Mitochondrial Genomic Evidence of Selective Constraints in Small-Bodied Terrestrial Cetartiodactyla.

Body size may drive the molecular evolution of mitochondrial genes in response to changes in energy requirements across species of different sizes. In this study, we perform selection pressure analysis and phylogenetic independent contrasts (PIC) to investigate the association between molecular evolution of mitochondrial genome protein-coding genes (mtDNA PCGs) and body size in terrestrial Cetartiodactyla. Employing selection pressure analysis, we observe that the average non-synonymous/synonymous substitution rate ratio (ω) of mtDNA PCGs is significantly reduced in small-bodied species relative to their medium and large counterparts. PIC analysis further confirms that ω values are positively correlated with body size (R2 = 0.162, p = 0.0016). Our results suggest that mtDNA PCGs of small-bodied species experience much stronger purifying selection as they need to maintain a heightened metabolic rate. On the other hand, larger-bodied species may face less stringent selective pressures on their mtDNA PCGs, potentially due to reduced relative energy expenditure per unit mass. Furthermore, we identify several genes that undergo positive selection, possibly linked to species adaptation to specific environments. Therefore, despite purifying selection being the predominant force in the evolution of mtDNA PCGs, positive selection can also occur during the process of adaptive evolution.

Analysis of the Complete Mitochondrial Genome of Pteronura brasiliensis and Lontra canadensis.

P. brasiliensis and L. canadensis are two otter species, which successfully occupied semi-aquatic habitats and diverged from other Mustelidae. Herein, the full-length mitochondrial genome sequences were constructed for these two otter species for the first time. Comparative mitochondrial genome, selection pressure, and phylogenetic independent contrasts (PICs) analyses were conducted to determine the structure and evolutionary characteristics of their mitochondrial genomes. Phylogenetic analyses were also conducted to confirm these two otter species' phylogenetic position. The results demonstrated that the mitochondrial genome structure of P. brasiliensis and L. canadensis were consistent across Mustelidae. However, selection pressure analyses demonstrated that the evolutionary rates of mitochondrial genome protein-coding genes (PCGs) ND1, ND4, and ND4L were higher in otters than in terrestrial Mustelidae, whereas the evolutionary rates of ND2, ND6, and COX1 were lower in otters. Additionally, PIC analysis demonstrated that the evolutionary rates of ND2, ND4, and ND4L markedly correlated with a niche type. Phylogenetic analysis showed that P. brasiliensis is situated at the base of the evolutionary tree of otters, and then L. canadensis diverged from it. This study suggests a divergent evolutionary pattern of Mustelidae mitochondrial genome PCGs, prompting the otters' adaptation to semi-aquatic habitats.

Toxicological evaluation of TBBPA by common carp (Cyprinus carpio) about the in vivo/vitro disturbance of the AHR pathway.

Tetrabromobisphenol A (TBBPA) is a widely used plastic additive with high bioaccumulation potential and toxicity on both humans and wildlife. Currently, research on its ecotoxicity and the underlying mechanism is limited. Using common carp (Cyprinus carpio), we evaluated the toxicity of TBBPA, especially focusing on its alteration of a key metabolism-related pathway aryl hydrocarbon receptor (AHR), using in vivo/vitro assays and in silico simulation. The 96 h LC50 of TBBPA of common carp was 4.2 mg/L and belonged to the acute toxic level II. The bioaccumulation potential of TBBPA follows the role of liver > gill > brain and varies between 3- and 14-day exposure. On the AHR pathway respect, as expected, the metabolism-related cyp1a1 and cyp1b1 were upregulated in the liver and brain. Ahr2, the receptor, was also upregulated in the brain under TBBPA exposure. The alteration of gene expression was tissue-specific while the difference between 3- or 14-day exposure was minor. AHR inhibition assay indicated the 2, 3, 7, 8-tetrachlorodibenzo-p-dioxin (TCDD)-induced AHR transactivation can be inhibited by TBBPA suggesting it is not a potent agonist but a competitive antagonist. In silico analysis indicated TBBPA can be successfully docked into the binding cavity with similar poses but still have AHR-form-specific interactions. Molecular dynamics simulation proved TBBPA can be more flexible than the coplanar ligand TCDD, especially in ccaAHR1b with greater root-mean-square deviation (RMSD), of which TCDD-induced transactivation seemed not to be blocked by TBBPA. This research increased the understanding of TBBPA toxicity and alteration of the AHR pathway, and pointed out the need to perform additional toxicology evaluation of emerging contaminants, especially on non-model species.

