A new species of the genus Xistra (Orthoptera: Tetrigoidea: Metrodorinae) with comments on the characters of mitochondrial genome Author Bai, Qiu-Xia 0009-0002-2038-3988 Key Laboratory of Ecology of Rare and Endangered Species and Environmental Protection (Guangxi Normal University), Ministry of Education, Guilin, Guangxi 541006, China; & Guangxi Key Laboratory of Rare and Endangered Animal Ecology, Guangxi Normal University, Guilin, Guangxi 541006, China; & College of Life Science, Guangxi Normal University, Guilin, Guangxi 541006, China Author Long, Min 0009-0005-1174-3533 Key Laboratory of Ecology of Rare and Endangered Species and Environmental Protection (Guangxi Normal University), Ministry of Education, Guilin, Guangxi 541006, China; & Guangxi Key Laboratory of Rare and Endangered Animal Ecology, Guangxi Normal University, Guilin, Guangxi 541006, China; & College of Life Science, Guangxi Normal University, Guilin, Guangxi 541006, China Author Deng, Wei-An Key Laboratory of Ecology of Rare and Endangered Species and Environmental Protection (Guangxi Normal University), Ministry of Education, Guilin, Guangxi 541006, China; & Guangxi Key Laboratory of Rare and Endangered Animal Ecology, Guangxi Normal University, Guilin, Guangxi 541006, China; & College of Life Science, Guangxi Normal University, Guilin, Guangxi 541006, China text Zootaxa 2024 2024-05-07 5447 3 373 384 http://dx.doi.org/10.11646/zootaxa.5447.3.3 journal article 10.11646/zootaxa.5447.3.3 1175-5326 11149908 11870D7A-C747-43DB-8158-D2D89CAFF00B Xistra zhengi Deng , sp. nov. (Figures 1,2) Description. Female. Body small, its surface interspersed with coarse protuberances and notches ( Fig. 1A ). Head. Head and eyes distinctly exserted above pronotal surface ( Fig. 1C ). Vertex narrow, in dorsal view, width of vertex between eyes 0.8 times width of compound eye; a little narrower in front; anterior margin of fastigium narrowly rounded and not surpassing anterior margin of eye ( Fig. 2C ); median carina visible anteriorly; lateral margins turned backward and slightly exserted above upper margin of eye; vertex uneven with paired fossulae. In lateral view, frontal costa nearly straight and slightly visible between eyes, protruded anteriorly and broadly rounded between antennal grooves ( Fig. 2B ). In frontal view, vertex with V-shaped concavity, bottom of which reaching one-fourth of upper inner margins of eyes; lateral margins of vertex slightly exerted above upper margin of eyes; frontal costa bifurcated above lateral ocelli, longitudinal furrow divergent between antennae, width of longitudinal furrow of frontal ridge narrower than antennal groove diameter ( Fig. 2A ). Antennae short, filiform, antennal grooves inserted between inferior margins of compound eyes ( Fig. 2B ), 15-segmented, the 9th and 10th segment are the longest, about 6–7 times longer than its width. Eyes globose, lateral (paired) ocelli located in slightly below the middle of compound eye height. FIGURE 1 A–D. Xistra zhengi Deng , sp. nov. , ♀, ♁. A—body of female, dorsal view; B—body of male, dorsal view; C—body of female, lateral view; D—body of male, lateral view (Scale bar = 1 mm). FIGURE 2 A–L. Xistra zhengi Deng , sp. nov. , A-J ♀, K-L ♁. A—head, frontal view; B—head and pronotum, dorsal view; C—head, lateral view; D—left fore femur, lateral view; E—left middle femur, lateral view; F—left hind femur, lateral view; G—left hind tibia, lateral view; H—left hind tarsi, lateral view; I—ovipositor of female, lateral view; J—female subgenital plate, ventral view; K—male subgenital plate, lateral view; L—male subgenital plate, ventral view (Scale bar = 1 mm). Thorax. Pronotum coarse, its surface granulose and with sporadic protuberances and concavities ( Fig. 1A ). In dorsal view, anterior margin of pronotum truncate; lateral carinae of prozona parallel; median carina of pronotum entire; before and behind humeral angles are depressed, behind middle of pronotum with sporadic protuberances and notches; obtuse humeral angles; without interhumeral carinae between shoulders; hind pronotal process narrow and long cone-shaped, slightly surpassing the apex of hind femur. In profile, median carina of pronotum slightly undulate before shoulders and straight behind shoulders. Lateral lobes of pronotum turning outwardly, posterior angles directed distinctly transversely and apex truncate. Posterior margins of lateral lobes of pronotum with ventral sinus and tegminal (upper) sinus. Tegmina long, ovate, apex rounded. Hind wings reaching or slightly surpassing apex of hind pronotal process. Legs. Fore and middle femora with carinated, margins finely serrated, dorsal margins of fore and middle femora nearly