PJZ_58_4_1539-1550

INTRODUCTION

Strong data must support conservation strategies to address the ongoing global biodiversity threat (Buxton et al., 2021). The entire mitochondrial genome of unidentified fish was employed to classify its species via a phylogenetic tree. Mitochondria are important in the context of invading species. One of the primary causes of biodiversity losses is invasive species. Invasive species are costly to manage and have increasingly significant economic impacts worldwide (Diagne et al., 2021). Thryssa vitrirostris (Gilchrist and Thompson, 1908) is found in coastal and estuarine waters. It belongs to the family Engraulidae and order Clupeiformes (Gopinath et al., 2024). It is distributed in various regions, including Madagascar, the Indian Ocean, the African coast, lfredo Port to the Persian Gulf, and the coasts of Pakistan (Pawase et al., 2020). In India, 18 species of Thryssa and 37 species worldwide have been reported. It is small, slender, and elongated anchovies. Globally, it plays an important role in ecology and improves the income of fishers (Hata and Lavoue, 2024). The morphological characteristics of the T. vitrirostris maxilla are that it is lengthy, the base of the first pectoral finray is extended, and the first supramaxilla is slightly elliptical. A dark bloch was observed behind the upper portion of the gill aperture; bright orange was found inside the gill cavity, 34 anal fins were present, and 27 keeled scutes were observed from the isthmus to the anus. The total number of lower gill rakers was 24, and serrae were found on the edge (Gopinath et al., 2024).

Fish and other vertebrates have extranuclear, closed, circular, double-stranded DNA molecules that make up their mitochondrial DNA (Gao et al., 2024; Satoh et al., 2016). It is made up of heavy and light strands, 13 PCGs, two rRNAs, and twenty-two tRNAs. In addition, a control region was observed in fish mtDNA, which typically has a size of 15–18 kb (Satoh et al., 2016; Brown, 2008). mtDNA differs from nuclear DNA because of its small size, lack of introns, rapid evolution rate, maternal inheritance, and number of copies. Consequently, it has emerged as a vital molecular marker in the fields of evolutionary genetics, species identification, molecular ecology, and fish conservation biology (Jo et al., 2022; Zhang et al., 2021, 2022). In recent years, a growing number of fish mitochondrial genome studies have been performed due to rapid advancements in high-throughput DNA sequencing technology along with statistical analysis. Many countries have utilized the COI gene to study populations, construct phylogeographies, and analyse the genetic diversity of fish. In our study, we examined the structural features and gene composition of the whole mt genome of T. vitrirostrisvia Sanger sequencing. Together with the mitochondrial sequence analysis of similar species, the protein-coding genome sequence was utilized to evaluate the evolutionary relationships of fish in the Engraulidae family.

MATERIALS AND METHODS

Sample collection

Specimens of Thryssa vitrirostriswere collected from a fishing harbour in South Tamil Nadu, India, in June 2020. It was transported on ice and stored at -20 °C at the Department of Bioinformatics DNA Barcode Research Centre, NMC College, India. The frozen tissue was dissected and stored in (v/v) 95% ethanol. The DNeasyQiagen Kit (Qiagen, Germany) was used to extract DNA following the manufacturer’s protocol. The concentration of isolated DNA was measured via a NanoDrop 2000 spectrophotometer (W: NanoDrop Technology, Minton, NC, USA).

DNA extraction and amplification

To analyse the entire mitochondrial (mt) genome, a primer walking technique was employed. First, depending on the primer designed for the fragments, the entire sequence was broken into smaller pieces. The general primer design concept was used in the preparation of the primer (19–21 bp). The PCR-amplified product was 800 bp long, and the GC content ranged from 45 to 55%. The designed primers were further checked via primer stat. To prevent mismatch, the primer has no hairpin structure to avoid the formation of a dimer between the primer and itself (Saha et al., 2024). Amplifications were performed via a PCR gradient thermal cycler in 50 µL PCR mixture. A DNA purification kit was utilized to purify the PCR products as described in the manufacturer’s instructions (Qiagen, Germany). The PCR products were sequenced at Eurofins Scientific (Bangalore, India). Chromatographic analysis was performed with ABI Sequence Editor 3.3 (USA).

Annotation and genome assembly

The sequences were manually aligned, corrected, and assembled viaClustalW software. Following splicing of the entire mtDNA sequence, the total length of the mt genome was then calculated. The beginning and ending locations of each gene were compared to the whole mtDNA sequences of Thryssa balaema, which are accessible in GenBank (http://www.ncbi.nlm.nih.gov/genbank). The beginnings and ends of each location of the genes were compared. OGDRAW was employed to annotate the gene structure of the mitochondria (Greiner et al., 2019). tRNA genes were identified, and a tRNA II hierarchical structure diagram was generated via the web application ARWEN (Laslett and Canbäck, 2008). MEGA 6.0 software was used to calculate the base composition along with codon usage (Tamura et al., 2013). In this study, “models-> Compute Codon Usage Bias” was used, and synonymous codon usage (RSCU) was computed. AClustal Omega online tool was employed, and the gene sequences were compared (Madeira et al., 2024). A CAP3 genome assembly program was used to analyse the data as described previously (http://doua.prabi.fr/software/caps) (Huang and Madan, 1999). The different mitochondrial genes were identified in NCBI databases via BLAST analyses (Altschul et al., 1990). All the mitochondrial DNA sequences were examined for base composition via the base manipulation suite (http://mobyle.Pasteur.fi-/cgibin/portal.py) (Néron et al., 2009). The AT and GC skews in the mtDNA, PCG, tRNA, rRNA, and control regions were calculated via the following formulae:

