Diagnostic methods for identification of root-knot nematodes species from Brazil

Cunha, Tiago Garcia da; Visôtto, Liliane Evangelista; Lopes, Everaldo Antônio; Oliveira, Claúdio Marcelo Gonçalves; God, Pedro Ivo Vieira Good

doi:10.1590/0103-8478cr20170449

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CROP PROTECTION • Cienc. Rural 48 (02) • 2018 • https://doi.org/10.1590/0103-8478cr20170449 copy

Diagnostic methods for identification of root-knot nematodes species from Brazil

Métodos diagnósticos usados na identificação de espécies do nematoide das galhas do Brasil

Authorship SCIMAGO INSTITUTIONS RANKINGS

ABSTRACT:

The accurate identification of root-knot nematode (RKN) species (Meloidogyne spp.) is essential for implementing management strategies. Methods based on the morphology of adults, isozymes phenotypes and DNA analysis can be used for the diagnosis of RKN. Traditionally, RKN species are identified by the analysis of the perineal patterns and esterase phenotypes. For both procedures, mature females are required. Over the last few decades, accurate and rapid molecular techniques have been validated for RKN diagnosis, including eggs, juveniles and adults as DNA sources. Here, we emphasized the methods used for diagnosis of RKN, including emerging molecular techniques, focusing on the major species reported in Brazil.

Key words:
DNA; esterase; Meloidogyne; molecular biology; morphological pattern

Authorship

Tiago Garcia da Cunha

Instituto de Ciências Agrárias, Universidade Federal de Viçosa (UFV), Campus Rio Paranaíba, Rio Paranaíba, MG, Brasil.Universidade Federal de Viçosa (UFV)BrazilRio Paranaíba, MG, BrazilInstituto de Ciências Agrárias, Universidade Federal de Viçosa (UFV), Campus Rio Paranaíba, Rio Paranaíba, MG, Brasil.

Liliane Evangelista Visôtto ^*E-mail: lvisotto@yahoo.com.br. *Corresponding author

Instituto de Ciências Biológicas e da Saúde, Universidade Federal de Viçosa (UFV), Campus Rio Paranaíba, 38810-000, Rio Paranaíba, MG, Brasil.Universidade Federal de Viçosa (UFV)BrazilRio Paranaíba, MG, BrazilInstituto de Ciências Biológicas e da Saúde, Universidade Federal de Viçosa (UFV), Campus Rio Paranaíba, 38810-000, Rio Paranaíba, MG, Brasil.

Everaldo Antônio Lopes

Claúdio Marcelo Gonçalves Oliveira

Instituto Biológico, Campinas, SP, Brasil.Instituto BiológicoBrasilCampinas, SP, BrasilInstituto Biológico, Campinas, SP, Brasil.

Pedro Ivo Vieira Good God

E-mail: lvisotto@yahoo.com.br. ^*Corresponding author

SCIMAGO INSTITUTIONS RANKINGS

Instituto Biológico, Campinas, SP, Brasil.Instituto BiológicoBrasilCampinas, SP, BrasilInstituto Biológico, Campinas, SP, Brasil.

Figures | Tables

Figures (2)
Tables (2)

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Figure 1
Anterior region of males of Meloidogyne incognita, M. enterolobii, M. paranaensis and M. javanica. DGO^* = Dorsal esophageal gland orifice.

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Figure 2
Phenotypic profile of the esterase enzyme of the main Meloidogyne species reported in Brazil. Colorless boxes represent weak bands, filled boxes represent strong bands. Rm: Migration ratio in relation to the slower band (Rm = 1.0) of M. javanica. E1, I1, I2, J3, A2, H1, P1, Co2, En2 and E3: codes used to identify the esterase bands pattern of M. exigua, M. incognita, M. incognita = M. polycephannulata, M. javanica, M. arenaria, M. hapla, M. paranaensis, M. coffeicola, M. enterolobii and M. ethiopica, respectively. Source: Adapted from CARNEIRO et al. (2016CARNEIRO, R.M.D.G. et al. Gênero Meloidogyne: diagnose através de eletroforese de isoenzimas e marcadores SCAR. In: OLIVEIRA, C.M.G.; SANTOS, M.A.; CASTRO, L.H.S. Diagnose de fitonematoides. Campinas: Millennium Editora, 2016. Cap.3, p.47-72.).

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Table 1
Universal and species-specific primers used in the identification of some Meloidogyne species through the polymerase chain reaction.

Thumbnail

Table 2
Primers for the identification of Meloidogyne species by real-time PCR (qPCR) and loop-mediated isothermal amplification (LAMP), including probes for qPCR.

Figure 1 Anterior region of males of Meloidogyne incognita, M. enterolobii, M. paranaensis and M. javanica. DGO^* = Dorsal esophageal gland orifice.

Figure 2 Phenotypic profile of the esterase enzyme of the main Meloidogyne species reported in Brazil. Colorless boxes represent weak bands, filled boxes represent strong bands. Rm: Migration ratio in relation to the slower band (Rm = 1.0) of M. javanica. E1, I1, I2, J3, A2, H1, P1, Co2, En2 and E3: codes used to identify the esterase bands pattern of M. exigua, M. incognita, M. incognita = M. polycephannulata, M. javanica, M. arenaria, M. hapla, M. paranaensis, M. coffeicola, M. enterolobii and M. ethiopica, respectively. Source: Adapted from CARNEIRO et al. (2016CARNEIRO, R.M.D.G. et al. Gênero Meloidogyne: diagnose através de eletroforese de isoenzimas e marcadores SCAR. In: OLIVEIRA, C.M.G.; SANTOS, M.A.; CASTRO, L.H.S. Diagnose de fitonematoides. Campinas: Millennium Editora, 2016. Cap.3, p.47-72.).

