Phylogeny of Apaturinae Butterflies (Lepidoptera: Nymphalidae) Based on Mitochondrial Cytochrome OxidaseⅠ Gene

The phylogenetic relationships of genera in the subfamily Apaturinae were examined using mtDNA sequence data from 1,471 bp of cytochrome oxidase subunit Ⅰ （COI）. The mitochondrial COI gene from a total of 16 species in 11 genera were sequenced to obtain mtDNA data, along with those of 4 species obtained from GenBank, to construct the MP and the NJ trees using Athyma jina, Penthema adelma, Polyura nepenthes, and Charaxes bernardus as outgroups. The transitions at the third codon positions of the COI data set were found saturated, but they were retained for analysis, because they contain the majority of the phylogenetic information. The impacts of equal weight assumptions for all characters in the parsimonious analysis were assessed by potential alternations in clades in response to different transition/transversion weighting schemes. The results indicated four distinct major groups in Apaturinae. Moreover, several well supported and stable clades were found in the Apaturinae. The study also identified undetermined taxon groups whose positions were weakly supported and were subject to changes under different weighting schemes. Within the Apaturinae, the clustering results are approximately identical to the classical morphological classification. The mtDNA data suggest the genus Mimathyma as a monophyletic group. Lelecella limenitoides and Dilipa fenestra have close relationship with very strong support in all phylogenetic trees. It also supports the taxonomic revision of removing several species from Apatura to other genera, namely Mimathyma schrenckii, M. chevana, M. nycteis, Chitoria subcaerulea, C. fasciola, C. pallas, and Helcyra subalba.

The Phylogenetic Relationships among Four Subspecies of the Genus Locusta Based on Sequences of Three Subunit of Cytochrome Oxidase

[Objective] The aim was to explore the phylogenetic relationships among four subspecies of the genus Locusta.[Method] The sequences of three subunits of cytochrome oxidase of Locusta migratoria tibetensis and Locusta migratoria manilensis were amplified and sequenced（COⅠ 1 539 bp,COⅡ 684 bp,CO Ⅲ 792 bp,with the total of 3 015 bp）.The corresponding sequenses of Locusta migratoria migratoria and Locusta migratoria migratorioides were obtained from GenBank and constructed a multiple alignment.Phylogenic trees of four subspecies of L.migratoria were constructed by Neighbor-Joining,Maximum-parsimony and Bayesian,respectively.[Result] The average content of A ＋ T in three subunits of four subspecies was 69.57%;the third site of codon showed the highest A ＋ T content,and the COⅠ had the highest A ＋ T content（87.6%）;The nucleotide substitution mainly occurred at the third site of codon,and the nucleotide replacement rate of CO Ⅱ was the highest.The second site of codon was conservative,so the replacement rate was in the range of 5.9%-15%.The start codon of COⅠ was CCG or ACG.Genetic distances among four subspecies were ranged from 0.001 to 0.076.The relationship between L.m.tibetensis and Locusta migratoria manilensis was the closest,followed by L.m.migratorioides and L.m.migratorioides,while the genetic distance between L.m.tibetensis and L.m.migratorioides was the largest.[Conclusion] The phylogenetic relationships among four subspecies of Locusta migratoria is L.m.tibetensis,L.m.manilensis,L.m.migratoria,L.m.migratorioides.

Phylogeny of Limenitidinae Butterflies (Lepidoptera: Nymphalidae) Inferred from Mitochondrial Cytochrome Oxidase I Gene Sequences

The phylogenetic analyses of the subfamily Limenitidinae are performed based on 1 471 bp of mtDNA cytochrome oxidase subunit I （COI） gene sequence data which were obtained from 21 individuals spanning 9 genera, along with those of 17 species obtained from GenBank, using Apatura iris, Aglais urticae, and Polyura dolon as outgroup species. Although the transitions at the third codon positions of the COI data set were highly saturated, they were still retained for analysis as they contain the majority of the phylogenetic information, and thus, the maximum pasimony （MP） under different weighting schemes and maximum likelihood （ML） trees were reconstructed in this study. The results showed that within this subfamily, the results based on the COI gene sequences are approximately identical to the traditional classification results. However, the clustering of Lexias pardalis and Tanaecia julii within the genus Euthalia as well as the clustering of Phaedyma aspasia within the genus Neptis with weak support are different from that of the current classification scheme made by Chinese scholars. The genus Limenitis is splited into two subclusters in the trees constructed by using MP and ML methods. These results support one of the strongest hypotheses for the tribe relationships within Limenitidinae.

