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The spotted pod borer, Maruca vitrata Fabricius is an important herbivore of major pulses and vegetable legumes in India and the chemical pesticides are major contributors for its management. In order to have an idea on other alternative management tools, the present studies were carried out on the availability of parasitoids and its genetic variation. Totally, four larval parasitoids viz., Bassus sp., Trathala flavoorbitalis Cameron, Phanerotoma hendecasisella Cameron and an undetermined Braconid wasp were recorded on M. vitrata larva. The occurrence of P. hendecasisella was reported for the first time from Tamil Nadu, India. The Bassus sp. was found to be dominant with the parasitism of 3.0 to 12.7% in different pulses and total parasitism of four parasitoids was maximum in pigeonpea (16.1 %). Total parasitism had a positive relationship with number of webbings on cowpea. The larval parasitoids Bassus sp. and braconid wasp (undetermined) yielded specific fragments (~800 bp) with mitochondrial COI primer. Presence of Wolbachia was confirmed in all four larval parasitoids with the amplicons size between 600 and 650 bp. Present study clearly indicated the close proximity of Bassus sp. on M. vitrata than other parasitoids studied. Hence, it gives way for further insights on suitability, mass culturing and development for sustainable management of this insect pest.
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Objective To investigate ecological isolation between Oncomelania hupensis snail populations in hilly regions and marshland and lake regions in Yuanjiang valley, Changde City, Hunan Province, and to unravel its underlying mechanisms. Methods Taoyuan County, Shimen County, Linli County and Lixian County in Changde City were selected as snail sampling sites in hilly regions, and Lixian County, Jinshi City, West Lake Administration District, Hanshou County and Dingcheng District were selected as snail sampling sites in marshland and lake areas. Cytochrome C oxidase 1 (cox 1) gene was amplified in snail samples and sequenced. The genetic sequences of O. hupensis snails were aligned using the software MEGA 11, and the haplotypes of O. hupensis snails were determined using the software DNASP 5.10.01. The phylogenetic tree was generated using Bayesian inference with the software MrBayes 3.2, and analysis of molecular variance (AMOVA) was performed to analyze the source of genetic divergence and estimate the genetic divergence index (FST) among snail populations with the software Arlequin 3.5.2.2. The genetic barrier among 11 O. hupensis snail populations was estimated using the Monmonier algorithm of adegenet toolkit in R package. The settings with “land in winter and water in summer” in the Yuanjian River section were divided into two categories according to the upstream and downstream, and the areas with “land in winter and water in summer” in the upstream and downstream were transformed into raster data, and then loaded into the software Fragstats 4 for analysis of landscape indicators. The trends in changes of digital elevation were extracted from the Yuanjiang River section based on the digital elevation model, and made three-dimensional visualization using the R package. Results The mitochondrial cox 1 gene were amplified in 165 O. hupensis snais from 11 sampling sites and sequenced, and a total of 152 valid gene sequences were obtained, with 46 haplotypes or 9 populations determined. No haplotype was shared in snails between Taoyuan County and Dingcheng District and Hanshou County along the downstream of the Yuanjiang River. The total area of settings with “land in winter and water in summer” was 617.66 hm2 in the upsteram of the Yuanjiang River, which consisted of 473 patches, with each patch measuring 1.31 hm2, the largest area index of 0.735 2, the landscape division index of 0.999 9, and the landscape shape index of 45.293 7. The total area of settings with “land in winter and water in summer” was 9 956.92 hm2 in the downstream of the Yuanjiang River, which consisted of 771 patches, with each patch measuring 12.91 hm2, the largest area index of 97.839 9, the landscape division index of 0.042 7, and the landscape shape index of 7.249 6. The area of settings with “land in winter and water in summer” was much larger in the downstream than that in the upstream of the Yuanjiang River, and the stronger landscape connectivity and non-remarkable alteration of riverbed elevation provided suitable habitats for snail breeding. Conclusion The hydrological and environmental characteristics of the upstream of the Yuanjiang River restrain the breeding and spread of O. hupensis, resulting in ecological isolation between Oncomelania hupensis in Taoyuan County and those in the downstream of Yuanjiang River.