Functional characterization of common carp (Cyprinus carpio) AHRs: Subform-specific sensitivity to dioxin and interspecies differences.

Dioxins are widely known to bioaccumulate in the body and produce a wide spectrum of toxic effects on both humans and wildlife. In addition, some novel sorts of compounds that were similar in structure and effect were gradually identified and termed dioxin-like compounds (DLCs). The toxicity of dioxins as well as DLCs is predominantly mediated by the dioxin receptor (aryl hydrocarbon receptor, AHR) in animals, which is usually differentially expressed and functionally distinct (especially the sensitivity to dioxins) among species, possibly resulting in species-specific variations in the toxicity of dioxins. Therefore, detailed functional exploration of the AHRs of a given species, such as the common carp (which is a vital wild and commercial species with a broad geological distribution) in the current study, will enable a comprehensive ecotoxicity evaluation. Through genome survey and phylogenetic analysis, we identified three AHRs (AHR1a, AHR1b, and AHR2) and two ARNTs (ARNT1 and ARNT2). AHR2 was observed to have greater expression abundance in the gill and brain, and may serve as the predominant subform. Those AHRs and ARNTs are functional, and the AHRs can be efficiently transactivated by the classical dioxin congener 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). We individually determined the EC50 values of AHR1a (0.41 ± 0.24 nM), AHR1b (12.80 ± 3.28 nM), and AHR2 (0.64 ± 0.49 nM), and found that: 1) The AHR sensitivities of common carp and zebrafish (phylogenetically close species) are relatively similar. AHR1a and the predominant form AHR2 have greater sensitivity to TCDD. 2) ARNT1 and ARNT2 do not produce different sensitivities, but with distinct induction fold, of a given AHR transactivation when cooperating as the partner; 3) Distinct AHR subforms of the same or distinct species can have even one or two orders of magnitude differences in sensitivity. In summary, the current study will add to the knowledge of AHR biology and help improve ecotoxicology research on dioxins and DLCs.

A chromosome-level genome assembly of the yellow-throated marten (Martes flavigula).

The yellow-throated marten (Martes flavigula) is a medium-sized carnivore that is widely distributed across much of Asia and occupies an extensive variety of habitats. We reported a high-quality genome assembly of this organism that was generated using Oxford Nanopore and Hi-C technologies. The final genome sequences contained 215 contigs with a total size of 2,449.15 Mb and a contig N50 length of 68.60 Mb. Using Hi-C analysis, 2,419.20 Mb (98.78%) of the assembled sequences were anchored onto 21 linkage groups. Merqury evaluation suggested that the genome was 94.95% complete with a QV value of 43.75. Additionally, the genome was found to comprise approximately 39.74% repeat sequences, of which long interspersed elements (LINE) that accounted for 26.13% of the entire genome, were the most abundant. Of the 20,464 protein-coding genes, prediction and functional annotation was successfully performed for 20,322 (99.31%) genes. The high-quality, chromosome-level genome of the marten reported in this study will serve as a reference for future studies on genetic diversity, evolution, and conservation biology.

Regional distribution of the plastic additive tris(butoxyethyl) phosphate in Nanyang Lake estuary, China, and toxic effects on Cyprinus carpio.