AT skew = (A−T)/(A+T); GC skew = (G−C)/(G+C) (Junqueira et al., 2004)

Phylogenetic analysis

The evolutionary position of T. vitrirostris was investigated via 13 PCGs in the mitochondrial genomes of 24 fish species from various families, including Engraulidae, Prestigasteridae, and Clupeadae (Table I). The maximum likelihood method was used to construct a phylogenetic tree, and Chirocentrus dorab was used as the outgroup (Ronquist et al., 2012). The best alternative model derived from the phylogenetic tree dataset was GTR+I+G. The maximum likelihood (ML) method was used to construct a phylogenetic tree through 1000000 bootstrap generations via MEGA11 (Tamura et al., 2021; Ronquist et al., 2012). We constructed a BI phylogenetic tree by discarding 25% of the ageing samples from the sequences, which were sampled and preserved every 500 generations for a total of 100,000 generations.

Table I. Complete mitochondrial genome of species used in this study.

S. No	Taxon (Species)	Acc. No	Size (bp)
1	Thryssa vitrirostris	PQ385613	16808
2	Thryssa mystax	NC_085785	17060
3	Thryssa baelama	NC_01426	16865
4	Thryssa hamiltonii	NC_036672	16737
5	Thryssa dussumieri	NC_035065	16920
6	Thryssa kammalensis	NC_029940	16968
7	Thryssa setirostris	NC_038196	16923
8	Coilia grayii	KP317088	16861
9	Coilia nasus	AP009135	16897
10	Coilia mystus	JX534238	17143
11	Stolephorus chinensis	AP011566	16593
12	Stolephorus commersonnii	NC_033521	16734
13	Ilisha africana	AP009140	16642
14	Pellona flavipinnis	AP009619	16742
15	Ilisha elongata	AP009141	16809
16	Sardinops melanostictus	AB032554	16881
17	Amblygaster sirm	NC_035064	17049
18	Escualosa thoracata	AP011601	16749
19	Sardinella albella	NC_016726	16652
20	Sardinella maderensis	AP009143	16651
21	Nematalosa nasus	NC_023824	16674
22	Nematalosa come	NC_021447	16644
23	Nematalosa japonica	NC-009586	16665
24	Chirocentrus dorab	AP006229	15989

RESULTS

Genome content

The whole mitogenome sequence of T. vitrirostris (accession number: PQ385613) was 16808 bp in length (Table II, Fig. 1). According to our findings, the A+T content of the whole mt genome was greater than the G+C content. A total of 13 PCGs were observed, and the longest gene was ND5 (1839 bp), whereas the smallest gene was the ATP8 (168 bp) gene. The length of the 12S rRNA was 951 bp, and the length of the 16S rRNA was1689 bp. The tRNA-Val gene separated the 12S and 16S rRNA genesand was positioned within tRNA-Phe and tRNA-Leu (Table II).

Protein-coding genes

The entire mt genome of T. vitrirostrisconsists of 22 tRNA genes, 13 PCGs, two rRNA genes, and one regulatory region (D-loop). The H-strand encodes all other

Table II. Gene profile and organization of the Thryssa vitrirostris.

Genes	From	To	Size (bp)	Amino acid	Start	Stop	GAP	St-and
tRNAphe	1	69	69				0	H
12S rRNA	70	1020	951				0	H
tRNA val	1021	1092	72				0	H
16S rRNA	1093	2781	1689				0	H
tRNALeu	2782	2856	75				0	H
ND1	2858	3832	975	324	ATG	TAA	1	H
tRNAIle	3833	3904	72				-1	H
tRNAGln	3904	3974	71				0	L
tRNA Met	3974	4042	69				0	H
ND2	4043	5087	1045	348	ATG	T--	2	H
tRNA Trp	5088	5159	72				1	H
tRNA Ala	5162	5230	69				1	L
tRNA Asn	5232	5304	73				31	L
tRNACys	5336	5401	66				0	L
tRNA Tyr	5402	5472	71				1	L
CO1	5474	7018	1545	514	GTG	TAA	0	H
tRNAser	7019	7089	71				5	L
tRNAASP	7095	7163	69				11	H
CO11	7175	7865	691	230	ATG	T	0	H
tRNALys	7866	7939	74				1	H
ATP8	7941	8108	168	55	ATG	TAA	8	H
ATP6	8099	8782	684	227	ATG	TAA	-1	H
CO111	8782	9566	785	261	ATG	TA	0	H
tRNAGly	9567	9638	72				0	H
ND3	9639	9987	349	116	ATG	T	0	H
tRNAArg	9988	10056	69				0	H
ND4L	10057	10353	297	98	ATG	TAA	5	H
ND4	10347	11727	1381	460	ATG	T	0	H
tRNAHis	11728	11796	69				1	H
tRNAser	11798	11864	67				0	H
tRNALeu	11865	11936	72				0	H
ND5	11937	13772	1836	611	ATG	TAA	3	H
ND6	13768	14289	522	173	ATG	AGG	0	L
tRNAGlu	14290	14358	69				5	L
CYTB	14364	15504	1141	380	ATG	T	0	H
tRNAThr	15505	15575	71				-1	H
tRNApro	15575	15645	70				0	L
Control region	15646	16808	1163				0	H