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Abstract Introduction: There is low evidence of genetic diversity and hybridization processes within Crocodylus acutus and C. moreletii populations. Objetive: To evaluate genetic diversity and some phylogenetic relationships in wild and captive populations of C. acutus and C. moreletii using the Barcode of Life Data System (COX1, cytochrome C oxidase subunit 1 gene). Methods: 28 individuals phenotypically like C. acutus located in the state of Guerrero, Oaxaca and Quintana Roo were sampled, as well as animals belonging to C. moreletii located in the states of Tabasco, Campeche, and Quintana Roo. 641 base pairs of nucleotide sequence from COX1 were used to obtain the haplotype and nucleotide diversity per population, and a phylogenetic and network analysis was performed. Results: Evidence of hybridization was found by observing C. moreletti haplotypes in animals phenotypically determined as C. acutus, as well as C. acutus haplotypes in animals classified as C. moreletti. Low haplotypic diversity was observed for C. acutus (0.455 ± 0.123) and for C. moreletii (0.505 ± 0.158). A phylogenetic tree was obtained in which the sequences of C. acutus and C. moreletii were grouped into two well-defined clades. Organisms identified phenotypically as C. acutus but with C. moreletii genes were separated into a different clade within the clade of C. moreletii. Conclusions: There are reproductive individuals with haplotypes different from those of the species. This study provides a small but significant advance in the genetic knowledge of both crocodile species and the use of mitochondrial markers, which in this case, the COX1 gene allowed the detection of hybrid organisms in wild and captive populations. Conservation efforts for both species of crocodiles should prevent the crossing of both threatened species and should require the genetic identification of pure populations, to design effective conservation strategies considering the possibility of natural hybridization in areas of sympatry.
Resumen Introducción: Existe poca evidencia de la diversidad genética y los procesos de hibridación dentro de las poblaciones de Crocodylus acutus y C. moreletii. Objetivo: Evaluar la diversidad genética y algunas relaciones filogenéticas en poblaciones silvestres y cautivas de C. acutus y C. moreletii utilizando el Sistema de Código de Barras de la vida (COX1, subunidad I del gen del citocromo C oxidasa). Métodos: Se muestrearon 28 individuos fenotípicamente similares a C. acutus ubicados en los estados de Guerrero, Oaxaca y Quintana Roo, así como animales pertenecientes a C. moreletii ubicados en los estados de Tabasco, Campeche y Quintana Roo. Se utilizaron 641 pares de bases de la secuencia de nucleótidos de la subunidad I del gen del citocromo C oxidasa para obtener el haplotipo y la diversidad de nucleótidos por población, y se realizó un análisis filogenético y de redes. Resultados: Se encontró evidencia de hibridación al observar haplotipos de C. moreletti en animales determinados fenotípicamente como C. acutus, así como haplotipos de C. acutus en animales clasificados como C. moreletti. Se observó una baja diversidad haplotípica para C. acutus (0.455 ± 0.123) y para C. moreletii (0.505 ± 0.158). Se obtuvo un árbol filogenético en el que las secuencias propias de C. acutus y C. moreletii se agruparon en dos grandes y bien definidos clados. Los organismos identificados fenotípicamente como C. acutus pero con genes de C. moreletii se separaron en un clado diferente dentro del clado de C. moreletii. Conclusiones: Existen individuos reproductores con haplotipos diferentes a los de la especie. Este estudio aporta un pequeño pero significativo avance en el conocimiento genético tanto de las especies de cocodrilos como del uso de marcadores mitocondriales, que, en este caso, el gen COX1 permitió la detección de organismos híbridos en poblaciones silvestres y cautivas. Los esfuerzos de conservación para ambas especies de cocodrilos deben evitar el cruce de ambas especies amenazadas y deben requerir la identificación genética de poblaciones puras, para diseñar estrategias de conservación efectivas considerando la posibilidad de hibridación natural en áreas de simpatría.
Subject(s)
Animals , Alligators and Crocodiles/genetics , Mexico , Electronic Data ProcessingABSTRACT
Objective To identify the species of trematodes isolated from laying ducks in Nanchang City using morphological and molecular approaches. Methods Trematodes were isolated from the hepatobiliary duct, gallbladder and large intestine of market-sold laying ducks in Nanchang City. Following morphological characterization, total DNA was extracted from all trematode specimens, and internal transcribed spacer region (ITS) and cytochrome C oxidase subunit 1 (Cox1) genes were amplified using PCR assay and sequenced. Sequence alignment was performed using the Blast software, and homology and phylogenetic analyses were done in the trematode isolates based on ITS and Cox1 gene sequences. Results The morphological characteristics of two trematode isolates from the large intestine of laying ducks were similar to those of Echinostoma revolutum and E. miyagawai, and the morphological characteristics of eight trematode samples isolated from the hepatobiliary duct and gallbladder of laying ducks were similar to those of Amphimerus anatis. The ITS and Cox1 gene sequences of the two trematode isolates from the large intestine of laying ducks had 99.3% and 98.9%-99.4% homology with E. miyagawai, and the phylogenetic analysis showed that two trematode isolates had the closest genetic relationship with E. miyagawai based on ITS and Cox1 gene sequences. The ITS gene sequences of eight trematode isolates from the hepatobiliary duct and gallbladder of laying ducks shared 95.1%-95.5% with Opisthorchis sudarikovi and Clonorchis sinensis, while the Cox1 gene sequences of eight trematode isolates from the hepatobiliary duct and gallbladder of laying ducks shared 86.3%-86.4% and 85.5%-85.7% with O. viverrini and O. sudarikovi. ITS gene sequence-based phylogenetic analysis showed that the duck-derived trematode isolates had the closest genetic relationship with C. sinensis, and Cox1 gene sequence-based phylogenetic analysis showed that the duck-derived trematode isolates had the closest genetic relationship with Metorchis orientalis and O. viverrini. Conclusions The trematode isolates from the large intestine of laying ducts in Nanchang City may be E. miyagawai, and the trematode isolates from the hepatobiliary duct and gallbladder may be an unidentified trematode species of the family Opisthorchiidae.
