Abstract:Most anurans primarily rely on sound communication for mate choice. To attract females, males might increase the number of syllables or signal complexity to enhance the attractiveness of calls. Working memory refers to memory that involves storing and manipulating information for relatively short time when conducting complex cognitive tasks. Now many studies focus on the functions of the vocal signals, while pay less attention to the effects of working memory on female mate choice and the evolution of complex sexual signals. In this study, phonotaxis experiments were performed to test the working memory of female serrate-legged small treefrogs (Kurixalus odontotarsus, Fig. 1) for different advertisement calls. The treefrogs can produce different calls of graded complexity that contains broadband A note and / or narrowband B note, such as simple advertisement call 5A (a call that contains five A notes) and complex combined calls 5A2B (a call that contains five A notes plus two B notes) / 5A5B (a call that contains five A notes plus five B notes) (Fig. 2). During the experiments, females were exposed to different sound stimulating pairs (5A vs. 5A2B, 5A vs. 5A5B) and then treated with different silent periods (0 s, 5 s, 10 s, 15 s, 30 s) before choosing (Fig. 3). If most females still preferred the speaker that had broadcasted more complex call before, we considered that this silent time was within females’ working memory for more complex call. In addition, the data were processed with generalized estimating equation (GEE) and exact binomial test in SPSS 25.0. As a result, we found that females’ working memory for 5A2B was about 15 s (Fig. 4) and for 5A5B was about 10 s (Fig. 6) compared with 5A, but the comparison between groups showed that there was no significant difference between females’ working memory for 5A2B and 5A5B. The latency of female choosing when broadcasting 5A vs. 5A2B before was significantly longer than that of broadcasting 5A vs. 5A5B before (Fig. 5, Fig. 7). In conclusion, this study demonstrates that working memory of female serrate-legged small treefrogs for complex vocal signals can influence the decision-making of female mate choice, but the effect of working memory on driving the evolution of complex mating signals may be species-specific.

Abstract:Garden plants provide important food resources for urban birds, and the configuration of garden plants will affect the species, number and community structure of birds in the city. The study on the relationship between urban birds and food plants can clarify the feeding relationship between birds and food plants and provide technical support for improving the diversity of urban birds. From 2017 to 2019, the species and quantity of food plants eaten by birds in Beijing were recorded by direct observation and field photo recognition. Six research sites were selected, and one transversal line was set for each study site. The length of transversal line was 2.52 km in Old Summer Palace, 3.12 km in The Summer Palace, 3.02 km in Beijing Botanical Garden, 2.76 km in Olympic Forest Park, 1.38 km in Beijing Forestry University and 1.6 km in Beijing Language and Culture University. A total of 49 species of plant-feeding birds and 56 species of food plants were recorded. The plants that attract the most bird species are Lonicera maackii, followed by Sabina chinensis, Diospyros kaki, Pinus tabuliformis, Ulmus pumila, etc., and the birds that feed on the most plant species are Azure-winged Magpie (Cyanopica cyana), followed by Light-vented Bulbul (Pycnonotus sinensis), Eurasian Tree Sparrow (Passer montanus), Brambling (Fringilla montifringilla), Oriental Magpie (Pica pica) and so on. The garden plants that attract the highest diversity of birds are L. maackii, D. kaki, Pinus tabuliformis, Ginkgo biloba, Populus tomentosa, etc., and the birds that feed on the highest diversity are Azure-winged Magpie, Light-vented Bulbul, Eurasian Tree Sparrow, Brambling, Oriental Magpie and so on. Birds feed on the source plants from October to May of the following year, and the peak period is from November to December, in which the peak of berry feeding is from November to December, the peak of eating nuts is from November to January of the following year, the peak of feeding on leaf buds is from November to December to March to April of the following year, and the peak of feeding on flowers is from March to April. The seasonality of birds feeding on food source plants in Beijing is obvious, so it is necessary to increase the proportion of garden plants that can provide food for birds for a long time in winter, such as Sabina vulgaris, Platycladus orientalis and other food resources to provide sufficient food resources for birds in late winter and early spring. As important food resources for birds living in cities, the assemblage of ornamental plants in gardens and parks plays a significant role in species composition and abundance of urban bird communities. Understanding the relationship between urban birds and edible plants can provide technical support for conservation of urban birds diversity, but remains poorly studied. From 2017 to 2019, species and abundance of plants that birds forage on were recorded in multiple sites in Beijing, by direct observation and field photo identification. The diversities of birds and plants were calculated using Shannon index, and the niche widths were calculated by Levins (1968) model. The chi - square test was used to assess the differences in the type of food that birds forage on among months. A total of 49 species of birds foraging on 56 species of plants were recorded. The plant foraged by most bird species was L. maackii, followed by S. chinensis, D. kaki, Pinus tabuliformis, U. pumila, etc. Among the bird species, Azure-winged Magpie, was recorded foraging on most plant species followed by Light-vented Bulbul, Eurasian Tree Sparrow, Brambling, Common Magpie (Pica pica), etc (Fig. 1). Plant-foraging in birds was concentrated from October to the following May, and peaked in November and December (Fig. 2), among which the main food resources change across months. Berries were mostly foraged from November to December, nuts from November to the following January, leaf buds from November to December and March to April, and the peak of flower-foraging occur from March to April (Fig. 3). Among the recorded plants, S. chinensis and Platycladus orientalis served as the main food resource for a long time in winter. Therefore, in order to increase number of birds in Beijing, the proportions of these plants should be increased to provide sufficient food resources in late winter and early spring when food resources are most scarce.