mitochondrial genes except for ND6, which is encoded by eight tRNA genes (Cys, Pro, Ser, Tyr, Gln, Ala, Asn, and Glu). The mitogenome order, direction, position, and gene coding strands were analysed. A total of 13 PCGs, including 67.93% of the mitochondrial genome, were11419 bp long. The overall composition of the base of the mt genome was A-30.15%, T-24.26%, G-16.84%, and C-28.75%. The AT and GC contents were 54.41% and 45.59%, respectively. The overall base compositions of A, T, G, and C were 30.15%, 24.26%, 16.84%, and 28.75%, respectively. A total of six PCGs ended with a stop codon, TAA, ND6 ended with AGG, and the other genes were incomplete with T and TA (ND2, COII, CO111, ND3, ND4, and CYTB). The maximum distance between tRNA-Asn and tRNA-Cys was 31 bp. In addition, 13 bp overlapped between ATP8 and ATP6 and between ND4Land ND4 (Table II). In this study, most of the GCskews in the PCGs were negative, with the exception of ND6, and the ATskews of CO1, CO3, ND6, and CYTB were negative. The analysed skews ranged from -0.51 (ND6) to 0.15 (ND2) (Fig. 3; Table III).

rRNA region

Our results revealed that the 12S and 16S rRNA gene lengths were 951 bp and 1689 bp, respectively. These genes are located within tRNA-Phe and tRNA-Leu and are separated by tRNA-Val. In the rRNA, the base compositions of A, T, G, and C were 33.3%, 18.79%, 20.98%, and 26.73%, respectively. The A+T value of 52.08% was slightly greater than the G+C content (47.92%). The two rRNAs encoded on the H-stand presented a negative G+C skew (AT skew=0.54, GC skew=-0.12) and positive A+T skew (Table IV). The A+T content and length of the two rRNAs were similar to those of other Engraulidaemito genomes.

Codon usage and amino acid composition

In this study, the observed RSCU value of 13 PCGs of T. vitrirostriswas 27, a value >1indicated positive bias in codon usage and a value <1 indicated negativebias in codon usage. The 11419 bp sequence encoded 3797 amino acids. With a total concentration of genome. However, at a concentration of 0.8%, cysteine (Cys) was the minimum prevalent amino acid in the T. vitrirostris mitochondrial metagenome (Table V).

Table III. Nucleotide composition of Protein coding genes of T. vitrirostris.

Gene	Length	A%	T%	G%	C%	AT%	GC%	ATskew	Gcskew
ND1	975	28.1	24.51	15.59	31.79	52.62	47.38	0.0682	-0.34
ND2	1045	29.86	22.01	12.82	35.31	51.87	48.13	0.1513	-0.47
CO1	1545	26.15	28.87	19.48	25.5	55.02	44.98	-0.049	-0.13
CO2	691	29.23	25.33	17.51	27.93	54.56	45.44	0.0715	-0.23
ATP8	168	33.93	26.19	12.5	27.38	60.12	39.88	0.1287	-0.37
ATP6	684	30.12	25.73	13.3	30.85	55.85	44.15	0.0786	-0.4
CO3	785	27.01	27.39	17.32	28.28	54.39	45.61	-0.007	-0.24
ND3	349	27.51	26.65	17.79	28.65	54.15	45.85	0.0159	-0.24
ND4L	297	27.27	27.27	15.82	29.63	54.55	45.45	0	-0.3
ND4	1381	28.46	25.13	14.77	31.64	53.58	46.42	0.0622	-0.36
ND5	1836	31.54	23.97	13.94	30.56	55.5	44.5	0.1364	-0.37
ND6	522	13.22	41.19	32.57	13.03	54.41	45.59	-0.514	0.429
CYTB	1141	26.56	26.91	16.3	30.24	53.46	46.57	-0.007	-0.3

Table IV. Nucleotide composition and codon position of the T. vitrirostris mitochondrial genome.