There is increasing concern regarding the toxicological effects of plastic additives on humans and aquatic organisms. This study investigated effects of the plastic additive tris(butoxyethyl) phosphate (TBEP) on Cyprinus carpio by measuring concentration distribution of TBEP in the Nanyang Lake estuary, as well as toxic effects of varying doses of TBEP exposure on carp liver. This also included measuring responses of superoxide dismutase (SOD), malondialdehyde (MDA), tumor necrosis factor-α (TNF-α), interleukin-1ß (IL-1ß), and cysteinyl aspartate-specific protease (caspase). Concentrations of TBEP in the polluted water environment (water company inlets, urban sewage pipes, etc.) in the survey area were as high as 76.17-3875.29 µg/L, and 3.12 µg/L in the river flowing through the urban area, and 1.18 µg/L in the estuary of the lake. In the subacute toxicity test, SOD activity in liver tissue with an increase in TBEP concentration was reduced significantly, while the MDA content continued to increase with an increase in TBEP concentration. Inflammatory response factors (TNF-α and IL-1ß) and apoptotic proteins (caspase-3 and caspase-9) gradually increased with increasing concentrations of TBEP. Additionally, reduced organelles, increased lipid droplets, swelling of mitochondria, and disorder of mitochondrial cristae structure were observed in liver cells of TBEP-treated carp. Generally, TBEP exposure induced severe oxidative stress in carp liver tissue, resulting in release of inflammatory factors and inflammatory response, mitochondrial structure changes, and the expression of apoptotic proteins. These findings benefit our understanding about the toxicological effects of TBEP in aquatic pollution.

Environmental factors and stochasticity affect the fungal community structures in the water and sediments of Hulun Lake, China.

Aquatic fungi form both morphologically and ecologically diverse communities. However, lake ecosystems are frequently overlooked as fungal habitats, despite the potentially important role of fungi in matter cycling and energy flow. Hulun Lake is a typical example of a seasonal glacial lake; however, previous studies have only focused on bacteria in this ecosystem. Therefore, in the current study, internal transcribed spacer ribosomal RNA (ITS rRNA) gene high-throughput sequencing was used to investigate the fungal communities in paired water and sediment samples from the Hulun Lake Basin in China. A significant difference was found between the fungal communities of the two sample types. Across all samples, we identified nine phyla, 30 classes, 78 orders, 177 families, and 307 genera. The dominant phyla in the lake were Ascomycota, Basidiomycota and Chytridiomycota. Our results show that both water and sediments have very high connectivity, are dominated by positive interactions, and have similar interaction patterns. The fungal community structures were found to be significantly affected by environmental factors (temperature, chemical oxygen demand, electrical conductivity, total phosphorus, and pH). In addition, the dispersal limitations of the fungi affected the structure of the fungal communities, and it was revealed that stochasticity is more important than deterministic mechanisms in influencing the structure and function of fungal communities. This study provides unique theoretical support for the study of seasonally frozen lake fungal communities and a scientific basis for the future management and protection of Hulun Lake.

Adaptability and Evolution of Gobiidae: A Genetic Exploration.

The Gobiidae family occupy one of the most diverse habitat ranges of all fishes. One key reason for their successful colonization of different habitats is their ability to adapt to different energy demands. This energy requirement is related to the ability of mitochondria in cells to generate energy via oxidative phosphorylation (OXPHOS). Here, we assembled three complete mitochondrial genomes of Rhinogobius shennongensis, Rhinogobius wuyanlingensis, and Chaenogobius annularis. These mitogenomes are circular and include 13 protein-coding genes (PCGs), two rRNAs, 22 tRNAs, and one non-coding control region (CR). We used comparative mitochondrial DNA (mtDNA) genome and selection pressure analyses to explore the structure and evolutionary rates of Gobiidae mitogenomics in different environments. The CmC model showed that the ω ratios of all mtDNA PCGs were <1, and that the evolutionary rate of adenosine triphosphate 8 (atp8) was faster in Gobiidae than in other mitochondrial DNA PCGs. We also found evidence of positive selection for several sites of NADH dehydrogenase (nd) 6 and atp8 genes. Thus, divergent mechanisms appear to underlie the evolution of mtDNA PCGs, which might explain the ability of Gobiidae to adapt to diverse environments. Our study provides new insights on the adaptive evolution of Gobiidae mtDNA genome and molecular mechanisms of OXPHOS.

Toxic effect of triphenyl phosphate (TPHP) on Cyprinus carpio and the intestinal microbial community response.

