Genetic intraspecific diversity of Meloidogyne javanica parasitizing vegetables in southern Iran

Number	Sampling area	N	E	Sample	Host plant	Molecular code
1	Boshehr province-Abdan	28°11´00.38˝	51°74´07.94˝	Soil and root	Tomato	MJ1
2	Boshehr province-Abdan	28°10´99.86˝	51°74´08.46˝	Soil and root	Tomato	MJ2
3	Boshehr province-Abdan	28°11´00.58˝	51°74´07.97˝	Soil and root	Tomato	MJ3
4	Boshehr province-Abdan	28°11´00.29˝	51°74´08.27˝	Soil and root	Tomato	MJ4
5	Boshehr province-Abdan	28°11´00.31˝	51°74´08.28˝	Soil and root	Tomato	MJ5
6	Boshehr province-Abdan	28°10´95.09˝	51°74´04.66˝	Soil and root	Tomato	MJ6
7	Boshehr province-Abdan	28°10´94.93˝	51°74´04.44˝	Soil and root	Tomato	MJ7
8	Boshehr province-Abdan	28°10´94.82˝	51°74´04.43˝	Soil and root	Tomato	MJ8
9	Boshehr province-Abdan	28°10´94.56˝	51°74´03.85˝	Soil and root	Tomato	MJ9
10	Boshehr province-Abdan	28°10´94.47˝	51°74´03.81˝	Soil and root	Tomato	MJ10
11	Boshehr province-Deir Port	27°91´15.43˝	51°98´02.27˝	Soil and root	Tomato	MJ11
12	Boshehr province-Deir Port	27°91´15.16˝	51°98´02.71˝	Soil and root	Tomato	MJ12
13	Boshehr province-Deir Port	27°91´17.39˝	51°98´03.58˝	Soil and root	Tomato	MJ13
14	Boshehr province-Deir Port	27°91´17.33˝	51°98´03.47˝	Soil and root	Tomato	MJ14
15	Boshehr province-Deir Port	27°91´44.84˝	51°98´43.36˝	Soil and root	Tomato	MJ15
16	Boshehr province-Bardkhon	28°07´54.24˝	51°48´89.26˝	Soil and root	Tomato	MJ16
17	Boshehr province-Bardkhon	28°07´51.78˝	51°48´90.21˝	Soil and root	Tomato	MJ17
18	Boshehr province-Bardkhon	28°36´13.93˝	51°48´90.22˝	Soil and root	Tomato	MJ18
19	Boshehr province-Bardkhon	28°07´51.25˝	51°48´84.44˝	Soil and root	Tomato	MJ19
20	Boshehr province-Bardkhon	28°07´51.92˝	51°48´82.48˝	Soil and root	Tomato	MJ20
21	Boshehr province-Bardkhon	28°07´52.13˝	51°48´82.42˝	Soil and root	Tomato	MJ21
22	Boshehr province-Bardkhon	28°07´52.15˝	51°48´82.19˝	Soil and root	Tomato	MJ22
23	Kerman province-Jiroft	28°41´09.00˝	57°40´13.81˝	Root	Mallow	MJ23
24	Kerman province-Jiroft	28°41´35.08˝	57°39´25.18˝	Root	Goosefoot	MJ24
25	Kerman province-Jiroft	28°42´22.57˝	57°43´05.86˝	Root	Eggplant	MJ25
26	Fars province-Beyza	29°55´30.53˝	52°26´28.41˝	Soil and root	Cucumber	MJ26
27	Fars province-Beyza	29°55´51.85˝	52°26´44.88˝	Soil and root	Cucumber	MJ27
28	Fars province-Beyza	29°55´59.49˝	52°26´51.00˝	Soil and root	Cucumber	MJ28
29	Fars province-Beyza	29°55´16.00˝	52°27´20.15˝	Soil and root	Cucumber	MJ29
30	Fars province-Beyza	29°55´11.72˝	52°27´27.93˝	Soil and root	Cucumber	MJ30
31	Fars province-khafr	28°57´44.57˝	53°11´58.46˝	Soil and root	Okra	MJ31
32	Fars province-khafr	28°57´35.80˝	53°12´32.31˝	Soil and root	Okra	MJ32
33	Fars province-khafr	28°57´34.97˝	53°12´28.38˝	Soil and root	Okra	MJ33
34	Fars province-khafr	28°58´56.08˝	53°11´47.87˝	Soil and root	Okra	MJ34
35	Fars province-Kazeron	29°33´23.23˝	51°45´56.27˝	Soil and root	Tomato	MJ35
36	Fars province-Kazeron	29°33´25.22˝	51°45´42.76˝	Soil and root	Tomato	MJ36
37	Fars province-Kazeron	29°33´42.68˝	51°45´08.64˝	Soil and root	Tomato	MJ37
38	Fars province-Fasa	29°00´46.56˝	53°37´42.27˝	Soil and root	Cucumber	MI1
39	Fars province-Fasa	29°00´46.56˝	53°37´42.27˝	Soil and root	Cucumber	MI2
40	Fars province-Fasa	29°00´46.56˝	53°37´42.27˝	Soil and root	Cucumber	MI3
41	Fars province-Fasa	29°00´46.56˝	53°37´42.27˝	Soil and root	Cucumber	MI4

1 Notes: MJ: M. javanica; MI: M. incognita.

Table 2.