	Length	A%	T%	G%	C%	AT%	GC%	ATskew	GCskew
Genome	16808	30.15	24.26	16.84	28.75	54.4	45.59	0.11	-0.261
PCG	11419	27.92	26.34	16.46	29.28	54.3	45.74	0.03	-0.28
1st codon position	3807	27	26	19.4	27.4	53	46.8	0.02	-0.171
2nd codon position	3806	26.2	29	15.4	29.6	55.2	45	-0.05	-0.356
3rd codon position	3806	30.6	24	14.6	30.6	54.6	45.2	1	-0.354
rRNAs	2640	33.3	18.79	20.98	26.93	52.1	47.92	0.54	-0.124
tRNAs	1482	30.09	23.41	20.38	26.11	53.5	46.49	0.12	0.1437
D-Loop	1163	31.99	31.04	15.74	21.24	63	36.97	0.02	-0.149

Table V. Codon usage of Thryssa vitrirostris.

Codon	Count	RSCU	Codon	Count	RSCU	Codon	Count	RSCU	Codon	Count	RSCU
UUU(F)	94	0.8	UCU(S)	29	0.78	UAU(Y)	39	0.7	UGU(C)	10	0.69
UUC(F)	133	1.2	UCC(S)	61	1.63	UAC(Y)	75	1.3	UGC(C)	19	1.31
UUA(L)	99	1	UCA(S)	73	1.96	UAA(*)	6	3.7	UGA(W)	96	1.63
UUG(L)	25	0.3	UCG(S)	6	0.16	UAG(*)	0	0	UGG(W)	22	0.37
CUU(L)	97	1	CCU(P)	37	0.69	CAU(H)	18	0.4	CGU(R)	4	0.21
CUC(L)	121	1.2	CCC(P)	77	1.43	CAC(H)	84	1.7	CGC(R)	16	0.85
CUA(L)	198	2	CCA(P)	92	1.7	CAA(Q)	94	1.8	CGA(R)	43	2.29
CUG(L)	66	0.7	CCG(P)	10	0.19	CAG(Q)	8	0.2	CGG(R)	12	0.64
AUU(I)	138	1	ACU(T)	43	0.52	AAU(N)	31	0.5	AGU(S)	12	0.32
AUC(I)	148	1	ACC(T)	##	1.7	AAC(N)	97	1.5	AGC(S)	43	1.15
AUA(M)	116	1.4	ACA(T)	##	1.54	AAA(K)	68	1.7	AGA(*)	0	0
AUG(M)	52	0.6	ACG(T)	19	0.23	AAG(K)	13	0.3	AGG(*)	1	0.31
GUU(V)	43	0.8	GCU(A)	61	0.69	GAU(D)	20	0.6	GGU(G)	29	0.49
GUC(V)	54	1	GCC(A)	##	1.43	GAC(D)	53	1.5	GGC(G)	49	0.82
GUA(V)	86	1.5	GCA(A)	##	1.65	GAA(E)	74	1.5	GGA(G)	113	1.89
GUG(V)	40	0.7	GCG(A)	21	0.24	GAG(E)	27	0.5	GGG(G)	48	0.8

tRNAs of the genome

In this study, 22 tRNA genes were detected in T. vitrirostris mt genes, which are commonly found in metazoans and range in length from 66 bp (tRNAcys) to 75 bp (tRNALeu). Fourteen tRNA genes were transcribed on the H-stand, whereas eight tRNA genes were directed to the L-stand. 26 G-U mismatches in the secondary structure of all arms of 12 of 22 tRNA genes, including RNA-Ala, tRNA-Tyr, tRNA-Gln, tRNA-Asn, tRNA-Trp, tRNA-Cys, tRNA-Asp, tRNA-SerUCN, tRNA-LeuCUN, tRNA-Lys, tRNA-Glu, and tRNA-Pro, created a weak link. A-C mismatches were discovered in the acceptor arm, TφC loop, and aniticodon arm of 11tRNAs, including tRNA-Phe, tRNA-LeuUUR, tRNA-Trp, tRNA-Ile, tRNA-His, and tRNA-LeuCUN, along with tRNA-Thr. In every arm, there were seven T-T mismatches (tRNA-Met, tRNA-Tyr, tRNA-Asn, tRNA-Thr, and tRNA-SerUCN) and four G-A mismatches in the acceptor arm (DHU) and anticodon arms (tRNA-Val, tRNA-Leu UUR, and tRNA-Tyr). One tRNA-Trp U-C mismatch was found in the anticodon arm. In the acceptor arm, DHU arm, TφC loop, and anticodon arm, there were five A-A mismatches of the tRNA-Ile, tRNA-Ser-AGY, tRNA-Thr, and tRNA-Met genes. The acceptor arm (tRNA-SerUCN) was 6 bp in length, and the other 21 tRNAs were 7 bp in length.The DHU arm was 4 bp long, and tRNA-Ile, tRNA-SerUCN, tRNA-Arg and tRNA-Ser-AGY were 3 bpin length. The TφCloops of tRNA-pheand tRNA-Lys were 4 bp in length. Ser-AGY was 6 bp in length, and the other tRNAs were 5 bp in length. The anticodon arm was 5 bp in length in all tRNAs (Fig. 4).

