Cytochromes b

细胞色素 b
  • 文章类型: Journal Article
    野生啮齿动物可以作为E.bieneusi的水库或载体,从而使寄生虫传播给家畜和人类。本研究旨在调查中国内蒙古自治区和辽宁省野生啮齿动物中E.bieneusi的流行情况。此外,为了评估基因型水平的人畜共患传播的可能性,对分离株进行了遗传分析.
    从中国两个省份共捕获了486只野生啮齿动物。进行聚合酶链反应(PCR)以扩增啮齿动物粪便DNA中的脊椎动物细胞色素b(cytb)基因,以检测其物种。通过rDNA的内部转录间隔区(ITS)区域的PCR扩增确定E.bieneusi的基因型。遗传特征和人畜共患潜力的检查需要应用相似性和系统发育分析。
    在四种确定的啮齿动物中,E.bieneusi的感染率为5.2%(n=89),黄鲸4.5%(n=96),小家鼠11.3%(n=106),褐家鼠为38.5%(n=195)。在486只啮齿动物中,平均感染率为17.4%。在确定的11种基因型中,已知9个:SHR1(在32个样品中检测到),D(30个样本),EbpA(9个样品),PigEbITS7(8个样品),HNR-IV(6个样品),IV型(5个样品),HNR-VII(2个样品),HNH7(1个样品),和HNPL-V(1个样品)。还发现了两种新的基因型,NMR-I和NMR-II,每个包含一个样本。通过系统发育分析将基因型分为第1组和第13组。
    根据初始报告,E.bieneusi在各自省和地区的野生啮齿动物中非常普遍,并且遗传多样性。这表明这些动物对于E.bieneusi的传播至关重要。携带人畜共患E.bieneusi的动物对当地居民构成重大危害。因此,有必要提高对这些啮齿动物带来的危险的认识,并减少其数量,以防止环境污染。
    UNASSIGNED: Wild rodents can serve as reservoirs or carriers of E. bieneusi, thereby enabling parasite transmission to domestic animals and humans. This study aimed to investigate the prevalence of E. bieneusi in wild rodents from the Inner Mongolian Autonomous Region and Liaoning Province of China. Moreover, to evaluate the potential for zoonotic transmission at the genotype level, a genetic analysis of the isolates was performed.
    UNASSIGNED: A total of 486 wild rodents were captured from two provinces in China. Polymerase chain reaction (PCR) was performed to amplify the vertebrate cytochrome b (cytb) gene in the fecal DNA of the rodents to detect their species. The genotype of E. bieneusi was determined via PCR amplification of the internal transcribed spacer (ITS) region of rDNA. The examination of genetic characteristics and zoonotic potential requires the application of similarity and phylogenetic analysis.
    UNASSIGNED: The infection rates of E. bieneusi in the four identified rodent species were 5.2% for Apodemus agrarius (n = 89), 4.5% for Cricetulus barabensis (n = 96), 11.3% for Mus musculus (n = 106), and 38.5% for Rattus norvegicus (n = 195). Infection was detected at an average rate of 17.4% among 486 rodents. Of the 11 identified genotypes, nine were known: SHR1 (detected in 32 samples), D (30 samples), EbpA (9 samples), PigEbITS7 (8 samples), HNR-IV (6 samples), Type IV (5 samples), HNR-VII (2 samples), HNH7 (1 sample), and HNPL-V (1 sample). Two novel genotypes were also discovered, NMR-I and NMR-II, each comprising one sample. The genotypes were classified into group 1 and group 13 via phylogenetic analysis.
    UNASSIGNED: Based on the initial report, E. bieneusi is highly prevalent and genetically diverse in wild rodents residing in the respective province and region. This indicates that these animals are crucial for the dissemination of E. bieneusi. Zoonotic E. bieneusi-carrying animals present a significant hazard to local inhabitants. Therefore, it is necessary to increase awareness regarding the dangers presented by these rodents and reduce their population to prevent environmental contamination.
