Zoological Research

525−547

Authors: Manuel Ruiz-García; Daniel Pinilla-Beltrán; Oscar E. Murillo-García; Christian Miguel Pinto; Jorge Brito; Joseph Mark Shostell5

The species within Xenarthra (sloths, anteaters, and armadillos) are quintessential South American mammals. Of the three groups, Vermilingua (anteaters) contains the fewest extant and paleontological species. Here, we sampled and sequenced the entire mitochondrial genomes (mitogenomes) of two Tamandua species ( Tamandua tetradactyla and T. mexicana ) ( n =74) from Central and South America, as well as Myrmecophaga tridactyla ( n =41) from South America. Within Tamandua , we detected three different haplogroups. The oldest (THI) contained many specimens with the T. tetradactyla morphotype (but also several with the T. mexicana morphotype) and originated in southeastern South America (currently Uruguay) before moving towards northern South America, where the THII haplogroup originated. THII primarily contained specimens with the T. mexicana morphotype (but also several with the T. tetradactyla morphotype) and was distributed in Central America, Colombia, and Ecuador. THI and THII yielded a genetic distance of 4%. THII originated in either northern South America or “ in situ ” in Central America with haplogroup THIII, which consisted of ~50% T. mexicana and 50% T. tetradactyla phenotypes. THIII was mostly located in the same areas as THII, i.e., Central America, Ecuador, and Colombia, though mainly in the latter. The three haplogroups overlapped in Colombia and Ecuador. Thus, T. tetradactyla and T. mexicana were not reciprocally monophyletic. For this reason, we considered that a unique species of Tamandua likely exists, i.e., T. tetradactyla . In contrast to Tamandua , M. tridactyla did not show different morphotypes throughout its geographical range in the Neotropics. However, two very divergent genetic haplogroups (MHI and MHII), with a genetic distance of ~10%, were detected. The basal haplogroup, MHI, originated in northwestern South America, whereas the more geographically derived haplogroup, MHII, overlapped with MHI, but also expanded into central and southern South America. Thus, Tamandua migrated from south to north whereas Myrmecophaga migrated from north to south. Our results also showed that temporal mitochondrial diversification for Tamandua began during the Late Pliocene and Upper Pleistocene, but for Myrmecophaga began during the Late Miocene. Furthermore, both taxa showed elevated levels of mitochondrial genetic diversity. Tamandua showed more evidence of female population expansion than Myrmecophaga . Tamandua experienced population expansion ~0.6–0.17 million years ago (Mya), whereas Myrmecophaga showed possible population expansion ~0.3–0.2 Mya. However, both taxa experienced a conspicuous female decline in the last 10 000–20 000 years. Our results also showed little spatial genetic structure for both taxa. However, several analyses revealed higher spatial structure in Tamandua than in Myrmecophaga . Therefore, Tamandua and Myrmecophaga were not subjected to the same biogeographical, geological, or climatological events in shaping their genetic structures.

562−573

Authors: Zu-Bing Cao1; Di Gao1; Hui-Qun Yin2; Hui Li1; Teng-Teng Xu1; Meng-Ya Zhang1; Xin Wang1; Qiu-Chen Liu1; Ye-Lian Yan1; Yang-Yang Ma1; Tong Yu1; Yun-Sheng Li1; Yun-Hai Zhang1;

Inositol requiring mutant 80 (INO80) is a chromatin remodeler that regulates pluripotency maintenance of embryonic stem cells and reprogramming of somatic cells into pluripotent stem cells. However, the roles and mechanisms of INO80 in porcine pre-implantation embryonic development remain largely unknown. Here, we show that INO80 modulates trophectoderm epithelium permeability to promote porcine blastocyst development. The INO80 protein is highly expressed in the nuclei during morula-to-blastocyst transition. Functional studies revealed that RNA interference (RNAi)-mediated knockdown of INO80 severely blocks blastocyst formation and disrupts lineage allocation between the inner cell mass and trophectoderm. Mechanistically, single-embryo RNA sequencing revealed that INO80 regulates multiple genes, which are important for lineage specification, tight junction assembly, and fluid accumulation. Consistent with the altered expression of key genes required for tight junction assembly, a permeability assay showed that paracellular sealing is defective in the trophectoderm epithelium of INO80 knockdown blastocysts. Importantly, aggregation of 8-cell embryos from the control and INO80 knockdown groups restores blastocyst development and lineage allocation via direct complementation of the defective trophectoderm epithelium. Taken together, these results demonstrate that INO80 promotes blastocyst development by regulating the expression of key genes required for lineage specification, tight junction assembly, and fluid accumulation

592−605

Authors: Sheng-Yu Luo; Cheng Liu; Jie Ding; Xin-Ming Gao; Jing-Qian Wang; Yi-Bo Zhang; Chen Du; Cong-Cong Hou; Jun-Quan Zhu; Bao Lou; Xiong-Fei Wu; Wei-Liang Shen