Retinal ganglion cells of high cytochrome oxidase activity in the rat

Retinal ganglion cells in the rat were studied using the heavy metal intensified oytochrome oxidase and horseradish peroxidase histochemieal methods. The results show that a population of large retinal ganglion cells was consistently observed with the eyto3hrome oxidase staining method in retinas of normal rats or rats which received unilateral thalamotomy at birrth. These oytochrome oxidase rich ganglion cells appeared to have large somata, 3-6 primary dendrites and extensive dendritic arbors, and are comparable to ganglion cells labeled by the wheat germ agglutinin conjugated to horseradish peroxidase (WGA-HRP). However, the morphological details of some of the cells revealed by the cytoahrome oxidase staining method are frequently better than those shown by the HRP histochemieal method. These results suggest that the mit03hondrial enzyme oytoohrome oxidase can be used as a simple but reliable marker for identifying and studying a population of retinal ganglion cells with high metabolie rate in the rat.

Microevolution of Mitochondrial Cytochrome oxidase subunit I in Drosophila melanogaster at ＂Evolution Canyon＂, Israel

We determined the sequence of mitochondrial Cytochrome oxidase subunit I （CO1） in two Drosophila melanogaster strains originating at ＂Evolution Canyon＂, Israel. CO1 nucleotide sequences from two iso-female strains, 2-1 and 6-1, were 1,534 and 1,543 base-pairs, respectively. In each strain, ATAA was used in initiation of translation. Exchange rates for nucleotide and amino acid sequences were larger in the 6-1 strain than in the 2-1 strain when Oregon-R was used as the standard. Non-synonymous exchange rate was larger than synonymous exchange rate among the three strains.

Natural variation in the cytochrome c oxidase subunit 5B OsCOX5B regulates seed vigor by altering energy production in rice

Seed vigor is a crucial trait for the direct seeding of rice.Here we examined the genetic regulation of seed vigor traits in rice,including germination index(GI)and germination potential(GP),using a genome-wide association study approach.One major quantitative trait locus,qGI6/qGP6,was identified simultaneously for both GI and GP.The candidate gene encoding the cytochrome c oxidase subunit 5B(OsCOX5B)was validated for qGI6/qGP6.The disruption of OsCOX5B caused the vigor traits to be significantly lower in Oscox5b mutants than in the japonica Nipponbare wild type(WT).Gene co-expression analysis revealed that OsCOX5B influences seed vigor mainly by modulating the tricarboxylic acid cycle process.The glucose levels were significantly higher while the pyruvic acid and adenosine triphosphate levels were significantly lower in Oscox5b mutants than in WT during seed germination.The elite haplotype of OsCOX5B facilitates seed vigor by increasing its expression during seed germination.Thus,we propose that OsCOX5B is a potential target for the breeding of rice varieties with enhanced seed vigor for direct seeding.

Identification of forensically important blow fly species （Diptera： Calliphoridae） in China by mitochondrial cytochrome oxidase I gene differentiation

Unambiguous and rapid sarcosaphagous insect species identification is an essential requirement for forensic investigations. Although some insect species are difficult to classify morphologically, they can be effectively identified using molecular methods based on similarity with abundant authenticated reference DNA sequences in local databases. However, local databases are still relatively incomplete in China because of the large land area with distinct regional conditions. In this study, 75 forensically important blow flies were collected from 23 locations in 16 Chinese provinces, and a 278-bp segment of the cytochrome oxidase subunit I gene of all specimens was successfully sequenced. Phylogenetic analysis of the sequenced segments showed that all Calliphorid specimens were properly assigned into nine species with relatively strong supporting values, thus indicating that the 278-bp cytochrome oxidase subunit one region is suitable for identification of Calliphorid species. The clear difference between intraspecific threshold and interspecific divergence confirmed the potential of this region for Calliphorid species identification, especially for distinguishing between morphologically similar species. Intraspecific geographic variations were observed in Lucilia sericata （Meigen, 1826） and Lucilia caesar （Linnaeus, 1758）.