Noncoding regions

The origin of light stand replication (OL) was found inside the WANCY region of tRNAs (tRNA-Trp, tRNA-Asn, tRNA-Ala, tRNA-Cys, and Tyr). The AT content of the OL region was 32.26%, while its GC content was 68.42%, indicating a clear preference for GCs.The control region of T. vitrirostris mito genome was located between tRNAProand tRNAPhe. The length of the D-loop1163bp. The overall nucleotide composition of G, A, T, and C were 31.99%, 31.04%, 17.74%, and 21.24%, respectively. It was rich in A+T (63.5%) than G+C content (36.5 %).

Phylogenetic relationships

Topological structures were generated via phylogenetic relationship analysis (Figs. 5, 6). The 13 PCGs of 24 Clupeoidei species, including three families (Engruulidae, Pristigasterridae, and Clupeidae), were constructed. For every internode, the topological relationships of the two phylogenetic analyses yielded high bootstrap support levels. We anticipate that the current findings will make research on the genetic structure, taxonomy and evolutionary relationships of Clupeoide much easier.

DISCUSSION

In the present study, Sanger sequencing was used to identify the mt genome of T. vitrirostris. The variation in mitochondrial DNA length is based on the varying lengths of regulatory regions generally found in the majority of vertebrates (Ketmaier and Bernardini, 2005; Randi et al., 1998; Skorupski, 2022). Each tRNA was able to fold into a typical cloverleaf secondary structure, in addition totRNA-Ser (AGY), which lacked the dihydrouracil arm. Consistent with other teleost mitogenomes, the 16,808 bp genome of T. vitrirostris contains 37 genes and a single control region. With the exception of ND6, all twelve GCskews and four ATskews were negative, suggesting that the H-strand had an excess of cytosine and adenine and that the nucleotide composition was asymmetrical. Combining DNA barcoding with phylogenetic analysis has been widely used to increase species identification and understand evolutionary relationships (Buxton et al., 2021). In addition to enhancing species variation and evolutionary biology, the combination of DNA barcoding and phylogenetic analysis also increases the accuracy of species identification. The mitochondrial genome contains nucleotide insertions, deletions, and tandem repetitions, and this show species variation (Peng et al., 2006). The phylogenetic construction of Thryssa revealed that several clusters were formed by members of the same species (Gopinath and Abdussamad, 2024).

The mitogenome order, direction, positions, and gene coding strands observed in this study were similar to those of the Engraulidae family (Zhanga et al., 2019; Chen et al., 2018; Zhao and Liu, 2016). The A+T content of the mt genome of Engraulidae observed in this study was greater than the G+C content reported previously (Zhang and Liu, 2016; Bo et al., 2013). The starting codons of PCGs (ATG and GTG) observed in this study were similar to those reported in other fish species (Gong et al., 2023; Nguyen et al., 2023; Xu et al., 2024; Zeng et al., 2024). In this study, some of the incomplete codons were observed, and this finding was corroborated by Ojala et al. (1981), who reported that posttranscriptional polyadenylation can transform such noncanonical stop codons into fully functional TAA stop codons. The RSCU value of 13 PCGs of T. vitrirostris was 27, with a value >1 indicating positive biases in codon usage, and no bias was observed, which was similar to the findings of a previous report (Sheikh et al., 2020). The nucleotide composition of metazoan genomes typically exhibits a pronounced strand bias that may be quantified as ATand GC skews (Hassanin et al., 2005; Perna and Kocher, 1995). Important indicators of nucleotide variations between the heavy and light chains were AT skew and GC skew values. The greater the variance between AT skew and GC skew is, the greater the absolute value (Perna and Kocher, 1995). In this study, most ATskews were negative. Since most mitogenomes have a conventional preference for positive ATskews and negative GCskews, the former is typically smaller in size than the latter (Fonseca et al., 2014).

In addition, totRNASer (AGY), twenty-one tRNA genes presented cloverleaf secondary structures (Zhang and Xian, 2016). Partial mismatches of RNA genes were observed in this study, and these results corroborated those of previous reports. These partially mismatched RNA genes are used to eliminate mutations and may be corrected through subsequent RNA editing without interfering with the transfer of amino acids (Tomita et al., 1996; Lynch, 1997). In this study, tRNA-Asn and tRNA-Cys were separated by 31 bp, similar to other vertebrates (Macey et al., 1997; Zhang et al., 2014). The control region of mtDNA is the quickest evolutionary sequence and has been used to study molecular systematics and population genetics (Liu et al., 2008; Xiao and Zang, 2000).