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  • 文章类型: Journal Article
    疟原虫属和血液变形杆菌属的禽血孢子虫是一组分布广泛的血液寄生虫,会对宿主的适应性产生负面影响。哥伦比亚拥有地球上最多样化的鸟类,但是关于血吸虫及其鸟类之间联系的知识很少且支离破碎。我们从属于27科和108种的255只鸟类(203名居民和52名新热带移民)中收集了血液样本。这项研究是在考卡河和马格达莱纳河的安第斯河谷之间的六个地区进行的。通过对线粒体基因cytb片段的形态和分子分析,在样品中鉴定了疟原虫和嗜血杆菌的寄生虫。9.3%(n=24)的疟原虫或血液变形杆菌阳性。在红眼Vireo中发现了疟原虫和变形杆菌的共感染。确定了17个血球谱系,其中五种是首次在常住鸟类中报告的(CommonGroundDove,Checker-throatedStiplethroat,热带金鸟,浅排鹅口疮,和Ruddy-breastedSeedeater)和一个在SummerTanager(新热带移民)中。研究结果证实了热带低地中存在的血吸虫的广泛多样性,以及新热带候鸟在沿着其迁徙路线在血吸虫上传播的可能作用。
    Avian haemosporidians of the genera Plasmodium and Haemoproteus are a group of widely distributed blood parasites that can negatively affect the fitness of their hosts. Colombia contains the greatest diversity of birds on the planet, but knowledge about the associations between haemosporidian and its avifauna is scarce and fragmented. We collected blood samples from 255 birds (203 residents and 52 neotropical migrants) belonging to 27 families and 108 species. The study was conducted in six localities in the inter-Andean valleys of the Cauca and Magdalena rivers. Parasites of the genera Plasmodium and Haemoproteus were identified in the samples by morphological and molecular analysis of a fragment of the mitochondrial gene cyt b. Among the samples, 9.3% (n = 24) were positive for Plasmodium or Haemoproteus. Co-infection with Plasmodium and Haemoproteus was found in Red-eyed Vireo. Seventeen haemosporidian lineages were identified, five of which were reported for the first time in resident birds (Common Ground Dove, Checker-throated Stipplethroat, Tropical Kingbird, Pale-breasted Thrush, and Ruddy-breasted Seedeater) and one in the Summer Tanager (neotropical migrant). The research results confirm the wide diversity of haemosporidian present in tropical lowlands and the possible role of neotropical migratory birds in dissemination on haemosporidian along their migratory routes.
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  • 文章类型: Journal Article
    Wild rodents are key carriers of various human pathogens, including Blastocystis spp. Our study aimed to assess the prevalence and genetic characteristics of Blastocystis among wild rodents in the Inner Mongolian Autonomous Region and Liaoning Province of China. From November 2023 to February 2024, 486 rodents were captured in these regions. Fresh feces were collected from the intestines of each rodent for the isolation of DNA and PCR amplification of the vertebrate cytochrome b (cytb) gene to identify rodent species. Subsequently, PCR analysis and sequencing of the partial small subunit of the ribosomal RNA (rRNA) gene were utilized to detect Blastocystis in all fecal samples. Of the total samples, 27.4% (133/486) were found to be Blastocystis positive. The results revealed the presence of four species of rodents infected with Blastocystis, 32.3% (63/195) in Rattus norvegicus, 15.1% (16/106) in Mus musculus, 20.2% (18/89) in Apodemus agrarius, and 37.5% (36/96) in Cricetulus barabensis. Sequence analysis confirmed the existence of five Blastocystis subtypes: ST1 (n = 4), ST2 (n = 2), the ST4 (n = 125, the dominant subtype), ST10 (n = 1), and a novel ST (n = 1). The identified zoonotic subtypes (ST1, ST2, ST4, and ST10) highlight the possible role played by wild rodents in the transmission of Blastocystis to humans, thereby elevating the chances of human infection. Meanwhile, the discovery of novel sequences also provides new insights into the genetic diversity of this parasite.