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Cytochrome c is a type of heme proteins that are widely distributed in living organisms. It consists of heme and apocytochrome c, and has potential applications in bioelectronics, biomedicine and pollutant degradation. However, heterologous overexpression of cytochrome c is still challenging. To date, expression of the cytochrome c from uncultured anaerobic methanotrophic archaea has not been reported, and nothing is known about the function of this cytochrome c. A his tagged cytochrome c was successfully expressed in E. coli by introducing a thrombin at the N-terminus of CytC4 and co-expressing CcmABCDEFGH, which is responsible for the maturation of cytochrome c. Shewanella oneidensis, which naturally has enzymes for cytochrome c maturation, was then used as a host to further increase the expression of CytC4. Indeed, a significantly higher expression of CytC4 was achieved in S. oneidensis when compared with in E. coli. The successful heterologous overexpression of CytC4 will facilitate the exploitation of its physiological functions and biotechnological applications.
Subject(s)
Anaerobiosis , Archaea/metabolism , Cytochromes c/metabolism , Escherichia coli/metabolism , Heme/metabolismABSTRACT
Parkin, an E3 ubiquitin ligase, plays a role in maintaining mitochondrial homeostasis through targeting damaged mitochondria for mitophagy. Accumulating evidence suggests that the acetylation modification of the key mitophagy machinery influences mitophagy level, but the underlying mechanism is poorly understood. Here, our study demonstrated that inhibition of histone deacetylase (HDAC) by treatment of HDACis activates mitophagy through mediating Parkin acetylation, leading to inhibition of cervical cancer cell proliferation. Bioinformatics analysis shows that Parkin expression is inversely correlated with HDAC2 expression in human cervical cancer, indicating the low acetylation level of Parkin. Using mass spectrometry, Parkin is identified to interact with two upstream molecules, acetylase acetyl-CoA acetyltransferase 1 (ACAT1) and deacetylase HDAC2. Under treatment of suberoylanilide hydroxamic acid (SAHA), Parkin is acetylated at lysine residues 129, 220 and 349, located in different domains of Parkin protein. In in vitro experiments, combined mutation of Parkin largely attenuate the interaction of Parkin with PTEN induced putative kinase 1 (PINK1) and the function of Parkin in mitophagy induction and tumor suppression. In tumor xenografts, the expression of mutant Parkin impairs the tumor suppressive effect of Parkin and decreases the anticancer activity of SAHA. Our results reveal an acetylation-dependent regulatory mechanism governing Parkin in mitophagy and cervical carcinogenesis, which offers a new mitophagy modulation strategy for cancer therapy.
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Objective:To identify 24 <italic>Rana</italic> species such as <italic>Rana dybowskii</italic> by mitochondrial cytochrome C oxidase subunit I （<italic>CO</italic>Ⅰ） gene-based DNA barcoding and build the neighbour-joining （NJ） tree for hierarchical cluster analysis， so as to provide a basis for the identification and classification of <italic>Rana</italic> species as well as the discovery of new species. Method:<italic>R. dybowskii</italic>， <italic>R. chensinensis</italic>， <italic>R. amurensis</italic>， <italic>R. culaiensi</italic>s， and <italic>R. huanrenesis</italic>， ten for each species， were collected for DNA extraction and polymerase chain reaction （PCR） amplification<italic> </italic>and sequencing. A total of 50 <italic>CO</italic>Ⅰ gene sequences were obtained. Then 163 <italic>CO</italic>Ⅰ gene sequences for 24 species of <italic>Rana</italic> and one <italic>CO</italic>Ⅰ gene sequence for <italic>Pelophylax</italic>，<italic> Odorrana</italic>， <italic>Nidirana</italic>， <italic>Hylarana</italic>， and <italic>Amolops</italic> were harvested from GenBank. After sequence alignment by MEGA X， the parsimony-informative sites of <italic>CO</italic>Ⅰ gene sequences were analyzed and the intraspecific and interspecific genetic distances were calculated， followed by the built of NJ tree and hierarchical cluster analysis. Result:The <italic>CO</italic>Ⅰ gene sequences of 24<italic> Rana</italic> species including <italic>R. dybowskii</italic> were 554 bp in length and there were 210 parsimony-informative sites in total. The intraspecific genetic distance of each species was smaller than 2%. Except that the interspecific genetic distance between <italic>R. sangzhiensis</italic> and <italic>R. zhengi</italic> was 0.004， the genetic distances between the other species ranged from 0.024 to 0.228. <italic>R. sangzhiensis</italic> and <italic>R. zhengi</italic> were clustered into one branch and some <italic>R. dybowskii</italic> and <italic>R. uenoi</italic> into one branch. There were two separate branches for <italic>R. chensinensis</italic> and the other species were all clustered independently. Conclusion:<italic>CO</italic>Ⅰ-based DNA barcoding enabled the identification of 24 species of <italic>Rana</italic> including <italic>R.dybowskii</italic>. The findings supported that <italic>R. sangzhiensis</italic>， <italic>R. zhengi</italic>， <italic>R. coreana</italic>， and <italic>R. kunyuensis</italic> were the same species. One branch of <italic>R. chensinensis </italic>might be one of the four undownloaded species in Ranidae or a new species. The results have demonstrated that <italic>CO</italic>Ⅰ-based DNA barcoding allows not only the identification of 24 species of Rana including <italic>R. dybowskii </italic>but also the classification of ranidae species and the discovery of new species or subspecies.
