Multiple concordant cytonuclear divergences and potential hybrid speciation within a species complex in Asia.

Various environmental factors impact the distribution, population structure, demography and evolutionary trajectory of a bird species, leading to genetic and morphological divergences between populations across its distribution. The Paradoxornis webbianus species complex is found throughout much of East Asia, where its geographically distinct populations exhibit dramatic morphological variation. This has resulted in a hotly debated taxonomy. This study intended to identify genetic divergence patterns and their underlying contributing factors for this species complex. We collected 243 birds, whose data was combined with those available in GenBank to perform phylogeographic analyses using one mitochondrial and six nuclear loci. Six mitochondrial clades were observed in the species complex, while individual-based Bayesian clustering using nuclear markers showed multiple congruent breaks. Overall, the six molecular lineages could be recognized as independent species under the lineage species concept in view of genetic divergence, clade-specific morphological changes and distribution: P. webbianus, P. w. bulomachus, P. alphonsianus, P. a. ganluoensis, P. brunneus brunneus and P. b. ricketti. The estimated divergence times range from 0.46 to 3.36 million years ago, suggesting it was likely impacted by paleoclimatic changes. Interestingly, P. alphonsianus carries two divergent mitochondrial lineages shared with P. webbianus and P. a. ganluoensis, respectively, and analyses based on nuclear loci found a similar pattern. We discussed the various hypotheses for this pattern and argued that P. alphonsianus was likely the result of hybridization between P. webbianus and P. a. ganluoensis. Further data on genome, transcriptome and breeding ecology are needed to address the hypothesis of hybrid speciation and its underlying mechanisms.

The absence of reproductive isolation between non-sister and deeply diverged mitochondrial lineages of the black-throated tit (Aegithalos concinnus) revealed by a multilocus genetic analysis in a contact zone.

BACKGROUND: A deep divergence of mitochondrial DNA is common in species delimitated by morphological traits. Several hypotheses can explain such variations, such as cryptic species, introgression, allopatric divergence and ancestral lineage. The black-throated tit harbors several deeply divergent mitochondrial lineages. Two lineages with 5% divergence, but having a high level of gene flow, have been detected in its subspecies, A. C. concinnus and A. c. talifuenses. In this study, we conducted a genetic analysis at a contact zone of these two lineages to identify whether these lineages either reflect a high level of intraspecific variation in mitochondrial loci or represent incipient speciation. Mitochondrial ND2 and 11 microsatellite loci were used to conduct phylogenetic and population structure analyses. RESULTS: ND2 haplotypes actually diverged into two groups within subspecies A. c. talifuenses; however, they formed a non-sister relationship when including all available GenBank ND2 sequences. Analyses of microsatellite data indicated no existing population structure and showed a pattern of isolation by distance. Individuals sampled at the contact zone were almost identified as F2 hybrids. CONCLUSIONS: Isolation for 2.4 Ma, as suggested by a previous study, appeared to be insufficient to develop robust reproductive barriers. Reproductive barriers were weak, or even absent between the divergent lineages, highlighting that incipient speciation was unlikely to be the case. Considering the results from previous studies, the divergent lineages may be better explained by secondary contact after allopatric isolation because of Pleistocene climate changes, but other hypotheses cannot be definitively ruled out because of the lack of representative samples from the other distribution region and its relatives. Considering that divergence in morphology was hardly observed and the particular split in genetics, the two subspecies might be better merged. The current findings also highlight the likely contribution of male-biased dispersal to male-biased gene flow among mitochondrial lineages; more efforts are needed to illustrate the evolutionary history of the black-throated tit.

Past hybridization between two East Asian long-tailed tits (Aegithalos bonvaloti and A. fuliginosus).