    UNASSIGNED: Enquête moléculaire sur les infections à Blastocystis chez des rongeurs sauvages de la région autonome de Mongolie intérieure et de la province du Liaoning, Chine : forte prévalence et dominance du sous-type ST4.
    UNASSIGNED: Les rongeurs sauvages sont des vecteurs clés de divers agents pathogènes humains, dont Blastocystis spp. Notre étude visait à évaluer la prévalence et les caractéristiques génétiques de Blastocystis chez les rongeurs sauvages de la région autonome de Mongolie intérieure et de la province chinoise du Liaoning. De novembre 2023 à février 2024, 486 rongeurs ont été capturés dans ces régions. Des matières fécales fraîches ont été collectées dans les intestins de chaque rongeur pour l’isolement de l’ADN et l’amplification par PCR du gène du cytochrome b des vertébrés (cytb) afin d’identifier les espèces de rongeurs. Par la suite, l’analyse PCR et le séquençage de la petite sous-unité partielle du gène de l’ARN ribosomal (ARNr) ont été utilisés pour détecter les Blastocystis dans tous les échantillons fécaux. Sur le total des échantillons, 27.4% (133/486) présentaient un résultat positif à Blastocystis. Les résultats ont révélé la présence de quatre espèces de rongeurs infectées par Blastocystis, 32.3% (63/195) chez Rattus norvegicus, 15.1% (16/106) chez Mus musculus, 20.2% (18/89) chez Apodemus agrarius et 37.5% (36/96) chez Cricetulus barabensis. L’analyse de séquence a confirmé l’existence de cinq sous-types de Blastocystis : ST1 (n = 4), ST2 (n = 2), ST4 (n = 125, le sous-type dominant), ST10 (n = 1) et un nouveau ST (n = 1). Les sous-types zoonotiques identifiés (ST1, ST2, ST4 et ST10) mettent en évidence le rôle possible joué par les rongeurs sauvages dans la transmission de Blastocystis à l’Homme, augmentant ainsi les risques d’infection humaine. Parallèlement, la découverte de nouvelles séquences fournit également de nouvelles informations sur la diversité génétique de ce parasite.
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  • 文章类型: Journal Article
    尽管在伊拉克很流行,在过去的10年中,没有发表任何报告来更新旧世界螺旋虫(OWSF)的分子流行病学,Chrysomyabezziana,在这个国家。在本研究中,来自伊拉克10个省的130只绵羊被发现感染了贝齐亚纳幼虫,其身份是基于细胞色素b(Cytb)基因的PCR确认,并成功对来自不同测试省份的23个分离株进行了测序。尽管大多数分离株(n=20)属于在伊拉克流行的常见单倍型,检测到两种新的单倍型。还建议伊拉克OWSF流行病学发生重大变化,自从发现了侵扰,第一次,在尼尼微省。本研究的分离株与以前从伊拉克和世界各地发表的分离株相结合,在搜索GenBank后收集的,并进行了各种遗传和种群结构分析。当进行纯化(阴性)选择测试时,这些分离株显示出很大的统计学显着价值。表明遗传变异的发生有限,检测到的高序列保守性(C=0.937)值证明了这一点。在我们搜索过程中发现了一些来自非洲的分离株,并聚集在亚洲分离株以外的独立谱系中。比较时,后者显示出不同的遗传变异模式。例如,从地理上分开的地区分离,例如,海湾阿拉伯国家和东南亚有明显的遗传差异。另一方面,来自地理邻近地区(海湾阿拉伯国家和伊朗)的分离株的遗传细分有限。在比较印度尼西亚群岛10个岛屿的分离株时,情况并非如此。分离了来自苏门答腊和Sumba的种群,并向其他种群显示出很高的遗传变异。相反,来自苏拉威西岛的人口,Lombok和Sumbawa显示出有限的遗传变异。这一点尤其重要,因为它可以帮助检测在各个区域建立不育昆虫技术的动力学,作为针对OWSF的有效控制策略。
    Despite being endemic in Iraq, no reports have been published in the past 10 years to update the molecular epidemiology of the Old World screwworm fly (OWSF), Chrysomya bezziana, in this country. In the present study, 130 sheep from 10 Iraqi governorates were found infected with C. bezziana larvae, whose identities were PCR-confirmed based on the cytochrome b (Cytb) gene, and 23 isolates from various tested governorates were successfully sequenced. Although most isolates (n = 20) belonged to the common haplotype circulating in Iraq, two new haplotypes were detected. Significant changes in OWSF epidemiology in Iraq were also suggested, since infestations