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Enzyme-catalysis self-assembled oligopeptide hydrogel holds great interest in drug delivery, which has merits of biocompatibility, biodegradability and mild gelation conditions. However, its application for protein delivery is greatly limited by inevitable degradation of enzyme on the encapsulated proteins leading to loss of protein activity. Moreover, for the intracellularly acted proteins, cell membrane as a primary barrier hinders the transmembrane delivery of proteins. The internalized proteins also suffer from acidic and enzymatic degradation in endosomes and lysosomes. We herein develop a protease-manipulated hybrid nanogel/nanofiber hydrogel for localized delivery of intracellularly acted proteins. The embedded polymeric nanogels (CytoC/aNGs) preserve activity of cytochrome
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Objective To investigate the effect of calycosin on cerebral ischemia/reperfusion injury and its mechanism. Methods Forty SPF male SD rats were randomly divided into sham group, model group, calycosin group (20 mg/kg), nimodipine group (0.7 mg/kg, positive control group). The occlusion model of middle cerebral artery in rats was established by modified thread occlusion method, and the environment of cerebral ischemia-reperfusion injury was simulated in vivo. Zea longa score was used to detect the neurological deficit of rats after ischemia-reperfusion injury, 2, 3, 5-triphenyltetranitrogen (TTC) was used to detect the volume of cerebral infarction, HE staining was used to detect the pathomorphological changes of nerve cells, Nissl staining was used to observe the changes of nissl bodies, TUNEL staining was used to detect the apoptosis of nerve cells, Western blotting was used to detect the expression of cytochrome C (Cyt C), apoptotic protease activating factor-1 (Apaf-1), Caspase-9 and Caspase-3. Results Compared with the sham group, the neurological deficit symptoms in the model group were significant (P<0.05), the volume of cerebral infarction increased significantly (P<0.05). Under the microscope, it was found that the nerve cells showed contraction of cell body, hyperchromatic and pyknosis of nucleus and poor growth state, the expression of nissl body reduced significantly (P < 0.05), the apoptotic nerve increased significantly (P< 0.05), the expression of Cyt C, Apaf-1, Caspase-9 and Caspase-3 increased significantly (P<0.05). Compared with the model group, the neurological deficit symptoms of calycosin group and nimodipine group reduced significantly (P<0.05), the volume of cerebral infarction reduced significantly (P<0.05). Under the microscope, the damage of nerve cells reduced significantly, the expression of nissl body increased significantly (P<0.05), the apoptotic nerve reduced significantly (P<0.05), the expression of Cyt C, Apaf-1, Caspase-9 and Caspase-3 decreased significantly (P<0.05). Conclusion Calycosin can significantly inhibit the apoptosis of nerve cells and reduce the cerebral ischemia-reperfusion injury. Its mechanism of action is related to the effective regulation of Cyt C/Apaf-1 apoptosis signaling pathway by calycosin.
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@#Hepatocellular carcinoma (HCC) is one of the leading causes of cancer-related deaths, characterized by highly hypoxic tumor microenvironment. Hypoxia-inducible factor-1α (HIF-1α) is a major regulator involved in cellular response to changes of oxygen levels, supporting the adaptation of tumor cells to hypoxia. Bruceine D (BD) is an isolated natural quassinoid with multiple anti-cancer effects. Here, we identified BD could significantly inhibit the HIF-1α expression and its subsequently mediated HCC cell metabolism. Using biophysical proteomics approaches, we identified inhibitor of β-catenin and T-cell factor (ICAT) as the functional target of BD. By targeting ICAT, BD disrupted the interaction of β-catenin and ICAT, and promoted β-catenin degradation, which in turn induced the decrease of HIF-1α expression. Furthermore, BD could inhibit HCC cells proliferation and tumor growth in vivo, and knockdown of ICAT substantially increased resistance to BD treatment in vitro. Our data highlight the potential of BD as a modulator of β-catenin/HIF-1α axis mediated HCC metabolism.
