Multigene Sequence-Based and Phenotypic Characterization Reveals the Occurrence of a Novel Entomopathogenic Nematode Species,  n. sp.

Aashaq Hussain Bhat; Ricardo A. R. Machado; Joaquín Abolafia; Tarique Hassan Askary; Vladimír Půža; Alba Nazaret Ruiz-Cuenca; Aasha Rana; Samy Sayed; Laila A. Al-Shuraym

Open Access

Multigene Sequence-Based and Phenotypic Characterization Reveals the Occurrence of a Novel Entomopathogenic Nematode Species, Steinernema anantnagense n. sp.

Aashaq Hussain Bhat

Ricardo A. R. Machado

Joaquín Abolafia

Tarique Hassan Askary

Vladimír Půža

Alba Nazaret Ruiz-Cuenca

Aasha Rana

Samy Sayed

and

Laila A. Al-Shuraym

| Jul 13, 2023

Journal of Nematology

Volume 55 (2023): Issue 1 (February 2023)

About this article

Cite

Published Online: Jul 13, 2023

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Received: Apr 06, 2023

DOI: https://doi.org/10.2478/jofnem-2023-0029

Keywords
Species description, nematodes, molecular characterization, phylogenetic systematics, taxonomy, biological control, sustainable agriculture

© 2023 Aashaq Hussain Bhat et al., published by Sciendo

This work is licensed under the Creative Commons Attribution 4.0 International License.

Line drawings of first- and second-generation adults and infective juveniles of Steinernema anantnagense n. sp. (A–C) First-generation female: (A) Anterior end; (B) Neck region; (C) Posterior end. (D–F) First-generation male: D) Anterior end; (E) Neck region; (F) Posterior end. (G–I) Second-generation female: (G) Anterior end; (H) Neck region; (I) Posterior end. (J–L) Second-generation male: (J) Anterior end; (K) Neck region; (L) Posterior end. (M–O) Infective juvenile: (M) Anterior end; (N) Neck; (O) Posterior end.

Light microscope micrographs of first-generation adults and IJ of Steinernema anantnagense n. sp. (A–C) Female: (A) Anterior end; (B) Neck region (arrow pointing to the excretory pore); (C) Posterior end. (D–F) Male: (D) Anterior end; (E) Neck region (arrow pointing to the excretory pore); (F) Posterior end (arrow pointing to the mid-ventral genital papillae). (G–I) Infective juvenile: (G) Anterior end; (H) Neck region (black arrow pointing to the excretory pore, white arrow pointing to the bacteria sac); (I) Posterior end.

Light microscope micrographs of second-generation adults and IJ of Steinernema anantnagense n. sp. (A–C) Second-generation female: (A) Anterior end; (B) Neck region; (C) Posterior end. (D–F) Second-generation male: D) Anterior end; (E) Neck region; (F) Posterior end (arrow pointing to the mid-ventral genital papillae). (G) Entire infective juvenile (arrow pointing to the bacteria sac).

Scanning electron microscope micrographs of first-generation adults of Steinernema anantnagense n. sp. (A, B) Lip region of female in frontal and lateral view, respectively (arrows pointing to the amphids); (C) Excretory pore of female; (D) Vulva in ventral view; (E) Female posterior end (arrow pointing to the anus). (F–H) Male posterior end in lateral, ventral and subventral view, respectively (GP: genital papilla, MP: mid-ventral papilla, ph: phasmid). (I, J) Lip region of IJ in frontal and lateral view, respectively (arrows pointing to the amphids); (K) Excretory pore of IJ; (L) Lateral field of IJ; (M, N) Posterior end of IJ in ventral and lateral view, respectively (arrows pointing to the phasmids).

Scanning electron microscope micrographs of second-generation adults and IJ of Steinernema anantnagense n. sp. (A, B) Lip region of female in frontal and lateral view, respectively (arrows pointing to the amphids); (C) Excretory pore of female; (D) Vulva in ventral view; (E, F) Female posterior end in lateral and ventral view, respectively (arrow pointing to the anus). (G, H) Lip region of male in frontal and lateral view, respectively (arrows pointing to the amphids); (I) Male excretory pore; (J–L) Male posterior end in lateral, ventral and subventral view, respectively (GP: genital papilla, MP: mid-ventral papilla, ph: phasmid).

Maximum-likelihood phylogenetic tree between the newly described Steinernema anantnagense n. sp. and other closely related species of Steinernema species based on nucleotide sequences of: (A) the Internal Transcribed Spacer (ITS1-5.8S-ITS2) rRNA, flanked by primers 18S and 26S, and (B) the D2-D3 expansion segments of the large subunit (28S) of rRNA flanked by primers D2F and 536. Numbers at nodes represent bootstrap values based on 100 replications. Bars represent average nucleotide substitutions per sequence position. NCBI accession numbers of the nucleotide sequences used for the analyses are shown next to the species names. The scale bar shows the number of substitutions per site.