Abstract:Reintroductions have been increasingly used in wildlife conservation. Understanding movement patterns in the early post-release period is critical for the survival and reestablishment of reintroduced animals. However, there was little information available on early post-release movement of reintroduced Eld’s deer (Cervus eldi hainanus). Using GPS collars for five reintroduced Eld’s deer in the Datian National Nature Reserve, early post-release movement of reintroduced Eld’s deer (2 captive-breeding animals, 3 wild animals) have been studied. We defined early post-release period as the first dry season (March – June) after release to control for potential seasonal effects on deer movements. Movements were compared between each 30-day period. Four time periods were included for captive-breeding deer and three time periods for wild deer as they were released in end of March. Movement and dispersal distance were processed by R package “adehabitatLT” in R3.5.1. Movement rates were compared between 30-day periods using one-way analysis of variance (ANOVA) with post hoc Tukey’s HSD test. Dispersal distance between 30-day periods and movement rates at different times of day were done using one-way analysis of variance (ANOVA) with hoc Bonferroni-Dunn test. The kernelUD method was used to construct 50% core and 95% full home range cumulative utilization distributions using R package “adehabitatHR in R3.5.1. Research results showed that released captive-breeding deer significantly increased movement rates and dispersal distance in fourth time periods (Fig. 1 and 3), and their 50% core and 95% full home range cumulative utilization distributions continued to grow after release (Fig. 4). All the information indicated that captive-breeding deer would continue to disperse and explore after release. Movement rates of released wild deer had no obvious changes (Fig. 1). However, their dispersal distance, 50% core and 95% full home range cumulative utilization distributions significantly reduced in third time periods (Fig. 3 and 4). That indicated that released wild deer returned to the area surrounding the release site after initial exploration. Released captive-breeding deer in general moved more during daytime from release to third time periods, but showed a distinct crepuscular bimodal activity patterns in fourth time periods (Fig. 2). Released wild deer had always been crepuscular bimodal activity patterns after release (Fig. 2). Results indicated that adaptability to new environment of released wild deer would be stronger than released captive-breeding animals.