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OBJECTIVE：To study the effects of Mahuang xixin fuzi decoction on Toll-like receptors （TLRs）response and cytochrome C oxidase （Cyt-CO）-mediated apoptosis regulation in mice with influenza disease of kidney-yang deficiency. METHODS：Totally 48 male Balb/c mice were randomly divided into normal group （n＝12）and modeling group （n＝36）. The modeling group was intraperitoneally injected with estradiol benzoate solution （8 mg/kg）and intranasally injected with influenza virus H 1N1（20 μL/mice）to establish the influenza disease compound model of kidney-yang deficiency. After modeling ，the mice were randomly divided into model group ，positive drug group （Oseltamivir phosphate capsules ，0.195 g/kg），Mahuang xixin fuzi decoction group （1.802 g/kg，by crude dru g），with 12 mice in each group. Each group was given relevant medicine intragastrically，normal group and model group were given， corresponding volume of normal saline intragastrically 20 mL/kg，once a day ，for consecutive 6 days. During admi-nistration，body weight and anal temperature of mice were mail：xsy407861520@163.com measured daily ；the percentage of initial body weight was calculated. After last medication ，the organ （spleen，thymus and lung ）indexes were calculated ；the pathological changes of lung tissue were observed. The viral load of influenza A virus H 1N1 in lung tissue was detected （reflected by M gene mRNA expression）；mRNA expressions of TLR3，TLR7，myeloid differentiation factor （MyD88）and Caspase- 3 in cardiac tissue as well as the activity of Cyt-CO and the content of cytochrome C （Cyt-C）were also determined. RESULTS ：Compared with normal group，initial body weight percentage and anal temperature of the model group continued to decrease （P＜0.05）；the spleen and thymus indexes were decreased significantly （P＜0.05），while lung index was increased significantly （P＜0.05）；the lung tissue lesions were serious. Viral load in lung tissue ，mRNA expressions of TLR 3，TLR7，MyD88 and Caspase- 3 in cardiac tissue as well as the content of Cyt-C were increased significantly （P＜0.05 or P＜0.01），while the activity of Cyt-CO in cardiac tissue was significantly decreased （P＜0.01）. Compared with model group ，initial body weight percentage and anal temperature of mice in Mahuang xixin fuzi decoction group showed an increasing trend from the fourth day of administration （P＜0.05 or P＜0.01）. The spleen and thymus indexes were increased significantly （P＜0.05），while the lung index was significantly decreased （P＜0.05）；the pathological injury of lung tissue was significantly improved ；viral load in lung tissue ，mRNA expressions of TLR 3 and Caspase- 3 as well as the content of Cyt-C in cardiac tissue were decreased significantly （P＜0.05 or P＜0.01），while the activity of Cyt-CO was increased significantly in cardiac tissue （P＜0.01）. CONCLUSIONS ：Mahuang xixin fuzi decoction can improve influenza disease of kidney-yang deficiency in mice ，the effect may related to inhibit TLRs response and apoptosis regulation pathway mediated by Cyt-CO.
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BACKGROUND Panstrongylus rufotuberculatus (Hemiptera-Reduviidae) is a triatomine species with a wide geographic distribution and a broad phenotypic variability. In some countries, this species is found infesting and colonising domiciliary ecotopes representing an epidemiological risk factor as a vector of Trypanosoma cruzi, etiological agent of Chagas disease. In spite of this, little is known about P. rufotuberculatus genetic diversity. METHODS Cytogenetic studies and DNA sequence analyses of one nuclear (ITS-2) and two mitochondrial DNA sequences (cyt b and coI) were carried out in P. rufotuberculatus individuals collected in Bolivia, Colombia, Ecuador and Mexico. Moreover, a geometric morphometrics study was applied to Bolivian, Colombian, Ecuadorian and French Guiana samples. OBJECTIVES To explore the genetic and phenetic diversity of P. rufotuberculatus from different countries, combining chromosomal studies, DNA sequence analyses and geometric morphometric comparisons. FINDINGS We found two chromosomal groups differentiated by the number of X chromosomes and the chromosomal position of the ribosomal DNA clusters. In concordance, two main morphometric profiles were detected, clearly separating the Bolivian sample from the other ones. Phylogenetic DNA analyses showed that both chromosomal groups were closely related to each other and clearly separated from the remaining Panstrongylus species. High nucleotide divergence of cyt b and coI fragments were observed among P. rufotuberculatus samples from Bolivia, Colombia, Ecuador and Mexico (Kimura 2-parameter distances higher than 9%). MAIN CONCLUSIONS Chromosomal and molecular analyses supported that the two chromosomal groups could represent different closely related species. We propose that Bolivian individuals constitute a new Panstrongylus species, being necessary a detailed morphological study for its formal description. The clear morphometric discrimination based on the wing venation pattern suggests such morphological description might be conclusive.