Phylogenetic reconstruction based on core genome sequences of Xenorhabdus bacterial strains. 1719910 nucleotide positions (1501 core genes) were used in the analysis. Numbers at the nodes represent SH-like branch supports. Bar represents 0.05 nucleotide substitutions per sequence position. Accession numbers of the genome sequences used for the reconstruction are shown in Table S3.

Comparison of morphometrics of the third-stage infective juveniles of Steinernema anantnagense n. sp. with other members of “Feltiae-Kushidai” clade. Measurements are in μm except n, ratio and percentage. Data for new species is in bold.

Species	Reference	Country	n	L	BD	EP	NR	NL	T	a	b	C	c′	D%	E%	H%
S. akhursti	Qiu et al. (2005)	China	20	770–835	33–35	55–60	83–95	115–123	68–75	23–26	6.6–7.2	10–12	3.3–3.7	45–50	73–86	49–56
S. anantnagense n. sp.	Present Study	India	20	749–834	32–42	45–62	54–71	120–143	49–66	19–24	5.5–6.7	12–16	1.6–2.1	35–48	74–113	20–36
S. africanum	Machado et al. (2022)	Rwanda	15	690–802	25–32	54–68	87–132	123–167	52–72	23–30	4.3–6.3	10–15	2.9–4.2	34–46	79–129	28–39
S. cholashanense	Nguyen et al. (2008)	China	20	727–909	26–35	59–65	72–97	110–138	60–80	24–34	6.1–7.2	10–14	3.5–5.0	46–53	76–91	33–47
S. citrae	Stokwe et al. (2011)	South Africa	20	623–849	23–28	49–64	83–108	118–137	63–81	25–34	5.1–7.1	13–14	13–17	39–58	85–132	37–50
S. feltiae	Nguyen et al. (2007)	Russia	25	766–928	22–32	58–67	108–117	130–143	81–89	27–34	5.8–6.8	9.4–11	4.5–5.1	44–50	67–81	37–51
S. hebeiense	Chen et al. (2006)	China	20	610–710	23–28	43–51	73–83	100–111	63–71	24–28	5.7–6.7	9.4–11	NA	40–50	65–80	32–50
S. ichnusae	Tarasco et al. (2008)	Italy	20	767–969	27–35	59–68	94–108	119–148	76–89	24–32	5.6–6.9	9–12	4.2–5.1	42–49	68–83	44–50
S. jollieti	Spiridonov et al. (2004)	USA	12	625–820	20–28	53–65	NA	115–135	60–73	25–34	4.9–6.4	9–12	NA	46–50	NA	46–60
S. kraussei	Nguyen et al. (2007)	Germany	25	797–1102	30–36	50–66	99–111	119–145	63–86	NA	NA	NA	NA	NA	NA	35–40
S. kushidai	Mamiya (1998)	Japan	20	424–662	22–31	42–50	70–84	106–120	44–59	19–25	4.9–5.9	10–13	NA	38–44	NA	NA
S. litorale	Yoshida (2004)	Japan	25	834–988	28–33	54–69	89–104	114–133	72–91	27–31	6.7–7.9	10–11.9	3.8–5.4	44–56	68–84	NA
S. nguyeni	Malan et al. (2016)	South Africa	20	673–796	22–28	47–58	74–86	101–121	61–73	27–33	6.2–7.4	10–12	2.8–4.8	43–57	70–86	20–31
S. oregonese	Liu and Berry (1996)	USA	20	820–1110	28–38	60–72	NA	116–148	64–78	24–37	6–8	12–16	NA	40–60	90–110	30–33
S. populi	Tian et al. (2022)	China	25	973–1172	33–41	70–86	98–113	134–159	55–72	28–33	6.8–7.5	15–20	2.4–3.3	47–61	105–140	26–44
S. puntauvense	Uribe-Lorío et al. (2007)	Costa Rica	19	631–728	31–38	20–30	46–69	81–103	51–59	17–23	7.1–7.9	11–13	NA	25–50	35–56	52–55
S. sandneri	Lis et al. (2021)	Poland	25	708–965	23–32	44–64	83–118	123–151	64–86	27–34	5.5–6.9	11–13	NA	36–45	63–86	23–40
S. sangi	Phan et al. (2001)	Vietnam	20	704–784	30–40	46–54	78–97	120–138	76–89	19–25	5.6–6.3	9–10	NA	36–44	56–70	44–52
S. silvaticum	Sturhan et al. (2005)	Germany	26	670–975	26–35	51–73	75–109	100–141	63–86	23–33	6.3–7.7	10–13	3.1–4.9	46–56	–	37–53
S. tielingense	Ma et al. (2012)	China	20	824–979	32–38	64–73	90–105	120–135	74–85	27–31	6.7–7.9	10–12	3.5–4.6	44–56	68–84	53–64
S. texanum	Nguyen et al. (2007)	USA	20	732–796	29–34	52–62	84–102	111–120	60–79	22–27	6.2–7.0	10–13	3.3–4.6	46–53	76–88	53–69
S. xinbinense	Ma et al. (2012)	China	20	635–744	28–31	46–53	75–90	109–125	65–78	21–25	5–7	8–11	3–5	40–47	65–78	30–42
S. xueshanense	Mrácek et al. (2009)	China	20	768–929	29–33	60–72	81–96	130–143	80–92	26–32	5.8–7.0	9–11	3.8–5.1	46–52	70–90	46–55
S. weiseri	Mrácek et al. (2003)	Czech Republic	20	586–828	24–29	43–65	72–92	95–119	49–68	25–33	5.7–7.2	10–14	3.2–4.1	44–55	NA	18–24