CONCLUSION

This research analysed the sequences of the entire mitogenome of T. vitrirostris (accession number: PQ385613), which was collected from the southern coast of India. The organization of the mitogenome structure and gene arrangement resembled those ofother fishes in the Engraulidae family. Like other teleost mitogenomes, the mitogenome is 16808 bp long and contains 37 genes and a regulatory region. The usual cloverleaf secondary structure can be formed by folding any tRNA. T. vitrirostris was shown to be closely related to T. hamiltonii and T. setirostris, forming a strong monophyletic group in this study. The present findings revealed T. vitrirostris taxonomy, species evolution and genetic diversity.

Declarations

Acknowledgement

Authors gratefully acknowledge Department of Zoology, Nesamony Memorial Christian College, Marthandam for laboratory facilities.

Funding

The study received no external funding.

IRB approval

Not applicable.

Ethical Statement

No live samples were used in the present investigation.

Generative AI or AI-assisted technology statement

The authors have declared that no generative AI or AI-assisted technologies were used to create this manuscript.

Statement of conflict of interest

The authors have declared no conflict of interest.

REFERENCES

Altschul, S.F., Gish, W., Miller, W., Myers, E.W. and Lipman, D.J., 1990. Basic local alignment search tool. J. mol. Biol., 215: 403-410. https://doi.org/10.1016/S0022-2836(05)80360-2

Bo, Z., Xu, T., Wang, R., Jin, X. and Sun, Y., 2013. Complete mitochondrial genome of the Osbeck’s grenadier anchovy Coiliamystus (Clupeiformes, Engraulidae). Mitochond. DNA, 24: 657-659. https://doi.org/10.3109/19401736.2013.772599

Brown, K.H., 2008. Fish mitochondrial genomics: sequence, inheritance and functional variation. J. Fish Biol., 72: 355-374. https://doi.org/10.1111/j.1095-8649.2007.01690.x

Buxton, R.T., Bennett, J.R., Reid, A.J., Shulman, C., Cooke, S.J., Francis, C.M., Nyboer, E.A., Pritchard, G., Binley, A.D., Avery-Gomm, S. and Ban, N.C., 2021. Key information needs to move from knowledge to action for biodiversity conservation in Canada. Biol. Conserv., 256: 108983. https://doi.org/10.1016/j.biocon.2021.108983

Chen, W., Jiang, H., Du, X., Gong, L., Liu, L., Liu, B. and Lü, Z., 2018. The complete mitochondrial genome of Thryssasetirostris (Engraulinae, Engranlidae, Clupeoidei) and phylogenetic studies of Clupeoidei. Mitochond. DNA Part B, 3: 326-327. https://doi.org/10.1080/23802359.2018.1437807

Diagne, C., Leroy, B., Vaissière, A.C., Gozlan, R.E., Roiz, D., Jarić, I., Salles, J.M., Bradshaw, C.J. and Courchamp, F., 2021. High and rising economic costs of biological invasions worldwide. Nat., 592: 571-576. https://doi.org/10.1038/s41586-021-03405-6

Fonseca, M.M., Harris, D.J. and Posada, D., 2014. The inversion of the control region in three mitogenomes provides further evidence for an asymmetric model of vertebrate mtDNA replication. PLoS One, 9: e106654. https://doi.org/10.1371/journal.pone.0106654

Gao, X., Li, Y., Zhang, R., Lv, Y., Wang, Y., Shi, J., Xie, J., Kong, C. and Li, L., 2024. The complete mitochondrial genome of a rare cavefish (Sinocyclocheilus cyphotergous) and comparative genomic analyses in Sinocyclocheilus. Pakistan J. Zool., 56: 2245. https://doi.org/10.17582/journal.pjz/20230213080243

Gong, Y., Yang, T., Su, C., Zhao, L. and Li, F., 2023. The complete mitochondrial genome of the Tanichthys flavianalis (Cypriniformes: Tanichthyidae) and its phylogeny. Mitochond. DNA Part B, 8: 1325-1329. https://doi.org/10.1080/23802359.2023.2282794

Gopinath, A. and Abdussamad, E.M., 2024. Phylogenetic analysis and redescription of the specimen of Thryssa stenosoma (Teleostei: Clupeiformes: engraulidae) collected from Puri, Odisha, Bay of Bengal. Int. Educ. Res., 10: 19-22.

Gopinath, A., Abdussamad, E.M., Tomy, E., Thomas, T. and Retheesh, T.B., 2024. Length-weight relationship of six species of Thryssa with emphasis on the biology of Thryssa mystax from Indian waters. J. Mar. Biol. Assoc., 66: 66-71. https://doi.org/10.6024/jmbai.2024.66.1.2439-10

Greiner, S., Lehwark, P. and Bock, R., 2019. Organellar Genome DRAW (OGDRAW) version 1.3. 1: expanded toolkit for the graphical visualization of organellar genomes. Nucl. Acids Res., 47: W59-W64. https://doi.org/10.1093/nar/gkz238

Hassanin, A., Leger, N.E.L.L.Y. and Deutsch, J., 2005. Evidence for multiple reversals of asymmetric mutational constraints during the evolution of the mitochondrial genome of Metazoa, and consequences for phylogenetic inferences. Syst. Biol., 54: 277-298. https://doi.org/10.1080/10635150590947843