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OBJECTIVES@#To identify the common sarcosaprophagous flies in the Yangtze River Delta based on mitochondrial cytochrome c oxidase subunit Ⅰ(COⅠ) gene sequence and verify the reliability of this method.@*METHODS@#Seven common genetically stable sarcosaprophagous flies in three families and six genera were collected from large domestic pig carcasses placed in the field and cultured in the laboratory for many generations. The whole genome DNA was extracted and the COⅠ gene fragment was amplified. The forward and reverse sequencing was followed by splicing. The base composition of the amplified fragment and the rate of interspecific evolutionary divergence were analyzed by software such as Mega 7.0.26. The phylogenetic tree of COⅠ gene sequence of common necrophagous flies in the Yangtze River Delta was established by neighbor joining (NJ) method and unweighted pair-group method with arithmetic means (UPGMA) method.@*RESULTS@#The average base composition of different flies was A(30.14%), T(38.23%), C(15.98%), G(15.65%). The rate of interspecific evolutionary divergence ranged from 2.2% to 15.3%, the lowest rate was between Chrysomya megacephala and Chrysomya pinguis, the highest rate was between Muscina stabulans and Boettcherisca peregrina.@*CONCLUSIONS@#COⅠ gene can be used to identify the common necrophagous flies in the Yangtze River Delta.
Subject(s)
Animals , Cadaver , Diptera/genetics , Phylogeny , Reproducibility of Results , RiversABSTRACT
Abstract Genetic and phylogenetic relationships among seven piranha species of the genera Serrasalmus and Pygocentrus from the Paraná-Paraguay, São Francisco and Tocantins River basins were evaluated in the present study by partial sequences of two mitochondrial genes, Cytochrome b and Cytochrome c Oxidase I. Phylogenetic analysis of Maximum-Likelihood and Bayesian inference were performed. Results indicated, in general, greater genetic similarity between the two species of Pygocentrus (P. nattereri and P. piraya), between Serrasalmus rhombeus and S. marginatus and between S. maculatus, S. brandtii and S. eigenmanni. Pygocentrus nattereri, S. rhombeus and S. maculatus showed high intraspecific genetic variability. These species have each one, at least two different mitochondrial lineages that, currently, occur in sympatry (S. rhombeus) or in allopatry (P. nattereri and S. maculatus). Species delimitation analysis and the high values of genetic distances observed between populations of S. rhombeus and of S. maculatus indicated that each species may corresponds to a complex of cryptic species. The non-monophyletic condition of S. rhombeus and S. maculatus reinforces the hypothesis. The geographic distribution and the genetic differentiation pattern observed for the piranha species analyzed herein are discussed regarding the geological and hydrological events that occurred in the hydrographic basins.
Resumo Relações genéticas e filogenéticas de sete espécies de piranhas dos gêneros Serrasalmus e Pygocentrus das bacias hidrográficas Paraná-Paraguai, São Francisco e Tocantins foram avaliadas com base em sequências parciais dos genes mitocondriais Citocromo b e Citocromo c Oxidase I. Foram realizadas análises filogenéticas de Máxima Verossimilhança e de inferência Bayesiana. Os resultados indicaram, em geral, maior similaridade genética entre as duas espécies de Pygocentrus (P. nattereri e P. piraya), entre Serrasalmus rhombeus e S. marginatus e entre S. maculatus, S. brandtii e S. eigenmanni. Pygocentrus nattereri, S. rhombeus e S. maculatus revelaram ter alta variabilidade genética intraespecífica. Essas espécies têm, cada uma, pelo menos duas linhagens mitocondriais que, atualmente, ocorrem em simpatria (S. rhombeus) ou alopatria (P. nattereri e S. maculatus). Análises de delimitação de espécies e os altos valores de distância genética observados entre as populações de S. rhombeus e de S. maculatus indicam que cada espécie pode, na verdade, corresponder a um complexo de espécies crípticas. A condição não-monofilética de S. rhombeus e S. maculatus reforça essa hipótese. A distribuição geográfica e o padrão de diferenciação genética observados para as espécies de piranhas analisadas são discutidos com relação aos eventos geológicos e hidrológicos que ocorreram nas bacias hidrográficas.
