalis and L. grandidieri, which is currently recognized as a synonym of L. dorsalis and compared their sequences with those of samples of known provenance from different IRSs. Results showed that the two type specimens of L. dorsalis and L. grandidieri had identical mitogenome sequences and clustered closely with samples collected in IRS V, indicating that the type locality could be fixed to IRS V. Consequently, L. dorsalis occurs in IRS V, and L. grandidieri and L. mittermeieri are junior synonyms of L. dorsalis. This finding demonstrates the value of type specimens for clarifying phylogeographic and taxonomic questions and clarifies the taxonomy of sportive lemurs in Northwest Madagascar.

450−460

Authors: Lin Zeng;He-Qun Liu;Xiao-Long Tu;Chang-Mian Ji;Xiao Gou;Ali Esmailizadeh;Sheng Wang;Ming-Shan Wang;Ming-Cheng Wang;Xiao-Long Li;Hadi Charati;Adeniyi;C. Adeola;Rahamon  Akinyele Moshood Adedokun;Olatunbosun Oladipo;Sunday Charles Olaogun;Oscar J. Sanke;Mangbon Godwin F.;Sheila Cecily Ommeh;Bernard Agwanda;Jacqueline Kasiiti Lichoti;Jian-Lin Han;Hong-Kun Zheng;Chang-Fa Wang;Ya-Ping Zhang; Laurent A. F. Frantz;Dong-Dong Wu

Over the last several hundred years, donkeys have adapted to high-altitude conditions on the Tibetan Plateau. Interestingly, the kiang, a closely related equid species, also inhabits this region. Previous reports have demonstrated the importance of specific genes and adaptive introgression in divergent lineages for adaptation to hypoxic conditions on the Tibetan Plateau. Here, we assessed whether donkeys and kiangs adapted to the Tibetan Plateau via the same or different biological pathways and whether adaptive introgression has occurred. We assembled a de novo genome from a kiang individual and analyzed the genomes of five kiangs and 93 donkeys (including 24 from the Tibetan Plateau). Our analyses suggested the existence of a strong hard selective sweep at the EPAS1 locus in kiangs. In Tibetan donkeys, however, another gene, i.e., EGLN1, was likely involved in their adaptation to high altitude. In addition, admixture analysis found no evidence for interspecific gene flow between kiangs and Tibetan donkeys. Our findings indicate that despite the short evolutionary time scale since the arrival of donkeys on the Tibetan Plateau, as well as the existence of a closely related species already adapted to hypoxia, Tibetan donkeys did not acquire adaptation via admixture but instead evolved adaptations via a different biological pathway.

469−477

Authors: Zhen-Zhen Chen;Jian-Ying Wang;Yu Kang;Qiao-Yan Yang;Xue-Ying Gu;Da-Long Zhi; Li Yan;Cheng-Zu Long;Bin Shen;Yu-Yu Niu

Mutations of PTEN-induced kinase I (PINK1) cause early-onset Parkinson’s disease (PD) with selective neurodegeneration in humans. However, current PINK1 knockout mouse and pig models are unable to recapitulate the typical neurodegenerative phenotypes observed in PD patients. This suggests that generating PINK1 disease models in non-human primates (NHPs) that are close to humans is essential to investigate the unique function of PINK1 in primate brains. Paired single guide RNA (sgRNA)/Cas9-D10A nickases and truncated sgRNA/Cas9, both of which can reduce off-target effects without compromising on-target editing, are two optimized strategies in the CRISPR/Cas9 system for establishing disease animal models. Here, we combined the two strategies and injected Cas9-D10A mRNA and two truncated sgRNAs into one-cell-stage cynomolgus zygotes to target the PINK1 gene. We achieved precise and efficient gene editing of the target site in three newborn cynomolgus monkeys. The frame shift mutations of PINK1 in mutant fibroblasts led to a reduction in mRNA. However, western blotting and immunofluorescence staining confirmed the PINK1 protein levels were comparable to that in wild-type fibroblasts. We further reprogramed mutant fibroblasts into induced pluripotent stem cells (iPSCs), which showed similar ability to differentiate into dopamine (DA) neurons. Taken together, our results showed that co-injection of Cas9-D10A nickase mRNA and sgRNA into one-cell-stage cynomolgus embryos enabled the generation of human disease models in NHPs and target editing by pair truncated sgRNA/Cas9-D10A in PINK1 gene exon 2 did not impact protein expression.