were detected, for the first time, in Nineveh governorate. Isolates of the present study were combined to those previously published from Iraq and worldwide, collected after searching the GenBank, and various genetic and population structure analyses were conducted. These isolates displayed a great statistically significant value when tested for the purifying (negative) selection, suggesting the limited occurrence of genetic variations, which was evidenced by the high sequence conservation (C = 0.937) value detected. A few isolates from Africa were revealed during our search, and clustered in a separate lineage other than that of the Asian isolates. The latter displayed different genetic variation patterns when compared. For example, isolates from geographically separate regions, e.g., the Gulf Arab countries and South-Eastern Asia had marked genetic differences. On the other hand, isolates from regions with close geographic proximity (the Gulf Arab countries and Iran) had limited genetic subdivision. This is not the case when comparing isolates from 10 islands in the Indonesian Archipelago. Populations from Sumatra and Sumba were isolated and displayed high genetic variations toward the other populations. On the contrary, populations from Sulawesi, Lombok and Sumbawa displayed limited genetic variations. This is particularly important, since it can help detecting the dynamics of establishing the sterile insect technique over various regions as an effective control strategy against the OWSFs.
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  • 文章类型: Journal Article
    狮子(Pantheraleo)在塑造和维护非洲脆弱的生态系统方面发挥着至关重要的生态作用。在考虑重新引入尝试时,保护工作应侧重于野生种群内的遗传变异性。我们研究了来自赞比亚和津巴布韦两个保护区的两组狮子,以确定它们的遗传组成,网站通常不知道的信息。在这项研究中,我们分析了17个标本的cytb和7个微卫星标记,以确定以前通过观察研究获得的家族关系和遗传多样性。然后,我们使用所有可用的完整有丝分裂基因组产生了标准化的单倍群系统发育,以及计算修正的分子钟.现代狮子谱系分化~151kya,分为两个亚种,都包含三个不同的单倍群。我们确认Pantheraleopersica不是亚种,而是北部P.l.leo的单倍群,离开非洲至少约31公里。所有狮子的祖先都存在~1.2Mya,可能在南非,后来离开非洲,分裂成两个洞穴狮子血统〜175kya。物种人口统计与主要的气候事件有关。鉴于气候变化的威胁,我们现在有了狮子进化的详细系统发育,并了解了它们的保护状况。
    Lions (Panthera leo) play a crucial ecological role in shaping and maintaining fragile ecosystems within Africa. Conservation efforts should focus on genetic variability within wild populations when considering reintroduction attempts. We studied two groups of lions from two conservation sites located in Zambia and Zimbabwe to determine their genetic make-up, information that is usually unknown to the sites. In this study, we analysed 17 specimens for cytb and seven microsatellite markers to ascertain family relationships and genetic diversity previously obtained by observational studies. We then produced a standardised haplogroup phylogeny using all available entire mitogenomes, as well as calculating a revised molecular clock. The modern lion lineage diverged ~151 kya and was divided into two subspecies, both containing three distinct haplogroups. We confirm that Panthera leo persica is not a subspecies, but rather a haplogroup of the northern P.l. leo that exited Africa at least ~31 kya. The progenitor to all lions existed ~1.2 Mya, possibly in SE Africa, and later exited Africa and split into the two cave lion lineages ~175 kya. Species demography is correlated to major climactic events. We now have a detailed phylogeny of lion evolution and an idea of their conservation status given the threat of climate change.