Subject(s)
Animals , Characiformes , Paraguay , Phylogeny , Brazil , Bayes Theorem , RiversABSTRACT
Objective: To study the prevalence and genotype of Enterobius (E.) vermicularis from adhesive tape samples in the cities of Shiraz and Khorramabad, Iran. Methods: A total of 1 000 adhesive tape samples from kindergartens in Shiraz (500 samples) and Khorramabad (500 samples) were collected and tested using a microscope to find E. vermicularis egg/s. A questionnaire was filled out for each sample. In order to characterize the genotype of E. vermicularis, the PCR-sequencing method of the mitochondrial cytochrome C oxidase subunit 1 (cox1) gene was used. Genomic DNA was extracted from the positive scotch tape samples of E. vermicularis. The cox1 gene was amplified by the polymerase chain reaction and sequenced. The sequence data were aligned using the BioEdit software and compared with the published sequences in GenBank. Phylogenetic analysis was performed using the maximum likelihood method. Results: The parasitological method showed that 15 out of the 500 samples from Shiraz (3.00%) and 12 out of the 500 samples from Khorramabad (2.40%) were infected with E. vermicularis eggs. BLAST analysis indicated that the sequenced isolates belonged to E. vermicularis genotype B while three different haplotypes were also identified. Conclusions: This is the first study on genotyping E. vermicularis in the cities of Shiraz and Khorramabad. Considering the public health importance of the disease, further studies are necessary to characterize the genotype of E. vermicularis in human populations from other regions of Iran.
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Objective: To explore genetic variations of Hypoderaeum conoideum collected from domestic ducks from 12 different localities in Thailand and Lao PDR, as well as their phylogenetic relationship with American and European isolates. Methods: The nucleotide sequences of their nuclear ribosomal DNA (ITS), mitochondrial cytochrome c oxidase subunit 1 (CO1), and NADH dehydrogenase subunit 1 (ND1) were used to analyze genetic diversity indices. Results: We found relatively high levels of nucleotide polymorphism in ND1 (4.02%), whereas moderate and low levels were observed in CO1 (2.11%) and ITS (0.96%), respectively. Based on these polymorphisms, the 20 ND1, 12 CO1, and 18 ITS haplotypes were classified, and several common haplotypes were observed in all samples. At least three major lineages, namely American, European and Asian lineages, have been classified by phylogenetic analyses based on ND1 sequences. Conclusions: Our report demonstrates that the ND1 gene is the most suitable genetic marker to explore genetic variation and phylogenetic relationship of Hypoderaeum conoideum. However, a combination of all loci for ND1, CO1 and ITS would be of great value toward further genetic investigation of this endemic worldwide parasite. Thus, comprehensive molecular genetic analyses of Hypoderaeum conoideum from its worldwide distribution is needed to further understanding of the evolutionary and systematic relationships of this parasite.
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Objective To analyze the sequences of the cytochrome C oxidase subunit I (Cox1) gene of various Echinococcus granulosus genotypes that are currently recorded in the GenBank database, so as to investigate the genetic variation and differentiation of the E. granulosus genotypes across the world. Methods The sequences of the Cox1 gene of various E. granulosus genotypes that are currently recorded in the GenBank database were collected, and the same sequences of the Cox1 gene identified from a region were excluded. The mutation sites among the Cox1 gene sequences were identified and a phylogenetic tree was created based on the Cox1 gene. Results Transversion mutation was the predominant type of mutation in the Cox1 gene of E. granulosus. The same Cox1 gene sequence was found in E. granulosus G1, G6 and G7 genotypes isolated from various geographical locations across the world, with the corresponding GenBank accession numbers of KY766891, MH300971 and MH301007, respectively. Phylogenetic analysis revealed that E. granulosus G10 genotype had a remarkable geographical aggregation. Conclusions E. granulosus G1, G6 and G7 genotypes have primitive Cox1 gene sequences. There is a geographical aggregation of the E. granulosus G10 genotype in the phylogenetic tree, which has a tendency towards reproductive isolation.
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Objective:To explore the effect of Anmeidan （AMD） on the learning and memory levels of sleep deprived rats through mitochondrial mediated hippocampal neuronal apoptosis pathway. Method:Forty-eight SD rats were randomly divided into blank group， model group， low， medium， high-dose AMD groups （4.86， 9.72， 19.44 g·kg-1·d-1） and estazolam group （0.1 mg·kg-1·d-1）. Insomnia model was prepared by self-made sleep deprivation box for 14 days. Morris water maze was used to detect learning and memory levels， enzyme-linked immunosorbent assay （ELISA） was used to detect the expressions of cytochrome C （Cyt-C）， cysteine aspartic acid protease-3 （Caspase-3） in hippocampus. Transmission electron microscopy （TEM） was used to observe the morphological structure of mitochondria in hippocampus. Protein and mRNA expressions of Cyt-C， Caspase-3， Bcl-2， Bax were detected by immunofluorescence （IF） and Real-time fluorescence quantitative polymerase chain reaction （Real-time PCR） respectively. Result:In the model group， the incubation period of the platform and the total distance of swimming and the time of first arriving platform were prolonged， the number of platform crossing and the time of target quadrant movement were reduced， protein and mRNA expressions of Bcl-2 dropped， protein and mRNA expressions of Bax increased （P<0.01）， and mitochondrial structure was abnormal with crista fracture， swelling and deformation. And protein and mRNA expressions of Cyt-C， Caspase-3 increased significantly （P<0.01）. Low， medium and high-dose AMD groups could improve levels of space exploration and navigation of SD rats （P<0.01）， increase protein and mRNA expressions of Bcl-2， decrease protein and mRNA expressions of Bax， improve the damage of mitochondria， and decrease the protein and mRNA expressions of Cyt-C， Caspase-3 （P<0.01）. Conclusion:AMD can improve the learning and memory levels of SD rats， the effect is related to the mitochondrial mediated hippocampal neuronal apoptosis pathway and decrease of Cyt-C and Caspase-3 expressions.