INTRODUCTION: Incomplete lineage sorting and hybridization are two major nonexclusive causes of haplotype sharing between species. Distinguishing between these two processes is notoriously difficult as they can generate similar genetic signatures. Previous studies revealed that the mitochondrial DNA (mtDNA) differentiation between two East Asian long-tailed tits (Aegithalos bonvaloti and A. fuliginosus) was extremely low, even lower than intraspecific differentiation in some other long-tailed tits. Using a combination of multilocus and coalescent analyses, we explored the causes of the anomalous lack of mtDNA differentiation between the two species. RESULTS: The mtDNA divergence between the two species was shallow, while the nuclear DNA (nuDNA) divergence was considerably deeper. The IMa analyses based on the mtDNA dataset suggested relatively high gene flow from A. fuliginosus to A. bonvaloti, while negligible gene flow in the opposite direction. In contrast to mtDNA, the migration rates at autosomal and Z-linked nuDNA loci were negligible or much lower. The NEWHYBRIDS analysis assigned all individuals except one to pure parental species with high posterior probability. The Bayesian skyline plot showed that both species underwent population expansions during the Last Glacial Maximum (LGM), and the ecological niche modelling suggested that their ranges overlapped more during the LGM than at present. CONCLUSIONS: We suggest that historical hybridization, in combination with selective sweep and/or genetic drift might be the main causes of the extremely low mtDNA differentiation between the two species. The hybridization probably occurred mainly between A. fuliginosus females and A. bonvaloti males. The LGM distribution expansion might have facilitated hybridization, while the post-LGM distribution contraction could have facilitated some mtDNA sorting. Ongoing hybridization between the two species might be very limited, but further studies with more samples from the contact zone are needed to test this conclusion.

Entomophagy and entomo-therapeutic practices in a mountainous territory in southeast Guangxi Zhuang Autonomous Region, China.

BACKGROUND: Although China has a long history of using insects as food and medicine and has developed numerous associated knowledge and practices, especially in its rural and mountainous areas, systematic surveys concerning this subject are limited. In-depth ethnobiological research is needed to compile a comprehensive database of edible and medicinal insects and record the associated knowledge of these food and medicinal resources. METHODS: Data on edible and medicinal insects and associated knowledge about them were collected by interviewing 216 local villagers in a mountainous territory in southeast Guangxi Zhuang Autonomous Region, China. RESULTS: Local villagers used at least 16 edible and 9 medicinal insects, of which 4 wasp species were used in both entomophagy and medicinal practices. Parapolybia varia, Polistes olivaceus, and Anomala chamaeleon were newly recorded edible insects in China. The wasps, Euconocephalus sp., Gryllotalpa orientalis, and Cyrtotrachelus longimanus, were preferred and culturally important edible insects. Populations of Euconocephalus sp. and G. orientalis were reported to have substantially decreased in recent years. Wasps and a bamboo bee were used to treat rheumatism, while cockroaches and antlions were used to treat common cold symptoms in infants. Insect-related knowledge was positively correlated with the interviewees' age. CONCLUSIONS: Villagers have accumulated considerable local and traditional knowledge of entomophagy and entomo-therapeutic practices. However, this knowledge is in danger of being lost, which highlights the urgent need to document this information. Edible insects enrich local diets, and a more sustainable supply (such as through insect farming) could maintain local entomophagy practices. Medicinal insects are a part of local folk medicine, and pharmacological and chemical techniques could be applied to identify various biologically active substances in these insects.

Comparison of the Gut Microbial Communities of Domestic and Wild Mallards (Anas platyrhynchos) Based on High-Throughput Sequencing Technology.

Mallards (Anas platyrhynchos) are currently one of the most popular species in rare bird breeding in several southern provinces of China, but there have been no studies comparing the gut microbial communities of domestic and wild mallards. In this study, 16S rRNA gene high-throughput sequencing technology was used to compare the composition and diversity of gut microbial communities in domestic and wild mallards. Alpha diversity analysis showed significant differences in gut microbial communities between the two groups of mallards, and the diversity and richness of gut microbial communities were significantly higher in wild mallards than in domestic mallards. Beta diversity analysis showed that the two groups of stool samples were mostly separated on the principal coordinate analysis (PCoA) plot. In domestic mallards, Firmicutes (68.0% ± 26.5%) was the most abundant bacterial phylum, followed by Proteobacteria (24.5% ± 22.9%), Bacteroidetes (3.1% ± 3.2%), Fusobacteria (2.2% ± 5.9%), and Actinobacteria (1.1% ± 1.8%). The dominant bacterial phyla in wild mallards were Firmicutes (79.0% ± 10.2%), Proteobacteria (12.9% ± 9.5%), Fusobacteria (3.4% ± 2.5%), and Bacteroidetes (2.8% ± 2.4%). At the genus level, a total of 10 dominant genera (Streptococcus, Enterococcus, Clostridium, Lactobacillus, Soilbacillus, Bacillus, Acinetobacter, Comamonas, Shigella, and Cetobacterium) with an average relative abundance greater than 1% were detected in the fecal samples of both groups. The average relative abundance of five potential pathogenic genera (Streptococcus, Enterococcus, Acinetobacter, Comamonas, and Shigella) was higher in domestic mallards than in wild mallards. The enrichment of pathogenic bacteria in the intestinal tract of domestic mallards should be of sufficient concern.