straight, ventral margins of fore and middle femora slightly undulated ( Fig. 2D, E ). Middle femur wider than visible part of tegmen in width. Hind femora robust and short, 3 times as long as wide; margins finely serrated ( Fig. 2F ); antegenicular denticlesand acute genicular denticles right angled. Outer side of hind tibia with 6–7 spines, inner side with 5-6 spines ( Fig. 2G ). First segment of posterior tarsi equal to third in length, first pulvillus and first segment of tarsus longer than the second and third, apices of three pulvilli obtuse ( Fig. 2H ). Abdomen. Ovipositor narrow and long, upper valve 4.7 times as long as wide, upper and lower valvulae with slender saw-like teeth ( Fig. 2H ). Length of subgenital plate nearly equal to its width, posterior margin of subgenital plate triangular projecting ( Fig. 2I ). Coloration. Body yellow-brown. Antennae yellow-brown, apical segments dark. Tegmina with anterior part dark and posterior part yellow-white. Hind wings black. Fore and mid legs yellow-brown, with two dark spots on femora and tibia. Hind femur with two irregular dark spots on outer side. Hind tibia yellow-brown, with three black rings ( Figs. 1A, C ). Sternites of thorax and abdomen dark brown, subgenital plate brown. Male. Similar to female, but body slender and smaller ( Figs. 1B, D ). Width of vertex between eyes 0.6-0.8 times width of compound eye. Middle femur much wider than visible part of tegmen in width. Subgenital plate short, coneshaped, apex biforked ( Figs. 2K, L ). Measurements (mm). Length of body ♀ 11.1–12.9 mm , ♁ 10.3–10.8 mm ; length of pronotum ♀ 10.3–11.2 mm , ♁ 8.7–9.7 mm ; length of hind femur ♀ 6.1–6.2 mm , ♁ 5.3–5.8 mm . Type material. Holotype : ♀ , China , Jiangxi prov. , Ganzhou (Yudu), 25°42ʹ21ʹʹ N , 115°25ʹ10ʹʹ E , 1073m alt., 19 August 2020 , collected by Wei-An DENG , CLSGNU . Paratypes . 4♁ 1♀ , same data, collected by Wei-An DENG , Jin Yue HE and Qiu Mei YU , CLSGNU . Diagnosis. New species can be easily distinguished from other species of the genus by pronotal surface granulose and with sporadic protuberances and concavities. The new species is similar to Xistra bannaensis Deng, Wang, Mao & Li, 2021 from which it differs in lateral margins of vertex slightly exerted above upper margin of eyes (lateral margins of vertex barely reaching the level of upper margin of eyes in X. bannaensis ); humero-apical carina absent (humero-apical carina conspicuous in X. bannaensis ); pronotal surface granulose and with sporadic protuberances and concavities (pronotal surface smooth and flat in X. bannaensis ); tegmina with anterior part dark and posterior part yellow-white (tegmina with upper part yellow and lower part brown in X. bannaensis ). Etymology. The new species was named in memory of Professor Zhemin Zheng (1932–2021), a famous Chinese orthopterologist and a founder of Chinese tetrigides study. Distribution. P. R. CHINA : Jiangxi . Characteristics of Newly Sequenced Mitogenomes. In this study, the mitogenome of Xistra zhengi Deng , sp. nov. was circular molecule, with total lengths of 18148 bp ( Table 2 ). The structure of this newly sequenced mitogenome contains 13 protein-coding genes (PCGs), 22 tRNAs, 2 rRNA ( rrnS and rrnL ), and control regions rich in A+T bases ( Table 3 ). In all mitochondrial genes, 14 genes (4 PCGs, 8 tRNAs and 2 rRNAs) were located on the minority strand, and 23 genes (9 PCGs and 14 tRNA genes) were located on the majority strand. Among the 13 protein-coding genes, trn1, nad4, nad4L, and nad5 were located on the N strand, while the other genes (cox1, cox2, cox3, nad2, nad3, nad6, cytb, atp6, and atp8) were located on the J strand. The gene arrangement of the newly sequenced mitochondrial genome was similar to those of other species of Tetrigidae (Lin, 2014) . The base composition of Xistra zhengi Deng , sp. nov. mitogenome is A (44.20%), T (28.30%), C (18.20%) and G (9.30%). The base composition was A+T-biased, and the proportion of A+T content was 72.50%. The AT-skew was 0.218, and the GC-skew was -0.325. This shows that the content of bases A is higher than that of T, and C is higher than G in the whole ( Table 3 ). The total length of the 13 PCGs is 11,107 bp. All of the 13 PCGs initiate with the standard start codon ATN. There are three types of stop codons, TAA (cox1, cox2, cox3, nad1, nad4l, nad6, atp8, atp6) and TAG (nad3, nad4, cytb), as well as T- (nad5). The relative synonymous codon usage (RSCU) values of the mitogenome were summerized ( Fig. 4 ). The codon distribution analysis showed that the two codons UUA (Leu2) and UCA (Ser2) were the most frequently used in this new species. TABLE 1. Species and Genbank accession numbers used in this study.

