肠舌状绦虫线粒体基因组特征及系统发育分析

doi:10.11843/j.issn.0366-6964.2024.12.035

摘要/Abstract

摘要：

本研究旨在获得肠舌状绦虫线粒体基因组全序列，了解其序列的结构特征，探究肠舌状绦虫系统发育相关信息。从麦穗鱼中获得肠舌状绦虫，提取基因组DNA，通过Illumina测序技术对肠舌状绦虫的线粒体全基因组进行测序、组装和注释，并进行生物信息学分析；从GenBank数据库中下载8个科34种绦虫的线粒体基因组序列，运用最大似然法和贝叶斯法构建系统发育树。结果显示，肠舌状绦虫线粒体基因组全长为13 655 bp，A+T含量为67.6%，其碱基组成具有明显的AT偏向性。肠舌状绦虫线粒体基因组包括12个蛋白编码基因(protein-coding genes, PCGs)、22个tRNA基因、2个rRNA基因和2个非编码控制区。在12个PCGs中，除cox3基因使用TTG作为起始密码子外，其余11个PCGs均使用ATG作为起始密码子；终止密码子方面，5个PCGs使用TAA，5个PCGs使用TAG，而cox3基因和nad3基因均没有终止密码子。PCGs使用密码子频率最高的是UUA，最低的是CGA。22个tRNA基因总长为1 272 bp，大多数tRNA能形成典型的三叶草结构，但是trnS1和trnR由于缺少二氢尿嘧啶臂无法形成典型的三叶草结构。nad2、nad6和nad4基因更适合作为肠舌状绦虫的分子标记。系统发育树结果表明，肠舌状绦虫与双线绦虫亲缘关系最近，并与双槽头属绦虫(Dibothriocephalus)形成姐妹群。本研究获得了肠舌状绦虫的线粒体全基因组，为研究肠舌状绦虫的分类学和系统学提供参考。

关键词: 肠舌状绦虫, 线粒体基因组, 结构特征, 系统发育

Abstract:

This experiment was conducted to obtain the complete mitochondrial genome sequence of Ligula intestinalis, revealed the sequence's structural features, and explored the phylogenetic information of L. intestinalis. Several L. intestinalis were obtained from Pseudorasbora parva and genomic DNA was extracted. The complete mitochondrial genome of L. intestinalis was sequenced, assembled, and annotated using Illumina sequencing technology after quality control of the DNA, followed by bioinformatic analysis. Maximum likelihood (ML) and bayesian inference (BI) analysis were used to construct the phylogenetic tree. The results showed that the complete mitochondrial genome of L. intestinalis was 13 655 bp in length, and its A+T content was 67.6%, exhibiting a clear AT bias. The complete mitochondrial genome was composed of 12 protein-coding genes (PCGs), 22 tRNA genes, two rRNA genes and two NCRs (non-coding regions). Among the 12 PCGs, 11 used ATG as the start codon, except for cox3, which had a TTG initial one. Among the termination codons, five out of twelve were identified as TAA, five as TAG, while neither the cox3 nor nad3 genes had termination codons. The total length of the 22 tRNA genes was 1 272 bp. Most tRNAs have a conventional cloverleaf structure, but trnS1 and trnR lack dihydrouridine arms of tRNA. The nad2, nad6 and nad4 genes are more suitable as molecular markers for L. intestinalis. The results of the phylogenetic tree showed that Ligula and Digramma were most closely related to each other, forming a sister group with Dibothriocephalus. The complete mitochondrial genome of L. intestinalis was obtained. This study will provide a reference for the study of the taxonomy and systematics of L. intestinalis.

Key words: Ligula intestinalis, mitogenome, structure characteristics, phylogeny

中图分类号:

S852.734

陈秀琴, 邱阳元, 吕庆博, 黄梅清. 肠舌状绦虫线粒体基因组特征及系统发育分析[J]. 畜牧兽医学报, 2024, 55(12): 5725-5737.

CHEN Xiuqin, QIU Yangyuan, LÜ Qingbo, HUANG Meiqing. Characteristics and Phylogenetic Analysis of Mitochondrial Genome in the Ligula intestinalis[J]. Acta Veterinaria et Zootechnica Sinica, 2024, 55(12): 5725-5737.