Molecular phylogeny of Pamphagidae （Acridoidea, Orthoptera） from China based on mitochondrial cytochrome oxidase II sequences

Phylogenetic relationships of Pamphagidae were examined using cytochrome oxidase subunit II （COIl） mtDNA sequences （684 bp）. Twenty-seven species of Acridoidea from 20 genera were sequenced to obtain mtDNA data, along with four species from the GenBank nucleotide database. The purpose of this study was analyzing the phylogenetic relationships among subfamilies within Pamphagidae and interpreting the phylogenetic position of this family within the Acridoidea superfamily. Phylogenetic trees were reconstructed using neighbor-joining （NJ）, maximurn parsimony （MP） and Bayesian inference （BI） methods. The 684 bp analyzed fragment included 126 parsimony informative sites. Sequences diverged 1.0%-11.1% between genera within subfamilies, and 8.8%-12.3% between subfamilies. Amino acid sequence diverged 0-6.1% between genera within subfamilies, and 0.4%-7.5% between subfamilies. Our phylogenetic trees revealed the monophyly of Pamphagidae and three distinct major groups within this family. Moreover, several well supported and stable clades were found in Pamphagidae. The global clustering results were similar to that obtained through classical morphological classification： Prionotropisinae, Thrinchinae and Pamphaginae were monophyletic groups. However, the current genus Filchnerella （Prionotropisinae） was not a monophyletic group and the genus Asiotmethis （Prionotropisinae） was a sister group of the genus Thrinchus （Thrinchinae）. Further molec- ular and morphological studies are required to clarify the phylogenetic relationships of the genera Filchnerella and Asiotmethis.

Hippocampal mitochondrial cytochrome C oxidase activity and gene expression in a rat model of chronic cerebral ischemia

The present study established a rat model of chronic cerebral ischemia using bilateral common carotid artery permanent ligation to analyze cytochrome C oxidase activity and mRNA expression in hippocampal mitochondria. Results showed significantly decreased cytochrome C oxidase activity and cytochrome C oxidase II mRNA expression with prolonged ischemia time. Further analysis revealed five mitochondrial cytochrome C oxidase II gene mutations, two newly generated mutations, and four absent mutational sites at 1 month after cerebral ischemia, as well as three mitochondrial cytochrome C oxidase III gene mutations, including two newly generating mutations, and one disappeared mutational site at 1 month after cerebral ischemia. Results demonstrated that decreased cytochrome C oxidase gene expression and mutations, as well as decreased cytochrome C oxidase activity, resulting in energy dysmetabolism, which has been shown to be involved in the DatholoQical Process of ischemic brain iniurv.

Gene damages of mitochondrial DNA encoding cytochrome oxidase of intestinal epithelial cells in hemorrhagic shock rats

ObJective： To investigate the detrimental effects of hemorrhagic shock on the structure and function of mitochondria DNA （mtDNA） encoding cytochrome oxidase genes in intestinal epithelial cells. Methods ： Wistar rats were used and divided into two groups： hemorrhagic shock group and control group. Hemorrhagic shock model of rats was utilized in this experiment. The mtDNA was extracted from the intestinal epithelial cells and amplified by polymerase chain reaction （ PCR ） with different primers of cytochrome oxidase （ COX Ⅰ, COX Ⅱ and COX Ⅲ）. The products of PCR were directly sequenced. Results ： Hemorrhagic shock could result in the point mutagenesis in mitochondrial genome encoding cytochrome oxidase （ COX I and COX II）. There were 4, 4, 22, 16, 35 point mutations in COX I from 5545 to 6838 bp in 5 shocked rats. There were five point mutations in COX II from 7191 to 7542 bp at the site of t7191c, t7212c, a7386g, a7483g, c7542g in 1 shocked rat. There was no mutation found in COX III. Conclusions： Hemorrhagic shock could significantly induce the damage of the gene of cytochrome oxidase encoded by mtDNA.