Pairwise distances in base pairs of the D2D3 fragment of 28S rRNA regions among closely related Steinernema species and Steinernema anantnagense n. sp. Data for new species is in bold.

Species (D2D3 rRNA)	S. anantnagense n. sp. OQ407498	S. akhursti KF289902	S. weiseri FJ165549	S. oregonense GU569055	S. puntauvense EF187018	S. feltiae AF331906	S. ichnusae EU421130	S. africanum OM423154	S. kushidai AF331897	S. tielingense GU994202	S. populi MZ367685	S. xueshanense FJ666053	S. kraussei KC631424	S. jollieti GU569051	S. cholashanense EF520284	S. texanum EF152569	S. xinbinense GU994204	S. citrae MF540676	S. silvaticum KC631426	S. sangi MF620997	S. nguyeni KR815816	S. monticola GU395647
S. anantnagense n. sp. OQ407498		5	12	13	13	13	15	16	17	18	18	18	19	19	23	24	25	25	27	30	31	35
S. akhursti KF289902	99		16	16	17	17	17	18	18	22	20	22	22	21	26	27	29	27	28	27	33	38
S. weiseri FJ165549	98	98		12	5	5	7	6	22	13	18	15	16	9	18	21	19	18	24	32	22	37
S. oregonense GU569055	98	98	98		11	11	13	14	24	10	27	17	8	17	13	22	14	25	19	33	29	38
S. puntauvense EF187018	98	98	99	99		0	4	5	23	10	23	16	13	10	15	20	16	17	21	30	21	40
S. feltiae AF331906	98	98	99	99	100		4	5	23	10	23	16	13	10	15	20	16	17	21	30	21	40
S. ichnusae EU421130	98	98	99	98	99	99		5	25	14	25	17	15	10	19	20	20	17	25	32	21	40
S. africanum OM423154	98	98	99	98	99	99	99		21	13	22	19	16	8	18	22	19	12	26	30	15	37
S. kushidai AF331897	98	98	97	97	97	97	97	97		26	30	26	26	29	28	36	29	30	34	32	33	42
S. tielingense GU994202	98	97	98	99	99	99	98	98	97		27	20	10	18	12	25	12	22	20	36	26	44
S. populi MZ367685	98	97	98	96	97	97	97	97	96	96		27	30	27	30	29	31	32	38	40	37	44
S. xueshanense FJ666053	98	97	98	98	98	98	98	98	97	97	96		20	22	20	25	22	30	28	41	33	45
S. kraussei KC631424	98	97	98	99	98	98	98	98	97	99	96	97		21	13	27	11	27	16	37	31	45
S. jollieti GU569051	98	97	99	98	99	99	99	99	96	98	96	97	97		23	24	22	20	31	35	24	42
S. cholashanense EF520284	97	97	98	98	98	98	98	98	96	98	96	97	98	97		27	10	27	19	38	30	45
S. texanum EF152569	97	96	97	97	97	97	97	97	95	97	96	97	96	97	96		32	32	37	39	35	45
S. xinbinense GU994204	97	96	98	98	98	98	97	98	96	98	96	97	99	97	99	96		30	17	39	34	47
S. citrae MF540676	97	96	98	97	98	98	98	98	96	97	96	96	96	97	96	96	96		37	42	11	49
S. silvaticum KC631426	96	96	97	98	97	97	97	97	95	97	95	96	98	96	98	95	98	95		41	41	50
S. sangi MF620997	96	96	96	96	96	96	96	96	96	95	95	94	95	95	95	95	95	94	95		46	46
S. nguyeni KR815816	96	96	97	96	97	97	97	98	96	97	95	96	96	97	96	95	95	99	95	94		51
S. monticola GU395647	95	95	95	95	95	95	95	95	94	94	94	94	94	94	94	94	94	93	93	94	93

Comparison of morphometrics of the first-generation females of Steinernema anantnagense n. sp. with other members of “Feltiae-Kushidai” clade. Measurements are in μm except n, ratio and percentage. Data for new species is in bold.