Abstract:From 2017 to 2020, 63 Ltl-6210 MC infrared cameras were grid-deployed in and around the Zhejiang Jiulongshan National Nature Reserve (Fig. 1) to study the Asiatic Black Bear (Ursus thibetanus) and other sympatric species. During the monitoring period, we obtained nine photographs and six videos of the activity of U. thibetanus from five infrared cameras and extracted seven effective pieces of information regarding U. thibetanus (Fig. 3). Based on our monitoring site data and records of U. thibetanus from literature, a total of 14 locations of U. thibetanus were confirmed in East China. Based on the habitat characteristics of U. thibetanus and correlation of climate factors, land use and seven habitat factors (mean diurnal range, Bio2; maximum temperature of the warmest month, Bio5; minimum temperature of the coldest month, Bio6; mean temperature of the wettest quarter, Bio8; precipitation of the driest month, Bio14; precipitation seasonality, Bio15; and precipitation of the wettest quarter, Bio16) (Fig. 2) were selected as the prediction background, and the potential distribution areas of U. thibetanus in East China were predicted through species distribution models. To develop an accurate projection for U. thibetanus, we used an ensemble modeling approach with 10 algorithms (artificial neural networks, ANN; classification tree analysis, CTA; flexible discriminant analysis, FDA; generalized additive model, GAM; generalized boosting model, GBM; generalized linear model, GLM; multiple adaptive regression splines, MARS; maximum entropy, MaxEnt; random forests, RF; and surface range envelope) using the package ‘biomod 2.’ We evaluated predictive performances of the 10 algorithms using two criteria parameters, true skill statistics (TSS > 0.8) and area under the curve (AUC > 0.9) (Fig. 4). Then, we selected nine algorithms (ANN, CTA, FDA, GAM, GBM, GLM, MARS, MaxEnt, and RF) to estimate the potential suitable distribution of U. thibetanus in East China. Our results showed that (1) 2﹣3 U. thibetanus are speculated to inhabit the nature reserve; (2) the minimum temperature of the coldest month (Bio6) and precipitation of the wettest quarter (Bio16) were the main environmental factors limiting the distribution of U. thibetanus (Fig. 5); and (3) there are three potential distribution areas of U. thibetanus in East China, including Zhejiang-Jiangxi, Zhejiang-Jiangxi-Anhui, and Zhejiang-Anhui (Fig. 6). In conclusion, these results can serve as basic information for strengthening the protective measures for U. thibetanus conservation in East China.

Abstract:The green turtle (Chelonia mydas) is a globally endangered species and listed as Class Ⅰ of protected animals in China. Habitat conservation is an important and effective conservation strategy for this species. However, the habits and habitat of juvenile green turtles in China are not clear yet. We six tagged juvenile green turtles aged from 2 to 14 years old from the Huidong National Sea Turtle Reserve of China with six satellite transmitters, including two American Wildlife Computer trackers (Spot5- AM-S181C, n = 1; Spot5-AM-S244A, n = 1), and four Telonics trackers (TAM-2639) (Table 1) between 2011 and 2015. We released them in the summer (n = 3) and in the winter (n = 3). The global ARGOS system was used to carry out an all-weather tracking and monitoring, and the geographic coordinate data were filtered and analyzed using the satellite tracking and analysis tool (STAT) software. We obtained a total of 397 effective locations for the four years (including 62 obtained from previous studies). GIS layers were then generated and overlaid with MapTool software to reveal the migration of juvenile green turtles. The SPSS software t-test was used to determine statistical difference between the two season data on individual size and minimum travel speeds The curve carapace lengths (CCL) in the turtle released in winter and in summer were with 48.9 ± 8.65 cm (n = 3), and 59.07 ± 7.64 cm (n = 3), respectively, with no difference between the two groups (t = 1.527, df = 4, P = 0.202). The average tracking period was 66 ± 47 d (n = 6), and the tracking distance was 1 653 ±1 585 km (n = 6). The average minimum travel speed in the summer group was 1.63 ± 0.22 km/h (n = 3), which is longer than those turtles in winter group (0.67 ± 0.12 km/h, n = 3; t = 6.726, df = 4, P = 0.003). They migrated to Taiwan in summer and to Hainan in winter, along and within the 140 m isobath of the continental shelf. The migration directions of all individuals were as same as the monsoon currents of the sea during the same period (Fig. 1). Homing behavior was observed only in the winter group (Fig. 2). The over-winter sites were mainly distributed in Con Co Island of Vietnam and the eastern part of Hainan Island, whereas the over-summer habitats were located between Fujian and Zhejiang, but all were within the 20 m isobath and with concentrated distribution of macro-algae, seagrass beds, or coral reefs. The activity area overlapped with the offshore fishing ground and other development areas, which was consistent with the results of previous research. The captive juvenile green turtles could migrate normally, forage, and adapt to the wild environment after being released, and they tended to migrate seasonally between the East China Sea and South China Sea. The foraging sites are primarily distributed along the coast of the continental shelf and affiliated islands of China, which further confirms the existence of migratory corridors for green turtles offshore of China. We suggest integrating and formulating marine national parks off the Beibu Gulf, Eastern Hainan, Guangdong, Fujian, Taiwan and Zhejiang Marine Nature Reserves, strengthening international and regional cooperation to protect the natal grounds, foraging grounds, migratory corridor, and other habitats of green turtles, to ensure effective protection of their life cycle.