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  • 文章类型: Journal Article
    N-(1,3-二甲基丁基)-N'-苯基-对苯二胺醌(6-PPDQ)是由6-PPD转化而来的新兴污染物。然而,6-PPDQ暴露对线粒体的影响和潜在机制尚不清楚。以秀丽隐杆线虫为动物模型,从L1幼虫到成年第1天,以0.1-10μg/L暴露于6-PPDQ。暴露于6-PPDQ(1和10μg/L)可以增加耗氧率并降低5'-三磷酸腺苷(ATP)含量,提示线粒体功能障碍的诱导。NADH脱氢酶(复合物I)和琥珀酸脱氢酶(复合物II)的活性受到抑制,伴随着gas-1、nuo-1和mev-1的表达减少。gas-1和mev-1的RNAi增强线粒体功能障碍并降低6-PPDQ暴露线虫的寿命。GAS-1和MEV-1并行起调节6-PPDQ毒性以减少寿命的作用。胰岛素肽和胰岛素信号通路在GAS-1和MEV-1的下游作用以控制6-PPDQ对寿命的毒性。此外,daf-16的靶向基因sod-2和sod-3的RNAi在减少寿命和引起活性氧(ROS)产生方面引起对6-PPDQ毒性的敏感性。因此,环境相关浓度(ERC)的6-PPDQ可能通过影响线粒体复合物I和II引起线粒体功能障碍,通过影响生物体中的胰岛素信号与寿命减少有关。
    N-(1,3-dimethylbutyl)-N\'-phenyl-p-phenylenediamine quinone (6-PPDQ) is an emerging pollutant transformed from 6-PPD. However, the effect of 6-PPDQ exposure on mitochondrion and underlying mechanism remains largely unclear. Using Caenorhabditis elegans as animal model, exposed to 6-PPDQ at 0.1-10 μg/L was performed form L1 larvae to adult day-1. Exposure to 6-PPDQ (1 and 10 μg/L) could increase oxygen consumption rate and decease adenosine 5\'-triphosphate (ATP) content, suggesting induction of mitochondrial dysfunction. Activities of NADH dehydrogenase (complex I) and succinate dehydrogenase (complex II) were inhibited, accompanied by a decrease in expressions of gas-1, nuo-1, and mev-1. RNAi of gas-1 and mev-1 enhanced mitochondrial dysfunction and reduced lifespan of 6-PPDQ exposed nematodes. GAS-1 and MEV-1 functioned in parallel to regulate 6-PPDQ toxicity to reduce the lifespan. Insulin peptides and the insulin signaling pathway acted downstream of GAS-1 and MEV-1 to control the 6-PPDQ toxicity on longevity. Moreover, RNAi of sod-2 and sod-3, targeted genes of daf-16, caused susceptibility to 6-PPDQ toxicity in reducing lifespan and in causing reactive oxygen species (ROS) production. Therefore, 6-PPDQ at environmentally relevant concentrations (ERCs) potentially caused mitochondrial dysfunction by affecting mitochondrial complexes I and II, which was associated with lifespan reduction by affecting insulin signaling in organisms.