Species	L	BD	EP	NR	NL	T	A	B	c	c′	V	ABD	D%	Mucron
S. akhursti	5625–9000	200–270	113–138	150–175	213–258	38–63	30*	32*	149*	0.6*	48–53	68–100	NA	P
S. anantnagense n. sp.	2327–4872	314–409	92–140	143–182	196–255	32–49	17–29	13–21	61–122	0.5–0.8	50–58	77–110	43–60	P
S. africanum	2469–4215	154–194	67–111	79–130	170–201	35–55	13–27	13–24	51–104	0.7–1.0	50–57	37–70	32–62	P
S. cholashanense	3232–6363	156–332	111–148	176–223	181–231	46–70	13–23	18–32	62–119	0.6–1.0	50–57	54–105	29–65	P
S. citrae	2038–4019	137–212	54–90	130–179	189–220	33–60	NA	NA	NA	NA	50–59	43–79	27–46	P
S. feltiae	3095–3774	170–254	68–97 **	70–97 **	197–304	39–70	14–20	12–17	49–88	0.7–1.2 *	44–57	47–62	40–54 *	P
S. hebeiense	3972–4254	142–245	48–95	88–123	133–158	133–158	17–25	21–29	67–129	0.5–0.9	50–57	45–65	36–66	A
S. ichnusae	4547–6186	242–323	106–156	NA	215–262	51–79	17–24	21–26	68–113	0.6–1.0	51–57	70–94	47–63	P
S. jollieti	3746–6030	219–298	96–136	NA	184–310	31–55	15–24	19–31	72–185	NA	44–56	NA	52	P
S. kraussei	2500–5400	153–288	66–99	127–146	178–205	33–59	17	22	88	NA	54	39–50	45	P
S. kushidai	2100–4700	54–59	78–105	111–144	204–255	30–45	NA	NA	NA	NA	54–59	54–84	37–46	A
S. litorale	3930–5048	175–215	65–105	130–165	185–213	25–60	21–26	20–26	78–157	0.5–0.9	0.5–0.9	55–75	33–57	P
S. nguyeni	2290–5361	130–216	49–98	84–139	137–194	20–67	15–30	15–30	53–165	0.6–1.1	52–63	130–216	30–56	A
S. oregonense	4400–6200	217–268	217–268	129–162	186–220	28–46	NA	NA	NA	NA	46–56	42–79	43–57	A
S. populi	4038–13762	217–531	90–178	150–213	213–278	41–88	18–36	19–50	75–182	0.5–0.9	45–60	60–157	36–65	A
S. puntauvense	3687–8335	181–221	51–85	123–146	141–206	41–66	NA	NA	NA	NA	51–55	57–102	25–45	P
S. sandneri	4244–5014	181–261	61–102	132–158	173–194	32–61	17–25	24–27	75–140	NA	49–57	62–122	36–54	P
S. sangi	4830–7200	270–360	80–121	140–170	216–240	36–62	NA	NA	NA	NA	43–530	84–140	35–51	P
S. silvaticum	1520–3290	50–175	50–175	50–175	121–188	33–79	15–41	10–18	34–80	1.0–1.8	44–57	26–53	33–79	A
S. texanum	2720–3623	130–202	78–107	111–135	160–189	30–52	NA	NA	NA	NA	50–55	50–71	NA	A
S. tielingense	4028–8538	200–307	82–103	111–144	186–263	40–69	17–32	21–45	72–158	0.5–0.9	49–54	56–92	32–49	A
S. xinbinense	3025–5121	159–200	70–87	106–141	167–192	30–53	19–25	17–26	79–123	0.5–0.8	46–57	50–67	38–45	P
S. xueshanense	4181–8181	182–343	117–148	NA	196–274	43–66	NA	NA	NA	NA	52–62	38–72	NA	P
S. weiseri	3780–5940	202–263	75–86	108–154	162–226	38–59	17–29	22–31	87–156	0.5–0.8	50–58	51–80	NA	P

Morphometrics of the IJs and adult generations of Steinernema anantnagense n. sp. (Steiner.7). All characters are in µm (except n, ratios and percentages) and given as mean ± s.d. (range).

Characters		First Generation		Second Generation		Infective Juveniles (paratypes)