图/表 9

表 1

表 2

用于构建进化树的绦虫线粒体基因组信息"

科 Family	属 Genus	种 Species	mtDNA GenBank登录号 GenBank accession No. of mtDNA	mtDNA长度/bp Length of mtDNA
带科 Taeniidae	泡尾带属 Hydatigera	克氏泡尾带绦虫 H. krepkogorski	NC_021142.1	13 792
		H. parva	NC_021141.1	13 482
	带属 Taenia	豆状带绦虫 T. pisiformis	NC_013844.1	13 387
		T. crocutae	NC_024591.1	13 711
		泡状带绦虫 T. hydatigena	NC_012896.1	13 492
	棘球属 Echinococcus	细粒棘球绦虫 E. granulosus	NC_044548.1	17 675
		马棘球绦虫 E. equinus	NC_020374.1	13 605
		多房棘球绦虫 E. multilocularis	NC_000928.2	13 738
		沃氏棘球绦虫 E. vogeli	NC_009462.1	13 750
裂头科 Diphyllobothriidae	双槽头属 Dibothriocephalus	阔节裂头绦虫 D. latum	NC_008945.1	13 608
		D. nihonkaiensis	NC_009463.1	13 747
	舌状属 Ligula	肠舌状绦虫 L. intestinalis	NC_039445.1	13 621
	复殖孔属 Diplogonoporus	D. balaenopterae 巨大殖孔绦虫	NC_017613.1 NC_017615.1	13 724 13 725
		D. grandis
	双线属 Digramma	双线绦虫 D. interrupta	NC_039446.1	13 685
	迭宫属 Spirometra	S. decipiens 刺猬绦虫	NC_026852.1 NC_011037.1	13 641 13 643
		S. erinaceieuropaei
膜壳科 Hymenolepididae	腔带属 Cloacotaenia	大腔带绦虫 C. megalops	NC_032295.1	13 887
	剑带属 Drepanidotaenia	矛形剑带绦虫 D. lanceolata	NC_028164.1	13 573
	膜壳属 Hymenolepis	H. diminuta 微小膜壳绦虫	NC_002767.1 NC_029245.1	1 3900 137 64
		H. nana
	假裸头属 Pseudanoplocephala	克氏假裸头绦虫 P. crawfordi	NC_028334.1	14 192
头槽科 Bothriocephalidae	Schyzocotyle	头槽绦虫 S. acheilognathi IHB0601	KX589243.1	14 046
		S. nayarensis	NC_030317.1	13 852
		S. ophiocephalina	NC_034715.1	13 816
头颊绦虫科 Capingentidae	Breviscolex	东方短结绦虫 B. orientalis	NC_035634.1	14 011
纽带绦虫科 Lytocestidae	Atractolytocestus Khawia	A. huronensis K. sinensis	NC_035635.1 NC_034800.1	15 130 13 759
裸头科 Anoplocephalidae	裸头属 Anoplocephala	大裸头绦虫 A. magna	NC_031801.1	13 759
		叶状大裸头绦虫 A. perfoliata	NC_028425.1	14 459
	莫尼茨属 Moniezia	扩展莫尼茨绦虫 M. expansa	NC_036219.1	13 934
		贝氏莫尼茨绦虫 M. benedeni	NC_036218.1	13 958
双壳科 Dipylidiida	复孔属 Dipylidium	犬复孔绦虫 D. caninum	NC_021145.1	14 296
双腔科 Dicrocoeliidae	窄体属 Lyperosomum	长尾窄体吸虫^* L. longicauda	NC_048467.1	14 567

表 2

图 1

表 3

表 4

图 2

图 3

表 5

图 4

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编号 No.	mtDNA GenBank登录号 GenBank accession No. of mtDNA	国家 Country	宿主 Host	mtDNA长度/bp Length of mtDNA
1	NC_039445.1	中国	青海湖裸鲤	13 621
2	MW602519.1	俄罗斯	拉多加环斑海豹	13 623
3	MW602520.1	俄罗斯	拉多加环斑海豹	13 589
4	OR756289.1	秘鲁	阿氏山鳉	13 657
5	PP109086.1	中国	麦穗鱼	13 655