Triphenyl phosphate (TPHP) is a kind of organophosphorus flame retardants, and its use is increasing annually. However, the toxic effect associated with exposure to it has not been adequately investigated. Therefore, in this study, we determined the toxic dose of TPHP in the economic fish species, Cyprinus carpio. Acute and subacute toxicity tests were conducted, and the enrichment of TPHP in the gills, brain, intestines, and liver were determined by Liquid Chromatography-Mass Spectrometry, and the response of carp gut microbial community to TPHP stress was determined using 16 S rRNA gene high-throughput sequencing. Results showed that the 96-h LC50 of TPHP in carp was 7 mg/L. At the 7 d, the order of TPHP absorption was as follows (from highest to lowest): gills > intestine > liver > brain, but at the 28 d and the purification period, the order of TPHP absorption was brain > gills > intestine > liver. TPHP exposure at 3.5 mg/L decreased α-diversity of the intestinal microbial community (p < 0.05), and altered community composition, in particular the relative abundance of dominant microbial populations. Functional profiles of the microbial communities predicted based on 16 S rRNA gene data showed upregulation in the degradation of exogenous substances and energy metabolism of the TPHP-treated groups (p < 0.05), suggesting that intestinal microbial taxa play a role in reducing TPHP toxicity. The results provide insights that could facilitate risk assessments of TPHP pollutants in aquatic environments and the management of associated water pollution.

Effects of shining pondweed (Potamogeton lucens) on bacterial communities in water and rhizosphere sediments in Nansi Lake, China.

Submerged macrophytes and microbial communities are important parts of lake ecosystems. In this study, the bacterial community composition in rhizosphere sediments and water from areas cultivated with (PL) and without (CK) shining pondweed (Potamogeton lucens Linn.) was investigated to determine the effects of P. lucens Linn. on the structure of the bacterial communities in Nansi Lake, China. Molecular techniques, including Illumina MiSeq and qPCR targeting of the 16S rRNA gene, were used to analyze the composition and abundance of the bacterial community. We found that bacterial alpha diversity was higher in PL water than in CK water, and the opposite trend was observed in sediment. In addition, 16S rRNA gene copy number in sediment was lower in PL than in CK. We found 30 (e.g., Desulfatiglans) and 29 (e.g., Limnohabitans) significantly different genera in sediment and water, respectively. P. lucens Linn. can change chemical properties in sediment and water and thereby affect the bacterial community. At the genus level, members of bacterial community clustered according to source (water/sediment) and area (PL/CK). Our study demonstrated that submerged macrophytes can affect the bacterial community composition in both sediment and water, suggesting that submerged macrophytes affect the transportation and cycling of nutrients in lake ecosystems.

Comparative Analyses of the Gut Microbiome of Two Fox Species, the Red Fox (Vulpes Vulpes) and Corsac Fox (Vulpes Corsac), that Occupy Different Ecological Niches.

The gut microbiome is integral for the host's living and environmental adaptation and crucially important for understanding host adaptive mechanisms. The red fox (Vulpes vulpes) dominates a wider ecological niche and more complicated habitat than that of the corsac fox (V. corsac). However, the adaptive mechanisms (in particular, the gut microbiome responsible for this kind of difference) are still unclear. Therefore, we investigated the gut microbiome of these two species in the Hulunbuir grassland, China, and evaluated their microbiome composition, function, and adaptive mechanisms. We profiled the gut microbiome and metabolism function of red and corsac foxes via 16S rRNA gene and metagenome sequencing. The foxes harbored species-specific microbiomes and functions that were related to ecological niche and habitat. The red fox had abundant Bacteroides, which leads to significant enrichment of metabolic pathways (K12373 and K21572) and enzymes related to chitin and carbohydrate degradation that may help the red fox adapt to a wider niche. The corsac fox harbored large proportions of Blautia, Terrisporobacter, and ATP-binding cassette (ABC) transporters (K01990, K02003, and K06147) that can help maintain corsac fox health, allowing it to live in harsh habitats. These results indicate that the gut microbiome of the red and corsac foxes may have different abilities which may provide these species with differing capabilities to adapt to different ecological niches and habitats, thus providing important microbiome data for understanding the mechanisms of host adaptation to different niches and habitats.