Abstract:Castration surgery was accomplished on muskrats (Ondatra zibethicus) to observe the changes in scent gland morphology and musk compositions after castration. Six adult male muskrats were randomly divided into castrated and control groups (3 in each group). The musk in scent glands was drained, and then both testes were removed under general anesthesia. Thirty days after the castration, the muskrats were anesthetized and killed, and the serum and the musk were collected. The testosterone (T) concentration in serum and muskrat musk was detected by enzyme-linked immunosorbent assay (ELISA). The scent glands were collected, and their lengths were measured. The paraffin sections of scent glands were stained by the H.E method, and the diameters of glandular cells were measured. And then, the compositions of muskrat musk were detected by gas chromatography-mass spectrometry (GC-MS). The comparison of differences between the two groups was analyzed with independent-samples T test method using SPSS 20.0. The concentration of testosterone in the serum of the castrated group was significantly lower than that of the control group (t = 5.270, P < 0.01), and the testosterone content of muskrat musk in the castrated group was also significantly lower than that of the control group (t = 3.229, P < 0.05) (Table 1). Morphologically, the lengths of muskrat scent glands in the castrated group and the control group were similar and there was no significant difference (t = 1.243, P > 0.05). Histologically, the Diameter of glandular cells of the castrated group was significantly reduced compared with the control group (t = 9.434, P < 0.01) (Fig. 2). The GC-MS analysis showed that both groups contained three rich compositions: macrocyclics, steroids and fatty acids. Among them, the content of macrocyclics in the castrated group was significantly higher (t =﹣3.084, P < 0.05), while the content of steroids was significantly reduced (t = 3.407, P < 0.05) (Table 2). Thus, the muskrat castration model was established, in which the morphology of the muskrat scent gland cells and the compositions of muskrat musk changed.

Abstract:Reproductive life-history traits of reptiles are the important areas of animal evolutionary biology, but such studies in viviparous snakes are still scarce in China. In the current study, we detected the annual variation in reproductive life-history traits in a viviparous snake, Hypsiscopus plumbea, which was widely distributed in southern China. From 2013 to 2015, we brought 117 pregnant snakes in Wenzhou to our laboratory in July each year (2013: n = 42; 2014: n = 46; 2015: n = 29), and measured the morphological traits of postpartum females and neonates after birth. We used one-way ANOVA or ANCOVA with post-hoc Tukey’s test to compare differences in life-history traits among years, and used linear regression to assess the relationship between two variables. Our results showed that females in 2013 had largest snout-vent length (Fig. 1a) and body mass (Fig. 1b) after birth. Body condition of postpartum females in 2015, which year was hot and dry (Table 1), was worse than those of other years (Fig. 1c). Both litter size and litter mass were significantly positively correlated to maternal snout-vent length (Fig. 2a, b), but no significant annual variation was found in litter size (Fig. 3a) or litter mass (Fig. 3b). The neonatal snout-vent length and body mass were not related to maternal snout-vent length. Neonates born in 2015 had largest snout-vent length (Fig. 3c) and body mass (Fig. 3d). No significant relationship was found between offspring size and litter size. In conclusion, annual variation in morphological traits of postpartum females and neonates in H. plumbea may be related to the local climate conditions, such as temperature and rainfall. Moreover, reproductive effort and offspring size in H. plumbea may have been optimized by natural selection.