	Male (holotype)	Male (paratypes)	Female (paratypes)	Male (paratypes)	Female (paratypes)
n	1	20	20	20	20	20
Body length (L)	1279	1618 ± 246 (1223–1899)	3765 ± 441 (2327–4872)	1068 ± 76 (899–1168)	2081 ± 200 (1761–2437)	789 ± 35 (749–834)
a (L/BD)	6.6	8.3 ± 1.1 (6.4–9.8)	23 ± 1.2 (17–29)	19 ± 2.6 (15.1–25.3)	14.7 ± 1.5 (12.1–17.4)	22 ± 1.9 (19–24)
b (L/NL)	7.0	9.2 ± 1.4 (7.0–11.3)	17.0 ± 2.4 (13–21)	7.1 ± 0.6 (5.9–8.2)	11.0 ± 1.0 (9.6–13.5)	6.0 ± 0.4 (5.5–6.7)
c (L/T)	35	49 ± 9.0 (34–64)	100 ± 18 (61–122)	43 ± 5.8 (31–52)	54 ± 6.6 (43–67)	13.8 ± 1.8 (12.2–16.4)
c’ (T/ABW)	1.2	1.1 ± 0.2 (0.8–1.6)	0.7 ± 0.1 (0.5–0.8)	0.9 ± 0.2 (0.6–1.4)	1.0 ± 0.1 (0.8–1.3)	1.8 ± 0.2 (1.6–2.1)
V (VA/L×100)		–	53 ± 1.7 (50–58)	–	53 ± 1.7 (50–57)	–
Max. body diam. (MBD)	193	194 ±11.3 (167–211)	434 ± 30 (314–409)	57 ± 7.3 (42–63)	143 ± 16 (123–173)	37 ± 3.6 (32–42)
Lip region width	10.2	10.1 ± 1.9 (8.2–12.2)	14.4 ± 1.9 (11.1–18.3)	8.4 ± 1.6 (6.1–12.1)	11.4 ± 1.7 (8.9–14.6)	4.6 ± 2.0 (3.7–5.8)
Stoma length	12.3	13.8 ± 1.6 (10.1–16.9)	16.4 ± 2.4 (12.4–22.1)	11.7 ± 1.1 (10.1–14.1)	14.1 ± 2.3 (11.1–18.1)	10.7 ± 2.0 (8.6–13.8)
Procorpus length	66	57 ± 4.1 (51–66)	65 ± 6.0 (55–76)	47 ± 3.9 (41–53)	54 ± 3.8 (49–65)	43 ± 5.4 (37–52)
Metacorpus length	33	34 ± 1.7 (30–36)	44 ± 4.6 (38–50)	30 ± 2.4 (25–35)	37 ± 2.4 (33–43)	26 ± 2.7 (23–29)
Isthmus length	40	42 ± 4.0 (37–50)	53 ± 6.4 (45–68)	37 ± 1.5 (35–40)	49 ± 3.7 (43–55)	36 ± 2.8 (31–39)
Bulb length (BL)	31	30 ± 2.2 (25–35)	43 ± 4.5 (33–50)	25 ± 2.7 (21–30)	35 ± 2.9 (30–39)	17 ± 1.8 (16–20)
Bulb width (EBW)	26	25 ± 1.6 (21–27)	35 ± 3.5 (27–41)	21 ± 1.9 (18–25)	28 ± 2.7 (25–33)	11 ± 1.7 (9.1–13.9)
Pharynx length (PL)	170	162 ± 5.9 (150–172)	206 ± 14 (182–232)	139 ± 5.0 (129–146)	175 ± 6.4 (164–191)	121 ± 8.0 (109–133)
Nerve ring – ant. end (NR)	109	112 ± 6.8 (103–129)	165 ± 12.1 (143–182)	91 ± 7.0 (75–104)	109 ± 11.9 (88–127)	63 ± 8.4 (54–71)
Excretory pore – ant. end (EP)	114	111 ± 10 (88–124)	112 ± 11.9 (92–140)	80 ± 9.2 (62–98)	92 ± 8.1 (82–112)	55 ± 6.7 (45–62)
Width at excretory pore (WEP)	58	61 ± 3.4 (49–65)	137 ± 11.3 (123–170)	35 ± 3.6 (30–42)	60 ± 6.7 (51–74)	22 ± 2.4 (19–25)
Neck length (stoma+pharynx, NL)	182	176 ± 6.4 (165–185)	223 ± 14 (196–255)	151 ± 5.0 (142–159)	189 ± 6.5 (177–204)	132 ± 8.3 (120–143)
Body width at neck base	79	86 ± 5.8 (76–96)	217 ± 23 (195–264)	41 ± 3.9 (36–50)	95 ± 8.5 (82–107)	29 ± 2.9 (24–32)
Testis reflexion	630	631 ± 37 (576–689)	–	522 ± 32 (481–568)	–	–
Vagina length	–	–	31 ± 3.1 (24–36)	–	24 ± 3.3 (19–30)	–
Body width at vulva	–	–	386 ± 53 (309–498)	–	164 ± 25 (124–193)	–
Vulva – ant. end (VA)	–	–	1989 ± 214 (1247–2252)	–	1105 ± 101 (961–1295)	–
Vulva – post. end (VP)	–	–	1776 ± 242 (1080–2038)	–	976 ± 110 (801–1161)	–
Rectum length	–	–	35 ± 4.0 (29–44)	–	21 ± 2.5 (17–24)	13.8 ± 1.9 (11–16)
Anal body diam. (ABD)	31	32 ± 3.9 (25–36)	86 ± 7.1 (77–110)	29 ± 4.1 (21–35)	41 ± 6.8 (31–53)	20 ± 1.7 (18–22)
Tail length (T)	37	34 ± 2.4 (29–39)	38 ± 4.2 (32–49)	26 ± 3.9 (21–35)	49 ± 3.9 (38–53)	58 ± 6.7 (49–66)
Hyaline part of tail (H)	–	–	–	–	–	16.3 ± 4.1 (11.8–23.7)
Spicule length (SL)	66	64 ± 4.6 (56–70)	–	44 ± 2.6 (40–49)	–	–
Gubernaculum length (GL)	34	36 ± 3.8 (31–43)	–	25 ± 2.3 (21–29)	–	–
Stoma length/lip region width	1.2	1.4 ± 0.2 (0.9–1.8)	1.2 ± 0.2 (0.9–1.8)	1.4 ± 0.3 (0.9–1.9)	1.3 ± 0.3 (0.9–1.8)	2.4 ± 0.5 (1.6–3.0)
Nerve ring % (NR/NL×100)	60	64 ± 5.3 (56–78)	74 ± 7.1 (64–86)	61 ± 4.6 (52–68)	58 ± 6.6 (46–67)	48 ± 7.8 (38–58)
Excretory pore % (EP/NL×100)	62	63 ± 6.1 (50–74)	50 ± 4.6 (42–60)	53 ± 6.6 (41–68)	49 ± 4.1 (43–58)	42 ± 4.4 (35–48)
Rectum% (R/ABD×100)	–	–	0.4 ± 0.1 (0.3–0.6)		0.5 ± 0.1 (0.3–0.8)	0.7 ± 0.1 (0.6–0.9)
D% (EP /NL×100)	62	63 ± 6.1 (49–74)	50 ± 4.9 (43–60)	53 ± 6.6 (41–68)	49 ± 4.1 (43–58)	42 ± 4.4 (35–48)
E% (EP/T×100)	311	333 ± 43 (256–403)	5.7 ± 1.2 (4.5–9.5)	319 ± 59 (219–460)	240 ± 36 (201–362)	96 ± 12.8 (74–113)
SW% (SL/ABD×100)	217	208 ± 39 (154–297)	–	2.3 ± 0.4 (1.8–3.4)
GS% (GL/SL×100)	51	57 ± 6.7 (46–70)	–	0.6 ± 0.1 (0.4–0.7)
H% (H/T ×100)	–	–	–	–	–	28 ± 5.8 (20–36)