All primers used for ISSR, RAPD and species-specific identification of Meloidogyne species during the present study.

ISSR		RAPD
(CCA)₅	5´-CCACCACCACCACCA-3'	OPA8	5’-GTGACGTAGG-3'
(GTG)₆	5'-GTGGTGGTGGTGGTGGTG-3'	OPA10	5'-GTGATCGCAG-3'
(ATG)₆	5'-ATGATGATGATGATGATG-3'	OPA13	5'-CAGCACCCAC-3'
(GAG)₄GC	5'-GAGGAGGAGGAGGC-3'	OPAD10	5'-AAGAGGCCAG-3'
(GACA)₄	5'-GACAGACAGACAGACA-3'	OPE18	5'-GGACTGCAGA-3'
(GA)₈C	5'-GAGAGAGAGAGAGAGAC-3'	OPP17	5'-TGACCCGCCT-3'
(AC)₈T	5'-ACACACACACACACACT-3'	OPB11	5'-GTAGACCCGT-3'
(CA)₈G	5'-CACACACACACACACAG-3'	OPB17	5'-AGGGAACGAG-3'
(CTC)₄GC	5'-CTCCTCCTCCTCGC-3'	OPD8	5'-GTGTGCCCCA-3'
(GAGA)₄GG	5'-GAGAGAGAGAGAGAGAGG-3'	OPC8	5'-TGGACCGGTG-3'
Species-specific primers
M. javanica	Fjav (5'-GGTGCGCGATTGAACTGAGC-3')	M. incognita	Finc (5'-CTCTGCCCAATGAGCTGTCC-3')
	Rjav (5'-CAGGCCCTTCAGTGGAACTATAC-3')		Rinc (5'-CTCTGCCCTCACATTAAG-3')
M. arenaria	Far (5'-TCGGCGATAGAGGTAAATGAC-3')	M. hapla	F (5'- TGACGGCGGTGAGTGCGA-3')
	Rar (5'-TCGGCGATAGACACTACAAACT-3')		R (5'-TGACGGCGGTACCTCATAG-3')

Table 3.

Average of the morphometrics of females, J2s and males of five populations of Meloidogyne javanica parasitizing vegetables in southern Iran.

	Females		Males		Second-stage juveniles (J2s)^†
Diagnostic characters	15	CV	10	CV	MJ1	MJ4	MJ8	MJ14	MJ15	50	CV
L (including neck)	686±127 (506-1,010)	18	1482±155 (1189-1,694)	10	463	457	449	482	469	464±19 (429-497)	4
Body width (W)	420±99 (274-568)	23	33±2.8 (29-38)	9	14	14	15	15	16	15±1.5 (12-18)	9
Neck length	259±87 (153-540)	33	−	−	−	−	−	−	−	−	−
Stylet	15±2 (13-18)	14	18±1.2 (16-19)	6	9.7	10.4	10.5	10.1	10.2	10±0.5 (9-12)	5
DGO	3.1±0.6 (2.0-3.9)	20	3.3±0.2 (3.1-3.5)	5	2.7	2.1	2.2	1.9	2.0	2.2±0.5 (1.5-3.7)	22
Excretory pore	190±30 (145-270)	16	−	−	−	−	−	−	−	−	−
Vulval slit length	21±3 (15-27)	15	−	−	−	−	−	−	−	−	−
Vulva-anus distance	16±4 (12-28)	25	−	−	−	−	−	−	−	−	−
Inter-phasmidial distance	78 ± 16 (58-108)	20	−	−	−	−	−	−	−	−	−
Tail length	−	−	14±2.1 (10-17)	15	51	55	55	58	57	55±5 (40-66)	9
Hyaline length	−	−	−	−	11	12	11	16	16	13±4 (7-20)	27
Pharynx (glands end)	−	−	473±50 (390-552)	11	164	202	197	201	201	193±23 (90-216)	12
Body length to width (a)	1.7±0.4 (1.2-2.6)	22	45±4.5 (37-51)	10	38	32	29	33	30	31±3 (25-39)	10
b	−	−	6.6±1.3 (4.8-8.1)	20	6.4	4.9	4.7	5.0	5.2	5.2±1.4 (4.3-12.4)	27
b'	−	−	3.1±0.6 (2.4-3.9)	18	3.0	2.3	2.3	2.4	2.3	2.5±0.5 (2.0-5.2)	22
c	−	−	113±26 (87-169)	23	9.4	8.4	8.2	8.3	8.2	8.5±0.8 (7.2-11.3)	10
c'	−	−	0.5±0.1 (0.3-1.0)	19	4.6	5.3	5.5	5.9	5.2	5.7±0.5 (4.7-7.2)	10
Spicules	−	−	23±5.8 (15-29)	24	−	−	−	−	−	−	−
Gubernaculum	−	−	13±2.8 (8-17)	20	−	−	−	−	−	−	−

1 Notes: Measurements are in µm. Data are the average of the measurements. ^†10 specimens were measured from each population.