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  • 文章类型: Journal Article
    线粒体基因组转录了13种编码氧化磷酸化所必需的众所周知的蛋白质的mRNA。我们在这里证明了细胞色素b(CYTB),复合物III中唯一的线粒体DNA编码转录本,还编码一种未识别的187个氨基酸长的蛋白质,CYTB-187AA,使用细胞质核糖体的标准遗传密码而不是线粒体遗传密码。在使用质谱和抗体验证由胞质翻译(mPACT)产生的mtDNA编码蛋白的存在后,我们表明CYTB-187AA主要位于线粒体基质中,并通过与溶质载体家族25成员3(SLC25A3)相互作用来调节ATP的产生,从而促进从初始到初始转变的多能状态.我们进一步建立了CYTB-187AA沉默的转基因敲入小鼠模型,并发现CYTB-187AA的减少通过减少卵巢卵泡的数量来损害雌性的生育能力。第一次,我们发现了线粒体mRNA的新mPACT模式,并证明了由mtDNA编码的第14种蛋白质的生理功能。
    The mitochondrial genome transcribes 13 mRNAs coding for well-known proteins essential for oxidative phosphorylation. We demonstrate here that cytochrome b (CYTB), the only mitochondrial-DNA-encoded transcript among complex III, also encodes an unrecognized 187-amino-acid-long protein, CYTB-187AA, using the standard genetic code of cytosolic ribosomes rather than the mitochondrial genetic code. After validating the existence of this mtDNA-encoded protein arising from cytosolic translation (mPACT) using mass spectrometry and antibodies, we show that CYTB-187AA is mainly localized in the mitochondrial matrix and promotes the pluripotent state in primed-to-naive transition by interacting with solute carrier family 25 member 3 (SLC25A3) to modulate ATP production. We further generated a transgenic knockin mouse model of CYTB-187AA silencing and found that reduction of CYTB-187AA impairs females\' fertility by decreasing the number of ovarian follicles. For the first time, we uncovered the novel mPACT pattern of a mitochondrial mRNA and demonstrated the physiological function of this 14th protein encoded by mtDNA.
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  • 文章类型: Journal Article
    绿猴Chlorocebussabaeus,L.1766,原产于西非,在16世纪被引入佛得角群岛。历史资料表明,由于佛得角作为大西洋奴隶贸易中的商业转口的重要性,建立这种引进物种的精确起源是一项挑战。从佛得角的野生和圈养绿猴个体收集非侵入性粪便样品。两个线粒体碎片,HVRI和cytb,用于确认该物种的分类学鉴定,并初步确定从非洲大陆引入群岛的地理起源。通过将这项研究的新序列与以前发表的序列进行比较,研究表明,佛得角个体在HVRI中具有独特的单倍型,同时也显示出与来自西北沿海非洲的一些人口的亲缘关系,提示可能的多种引入来源和不确定的最可能来源。后者与历史信息一致,但也可能是由于仅使用mtDNA作为遗传标记和物种的扩散特征所致。讨论了该方法的局限性,并提出了未来的研究方向。
    The green monkey Chlorocebus sabaeus, L. 1766, native to West Africa, was introduced to the Cabo Verde Archipelago in the 16th century. Historical sources suggest that, due to the importance of Cabo Verde as a commercial entrepôt in the Atlantic slave trade, establishing the precise place of origin of this introduced species is challenging. Non-invasive fecal samples were collected from feral and captive green monkey individuals in Cabo Verde. Two mitochondrial fragments, HVRI and cyt b, were used to confirm the taxonomic identification of the species and to tentatively determine the geographic origin of introduction to the archipelago from the African continent. By comparing the new sequences of this study to previously published ones, it was shown that Cabo Verde individuals have unique haplotypes in the HVRI, while also showing affinities to several populations from north-western coastal Africa in the cyt b, suggesting probable multiple sources of introduction and an undetermined most probable origin. The latter is consistent with historical information, but may also have resulted from solely using mtDNA as a genetic marker and the dispersal characteristics of the species. The limitations of the methodology are discussed and future directions of research are suggested.
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  • 文章类型: Journal Article
    携带氧化还原辅因子的膜蛋白是呼吸链复合物的关键亚基,然而,它们折叠和成熟的确切路径仍然知之甚少。这里,使用冷冻EM和通过Alphafold2进行结构预测,我们生成了细胞色素b(Cytb)的早期组装中间体模型,复合体III的中央亚单位。第一组装中间体的预测结构表明Cytb与组装因子Cbp3-Cbp6的结合如何施加开放构型以促进其血红素辅因子的获得。此外,第二个中间体的结构预测表明血红素如何通过组装因子Cbp4的结合而稳定,伴随着Cbp3-Cbp6和Cytb之间的接触减弱,准备从组装因子中释放完全溶血的蛋白质。
    Membrane proteins carrying redox cofactors are key subunits of respiratory chain complexes, yet the exact path of their folding and maturation remains poorly understood. Here, using cryo-EM and structure prediction via Alphafold2, we generated models of early assembly intermediates of cytochrome b (Cytb), a central subunit of complex III. The predicted structure of the first assembly intermediate suggests how the binding of Cytb to the assembly factor Cbp3-Cbp6 imposes an open configuration to facilitate the acquisition of its heme cofactors. Moreover, structure predictions of the second intermediate indicate how hemes get stabilized by binding of the assembly factor Cbp4, with a concomitant weakening of the contact between Cbp3-Cbp6 and Cytb, preparing for the release of the fully hemylated protein from the assembly factors.