Regulation of cytochrome c oxidase contributes to health and optimal life

The generation of cellular energy in the form of ATP occurs mainly in mitochondria by oxidative phosphorylation.Cytochrome c oxidase(CytOx),the oxygen accepting and rate-limiting step of the respiratory chain,regulates the supply of variable ATP demands in cells by“allosteric ATP-inhibition of CytOx.”This mechanism is based on inhibition of oxygen uptake of CytOx at high ATP/ADP ratios and low ferrocytochrome c concentrations in the mitochondrial matrix via cooperative interaction of the two substrate binding sites in dimeric CytOx.The mechanism keeps mitochondrial membrane potentialΔΨm and reactive oxygen species(ROS)formation at low healthy values.Stress signals increase cytosolic calcium leading to Ca^2+-dependent dephosphorylation of CytOx subunit I at the cytosolic side accompanied by switching off the allosteric ATPinhibition and monomerization of CytOx.This is followed by increase ofΔΨm and formation of ROS.A hypothesis is presented suggesting a dynamic change of binding of NDUFA4,originally identified as a subunit of complex I,between monomeric CytOx(active state with highΔΨm,high ROS and low efficiency)and complex I(resting state with lowΔΨm,low ROS and high efficiency).

Enzymatic Activity of Escherichia Coil Lactate Dehydrogenase and Cytochrome c Oxidase

The catalytic activity of two common bacterial enzymes, lactate dehydrogenase （LDH） and cytochrome c oxidase （COX） from Escherichia coli was examined following bacterial exposure to microwave （MW） radiation under well-defined experimental conditions. The experiments were conducted in a specialized microwave processing apparatus, with an exposure frequency of 18 GHz, and a temperature profile that was restricted to below 40℃ to avoid thermal degradation of the bacteria. The absorbed power was calculated to be 1,500 kW/m3 and the electric field was determined to be 300 Wm. Both values were theoretically confirmed using Computer Simulation Technology （CST） Microwave Studio 3D Electromagnetic Stimulation Software. Results showed that the activity of both enzymes was increased following MW radiation compared to negative controls and thermally treated samples subjected to similar temperature profiles. It is suggested that the increase in COX and LDH enzyme activity could not be explained by conventional heating alone, but was rather a result of micro-thermal effects that incorporated ＇undetectable＇ thermal mechanisms.

Construction of Cox7a2 fluorescent vector and its effect on cytochrome C oxidase activity in mouse Sertoli cell line TM4

Objective To construct Cox7a2 fluorescent vector and study its effect on cytochrome C oxidase ( COX) activity in mouse Sertoli cell line TM4. Methods The coding region of CoxTa2 was amplified from mouse Sertoli cell line TM4 by RT-PCR. PCR product was

Benthodytes occidentpalauta sp.nov.,a new species of deep-sea holothuroid(Elasipodida:Psychropotidae)from the west of Kyushu-Palau Ridge in the Western Pacific Ocean

Benthodytes occidentpalauta sp.nov.was collected from the Kyushu-Palau Ridge at a depth of 5481 m in 2021.This new species is characterized by a gelatinous body wall,violet skin,six pairs of dorsal papillae,and a rough mid-ventral surface without tube feet.The dorsal deposits are rod-shaped and tripartite.Two types of papillae deposits as crosses with four arms with central bipartite apophyses.Ventral deposits are rods.Tentacle ossicles are rod-shaped with end protrusions.Gonad deposits are rodshaped,tripartite,and cross-shaped.The phylogenetic analyses based on cytochrome oxidase subunit 1(COI)and 16S individually and a concatenated dataset of COI and 16S genes of this species support that B.occidentpalauta sp.nov.belongs to Benthodytes.