High-altitude adaptation in vertebrates as revealed by mitochondrial genome analyses.

The high-altitude environment may drive vertebrate evolution in a certain way, and vertebrates living in different altitude environments might have different energy requirements. We hypothesized that the high-altitude environment might impose different influences on vertebrate mitochondrial genomes (mtDNA). We used selection pressure analyses and PIC (phylogenetic independent contrasts) analysis to detect the evolutionary rate of vertebrate mtDNA protein-coding genes (PCGs) from different altitudes. The results showed that the ratio of nonsynonymous/synonymous substitutions (dN/dS) in the mtDNA PCGs was significantly higher in high-altitude vertebrates than in low-altitude vertebrates. The seven rapidly evolving genes were shared by the high-altitude vertebrates, and only one positive selection gene (ND5 gene) was detected in the high-altitude vertebrates. Our results suggest the mtDNA evolutionary rate in high-altitude vertebrates was higher than in low-altitude vertebrates as their evolution requires more energy in a high-altitude environment. Our study demonstrates the high-altitude environment (low atmospheric O2 levels) drives vertebrate evolution in mtDNA PCGs.

Analysis of gut microbiota in three species belonging to different genera (Hemitragus, Pseudois, and Ovis) from the subfamily Caprinae in the absence of environmental variance.

This study aimed to identify the effects of host species on the gut microbial flora in three species (Hemitragus jemlahicus, Pseudois nayaur, and Ovis orientalis) from the subfamily Caprinae, by excluding the impact of environment factors. We investigated the differences in intestinal flora of three species belonging to Caprinae, which were raised in identical conditions. Fecal samples were collected from tahr, mouflon, and bharal, and the V3-V4 region of the 16S ribosomal RNA gene was analyzed by high-throughput sequencing. The analysis of 16S rRNA gene sequences reveals that fecal samples were mainly composed of four phyla: Firmicutes, Bacteroidetes, Spirochaetes, and Proteobacteria. The most abundant phyla included Firmicutes and Bacteroidetes accounting for >90% of the bacteria, and a higher Firmicutes/Bacteroidetes ratio was observed in tahrs. Moreover, significant differences existed at multiple levels of classifications in the relative abundance of intestinal flora, differing greatly between species. Phylogenetic analyses based on 16S rRNA gene indicated that mouflon is closely related to bharal, and it is inconsistent with previous reports in the species evolutionary relationships. In this study, we demonstrated that the gut microbiota in tahr had a stronger ability to absorb and store energy from the diet compared with mouflon and bharal, and the characteristics of host-microbiome interactions were not significant.

Global view on virus infection in non-human primates and implications for public health and wildlife conservation.

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 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.

FGF gene family characterization provides insights into its adaptive evolution in Carnivora.

Fibroblast growth factors (FGFs) encoded by the FGF gene family can regulate development and physiology in animals. However, their evolutionary characteristics in Carnivora are largely unknown. In this study, we identified 660 sequences of three types of FGF genes from 30 unannotated genomes of Carnivora animals (before 7th May 2020), and the FGF genes from 52 Carnivora species were analyzed through the method of comparative genomics. Phylogenetic and selective pressure analyses were carried out based on the FGF genes of these 52 Carnivora species. The phylogenetic analysis results demonstrated that the FGF gene family was divided into 10 subfamilies and that FGF5 formed one clade rather than belonging to the subfamilies of FGF4 and FGF6. The evolutionary analysis results showed that the FGF genes were prominently subjected to purifying selection and were highly conserved in the process of Carnivora evolution. We also carried out phylogenetic comparative analyses, which indicated that the habitat was one of the factors that shaped the evolution of Carnivora FGF genes. The FGF1 and FGF6 genes were positively selected in the Carnivora animals, and positive selection signals were detected for the FGF19 gene in semiaquatic Carnivora animals. In summary, we clarified the phylogenetic and evolutionary characteristics of Carnivora FGF genes and provided valuable data for future studies on evolutionary characterization of Carnivora animals.