The evolution of bitter taste receptor gene in primates: Gene duplication and selection.

Bitter taste perception plays an important role in preventing animals from digesting poisonous and harmful substances. In primates, especially the Cercopithecidae species, most species feed on plants; thus, it is reasonable to speculate that most of the bitter taste receptor genes (T2Rs) of primates are under purifying selection to maintain the functional stability of bitter taste perception. Gene duplication has happened in T2Rs frequently, and what will be the fate of T2Rs copies is another question we are concerned about. To answer these questions, we selected the T2Rs of primates reported in another study and conducted corresponding selective pressure analyses to determine what kind of selective pressure was acting on them. Further, we carried out selective pressure analyses on gene copies and their corresponding ancestors by considering several possible situations. The results showed that among the 25 gene groups examined here, 15 groups are subject to purifying selection and others are under relaxed selection, with many positively selected sites detected. Gene copies existed in several groups, but only some groups (clade1_a1-b2, clade1_c-c2, clade1_d1-d3, clade1_f1-f2, T2R10, T2R13, and T2R42) have positively selected sites, inferring that they may have some relation to functional divergence. Taken together, T2Rs in primates are under diverse selective pressures, and most gene copies are subject to the same selective pressures. In such cases, the copies may be just to keep the function conservative, and more copies can increase the quantity of the bitter taste receptor, raise the efficiency of bitter substance recognition, and finally enhance the fitness of feeding during the evolutionary course of primates. This study can improve our understanding of T2Rs evolution in primates.

Incorporating local stakeholders' voices and knowledge into conservation decisions: a case study on the Chinese Hwamei (Garrulax canorus Linnaeus, 1758) in Taijiang, Guizhou, China.

BACKGROUND: The Chinese Hwamei (Garrulax canorus Linnaeus, 1758) is a widely distributed species and has long been kept as a pet, especially by the ethnic communities in Southwest China. According to conservation experts' suggestions, it has been designated as a second-level national key protected species in February, 2021 to protect this bird, indicating that keeping it at home is no longer permitted in China. However, a key factor to ensure effectiveness and success of conservation initiatives is local stakeholders' acceptance and support. METHODS: Interviews and focus group discussions were used to document the policy outcomes and the views of 108 local bird-keepers in a county in Guizhou province. RESULTS: Despite awareness about the illegality of the practice, the bird was still commonly caged both in rural and urban regions. To justify their unwillingness to stop keeping these birds, the interviewees presented many arguments, such as benefits for the community members' health, cultural heritage and contributions to local livelihoods. Fewer than 30% of the bird-keepers believed that the practice of self-keeping has reduced the wild population. Most argued the decline was mainly generated by the harvesting and keepers with monetary interests. They suggested enforcement should target those people and bird markets, as well as the harvesting methods. They also recommended restricting the number of birds allowed to be kept by one keeper, establishing protected areas and a harvesting ban period. The study participants demonstrated considerable local ecological knowledge about approaches for managing the species' use. CONCLUSIONS: Due to the benefits for the people and the bird's large distribution, I argued that a conservation goal to lower the harvesting and keeping rates would be more appropriate than a strict ban on keeping them. Such a policy would be more feasible and culturally acceptable because it is built on keepers' support and suggestions. It is necessary to monitor the effects of bird keeping on the wild population. Overall, this qualitative study demonstrated the advantage of factoring in local voices in conservation decisions.

Students' knowledge of and conservation attitude toward the black-necked crane (Grus nigricollis) in Guizhou, China: insights for conservation.