NC_071822

NC_082935

NC_063117

MH791445

MT162547

TABLE 2. Organization of the Xistra zhengi Deng , sp. nov. mitogenome.

Genes	Strand	Anticodon	Location	Length(bp)	Ovl/nc	Start/Stop Codon
trnI	J	GAT	1–66	66	0
trnQ	N	TTG	67–135	69	-1
trnM	J	CAT	135–201	67	14
nad2	J	216–1211	996	-2	ATG/TAA
trnW	J	TCA	1210–1274	65	-8
trnC	N	GCA	1267–1327	61	0
trnY	N	GTA	1328–1389	62	-3
cox1	J	1387–2925	1539	-5	ATC/TAA
trnL2	J	TAA	2921–2983	63	2
cox2	J	2986–3663	678	-2	ATG/TAA
trnK	J	CTT	3662–3723	62	1
trnD	J	GTC	3725–3791	67	3
atp8	J	3795–3953	159	-7	ATG/TAA
atp6	J	3947–4618	672	-1	ATG/TAA

......continued on the next page TABLE 2. (Continued)

Genes	Strand	Anticodon	Location	Length(bp)	Ovl/nc	Start/Stop Codon
cox3	J	4618–5409	792	-1	ATG/TAA
trnG	J	TCC	5409–5471	63	0
nad3	J	5472–5825	354	-2	ATA/TAG
trnA	J	TGC	5824–5888	65	-1
trnR	J	TCG	5888–5953	66	1
trnN	J	GTT	5955–6017	63	0
trnS1	J	GCT	6018–6084	67	0
trnE	J	TTC	6085–6148	64	-2
trnF	N	GAA	6147–6207	61	6
nad5	N	6214–7933	1720	1	ATG/T(AA)
trnH	N	GTG	7935–7999	65	-1
nad4	N	7999–9324	1326	-7	ATG/TAG
nad4l	N	9318–9608	291	2	ATT/TAA
trnT	J	TGT	9611–9673	63	0
trnP	N	TGG	9674–9738	65	10
nad6	J	9749–10249	501	-1	ATT/TAA
cob	J	10249–11385	1137	-2	ATG/TAG
trnS2	J	TGA	11384–11448	65	543
nad1	N	11992–12933	942	0	ATT/TAA
trnL1	N	TAG	12934–12996	63	1
rrnL	N	12998–14290	1293	3
trnV	N	TAC	14294–14360	67	-3
rrnS	N	14358–15086	729	0
CR	15087–18148	3062