Biomarker responses and histological damage in the gill, liver, and gonad of Cyprinus carpio with benzo(a)pyrene exposure.

The risk of polycyclic aromatic hydrocarbon exposure in aquatic organisms is a global concern. In this study, we investigated the toxic effects of different doses of benzo(a)pyrene (BaP) on Cyprinus carpio in microcosms from the following aspects: superoxide dismutase (SOD) and peroxidase (POD) activity, malondialdehyde (MDA) content in the gill, liver, and gonad; glutathione s-transferase (GST), aromatic hydroxylase (AHH), and 7-ethoxyresorufin-O-deethylase (EROD) activity in the liver; and altered tissue and cellular structures of the gill, liver,and gonad. SOD and POD activity in the gill, liver, and gonad increased in low-dose BaP groups and significantly decreased with an increase in BaP. MDA content increased continuously with an increase in BaP in the gill, liver, and gonad. The activity of enzymes related to detoxification, specifically GST, AHH, and EROD, gradually increased in the liver with an increase in BaP. Upon exposure to BaP, gill hypertrophy, bulging, necrosis, and cavitation occurred, gonadal cells became larger, with an increase in pyknotic or vacuolar nuclei, bulging and cavitation of organelles, and cytoplasm leakage, and nuclear membrane lysis was observed in the liver. Collectively, BaP exposure changed the SOD and POD activity in the gill, liver, and gonad of carp with increases in MDA content, increased GST, AHH, and EROD activity in liver, and damaged the tissue and cellular structures of the gill, liver, and gonad, revealing the toxic effects of BaP exposure on carp.

Comparison of bacterial and archaeal communities in two fertilizer doses and soil compartments under continuous cultivation system of garlic.

Soil microbial communities are affected by interactions between agricultural management (e.g., fertilizer) and soil compartment, but few studies have considered combinations of these factors. We compared the microbial abundance, diversity and community structure in two fertilizer dose (high vs. low NPK) and soil compartment (rhizosphere vs. bulk soils) under 6-year fertilization regimes in a continuous garlic cropping system in China. The soil contents of NO3- and available K were significantly higher in bulk soil in the high-NPK. The 16S rRNA gene-based bacterial and archaeal abundances were positively affected by both the fertilizer dose and soil compartment, and were higher in the high-NPK fertilization and rhizosphere samples. High-NPK fertilization increased the Shannon index and decreased bacterial and archaeal richness, whereas the evenness was marginally positively affected by both the fertilizer dose and soil compartment. Soil compartment exerted a greater effect on the bacterial and archaeal community structure than did the fertilization dose, as demonstrated by both the nonmetric multidimensional scaling and redundancy analysis results. We found that rhizosphere effects significantly distinguished 12 dominant classes of bacterial and archaeal communities, whereas the fertilizer dose significantly identified four dominant classes. In particular, a Linear Effect Size analysis showed that some taxa, including Alphaproteobacteria, Rhizobiales, Xanthomonadaceae and Flavobacterium, were enriched in the garlic rhizosphere of the high-NPK fertilizer samples. Overall, the fertilizer dose interacted with soil compartment to shape the bacterial and archaeal community composition, abundance, and biodiversity in the garlic rhizosphere. These results provide an important basis for further understanding adaptive garlic-microbe feedback, reframing roots as a significant moderating influence in agricultural management and shaping the microbial community.

The complete mitochondrial genome of Sorex minutissimus (Eulipotyphla: Soricidae).

In this study, the complete mitochondrial genome of Sorex minutissimus was sequenced and deposited to GeneBank for the first time using muscle tissue. This mitochondrial genome is a circular molecule of 16,700 bp in length and sequence analysis showed it contains 2 rRNA genes, 22 tRNA genes, 13 protein-coding genes, rep_origin, and D_loop. Phylogenetic analysis on the basis of 12 protein-coding genes except ND6 of 13 Soricidae species' mitochondrial genomes using ML and BI demonstrated that S. minutissimus and other Sorex species were clustered into same clade.