Abstract:Four species of hypotrichous ciliates isolated from farmland soils in Gansu, Cyrtohymena citrina (Fig. 1 and 2), Gastrostylides dorsicirratus (Fig. 3 and 4), Vermioxytricha arenicola (Fig. 5﹣7) and Hemisincirra sp. (Fig. 8 and 9), were studied morphologically using non-flooded Petri dish method, live observation and protargol staining methods. Among the species, C. citrine, G. dorsicirratus and V. arenicola are new record species of ciliates in China and H. sp. is kept as an unnamed form owing to the lack of some key taxonomic data and morphogenetic as well molecular information, which are required for accurate identification. Their morphological character were described in details and discussed.

Abstract:In multi-male, multi-female primate societies, although the sexual behaviors of adult males and females are not confined to the female's fertile period, it is believed that sexual behaviors are most active during the fertile period. For primate species in which females conceal their estrus, the relationship between sexual behaviors of male and female individuals and the female menstrual cycle is unclear. In this study, Tibetan Macaques (Macaca thibetana) were used to investigate the relationship between sexual behaviors and menstrual cycle stages in female primates with concealed estrus. During the breeding season of Tibetan Macaques, 490 fecal samples were collected from 6 adult female macaques at the Tongling Zoo in Anhui Province from October 2, 2019 to January 14, 2020 (Fig. 1). The concentration of estradiol hormone in feces was determined by radioimmunoassay. The sexual behavior data of target individuals (4 females and 4 males) were collected by focus animal sampling method and behavior sampling method. The Spearman correlation coefficient was used to test the correlation between estrogen and sexual behavior. The independent sample t-test was used to analyze the distribution differences of preceptive indicators (female presentation), attractive indicators (male sexual-inspection) and copulations in female Tibetan Macaques during three stages of the menstrual cycle (pre-fertile phase: 5 days before fertile phase; fertile phase: 2 days when ovulation is most likely plus 3 preceding days to account for sperm life span; post-fertile phase: 5 days after fertile phase). The results showed that the estradiol concentration of female Tibetan macaques fluctuated periodically during the breeding season, with a cycle length of 31.1 ± 1.8 d (n = 7, range: 23﹣37 d); During the menstrual cycle, estradiol concentration was negatively correlated with female presentation (r =﹣0.616, n = 15, P = 0.015), but had no significant correlation with copulation and male sexual-inspection. The average frequency of female presentation was mainly concentrated in the pre-fertile phase (T =﹣1.215, df = 12, P = 0.044). There was no significant difference between copulation and male sexual-inspection in the three stages of the menstrual cycle. The sexual behavior of Tibetan macaques was not concentrated in the fertile period of female, suggesting that estrogen has no strict control over the sexual behavior of Tibetan macaques and that male Tibetan macaques can not accurately identify the fertile period of female.