BACKGROUND: The persistence of threatened and protected wildlife depends not only on habitat suitability but also remarkably on local communities' acceptance. The black-necked crane (Grus nigricollis) is a flagship species for conservation on the plateau in western China. However, the human dimension has been completely ignored in the decision-making process for conservation. METHODS: In this study, a questionnaire survey aiming to assess knowledge of and conservation attitude toward this bird was carried out among 1042 students of 7th and 9th grade from 12 schools in Weining county, Guizhou province, which has a large wintering population in an urban wetland. Logistic regression was used in the generalized linear model to identify the determinants that significantly affect students' knowledge of and conservation attitude toward this species. RESULTS: Most students have positive attitudes toward conservation, which is significantly affected by awareness, knowledge of this bird and grade. However, they have somewhat limited knowledge of this bird's biology and ecosystem services (nature's contributions to people). Knowledge was found to link with observation of the bird and grade, while observation related to the experience of visiting the wetland. Social media is the most cited resource to obtain knowledge on this bird. CONCLUSION: It is suggested that local conservation experts could help introduce more information on the black-necked crane in the schools and help conduct outdoor education activities in and around the wetland. Traditional knowledge and culture could also be incorporated into the conservation awareness enhancing program. This study focuses on the human dimension for conserving the black-necked crane in China, showing significant implications in the design and application of effective measurements to improve students' perception and attitude toward its conservation. Future assessments should include other local populations, such as farmers, fishers, and urban citizens.

An unstructured phylogeographic pattern with extensive gene flow in an endemic bird of South China: collared Finchbill (Spizixos semitorques).

Recent phylogeographical studies indicated that glacial oscillations played a key role on the phylogeographic pattern of extant species. As most studies have previously been carried out on heavily ice-covered regions, such as in European and North American regions, potential effects of climatic oscillations on species that are distributed on ice-free regions are less known. To address this, we investigated the phylogeographic pattern of an avian species endemic to South China, which was not glaciated during the Pleistocene glaciations. By using 2142 bp mitochondrial DNA, we identified 89 haplotypes defined by 39 polymorphic sites. A combination of high haplotype diversity (0.786-1.00) and low nucleotide diversity (0.00132-0.00252) was detected among geographic populations. Explicit genetic divergence was observed between S. s. semitorques and S. s. cinereicapillus but not detected among geographic populations of S. s. semitorques. Divergence time of the two subspecies was dated back to 87 Kyr which is congruent with the interglacial MIS 5. A weak phylogeographic structure due to strong gene flow among geographic populations was identified in this species, suggesting complex topography of South China has not formed barriers for this species.

Profound climatic effects on two East Asian black-throated tits (Ave: Aegithalidae), revealed by ecological niche models and phylogeographic analysis.

Although a number of studies have assessed the effects of geological and climatic changes on species distributions in East Asian, we still have limited knowledge of how these changes have impacted avian species in south-western and southern China. Here, we aim to study paleo-climatic effects on an East Asian bird, two subspecies of black-throated tit (A. c. talifuensis-concinnus) with the combined analysis of phylogeography and Ecological Niche Models (ENMs). We sequenced three mitochondrial DNA markers from 32 populations (203 individuals) and used phylogenetic inferences to reconstruct the intra-specific relationships among haplotypes. Population genetic analyses were undertaken to gain insight into the demographic history of these populations. We used ENMs to predict the distribution of target species during three periods; last inter-glacial (LIG), last glacial maximum (LGM) and present. We found three highly supported, monophyletic MtDNA lineages and different historical demography among lineages in A. c. talifuensis-concinnus. These lineages formed a narrowly circumscribed intra-specific contact zone. The estimated times of lineage divergences were about 2.4 Ma and 0.32 Ma respectively. ENMs predictions were similar between present and LGM but substantially reduced during LIG. ENMs reconstructions and molecular dating suggest that Pleistocene climate changes had triggered and shaped the genetic structure of black-throated tit. Interestingly, in contrast to profound impacts of other glacial cycles, ENMs and phylogeographic analysis suggest that LGM had limited effect on these two subspecies. ENMs also suggest that Pleistocene climatic oscillations enabled the formation of the contact zone and thus support the refuge theory.