Comparison of morphometrics of the first-generation males of Steinernema anantnagense n. sp. with other members of “Feltiae-Kushidai” clade. Measurements are in μm except n, ratio and percentage. Data for new species is in bold.

Species	n	L	BD	EP	NR	NL	T	SL	GL	a	B	c	c′	D%	SW%	GS%	Mucron
S. akhursti	20	1350–1925	115–150	93–113	120–163	168–205	30–40	85–100	58–68	NA	NA	NA	NA	52–61	140–200	65–77	P
S. anantnagense n. sp.	20	1223–1899	167–211	88–124	103–129	165–185	29–39	56–70	31–43	6–10	7–11	34–64	0.8–1.6	49–74	154–297	46–70	P
S. africanum	15	977–1400	65–131	69–109	79–104	132–147	34–46	65–76	32–49	9–12	7–12	25–34	0.9–1.1	52–74	144–197	49–68	P
S. cholashanense	20	1070–1778	73–204	75–135	91–126	135–173	29–43	60–71	32–45	9–24	8–11	36–51	0.6–0.9	50–85	92–144	61–85	P
S. citrae	20	1028–1402	87–113	64–92	92–119	123–155	17–31	57–80	32–59	NA	NA	NA	NA	47–67	156–233	48–89	P
S. feltiae	25	1414–1817	60–90	110–126	NA	164–180	37–43	65–77	34–47	NA	NA	NA	NA	51–64	99–130	52–61	P
S. hebeiense	20	1036–1450	74–98	58–73	78–93	118–132	24–35	51–63	38–50	12–17	8–11	30–49	0.6–0.9	48–59	120–170	60–90	A
S. ichnusae	20	1151–1494	73–204	94–108	NA	135–173	33–48	64–67	43–46	20–29	7–9	29–39	0.8–0.9	59–65	120–162	64–69	A
S. jollieti	12	1296–1952	98–135	83–110	NA	110–168	24–38	55–70	45–60	12–19	8–14	53–86	NA	53–83	NA	NA	A
S. kraussei	20	1200–1600	110–144	73–99	95–122	137–178	36–44	42–53	29–37	11	9 NA	NA	NA	NA	NA	NA	P
S. kushidai	20	1200–1900	75–156	71–105	120–137	156–189	30–40	48–72	39–60	NA	NA	NA	NA	42–59	NA	NA	A
S. litorale	25	1230–1514	82–111	77–107	94–128	133–163	26–41	67–89	44–64	12–16	8–10	33–56	0.6–0.9	34–56	154–200	62–81	P
S. nguyeni	20	818–1171	58–106	47–71	70–103	112–144	18–25	58–75	30–55	11–15	7–10	38–53	0.6–0.8	38–57	185–279	46–81	P
S. oregonense	20	1560–1820	105–161	95–139	101–133	139–182	24–32	65–73	52–59	NA	NA	NA	0.6 NA	64–75	NA	NA	A
S. populi	25	1258–1514	66–95	95–121	107–143	131–177	39–68	57–77	38–60	15–20	8–10	20–33	0.8–1.5	59–78	107–160	58–82	P/A
S. puntauvense	19	1010–1931	101–139	68–114	104–128	130–159	28–40	71–81	30–40	NA	NA	NA	NA	45–85	140–200	55–75	P
S. sandneri	25	1206–1635	124–178	64–92	112–138	148–170	35–46	53–65	39–50	9–11	8–10	31–42	NA	42–59	97–127	61–83	P
S. sangi	20	1440–2325	120–225	67–99	109–166	150–221	27–42	58–80	34–46	NA	NA	NA	NA	42–63	120–160	50–70	P
S. silvaticum	26	975–1270	52–78	71–92	90–126	116–168	20–47	42–64	30–43	14–20	8–9	24–55	0.8–1.4	45–63	NA	NA	P
S. tielingense	20	1430–2064	111–159	94–133	96–132	145–173	22–33	79–98	49–70	11–18	9–13	57–85	0.3–0.6	64–78	176–212	59–82	A
S. texanum	20	1197–1406	81–116	79–100	94–114	123–147	19–30	55–66	39–53	NA	NA	NA	NA	58–73	127–203	62–84	A
S. xinbinense	20	1133–1440	90–126	57–75	91–120	138–159	30–41	49–62	30–41	11–13	7–9	31–39	0.7–1.0	41–50	114–156	54–72	P
S. xueshanense	20	1313–2040	97–159	113–137	NA	151–175	29–48	66–91	41–60	NA	NA	NA	NA	73–87	93–172	58–95	A
S. weiseri	20	990–1395	84–138	57–84	94–115	134–154	19–32	62–72	46–57	9–12	7–10	36–64	0.6–0.9	39–60	150–240	70–85	A