The large yellow croaker (Larimichthys crocea), which is an economically important mariculture fish in China, is often exposed to environmental hypoxia. Reactive oxygen species (ROS) homeostasis is essential for the maintenance of normal physiological conditions in an organism. Direct evidence that environmental hypoxia leads to ROS overproduction is scarce in marine fish. Furthermore, the sources of ROS overproduction in marine fish under hypoxic stress are poorly known. In this study, we investigated the effects of hypoxia on redox homeostasis in L. crocea and the impact of impaired redox homeostasis on fish. We first confirmed that hypoxia drove ROS production mainly via the mitochondrial electron transport chain and NADPH oxidase complex pathways in L. crocea and its cell line (large yellow croaker fry (LYCF) cells). We subsequently detected a marked increase in the antioxidant systems of the fish. However, imbalance between the pro-oxidation and antioxidation systems ultimately led to excessive ROS and oxidative stress. Cell viability showed a remarkable decrease while oxidative indicators, such as malondialdehyde, protein carbonylation, and 8-hydroxy-2 deoxyguanosine, showed a significant increase after hypoxia, accompanied by tissue damage. N-acetylcysteine (NAC) reduced ROS levels, alleviated oxidative damage, and improved cell viability in vitro. Appropriate uptake of ROS scavengers (e.g., NAC and elamipretide Szeto-Schiller-31) and inhibitors (e.g., apocynin, diphenylene iodonium, and 5-hydroxydecanoate) may be effective at overcoming hypoxic toxicity. Our findings highlight previously unstudied strategies of hypoxic toxicity resistance in marine fish.

614−619

Authors: Wen-Ting Wan; Zhi-Wei Dong; Yan-Dong Ren; Jie Yang; Xiang-Yu Pan; Jin-Wu He; Zhou Chang; Wei Liu; Gui-Chun Liu; Ruo-Ping Zhao; Ping Hu; Chu- Yang Mao; Jun Li; Wen Wang; Xue-Yan Li

Butterflies are diverse in virtually all aspects of their ontogeny, including morphology, life history, and behavior. However, the developmental regulatory mechanisms underlying the important phenotypic traits of butterflies at different developmental stages remain unknown. Here, we investigated the developmental regulatory profiles of butterflies based on transposase accessible chromatin sequencing (ATAC-seq) at three developmental stages in two representative species (Papilio xuthus and Kallima inachus). Results indicated that 15%–47% of open chromatin peaks appeared in associated genes located 3 kb upstream (i.e., promoter region) of their transcription start site (TSS). Comparative analysis of the different developmental stages indicated that chromatin accessibility is a dynamic process and associated genes with differentially accessible (DA) peaks show functions corresponding to their phenotypic traits. Interestingly, the black color pattern in P. xuthus 4th instar larvae may be attributed to promoter peak-related genes involved in the melanogenesis pathway. Furthermore, many longevity genes in 5th instar larvae and pupae showed open peaks 3 kb upstream of their TSS, which may contribute to the overwintering diapause observed in K. inachus adults. Combined with RNA-seq analysis, our data demonstrated that several genes enriched in the melanogenesis and longevity pathways also exhibit higher expression, confirming that the expression of genes may be closely related to their phenotypic traits. This study offers new insights into larval cuticle color and adult longevity in butterflies and provides a resource for investigating the developmental regulatory mechanisms underlying butterfly ontogeny.

626−632

Authors: Zhi-Jin Liu; Xue-Kun Qian; Min-Heng Hong1; Jia- Li Zhang; Da-Yong Li; Tian-Han Wang; Zuo-Min Yang; Li-Ye Zhang; Zi-Ming Wang; Hua-Jian Nie; Ke- Yue Fan; Xiong-Fei Zhang; Meng-Meng Chen; Wei- Lai Sha; Christian Roos; Ming Li

Viruses can be transmitted from animals to humans (and vice versa) and across animal species. As such, host-virus interactions and transmission have attracted considerable attention. Non-human primates (NHPs), our closest evolutionary relatives, are susceptible to human viruses and certain pathogens are known to circulate between humans and NHPs. Here, we generated global statistics on virus infections in NHPs (VI-NHPs) based on a literature search and public data mining. In total, 140 NHP species from 12 families are reported to be infected by 186 DNA and RNA virus species, 68.8% of which are also found in humans, indicating high potential for crossing species boundaries. The top 10 NHP species with high centrality in the NHP-virus network include two great apes (Pan troglodytes, Pongo pygmaeus) and eight Old World monkeys (Macaca mulatta, M. fascicularis, M. leonina, Papio cynocephalus, Cercopithecus ascanius, C. erythrotis, Chlorocebus aethiops, and Allochrocebus lhoesti). Given the wide distribution of Old World monkeys and their frequent contact with humans, there is a high risk of virus circulation between humans and such species. Thus, we suggest recurring epidemiological surveillance of NHPs, specifically Old World monkeys that are in frequent contact with humans, and other effective measures to prevent potential circulation and transmission of viruses. Avoidance of false positives and sampling bias should also be a focus in future work.