TABLE 3. Nucleotide composition of mitochondrial genome of Xistra zhengi , sp. nov.

Genes or partitions	A (%)	T (%)	G (%)	C (%)	A+T (%)	AT-skew	GC-skew
Whole genome	44.20	28.30	9.30	18.20	72.50	0.218	-0.325
PCGs	30.40	38.90	14.80	15.90	69.30	-0.123	-0.036
PCGs-1st	33.70	33.80	19.20	13.30	67.50	-0.002	0.181
PCGs-2st	20.60	45.20	15.10	19.10	65.80	-0.374	-0.116
PCGs-3st	37.10	37.90	9.90	15.10	75.00	-0.011	-0.207
tRNA	36.20	35.80	16.30	11.80	72.00	0.005	0.161
rRNA	26.60	48.20	17.10	8.20	74.80	-0.289	0.350
CR	51.40	29.33	4.90	14.37	80.73	0.273	-1.831

FIGURE 3. Circular map of the mitogenome of Xistra zhengi Deng , sp. nov. FIGURE 4. The relative synonymous codon usage (RSCU) in the mitogenome of Xistra zhengi , sp. nov. The lengths of the 22 tRNA genes range from 61 to 69 bp. Two rRNA genes ( rrnL and rrnS ), separated by tRNA-Val , are located between tRNA-Leu1 and A+T-rich region. The lengths of rrnL and rrnS are 1,293 bp and 729 bp, respectively. The A+T-rich region of 3,062 bp, with an A+T content of 80.73%, is located between rrnS and tRNA-Ile . The secondary structures of tRNAs were illustrated in Figure 5 . All tRNA genes could be folded into the typical clover-leaf structure. The mismatch of A–A occurred in trnG ; A–G occurred in trnW and trnY ; A–C occurred in trnG , trnM and trnS1 ; and C–U only occurred in trnA ( Fig. 5 ). The mismatch of U–U existed in trnI , trnE , trnL2 , and trnY . The G–U mismatch occurred most frequently, but it did not appear in trnA , trnD , trnE , trnI , trnK , trnL2 , trnN , trnR , trnS2 , trnW , or trnY , and there were 30 G–U mismatches. Phylogenetic Analysis . Bayesian Inference (BI) and Maximum Likelihood (ML) tree were constructed using 13 PCGs from mitogenomes of 16 species of Tetrigoidea and one outgroup ( Mirhipipteryx andensis of Tridactyloidea) to validate the phylogenetic position of Xistra zhengi Deng , sp. nov. ( Table 1 ). ML and BI trees had the same topological structures ( Fig. 6 ). The results of the phylogenetic tree showed that the new species was placed in Metrodorinae , and formed a separate clade from other species of Metrodorinae . The results of phylogenetic tree constructed by ML and BI methods were as follows: (Batrachideinae + (Tripetalocerinae + ( Metrodorinae + ( Metrodorinae + Thoradontini)))).The phylogenetic tree in this study appears to support the placement of the genus Thoradonta within the Metrodorinae subfamily ( Fig. 6 ), however, the available data is insufficient to confirm this placement conclusively ( Adžić et al. , 2020 ; Qin et al. , 2023 ). More molecular and morphological data are necessary to provide a more robust understanding of the evolutionary relationships within this group. By incorporating more sequences and specimens, we can achieve a more comprehensive phylogeny that will aid in the accurate classification of Thoradonta and other Tetrigidae genera.