Epiphytic zooplankton community profiles in a typical urban wetland as revealed by DNA metabarcoding

Zooplankton,a crucial component of urban wetland,are one of the effective bioindicators for monitoring the feeding stocks of organisms at higher trophic levels and assessing the ecological quality of ecosystems.However,information about the characteristics of epiphytic zooplankton community structure resulted from traditional methods is limited and hindered by the large amount of detritus and sludge attached to the macrophytes.We investigated the epiphytic zooplankton communities associated with macrophytes(Vallisneria,Nymphaea,and Thalia dealbata)in a subtropical wetland using as DNA markers of the 18 S rRNA gene and the mitochondrial cytochrome c oxidase subunit I(COI)gene.A total of 241 OTUs of zooplankton were obtained from COI amplicons,including 194 OTUs of Rotifera,22 of Cladocera,and 25 of Copepoda,while only 62 OTUs of zooplankton were obtained from 18 S rDNA amplicons including 34 OTUs of Rotifera and 28 of Copepoda.The zooplankton communities associated with the three macrophytes were similar,but they differed significantly from those in the open waters.However,there were no significant temporal differences among the zooplankton communities.Epiphytic zooplankton communities were dominated by littoral zooplankton such as Testudinella,Lecane,and Philodina.Microzooplankton,especially littoral species,utilize macrophytes as food sources and as refuges against predation.This further led to an increase inαandβdiversity of zooplankton communities in urban wetlands.Our result suggests that the joint use of multiple molecular markers could improve the taxonomic resolution and generate a comprehensive biodiversity profile of zooplankton.

MitochondrialtRNA^(leu(UUR)) Gene Mutation and the DecreasedActivity of Cytochrom e c Oxidase in Preeclam psia

To explore the roles of mitochondria tRNA leu(UUR) gene mutation at nucleotide 3243 and the activity of cytochrome c oxidase in pathogenesis of preeclampsia, 57 patients with preeclampsia and 60 normotension pregnancy women were screened for tRNA leu(UUR) nt3243 A→G mutation with the method of polymers chain reaction (PCR) and restriction fragment length polymorphism. Cytochrome c oxidase activity was determined by measuring the rate of cyanide sensitive oxidation of reduced cytochrome c using luminosity photographer. The results showed that cytochrome c oxidase activity was significantly lower in the preeclampsia group (0.30±0.39/min, n = 32) than that in the controls (0.73±0.54/min, n = 26, P <0.01). The mitochondria DNA mutation at position 3243 was not found in our series. The results suggested that the decreased activity of cytochrome c oxidase might impair the energy production, leading to the mitochondria dysfunction and placenta dysfunction in preeclampsia patients. Mitochondria dysfunction may be involved in the pathogenesis of preeclampsia. The mutation of mitochondria DNA may not be the common contributor of preeclampsia in our series.

DNA barcoding of fishes from Zhoushan coastal waters using mitochondrial COI and 12S rRNA genes

Accurate species identification is a key component of biodiversity research.DNA barcoding is an effective molecular method used for fish species identification.We aimed to study the DNA barcoding of fish in Zhoushan coastal waters,explore the differences and applicability of two gene fragments(12S rRNA and COI)of DNA barcoding in fish species identification,and established a comprehensive fish barcoding reference database.Two hundred and eighty-seven captured fish samples from Zhoushan coastal waters were identified using morphological characteristics and DNA barcoding.A total of 26412S rRNA sequences(belonging to eight orders,31 families,55 genera,and 66 species)and 188 COI sequences(belonging to seven orders,30 families,48 genera,and 58 species)were obtained.The lengths of the 12S rRNA sequences ranged from 165 to 178 bp,and the guanine-cytosine(GC)content was 45.37%.The average 12S rRNA interspecific and intraspecific genetic distances(K2P)were 0.10%and 26.66%,respectively.The length of the COI sequence ranged 574–655 bp,and the content of GC was 45.97%.The average 12S rRNA interspecific and intraspecific genetic distances(K2P)were 0.16%and 27.45%,respectively.The minimum interspecific genetic distances of 12S rRNA and COI(1.23%and 1.86%)were both greater than their maximum intraspecific genetic distances(2.42%and 8.66%).Three molecular analyses(NJ tree,ABGD,and GMYC)were performed to accurately identify and delineate species.Clustering errors occurred when the 12S rRNA sequences were delimited using the NJ tree method,and the delimitation results of ABGD and GMYC are consistent with the final species identification results.Our results demonstrate that DNA barcoding based on 12S rRNA and COI can be used as an effective tool for fish species identification,and 12S rRNA has good application prospects in the environmental DNA(eDNA)metabarcoding of marine fish.