Abstract:Small mammals in the wild often show physiological and behavioral changes in response to seasonal environmental variations, such as food, temperature, and photoperiod. To investigate the physiological and behavioral adaptation strategies of Eothenomys miletus in different regions of Hengduan Mountain regions in response to different sugar foods, E. miletus in Jianchuan and Xianggelila were fed a high-sugar diet for 28 d and returned to a standard food for another 28 d, and then their body masses, food intakea, resting metabolic rates (RMR) and activity behaviors were measured. Moreover, serum leptin levels, hypothalamic neuropeptide gene expressions and body compositions were measured on day 0, 28 and 56. Food intake was measured by food balance method, hypothalamic neuropeptide gene expression was measured by real-time fluorescence quantitative polymerase chain reaction (RT-PCR). Data were analyzed by two-way ANOVA or two-way ANCOVA, and associations were judged by Pearson-correlation analysis. The results showed that high-sugar diet could significantly increase the body mass (F1, 907 = 8.11, P < 0.01) (Fig. 1) and food intake (F1, 907 = 1034.94, P < 0.01) (Fig. 2) of the E. miletus in the two regions, but had no significant effect on RMR and activity behavior. After refeeding standard food, the body mass of Xianggelila E. miletus could be restored to the level of standard diet group, while the body mass of Jianchuan E. miletus was still higher. Leptin was positively correlated with body mass (r = 0.80, P < 0.01) (Fig. 5), but not with neuropeptide expression (P > 0.05). In addition, the physiological characteristics of the E. miletus in the two regions also showed regional differences: The E. miletus in Xianggelila had lower body mass (F1, 907 = 842.02, P < 0.01) (Fig. 1) than those in Jianchuan region, but RMR (F1, 907 = 6779.51, P < 0.01) (Fig. 3) and activity behavior (F1, 907 = 79.89, P < 0.01) (Fig. 4) were higher than in Jianchuan area, the higher food intake (F1, 907 = 49.96, P < 0.01) (Fig. 2) may be related to the higher expression of neuropeptide Y (F1, 36 = 4.672, P < 0.05) (Table 3). All of the results showed that body mass of E. miletus in two regions increased when they were exposed to the high-sugar food, but change of body mass in the two regions was significantly different after refeeding, showing a great difference between E miletus in the two regions. Leptin and NPY expression levels may play an important role in body mass regulation and energy balance in E. miletus. Location-determined environmental factors (food resources, temperature, and altitude) may be critical for determining phenotypic differences between biological regions and their adaptation to extreme environmental conditions.

Abstract:Seasonal adjustments of metabolic thermogenesis is common in endotherms, which is important to energy allocation, survival, reproduction and other aspects of these animals. To understand the response of thermoregulation to climate change, we used a continuous flow respirometry system to determine thermoregulatory characteristics including body temperature, basal metabolic rate, thermal neutral zone and thermal conductance in the winter-acclimatized Common Rosefinch Carpodacus erythrinus, which were captured in breeding ground and in captivity for 3 years (2018﹣2020). The results analyzed by one-way ANOVA showed that their body temperatures (Tb) maintained at 42.9 ± 0.15 ℃, when the air temperature (Ta) ranged between 10﹣25 ℃; and Tb increased when the Ta upper this range, and decreased to 41.6 ± 0.13 ℃ at the Ta of 5 ℃ (Fig. 1). The resting metabolic rate increased significantly with the descending of Ta when the Ta lower than 20 ℃, and when Ta exceeded 35 ℃. Metabolic rate maintained lowest and stable when the Ta ranged between 20-35 ℃ (Fig. 2), therefore, this temperature range should be the thermal neutral zone (TNZ) in this bird, with the basal metabolic rate (BMR) of 6.5 ± 0.3 ml/(g·h). Thermal conductance increased with the upgrade of Ta (Fig. 3) and with the least value of 0.25 ± 0.02 ml/(g·h·℃). Compared to the previously reports, our results indicated that the Common Rosefinch changed their TNZ wider and increased metabolic thermogenesis to maintain their higher Tb in winter, and could survival in the temperate conditions with abundant food.