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  • 文章类型: Journal Article
    热带Theileriosis是由tick传播的原生动物寄生虫Theileriaannulata引起的致命的牛白血病样疾病。在流行地区,牛肉和牛奶的生产经济受到严重影响。羟基萘醌-丁巴伐醌(BPQ)是目前用于治疗临床Theileriosis的唯一可用药物,而BPQ抗性正在流行地区出现并蔓延。这里,我们在体外长期暴露于BPQ并监测耐药寄生虫的出现。与野生型寄生虫相比,存活的寄生虫显示BPQIC50显着增加。来自两个独立克隆系的耐药寄生虫具有相同的单突变,M128I,在编码环抱毛囊细胞色素B(Tacytb)的基因中。这种体外产生的突变以前在抗性领域分离株中没有报道过,但让人联想到甲硫氨酸到异亮氨酸突变在抗阿托夫酮的疟原虫和巴贝虫。M128I突变似乎对寄生虫适应性(增殖和分化为裂殖子)没有任何有害作用。为了深入了解耐药性是否可能是由于药物与TaCytB的结合改变而引起的,我们在计算机上生成了野生型TaCytB的3D模型,并将BPQ与预测的3D结构对接。潜在的结合位点聚集在包括Q01位点的蛋白质结构的四个区域中。预计Q01位点的结合药物会与α螺旋相反,其中包括M128,这表明该位置的氨基酸变化可能会改变药物结合。体外产生的BPQ抗性T.annulata是确定各种预测的对接位点对BPQ抗性的贡献的有用工具,并且还将允许测试针对Theleriosis的新型药物克服BPQ抗性的潜力。
    Tropical theileriosis is a fatal leukemic-like disease of cattle caused by the tick-transmitted protozoan parasite Theileria annulata. The economics of cattle meat and milk production is severely affected by theileriosis in endemic areas. The hydroxynaphtoquinone buparvaquone (BPQ) is the only available drug currently used to treat clinical theileriosis, whilst BPQ resistance is emerging and spreading in endemic areas. Here, we chronically exposed T. annulata-transformed macrophages in vitro to BPQ and monitored the emergence of drug-resistant parasites. Surviving parasites revealed a significant increase in BPQ IC50 compared to the wild type parasites. Drug resistant parasites from two independent cloned lines had an identical single mutation, M128I, in the gene coding for T. annulata cytochrome B (Tacytb). This in vitro generated mutation has not been reported in resistant field isolates previously, but is reminiscent of the methionine to isoleucine mutation in atovaquone-resistant Plasmodium and Babesia. The M128I mutation did not appear to exert any deleterious effect on parasite fitness (proliferation and differentiation to merozoites). To gain insight into whether drug-resistance could have resulted from altered drug binding to TaCytB we generated in silico a 3D-model of wild type TaCytB and docked BPQ to the predicted 3D-structure. Potential binding sites cluster in four areas of the protein structure including the Q01 site. The bound drug in the Q01 site is expected to pack against an alpha helix, which included M128, suggesting that the change in amino acid in this position may alter drug-binding. The in vitro generated BPQ resistant T. annulata is a useful tool to determine the contribution of the various predicted docking sites to BPQ resistance and will also allow testing novel drugs against theileriosis for their potential to overcome BPQ resistance.
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