Pairwise distances in base pairs of the ITS rRNA regions among closely related Steinernema species and Steinernema anantnagense n. sp. Data for new species is in bold.

Species (ITS rRNA)	S. anantnagense n. sp. OQ407490	S. akhursti DQ375757	S. kushidaiAB243440	S. cholashanense EF431959	S. oregonense AY230180	S. sangi AY355441	S. texanum EF152568	S. xueshanense FJ666052	S. populi MZ367621	S. jollieti AY171265	S. xinbinense JN171593	S. weiseri KJ696685	S. tielingense GU994201	S. africanum ON041031	S. kraussei AY230175	S. citrae EU754718	S. silvaticum AY230162	S. litorale AB243441	S. ichnusae EU421129	S. nguyeni KP325084	S. feltiae AY230169	S. hebeiense DQ105794	S. monticola AB698756
S. anantnagense n. sp. OQ407490		19	53	72	72	72	72	73	74	74	80	83	85	86	86	87	87	90	91	92	101	116	117
S. akhursti DQ375757	97		51	75	74	73	77	76	65	75	82	86	89	89	90	87	91	93	95	94	102	119	115
S. kushidai AB243440	92	93		95	89	96	103	90	94	89	97	104	101	103	108	110	104	107	110	110	122	128	130
S. cholashanense EF431959	89	88	85		61	74	58	58	106	69	68	72	71	72	83	70	82	73	74	84	75	112	116
S. oregonense AY230180	89	88	86	91		47	51	33	99	19	31	43	46	39	52	42	45	56	49	52	51	94	115
S. sangi AY355441	89	88	85	88	93		57	58	98	57	54	43	45	60	53	60	61	48	48	54	47	88	103
S. texanum EF152568	89	88	83	91	92	91		53	95	61	60	66	67	59	75	55	69	74	69	77	68	110	118
S. xueshanense FJ666052	89	88	86	91	95	91	92		99	41	43	46	54	47	60	44	54	57	53	60	54	86	114
S. populi MZ367621	89	90	86	83	84	84	85	84		105	103	110	111	112	109	113	115	117	116	112	124	145	123
S. jollieti AY171265	88	88	86	89	97	91	91	94	83		48	52	54	55	64	55	58	63	56	62	58	95	115
S. xinbinense JN171593	88	87	85	90	96	92	91	94	84	93		45	50	32	58	40	38	58	57	57	50	92	116
S. weiseri KJ696685	87	86	83	89	94	94	90	93	82	92	93		17	59	32	60	65	26	27	34	27	80	114
S. tielingense GU994201	87	86	84	89	93	93	90	92	82	92	93	98		63	46	65	66	30	28	47	32	88	117
S. africanum ON041031	87	86	84	89	94	91	91	93	82	92	95	91	91		64	45	35	72	68	64	62	98	124
S. kraussei AY230175	86	86	82	87	92	92	88	91	82	90	91	95	93	90		65	69	44	46	19	40	89	120
S. citrae EU754718	86	86	82	89	94	91	92	94	82	92	94	91	90	93	90		55	73	70	64	60	98	118
S. silvaticum AY230162	86	86	83	87	93	91	89	92	81	91	95	90	90	95	90	92		76	70	68	68	101	126
S. litorale AB243441	86	85	82	89	92	93	88	91	80	90	91	96	96	89	93	89	88		38	45	34	79	118
S. ichnusae EU421129	86	85	82	88	93	93	89	92	81	92	92	96	96	90	93	89	89	94		45	25	87	122
S. nguyeni KP325084	85	85	82	87	92	92	88	91	81	90	91	95	93	90	97	90	90	93	93		41	86	117
S. feltiae AY230169	84	84	80	88	92	93	89	92	79	91	92	96	95	91	94	91	90	95	96	94		79	119
S. hebeiense DQ105794	81	80	78	82	85	86	82	86	75	85	86	88	86	84	86	85	84	88	86	87	88		135
S. monticola AB698756	78	79	76	78	79	81	78	79	77	79	79	79	78	77	78	78	77	78	77	78	78	74