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BACKGROUND: The previous research of our group has shown that DAIa2GIP, a new analogue of glucose dependent insulin stimulating polypeptide, has a protective effect on chondrocytes, but its mechanism is not clear. OBJECTIVE: To observe the effect of DAIa2GIP on interleukin-1β (IL-1β) induced apoptosis of chondrocytes, and to explore its molecular biological mechanism. METHODS: The costal chondrocytes of Sprague-Dawley rats were extracted and identified by SABC immunohistochemistry. The third generation cells were divided into six groups: (1) normal control group; (2) IL-1β induction group; (3) DAIa2GIP+IL-1β induction group; (4) DAIa2GIP+IL-1β+pro3GIP (GIPR antagonists) induction group; (5) DAIa2GIP induction group; (6) pro3GIP induction group. After 48 hours of drug treatment, mitochondrial membrane potential was tested, cell apoptosis was detected using Annexin-V FITC Kit (phosphatidylserine eversion analysis), calcium overload of cells was determined under laser confocal microscope, and cytochrome C content were measured by ELISA. The study protocol was approved by the Animal Ethics Committee of Shanxi Medical University. RESULTS AND CONCLUSION: SABC immunohistochemical staining showed that collagen II could be developed as chondrocytes; the maintenance degree of mitochondrial membrane potential was significantly higher in the IL-1β+DAIa2GIP incubation group than the IL-1β induction group, but the potential in both groups was lower than that in the normal control group. The apoptotic rate of chondrocytes in the IL-1β+DAIa2GIP incubation group was significantly lower than that in the IL-1β induction group, but the apoptotic rate in both groups was higher than that in the normal control group. The degree of calcium overload was compared with fluorescence intensity, and the result showed that the IL-1β+DAIa2GIP incubation group had a significantly higher intensity than the IL-1β induction group, and the fluorescence intensity in both groups was higher than that in the normal control group. The release of chondrocyte mitochondrial cytochrome C in the IL-1β+DAIa2GIP incubation group was significantly lower than that in the IL-1β induction group (P < 0.01), and the release amount in both groups was significantly higher than that in the normal control group (P < 0.01). The above four indicators showed no significant difference between the IL-1β induction group and the DAIa2GIP+IL-1β+pro3GIP group, but the values in both groups were lower than those in the control group. There was no significant difference among the normal control group, DAIa2GIP induction group, and pro3GIP induction group. To conclude, DALa2GIP can effectively antagonize IL-1β induced mitochondrial dysfunction of rat chondrocytes, thus antagonizing chondrocyte apoptosis. In this process, DALa2GIP can effectively reduce the degree of calcium overload and the release of cytochrome C.
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Objective: To study the prevalence and genotype of Enterobius (E.) vermicularis from adhesive tape samples in the cities of Shiraz and Khorramabad, Iran. Methods: A total of 1 000 adhesive tape samples from kindergartens in Shiraz (500 samples) and Khorramabad (500 samples) were collected and tested using a microscope to find E. vermicularis egg/s. A questionnaire was filled out for each sample. In order to characterize the genotype of E. vermicularis, the PCR-sequencing method of the mitochondrial cytochrome C oxidase subunit 1 (cox1) gene was used. Genomic DNA was extracted from the positive scotch tape samples of E. vermicularis. The cox1 gene was amplified by the polymerase chain reaction and sequenced. The sequence data were aligned using the BioEdit software and compared with the published sequences in GenBank. Phylogenetic analysis was performed using the maximum likelihood method. Results: The parasitological method showed that 15 out of the 500 samples from Shiraz (3.00%) and 12 out of the 500 samples from Khorramabad (2.40%) were infected with E. vermicularis eggs. BLAST analysis indicated that the sequenced isolates belonged to E. vermicularis genotype B while three different haplotypes were also identified. Conclusions: This is the first study on genotyping E. vermicularis in the cities of Shiraz and Khorramabad. Considering the public health importance of the disease, further studies are necessary to characterize the genotype of E. vermicularis in human populations from other regions of Iran.