Hata, H. and Lavoué, S., 2024. Resurrection and redescriptions of nominal species previously regarded as synonyms of Thrissina mystax (Bloch and Schneider, 1801) (Teleostei: Clupeiformes: Engraulidae). J. Fish Biol., 104: 1445-1467. https://doi.org/10.1111/jfb.15684

Huang, X. and Madan, A., 1999. CAP3: A DNA sequence assembly program. Genome Res., 9: 868-877. https://doi.org/10.1101/gr.9.9.868

Jo, T., Takao, K. and Minamoto, T., 2022. Linking the state of environmental DNA to its application for biomonitoring and stock assessment: Targeting mitochondrial/nuclear genes, and different DNA fragment lengths and particle sizes. Environ. DNA, 4: 271-283. https://doi.org/10.1002/edn3.253

Junqueira, A.C.M., Lessinger, A.C., Torres, T.T., da Silva, F.R., Vettore, A.L., Arruda, P. and Espin, A.M.L.A., 2004. The mitochondrial genome of the blowfly Chrysomya chloropyga (Diptera: Calliphoridae). Gene, 339: 7-15. https://doi.org/10.1016/j.gene.2004.06.031

Ketmaier, V. and Bernardini, C., 2005. Structure of the mitochondrial control region of the Eurasian otter (Lutralutra; Carnivora, Mustelidae): Patterns of genetic heterogeneity and implications for conservation of the species in Italy. J. Hered., 96: 318-328. https://doi.org/10.1093/jhered/esi037

Laslett, D. and Canbäck, B., 2008. ARWEN: A program to detect tRNA genes in metazoan mitochondrial nucleotide sequences. Bioinformatics, 24: 172-175. https://doi.org/10.1093/bioinformatics/btm573

Liu, H.Y., Yu, L.N. and Zhang, F.R., 2008. Molecular structure and application progress of fish mitochondrial DNA control region. Reservoir. Fish, 28: 4-8.

Lynch, M., 1997. Mutation accumulation in nuclear, organelle, and prokaryotic transfer RNA genes. Mol. Biol. Evol.,14: 914-925. https://doi.org/10.1093/oxfordjournals.molbev.a025834

Macey, J.R., Larson, A., Ananjeva, N.B., Fang, Z. and Papenfuss, T.J., 1997. Two novel gene orders and the role of light-strand replication in rearrangement of the vertebrate mitochondrial genome. Mol. Biol. Evol., 14: 91-104. https://doi.org/10.1093/oxfordjournals.molbev.a025706

Madeira, F., Madhusoodanan, N., Lee, J., Eusebi, A., Niewielska, A., Tivey, A.R., Lopez, R. and Butcher, S., 2024. The EMBL-EBI job dispatcher sequence analysis tools framework in 2024. Nucl. Acids Res., 52: W521-W525. https://doi.org/10.1093/nar/gkae241

Néron, B., Ménager, H., Maufrais, C., Joly, N., Maupetit, J., Letort, S., Carrere, S., Tuffery, P. and Letondal, C., 2009. Mobyle: A new full web bioinformatics framework. Bioinformatics, 25: 3005-3011. https://doi.org/10.1093/bioinformatics/btp493

Nguyen, P.T., Lee, S., Jeong, J., Kim, J., Han, D.W., Kim, I.C., Lee, J.H., Park, J. and Kim, J.H., 2023. Complete mitochondrial genome of Trematomus newnesi (Perciformes, Nototheniidae). Mitochond. DNA B, 8: 1196-1199. https://doi.org/10.1080/23802359.2023.2194456

Ojala, D., Montoya, J. and Attardi, G., 1981. tRNA punctuation model of RNA processing in human mitochondria. Nature, 290: 470-474. https://doi.org/10.1038/290470a0

Pawase, S.V., Nirmale, V.H., Bhosale, B.P., Pawar, R.A., Sawant, M.S. and Kende, D.R., 2020. Study on biology of Thryssa dussumieri (Valenciennes, 1848) from the coast of Ratnagiri, Maharashtra, India. Int. J. mol. Sci., 49: 87-94. https://doi.org/10.1016/j.gene.2006.04.014

Peng, Z., Wang, J. and He, S., 2006. The complete mitochondrial genome of the helmet catfish Cranoglanis bouderius (Siluriformes: Cranoglanididae) and the phylogeny of otophysan fishes. Gene, 376: 290-297.