Species-specific 670 bp band in the populations of *Meloidogyne javanica* recovered from southern Iran. (For population codes, see Table 1).

Species-specific 1200 bp band in the populations of *Meloidogyne incognita* recovered from southern Iran. (For population codes, see Table 1).

Table 4.

Genetic diversity estimates of Meloidogyne javanica populations recovered from southern Iran by RAPD and ISSR analyses.

	ISSR			RAPD
Population	%P^†	H	I	%P	H	I
Abdan (10)^††	25.61	0.110	0.158	36.11	0.150	0.218
Deir (5)	20.73	0.092	0.130	30.00	0.132	0.189
Bardkhon (7)	24.39	0.097	0.141	30.56	0.118	0.173
Jiroft (3)	10.37	0.039	0.058	17.78	0.079	0.113
Beiza (5)	21.34	0.088	0.128	22.78	0.095	0.137
Khafr (4)	18.29	0.076	0.110	16.11	0.064	0.093
Kazeron (3)	12.80	0.057	0.081	18.89	0.079	0.114
Mean	19.08	0.080	0.115	24.60	0.103	0.148

1 Notes: ^†P(%): percentage of polymorphic loci; H: the average expected heterozygosity; I: Shannon’s information index; ^††the data between parentheses are the number of populations collected from the given region.

RAPD (left) and ISSR (right) patterns for 19 populations of *Meloidogyne javanica* generated with the primer (CCA)₅ and OPAD10. DNA Ladder = 200 bp. (For population codes, see Table 1).

Table 5.

Nei’s (1978) unbiased genetic distance between pairs of Meloidogyne javanica populations detected by RAPD and ISSR analyses.

Population^†	Abdan	Deir	Bardkhon	Jiroft	Beiza	Khafr	Kazeron
Abdan	–	0.032	0.070	0.074	0.091	0.084	0.088
Deir	0.082	–	0.054	0.076	0.068	0.088	0.091
Bardkhon	0.090	0.076	–	0.046	0.045	0.039	0.059
Jiroft	0.069	0.127	0.119	–	0.071	0.040	0.025
Beiza	0.043	0.080	0.119	0.108	–	0.073	0.071
Khafr	0.080	0.055	0.143	0.134	0.075	–	0.051
Kazeron	0.120	0.071	0.050	0.156	0.132	0.162	–

1 Note: ^†Nei’s genetic distance matrices detected by RAPD (above diagonal) and ISSR (below diagonal).

UPGMA cluster diagram based on Dice coefficient estimated from 180 RAPD markers for 37 populations of *Meloidogyne javanica* (For population codes, see Table 1).

UPGMA cluster diagram based on Dice coefficient estimated from 164 ISSR markers for 37 populations of the *Meloidogyne javanica* (For population codes, see Table 1).

Table 6.

Analysis of molecular variance (AMOVA) of Meloidogyne javanica populations based on RAPD and ISSR data.

Marker	Source of variation	Degree of freedom	Sum of squares	Variance components	Percentage of variation	Fixation index (F _ST)^†	Number of migrants (Nm)
	Between populations	6	148.88	2.63*	19
RAPD	Within populations	30	340.85	11.36*	81	0.188	1.079
	Total	36	489.73	13.99
	Between populations	6	90.68	1.34*	14
ISSR	Within populations	30	247.80	8.26*	86	0.140	1.535
	Total	36	338.49	9.60

1 Note: ^* p < 0.01, ^† p < 0.001

A Mantel test showing correlation between RAPD and ISSR datasets of the populations of *Meloidogyne javanica* recovered from southern Iran.

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DOI: https://doi.org/10.21307/jofnem-2020-035 | Journal eISSN: 2640-396X | Journal ISSN: 0022-300X

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Language: English

Page range: 1 - 13

Submitted on: Oct 30, 2019

Published on: Apr 23, 2020

Published by: Society of Nematologists, Inc.

In partnership with: Paradigm Publishing Services

Publication frequency: 1 issue per year

Keywords:

Diversity,

ISSR,

RAPD,

root-knot nematodes,

species-specific primer

Related subjects:

Life sciences,

Life sciences, other

© 2020 Reza Ghaderi, Ali Asghar Dehghan, Abbas Mokaram Hesar, Akbar Karegar, published by Society of Nematologists, Inc.
This work is licensed under the Creative Commons Attribution 4.0 License.

Volume 52 (2020): Issue 1 (January 2020)

Genetic intraspecific diversity of Meloidogyne javanica parasitizing vegetables in southern Iran

Figures & Tables

Table 1.

Table 2.

Table 3.

Figure 1:

Figure 2:

Table 4.

Figure 3:

Table 5.

Figure 4:

Figure 5:

Table 6.

Figure 6:

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