Abstract:Hynobius maoershanensis and Pachytriton intexpectatus are endemic to Guangxi Maoershan National Nature Reserve, China. Their living environments and distribution altitudes are different. H. maoershanensis is inhabited in an alpine swamp at an altitude of 2 000 m with small population size, but P. intexpectatus lives in mountain streams with gentle currents at an altitude of 1 140﹣1 800 m. This study was designed to explore the differences of digestive tracts between H. maoershanensis and P. intexpectatus, and how the 5-hydroxytryptamine cells of digestive tract adapt to their feeding habits and living environments. Anatomical, histological and envision two-step immunohistochemistry staining techniques were used. The total length, head length, body weight, snout-vent length, digestive tract length, and the thickness of mucosa and muscle layers were measured. Statistical analysis and inspection were performed using the Kolmogorov-Simirnov test, Kruskal-Wallis test and Mann-Whitney U test (Microsoft Excel 2016 and SPSS 26.0 software). Results showed that the digestive tracts of H. maoershanensis and P. intexpectatus were composed of esophagus, stomach, duodenum, ileum and rectum (Fig. 1). The digestive tract of P. intexpectatus was longer than that of H. maoershanensis. The ratios of intestine length and snout-vent length in H. maoershanensis and P. intexpectatus were 0.57 ± 0.03 (n = 5) and 0.84 ± 0.03 (n = 5), respectively. The histological structure of the digestive tracts in H. maoershanensis and P. intexpectatus could be divided into mucosa, submucosa, muscularis and adventitia (Fig. 2). There were many glands, such as pyloric glands (Fig. 2e﹣h). The thicknesses of mucosa layer in the stomach body of H. maoershanensis was the highest (712.82 ± 37.67 μm, n = 5), while that of P. intexpectatus was in the cardia (403.24 ± 55.81 μm, n = 5). The thickness of muscular layer of the digestive tract was the highest in the pylorus, with that of P. intexpectatus thicker than in H. maoershanensis. Both open and closed 5-HT cells were distributed in the digestive tracts of H. maoershanensis and P. intexpectatus (Fig. 3), while the distributional patterns of 5-HT cells along the digestive tracts were different. 5-HT cells showed the highest density in the rectum and the least density in the esophagus of H. maoershanensis. But in P. intexpectatus, the highest count was found in the duodenum but the least count in the rectum. In conclusion, the morphological and histological structures of the digestive tracts of H. maoershanensis and P. intexpectatus are similar. The ratios of intestine length to snout-vent length of both were not more than 1, which is consistent with the characteristics of the short digestive tract of carnivorous animals. The 5-HT cells in the digestive tracts of both have endocrine and exocrine functions, but the distributive patterns of these cells have their own characteristics, which may be related to their different feeding habits and living environments.

Abstract:In order to explore the effect of red light on the formation of skin pigment cells, CRISPR/Cas9 gene editing technology was used to construct a zebrafish line with homozygous deletion of the red light opsin 2 gene opn1lw2. A red LED lamp with a light intensity of 800 ± 100 lx was used to irradiate 15-day-old AB strain wild-type zebrafish (Danio rerio) and opn1lw2-/- strain zebrafish for 60 days, the number of melanocytes in the dorsum skin of wild-type and mutant zebrafish were counted with Photoshop CS5，and the data were analysed using GrahPad Prsim6. The results showed that the number of melanocytes in the dorsum skin of wild-type zebrafish was significantly more than that of the mutant (Fig. 2a﹣c). Real-Time quantitative PCR (RT-PCR) analysis found that the expression of the melanocyte marker gene kit in the dorsum skin of wild-type zebrafish was significantly higher than that of the mutant (Fig. 2d). There was no significant difference in the expression of the xanthophore marker gene csf1ra and the iridophores marker gene pnp4a of the dorsum skin between wild zebrafish and the opn1lw2-/- mutant (Fig. 3c, d), indicating that red light can regulate the formation of melanocytes in the dorsum skin of zebrafish through the opn1lw2, but does not affect the formation of skin xanthophores and iridophores; moreover, it regulates the differentiation of melanocytes. The expression of the precursor gene pomca of α-MSH in the dorsum skin of opn1lw2-/- zebrafish was significantly lower than that of the wild type (Fig. 2e), indicating that red light regulates the expression of pomca gene through opn1lw2 to induce the formation of melanocyte; RT-PCR detection found that the expression of retinal dehydrogenase raldh3 in wild-type zebrafish skin was significantly higher than that of mutant opn1lw2-/- (Fig. 4b), while there was no difference in expression of raldh2 encoding retinal dehydrogenase (Fig. 4a), indicating that opn1lw2 may selectively mediate the red light-induced synthesis of retinoic acid to regulate the formation of melanocytes. These results are importantfor understanding the formation of pigment cells in fish skin induced by red light irradiation.