Perna, N.T. and Kocher, T.D., 1995. Patterns of nucleotide composition at fourfold degenerate sites of animal mitochondrial genomes. J. mol. Evol., 41: 353-358. https://doi.org/10.1007/BF01215182

Randi, E. and Lucchini, V., 1998. Organization and evolution of the mitochondrial DNA control region in the avian genus Alectoris. J. mol. Evol., 47: 449-462. https://doi.org/10.1007/PL00006402

Ronquist, F., Teslenko, M., Van Der Mark, P., Ayres, D.L., Darling, A., Höhna, S., Larget, B., Liu, L., Suchard, M.A. and Huelsenbeck, J.P., 2012. MrBayes 3.2: Efficient Bayesian phylogenetic inference and model choice across a large model space. Syst. Boil., 61: 539-542. https://doi.org/10.1093/sysbio/sys029

Saha, S., haoJia, C., Song, N., Suzuki, T., Baki, M.A., Yang, T. and Gao, T., 2024. Comparative analyses of complete mitochondrial genomes of four sillaginids fish (Perciformes: Sillaginidae) and phylogenetic implications. Bangladesh J. scient. indust. Res., 59: 87-104. https://doi.org/10.3329/bjsir.v59i2.71326

Satoh, T.P., Miya, M., Mabuchi, K. and Nishida, M., 2016. Structure and variation of the mitochondrial genome of fishes. BMC Genom., 17: 1-20. https://doi.org/10.1186/s12864-016-3054-y

Sheikh, A., Al-Taher, A., Al-Nazawi, M., Al-Mubarak, A.I. and Kandeel, M., 2020. Analysis of preferred codon usage in the coronavirus N genes and their implications for genome evolution and vaccine design. J. Virol. Methods, 277: 113806. https://doi.org/10.1016/j.jviromet.2019.113806

Skorupski, J., 2022. Characterisation of the complete mitochondrial genome of critically endangered Mustelal utreola (Carnivora: Mustelidae) and its phylogenetic and conservation implications. Genes, 13: 125. https://doi.org/10.3390/genes13010125

Tamura, K., Stecher, G. and Kumar, S., 2021. MEGA11: Molecular evolutionary genetics analysis version 11. Mol. Biol. Evol., 38: 3022-3027. https://doi.org/10.1093/molbev/msab120

Tamura, K., Stecher, G., Peterson, D., Filipski, A. and Kumar, S., 2013. MEGA6: Molecular evolutionary genetics analysis version 6.0. Mol. Biol. Evol., 30: 2725-2729. https://doi.org/10.1093/molbev/mst197

Tomita, K., Ueda, T. and Watanabe, K., 1996. RNA editing in the acceptor stem of squid mitochondrial tRNATyr. Nucl. Acids Res., 24: 4987-4991. https://doi.org/10.1093/nar/24.24.4987

Xiao, W.H. and Zhang, Y.P., 2000. Genetics and evolution of mitochondrial DNA in fish. Acta Hydrobiol. Sin., 24: 384-391. https://doi.org/10.3724/issn1000-3207-2000-4-384-v

Xu, C.H., Song, L., Wang, Q. and Chen, X.J., 2024. Complete mitochondrial genome of Devarios hanensis (Cypriniformes: Danionidae: Danioninae): Genome characterization and phylogenetic consideration. Mitochond. DNA B, 9: 797-801. https://doi.org/10.1080/23802359.2024.2363367

Zeng, Q., Chen, Y.J., Liu, M. and Wang, C., 2024. Characteristics of the complete mitochondrial genome of Gerres limbatus (Cuvier, 1830) (Perciformes: Gerreidae). Mitochond. DNA B, 9: 419-422. https://doi.org/10.1080/23802359.2024.2333571

Zhang, B., Sun, Y. and Shi, G., 2014. The complete mitochondrial genome of the four finger threadfin Eleutheronema tetradactylum (Perciforms: Polynemidae) and comparison of light strand replication origin within Percoidei. Mitochond. DNA, 25: 411–413. https://doi.org/10.3109/19401736.2013.809433

Zhang, H. and Xian, W., 2016. The complete mitochondrial genome of the larvae Osbeck’s grenadier anchovy Coilia mystus (Clupeiformes, Engraulidae) from Yangtze estuary. Mitochond. DNA A, 27: 966-967. https://doi.org/10.3109/19401736.2014.926508

Zhang, J., Zhang, N., Li, Y., Zhang, R. and Lin, L., 2019. Complete mitochondrial DNA genome of Thryssa dussumieri (Valenciennes, 1848). Mitochond. DNA B, 4: 1314-1315. https://doi.org/10.1080/23802359.2019.1591208

Zhang, R., Deng, L., Lv, X. and Tang, Q., 2022. Complete mitochondrial genomes of two catfishes (Siluriformes, Bagridae) and their phylogenetic implications. Zookeys, 1115: 103. https://doi.org/10.3897/zookeys.1115.85249

Zhang, R., Tang, Q. and Deng, L., 2021. The complete mitochondrial genome of Microphysogobio elongatus (Teleostei, Cyprinidae) and its phylogenetic implications. ZooKeys, 1061: 57. https://doi.org/10.3897/zookeys.1061.70176

Zhao, L. and Liu, Q., 2016. Complete mitochondrial genome sequence of Thryssa kammalensis (Clupeiformes, Engraulidae). Mitochond. DNA B, 1: 302-303. https://doi.org/10.1080/23802359.2016.1167637