482−486

Authors: Jia-Liang Yang;Tong-Dan Zou;Fang Yang;Zheng- Lin Yang;Hou-Bin Zhang

Retinitis pigmentosa (RP) is an inherited retinal degenerative disease that begins with defective rod photoreceptor function, followed by impaired cone function, and complete blindness in its late stage. To date, however, there is no effective treatment for RP. By carrying a nonsense mutation in the Pde6b gene, rd1 mice display elevated cGMP in conjunction with higher intracellular Ca²⁺ in their rod photoreceptors, resulting in fast retinal degeneration. Ca²⁺ has been linked to activation of the mammalian target of rapamycin (mTOR) pathway. The mTOR pathway integrates extracellular and intracellular signals to sense the supply of nutrients and plays a central role in regulating protein and lipid synthesis as well as apoptosis and autophagy. In the present study, we showed that mTOR and phosphorylated mTOR (p-mTOR, activated form of mTOR) are up-regulated in rd1 photoreceptors at postnatal day 10 (P10), a pre-degenerative stage. Moreover, the downstream effectors of mTOR, such as pS6K and S6K, are also increased, suggesting activation of the mTOR signaling pathway. Intravitreal administration of rapamycin, a negative regulator of mTOR, inhibits the mTOR pathway in rd1 photoreceptors. Consequently, the progression of retinal degeneration is slower and retinal function is enhanced, possibly mediated by activation of autophagy in the photoreceptors. Taken together, these results highlight rapamycin as a potential therapeutic avenue for retinal degeneration.

492−501

Authors: Yu-Hang Liao;Chao-Wei Zhou;Wei-Zhen Liu;Jing-Yi Jin;Dong-Ye Li;Fei Liu;Ding-Ding Fan;Yu Zou;Zen-Bo Mu;Jian Shen;Chun-Na Liu;Shi-Jun Xiao;Xiao-Hui Yuan;Hai-Ping Liu

Fish morphological phenotypes are important resources in artificial breeding, functional gene mapping, and population-based studies in aquaculture and ecology. Traditional morphological measurement of phenotypes is rather expensive in terms of time and labor. More importantly, manual measurement is highly dependent on operational experience, which can lead to subjective phenotyping results. Here, we developed 3DPhenoFish software to extract fish morphological phenotypes from three-dimensional (3D) point cloud data. Algorithms for background elimination, coordinate normalization, image segmentation, key point recognition, and phenotype extraction were developed and integrated into an intuitive user interface. Furthermore, 18 key points and traditional 2D morphological traits, along with 3D phenotypes, including area and volume, can be automatically obtained in a visualized manner. Intuitive fine-tuning of key points and customized definitions of phenotypes are also allowed in the software. Using 3DPhenoFish, we performed high-throughput phenotyping for four endemic Schizothoracinae species, including Schizopygopsis younghusbandi, Oxygymnocypris stewartii, Ptychobarbus dipogon, and Schizothorax oconnori. Results indicated that the morphological phenotypes from 3DPhenoFish exhibited high linear correlation (>0.94) with manual measurements and offered informative traits to discriminate samples of different species and even for different populations of the same species. In summary, we developed an efficient, accurate, and customizable tool, 3DPhenoFish, to extract morphological phenotypes from point cloud data, which should help overcome traditional challenges in manual measurements. 3DPhenoFish can be used for research on morphological phenotypes in fish, including functional gene mapping, artificial selection, and conservation studies. 3DPhenoFish is an open-source software and can be downloaded for free at https://github.com/lyh24k/3DPhenoFish/tree/master.

514−524

Authors: De-Zhe Qin;Hui Cai;Chen He;Dong-Hui Yang;Jing Sun;Wen-Lai He;Ba-Lun Li;Jin-Lian Hua;Sha Peng

Normal spermatogenic processes require the scrotal temperature to be lower than that of the body as excessive heat affects spermatogenesis in the testes, reduces sperm quality and quantity, and even causes infertility. Endoplasmic reticulum stress (ERS) is a crucial factor in many pathologies. Although several studies have linked ERS to heat stress, researchers have not yet determined which ERS signaling pathways contribute to heat-induced testicular damage. Melatonin activates antioxidant enzymes, scavenges free radicals, and protects the testes from inflammation; however, few studies have reported on the influence of melatonin on heat-induced testicular damage. Using a murine model of testicular hyperthermia, we observed that heat stress causes both ERS and apoptosis in the testes, especially in the spermatocytes. These observations were confirmed using the mouse spermatocyte cell line GC2, where the Atf6 and Perk signaling pathways were activated during heat stress. Knockout of the above genes effectively reduced spermatocyte damage caused by heat stress. Pretreatment with melatonin alleviated heat-induced apoptosis by inhibiting the Atf6 and Perk signaling pathways. This mitigation was dependent on the melatonin receptors. In vivo experiments verified that melatonin treatment relieved heat-induced testicular damage. In conclusion, our results demonstrated that ATF6 and PERK are important mediators for heat-induced apoptosis, which can be prevented by melatonin treatment. Thus, our study highlights melatonin as a potential therapeutic agent in mammals for subfertility/infertility induced by testicular hyperthermia.