Abstract:The China-Vietnam border region is a well-known biodiversity hotspot, harboring a striking diversity of species and endemism. New species and new records have been reported on both sides of the border in recent years. Although Paramesotriton deloustali was first recorded in China in 2017, only the complete mitochondrial sequence was provided, and no other information was available, including its detailed distribution in China and morphological characteristics. Here we describe P. deloustali from China based on six newly collected specimens (four males and two females) from Dawishan Reserve (22°37′ N, 103°52′ E, altitude 350 m) in Hekou County, Honghe Hani and Yi Autonomous Prefecture, Yunnan Province (Table 1). The morphological characters of the Chinese specimens align closely with the type series from Vietnam (Fig. 1, 2), and the phylogenetic analyses based on the cytochrome coxidase subunit I (COI) gene revealed a low genetic differentiation (p-distance is 0.6%) between Chinese population and those from its type locality at Tam Dao, Vinh Phuc Province, northern Vietnam. Additionally, our newly collected specimens have low genetic divergence (p-distance is 0.5%) from the sequences of the former record in Yunnan (Fig. 3). P. deloustali is currently recorded in the China-Vietnam border area only, and it is found in a narrow area with unknown population size and trends. We recommend further international collaboration between China and Vietnam to understand the distribution and conservation situation of this species better.

Abstract:An unidentified snake was collected at a freshly reclaimed hillside in Qianwanguan Township, Leibo County, Sichuan Province (103°50′ E, 28°15′ N) during a reptile diversity survey in Sichuan Province in late July 2020. We conducted Bayesian and maximum likelihood phylogenetic analyses basis on a mitochondrial cytochrome b (Cyt b) DNA sequence to ascertain the phylogenetic position of this specimen (p-distance ≥ 3.6%). Furthermore, the newly specimen can be diagnosed as Psammodynastes pulverulentus by a combination of pholidosis characters, which is the new snakes fauna in Sichuan Province. On the basis of 17 individuals of P. pulverulentus from Sichuan, Guangxi, Guangdong, Yunnan, Hainan and Namibia, we recognize for the first time a complex intraspecific phylogeny of P. pulverulentus, and to better solve this issue, more detailed samples and more markers are needed.

Abstract:Here we introduce a new distribution location of Elysia leucolegnote in Hainan province, China. We have identified this species by its external characters and molecular approaches, recorded its behavior in its habitat location. We used the targeted PCR method to amplify the cytochrome oxidase subunit I (COI) gene. Based on the sequence of COI, a phylogenetic tree of the Elysia genus species reported in China has been constructed. This sea slug has a body length from 5﹣13 mm, the body size increases from May to November. Small herds of E. leucolegnote with 1 to 100 individuals can be easily found in the puddles of the mangrove swamp at low tide or in the crab holes for shading when these were exposed to air (Fig. 1); The external characters of E. leucolegnote are identical to the original reports, the adult sea slug is green in color with white-edged parapodia and white spots (Fig. 2); The sequence of COI gene of the sea slugs collected in Hainan conformed this species is E. leucolegnote. We uploaded the sequence of cytochrome oxidase subunit I (COI) of mitochondrial DNA (GenBank accession number MT933184). And in phylogenetic tree (neighbor-joining, NJ) 100% degree of confidence of branch shows our sea slugs were E. leucolegnote (Fig. 3). We suggest that E. leucolegnote may be potentially used as a new model system for studying chloroplast endocytosis and endosymbiosis. Therefore, this is also a unique system that may be employed in gene expression and regulation in chloroplasts. Furthermore, this model system may also be utilized in research studies on animal cell immunity.

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