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Multigene Sequence-Based and Phenotypic Characterization Reveals the Occurrence of a Novel Entomopathogenic Nematode Species, Steinernema anantnagense n. sp.

Article Category: Research Paper

Published Online: Jul 13, 2023

Page range: -

Received: Apr 06, 2023

DOI: https://doi.org/10.2478/jofnem-2023-0029

Keywords
Species description, nematodes, molecular characterization, phylogenetic systematics, taxonomy, biological control, sustainable agriculture

© 2023 Aashaq Hussain Bhat et al., published by Sciendo

This work is licensed under the Creative Commons Attribution 4.0 International License.

Figure 1:

Figure 2:

Figure 3:

Figure 4:

Figure 5:

Figure 6:

Figure 7:

Figure 8:

Comparison of morphometrics of the third-stage infective juveniles of Steinernema anantnagense n. sp. with other members of “Feltiae-Kushidai” clade. Measurements are in μm except n, ratio and percentage. Data for new species is in bold.

Pairwise distances in base pairs of the D2D3 fragment of 28S rRNA regions among closely related Steinernema species and Steinernema anantnagense n. sp. Data for new species is in bold.

Comparison of morphometrics of the first-generation females of Steinernema anantnagense n. sp. with other members of “Feltiae-Kushidai” clade. Measurements are in μm except n, ratio and percentage. Data for new species is in bold.

Morphometrics of the IJs and adult generations of Steinernema anantnagense n. sp. (Steiner.7). All characters are in µm (except n, ratios and percentages) and given as mean ± s.d. (range).

Comparison of morphometrics of the first-generation males of Steinernema anantnagense n. sp. with other members of “Feltiae-Kushidai” clade. Measurements are in μm except n, ratio and percentage. Data for new species is in bold.

Pairwise distances in base pairs of the ITS rRNA regions among closely related Steinernema species and Steinernema anantnagense n. sp. Data for new species is in bold.

Multigene Sequence-Based and Phenotypic Characterization Reveals the Occurrence of a Novel Entomopathogenic Nematode Species, Steinernema anantnagense n. sp.

Article Category: Research Paper

Published Online: Jul 13, 2023

Page range: -

Received: Apr 06, 2023

DOI: https://doi.org/10.2478/jofnem-2023-0029

KeywordsSpecies description, nematodes, molecular characterization, phylogenetic systematics, taxonomy, biological control, sustainable agriculture

© 2023 Aashaq Hussain Bhat et al., published by Sciendo

This work is licensed under the Creative Commons Attribution 4.0 International License.

Figure 1:

Figure 2:

Figure 3:

Figure 4:

Figure 5:

Figure 6:

Figure 7:

Figure 8:

Comparison of morphometrics of the third-stage infective juveniles of Steinernema anantnagense n. sp. with other members of “Feltiae-Kushidai” clade. Measurements are in μm except n, ratio and percentage. Data for new species is in bold.

Pairwise distances in base pairs of the D2D3 fragment of 28S rRNA regions among closely related Steinernema species and Steinernema anantnagense n. sp. Data for new species is in bold.

Comparison of morphometrics of the first-generation females of Steinernema anantnagense n. sp. with other members of “Feltiae-Kushidai” clade. Measurements are in μm except n, ratio and percentage. Data for new species is in bold.

Morphometrics of the IJs and adult generations of Steinernema anantnagense n. sp. (Steiner.7). All characters are in µm (except n, ratios and percentages) and given as mean ± s.d. (range).

Comparison of morphometrics of the first-generation males of Steinernema anantnagense n. sp. with other members of “Feltiae-Kushidai” clade. Measurements are in μm except n, ratio and percentage. Data for new species is in bold.

Pairwise distances in base pairs of the ITS rRNA regions among closely related Steinernema species and Steinernema anantnagense n. sp. Data for new species is in bold.

Keywords
Species description, nematodes, molecular characterization, phylogenetic systematics, taxonomy, biological control, sustainable agriculture