基因或区域 Gene or region	位置/nt Position	长度/bp Size	反密码子 Anticodon	起始密码子 Start codon	终止密码子 Stop codon	基因间隔/bp Intergenic length
cox3	1~642	642	-	GTG	-
trn-His(H)	644~710	67	GTG	-	-	1
cytb	714~1820	1 107	-	ATG	TAA	3
nad4L	1 822~2 082	261	-	ATG	TAA	1
nad4	2 043~3 293	1 251	-	ATG	TAG	-40
trn-Gln(Q)	3 297~3 352	56	TTG	-	-	3
trn-Phe(F)	3 353~3 419	67	GAA	-	-	0
trn-Met(M)	3 416~3 482	67	CAT	-	-	-4
atp6	3 486~3 995	510	-	ATG	TAA	3
nad2	3 998~4 876	879	-	ATG	TAG	2
trn-Val(V)	4 878~4 941	64	TAC	-	-	1
trn-Ala(A)	4 950~5 010	61	TGC	-	-	8
trn-Asp(D)	5 014~5 077	64	GTC	-	-	3
nad1	5 078~5 968	891	-	ATG	TAG	0
trn-Asn(N)	5 968~6 033	66	GTT	-	-	-1
trn-Pro(P)	6 041~6 105	65	TGG	-	-	7
trn-Ile(I)	6 114~6 178	65	GAT	-	-	8
trn-Lys(K)	6 195~6 258	64	CTT	-	-	16
nad3	6 260~6 613	354	-	ATG	-	1
trn-Ser(S1)	6 606~6 664	59	GCT	-	-	-8
trn-Trp(W)	6 667~6 729	63	TCA	-	-	2
cox1	6 738~8 303	1 566	-	ATG	TAG	8
trn-Thr(T)	8 294~8 355	62	TGT	-	-	-10
rrnL	8 356~9 321	966	-	-	-	0
trn-Cys(C)	9 322~9 385	64	GCA	-	-	0
rrnS	9 386~10 101	716	-	-	-	0
cox2	10 129~10 698	570	-	ATG	TAA	27
trn-Glu(E)	10 700~10 763	64	TTC	-	-	1
nad6	10 769~11 227	459	-	ATG	TAG	5
trn-Tyr(Y)	11 231~11 296	66	GTA	-	-	3
NCR1	11 297~11 521	225	-	-	-	-
trn-Leu(L1)	11 522~11 588	67	TAG	-	-	225
trn-Ser(S2)	11 599~11 664	66	TGA	-	-	10
trn-Leu(L2)	11 676~11 739	64	TAA	-	-	11
trn-Arg(R)	11 740~11 794	55	ACG	-	-	0
nad5	11 798~13 366	1 569	-	ATG	TAA	3
NCR2	13 367~13 586	220	-	-	-	-
trn-Gly(G)	13 587~13 652	66	TCC			220

区域 Regions	大小/bp Size (bp)	T (U)含量/% T (U)%	C含量/% C%	A含量/% A%	G含量/% G%	AT含量/% AT%	AT偏斜度 AT skew	GC偏斜度 GC skew
PCGs	10 059	45.7	12.4	21.4	20.5	67.1	-0.362	0.245
1st condon	3 353	41.1	11.4	23.3	24.2	64.4	-0.277	0.360
2nd condon	3 353	47.2	15.4	17.6	19.8	64.8	-0.458	0.124
3rd condon	3 353	48.6	10.5	23.4	17.5	72.0	-0.351	0.252
atp6	510	47.3	13.3	19.6	19.8	66.9	-0.413	0.195
cox1	1 566	44.8	13.0	21.6	20.5	66.4	-0.349	0.223
cox2	570	39.8	13.0	24.6	22.6	64.4	-0.237	0.271
cox3	642	47.2	12.1	19.5	21.2	66.7	-0.416	0.271
cytb	1 107	43.7	13.9	22	20.3	65.7	-0.33	0.187
nad1	891	45.9	10.7	20.5	22.9	66.4	-0.382	0.365
nad2	879	50.5	9.4	20.6	19.5	71.1	-0.421	0.346
nad3	354	48.9	11.0	21.5	18.6	70.4	-0.39	0.257
nad4	1 251	46.6	14.3	18.9	20.1	65.5	-0.422	0.169
nad4L	261	50.2	8.8	25.7	15.3	75.9	-0.323	0.27
nad5	1 569	43.3	13.1	23.2	20.5	66.5	-0.302	0.221
nad6	459	47.5	10.5	21.1	20.9	68.6	-0.384	0.333
rrnL	966	39.8	12.0	28.9	19.4	68.7	-0.158	0.234
rrnS	716	37.6	13.1	29.7	19.6	67.3	-0.116	0.197
rRNAs	1 682	38.8	12.5	29.3	19.4	68.1	-0.141	0.218
tRNAs	1 272	37.7	12.7	28.8	20.8	66.5	-0.135	0.239
NCR	445	46.97	7.19	29.89	15.96	76.85	-0.222	0.379
Full genome	13 655	44.1	12.3	23.5	20.2	67.6	-0.305	0.245

参数 Parameter	基因名称Gene name
参数 Parameter	cox3	cytb	nad4L	nad4	atp6	nad2	nad1	nad3	cox1	cox2	nad6	nad5	rrnS	rrnL
总位点数^* Total number of sites	642	1107	261	1251	510	879	891	346	1 566	570	459	1 569	711	966
保守位点数 Invariable sites	522	926	226	997	425	717	740	278	1 325	487	363	1 239	664	885
变异位点数 Variable sites	120	181	35	254	85	162	151	68	241	83	96	330	47	81
简约突变数 Parsimony informative sites	42	79	17	107	27	60	62	22	85	30	31	137	17	32
单现突变数 Singleton variable sites	74	102	18	135	58	102	89	46	156	53	65	193	28	49
变异位点比例% Ratio of variable sites/%	18.69	16.35	0.44	20.30	16.67	22.59	16.94	19.65	15.39	14.56	20.92	21.03	6.61	8.39

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