Benthodytes tetrapapillata sp.nov.,a new elasipodid sea cucumber(Elasipodida:Psychropotidae)from a seamount in the Western Pacific Ocean

Benthodytes tetrapapillata sp.nov.was collected from a seamount located on the Caroline Ridge at a depth of 2289 m,during the cruise of R/V Kexue in June 2019.We provided detailed descriptions of external and deposits morphology.The phylogenetic analyses based on cytochrome c oxidase I(COI)and a concatenated dataset of 16S and COI genes showed that the new species belonged to Benthodytes that is not monophyletic.Both details of morphological comparisons and molecular analyses confirmed that Benthodytes tetrapapillata sp.nov.is a new psychropotid species.A state of main morphological characters in valid species of Benthodytes is also provided in this study.

Genetic diversity and population structure of the sea star Asterias amurensis in the northern coast of China

The sea star Asterias amurensis is widely viewed as a severe“marine pest”because of its broad feeding habits.Over the past few decades,A.amurensis undergoes massive and sporadic population outbreaks worldwide,causing extensive economic and ecological losses to the local aquaculture industry and marine ecosystem.Understanding the genetic diversity and population structure of A.amurensis can provide vital information for resource management.By analyzing the polymorphism of the mitochondrial cytochrome C oxidase subunit I(COI)gene and ten simple sequence repeat(SSR)microsatellites markers,the genetic diversity and population structure of A.amurensis of four populations along the northern coast of China was uncovered.A total of 36 haplotypes were identified,and a main haplotype was found in four populations.The Qingdao(QD)population displayed the highest genetic diversity among all the populations.The AMOVA and pairwise F_(st)showed that there was small but statistically significant population differentiation among the four populations,especially between QD and Weihai(WH).Moreover,the principal component analysis(PCA)and admixture analysis showed that several individuals in Yantai(YT)and Dalian(DL)had little genetic association with other individuals.Overall,this study provided useful information of the genetic diversity and population structure of A.amurensis and will contribute to the resource management of A.amurensis in China.

Phylogenetic relationships of some Sylviidae species based on complete mtDNA cyt b and partial COI sequence data

Members of the passerine family Sylviidae are distributed widely around the Old World and the phylogenetic relationships of many species still remain controversial.In this study,we investigated the phylogeny and relationships among 37 species of 11 genera by analyzing DNA sequences obtained from the complete mitochondrial cytochrome b (cyt b) and partial cytochrome oxidase I (COI) genes.The data were analyzed by maximum-likelihood analysis and Bayesian inference.The results demonstrate that the current genus Phylloscopus is non-monophyletic,while Seicercus is synonymized with Phylloscopus or is a group within the genus Phylloscopus.We may conclude that within Phylloscopus there are close relations between P.magnirostris and P.borealis;among P.proregulus,P.yunnanensis,P.pulcher and P.maculipennis,as well as among P.occisinensis,P.schwarzi,P.armandii,P.collybita and P.fuscatus.Monophyly of the genus Cettia could not be corroborated;it is closely related to the genera Abroscopus and Tesia.However monophyly of Acrocephalus is supported.Furthermore,the close relationships between Sylvia and Zosterops and among Prinia,Orthotomus and Cisticola are also supported.