637−649

Authors: Yong Cui; Zu-Lian Liu; Cen-Cen Li; Xiang-Min Wei; Yong-Jian Lin; Lang You; Zi-Dan Zhu; Hui-Min Deng; Qi-Li Feng; Yong-Ping Huang; Hui Xiang

The insect brain is the central part of the neurosecretory system, which controls morphology, physiology, and behavior during the insect’s lifecycle. Lepidoptera are holometabolous insects, and their brains develop during the larval period and metamorphosis into the adult form. As the only fully domesticated insect, the Lepidoptera silkworm Bombyx mori experienced changes in larval brain morphology and certain behaviors during the domestication process. Hormonal regulation in insects is a key factor in multiple processes. However, how juvenile hormone (JH) signals regulate brain development in Lepidoptera species, especially in the larval stage, remains elusive. We recently identified the JH receptor Methoprene tolerant 1 (Met1) as a putative domestication gene. How artificial selection on Met1 impacts brain and behavioral domestication is another important issue addressing Darwin’s theory on domestication. Here, CRISPR/Cas9-mediated knockout of Bombyx Met1 caused developmental retardation in the brain, unlike precocious pupation of the cuticle. At the whole transcriptome level, the ecdysteroid (20-hydroxyecdysone, 20E) signaling and downstream pathways were overactivated in the mutant cuticle but not in the brain. Pathways related to cell proliferation and specialization processes, such as extracellular matrix (ECM)-receptor interaction and tyrosine metabolism pathways, were suppressed in the brain. Molecular evolutionary analysis and in vitro assay identified an amino acid replacement located in a novel motif under positive selection in B. mori, which decreased transcriptional binding activity. The B. mori MET1 protein showed a changed structure and dynamic features, as well as a weakened co-expression gene network, compared with B. mandarina. Based on comparative transcriptomic analyses, we proposed a pathway downstream of JH signaling (i.e., tyrosine metabolism pathway) that likely contributed to silkworm larval brain development and domestication and highlighted the importance of the biogenic amine system in larval evolution during silkworm domestication.

660−665

Authors: Sheng-Yong Xu; Zhi-Chuang Lu; Bin-Bin Cao; Dong- Dong Han; Shan-Shan Cai; Zhi-Qiang Han; Tian-Xiang Gao

Flatheads are valuable commercial fish species endemic to the Indo-West Pacific. Due to their economic value and unique biological traits, such as metamorphosis and camouflage, they serve as ideal marine organisms for studies on demersal adaptation and evolution. The brown-spotted flathead (Platycephalus sp.1) is the most widely distributed in the northwestern Pacific. Despite the lack of a valid scientific name, it has been long recognized and exploited in the marine fisheries of China, Japan, and Korea. In the current study, we applied Illumina, PacBio, and Hi-C sequencing to assemble a chromosome-scale genome for this species. The assembled genome was 660.63 Mb long with a scaffold N50 of 28.65 Mb and 100% of the contigs were anchored onto 24 chromosomes. We predicted 22 743 protein-coding genes, 94.8% of which were functionally annotated. Comparative genomic analyses suggested that Platycephalus sp.1 diverged from its common ancestor with Gasterosteus aculeatus ~88.4 million years ago. The expanded gene families were significantly enriched in immune, biosynthetic, and metabolic pathways. Furthermore, three shared Gene Ontology (GO) terms and 377 common positively selected genes were identified between flathead and flatfish species, suggesting that these genes may contribute to demersal adaptation in flatheads. The assembled genomic data provide a valuable molecular resource for further research on the biological and adaptive evolution of flathead species.

671−674

Authors: Yuan-Ting Guo; Jia Zhang; Dong-Ming Xu; Li-Zhou Tang; Zhen Liu;

All extant species in the rodent family Spalacidae are subterranean and have evolved various traits for underground life. However, the phylogenomic relationships among its three subfamilies (Myospalacinae, Spalacinae, and Rhizomyinae) and the molecular basis underlying their adaptations to underground life remain poorly understood. Here, we inferred the phylogenomic relationships among these subfamilies based on de novo sequencing the genome of the hoary bamboo rat (Rhizomys pruinosus). Analyses showed that ~50% of the identified 11 028 one-to-one orthologous protein-coding genes and the concatenated sequences of these orthologous genes strongly supported a sister relationship between Myospalacinae and Rhizomyinae. The three subfamilies diversified from each other within ~2 million years. Compared with the non-subterranean controls with similar divergence dates, the spalacids shared more convergent genes with the African subterranean mole-rats at the genomic scale due to more rapid protein sequence evolution. Furthermore, these convergent genes were enriched in the functional categories of carboxylic acid transport, vascular morphogenesis, and response to oxidative stress, which are closely associated with adaptations to the hypoxic-hypercapnic underground environment. Our study presents a well-supported phylogenomic relationship among the three subfamilies of Spalacidae and offers new insights into the molecular adaptations of spalacids living underground.