Table 1 Universal and species-specific primers used in the identification of some Meloidogyne species through the polymerase chain reaction.

Target	Code	Sequence (5’→3’)	Amplicon Size (bp)	Citation
Universal (5S - 18S rDNA)	194	TTAACTTGCCAGATCGGACG	720	BLOK et al. (1997BLOK, V.C. et al. Comparison of sequences from the ribosomal DNA intergenic region of Meloidogyne mayaguensis and other major tropical root knot nematodes. Journal of Nematology, v.29, p.16-22, 1997. Available from: http://www.documentation.ird.fr/hor/fdi:010048922>. Accessed: Dec. 03, 2016. http://www.documentation.ird.fr/hor/fdi:... )
	195	TCTAATGAGCCGTACGC
Universal (D2/D3 28S rDNA)	D2	ACAAGTACCGTGAGGGAAAGT	500	NUNN (1992NUNN, G. Nematode molecular evolution. An investigation of evolutionary patterns among nematodes based upon DNA sequences. 1992. Ph.D. dissertation, University of Nottingham, UK, 1992. Available from: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.334855>. Accessed: Mar. 15, 2017. http://ethos.bl.uk/OrderDetails.do?uin=u... )
	D3	TGCGAAGGAACCAGCTACTA
Universal (COII 16S mtRNA)	C2F3	GGTCAATGTTCAGAAATTTGTGG	-	POWERS & HARRIS (1993POWERS, T.O.; HARRIS, T.S.A polymerase chain reaction method for identification of five major Meloidogyne species. Journal of Nematology , v.25, p.1-6, 1993. Available from: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2619349/>. Accessed: Jan. 14, 2017. https://www.ncbi.nlm.nih.gov/pmc/article... )
	1108	TACCTTTGACCAATCACGCT
M. arenaria (SCAR)	Far	TCGGCGATAGAGGTAAATGAC	420	ZIJLSTRA (2000aZIJLSTRA, C. et al. Identification of Meloidogyne incognita, M. javanica and M. arenaria using sequence characterized amplified regions (SCAR) based PCR assays. Nematology , v.2, n.8, p.847-853, 2000a. Accessed: Dec. 03, 2017. doi: 10.1163/156854100750112798. https://doi.org/10.1163/1568541007501127... )
	Rar	TCGGCGATAGACACTACAAACT
M. arenaria (SCAR)	-	TCGAGGGCATCTAATAAAGG	950	DONG et al. (2001DONG, K. et al. Development of PCR primers to identify species of root-knot nematodes: Meloidogyne arenaria, M. hapla, M. incognita and M. javanica. Nematropica, v.31, n.2, p.271-280, 2001. Available from: http://journals.fcla.edu/nematropica/article/viewFile/69633/67293>. Accessed: Mar. 03, 2017. http://journals.fcla.edu/nematropica/art... )
	-	GGGCTGAATAATCAAAGGAA
M. enterolobii (SCAR)	-	GAAATTGCTTTATTGTTACTAAG	322	BLOK et al. (2002BLOK, V.C. et al. Mitochondrial differences distinguishing Meloidogyne mayaguensis from the major species of tropical root-knot nematodes. Nematology, v.4, n.7, p.773-781, 2002. Accessed: Mar. 09, 2017. doi: 10.1163/156854102760402559. https://doi.org/10.1163/1568541027604025... )
	-	TAGCCACAGCAAAATAGTTTTC
M. enterolobii (SCAR)	MK7-F	GATCAGAGGCGGGCGCATTGCGA	520	TIGANO et al. (2010TIGANO, M.S. et al. Genetic diversity of the root-knot nematode Meloidogyne enterolobii and development of a SCAR marker for this guava-damaging species. Plant Pathology , v.59, n.6, p.1054-1061, 2010. Available from: https://www.researchgate.net/publication/229721445_Genetic_diversity_of_the_root-knot_nematode_Meloidogyne_enterolobii_and_development_of_a_SCAR_marker_for_this_guava-damaging_species>. Accessed: Nov. 13, 2017. doi: 10.1111/j.1365-3059.2010.02350.x. https://www.researchgate.net/publication... )
	MK7-R	CGAACTCGCTCGAACTCGAC
M. enterolobii (SCAR)	Me-F	AACTTTTGTGAAAGTGCCGCTG	236	LONG et al. (2006LONG, H. et al. Development of a PCR diagnostic for the root-knot nematode Meloidogyne enterolobii. Acta Phytopathologica Sinica, v.36, p.109-115, 2006. Accessed: Mar. 05, 2017.)
	Me-R	TCAGTTCAGGCAGGATCAACC
M. ethiopica (SCAR)	meth-F	ATGCAGCCGCAGGGAACGTAGTTG	350	CORREA et al. (2014CORREA, V.R. et al. Genetic diversity of the root-knot nematode Meloidogyne ethiopica and development of a species-specific SCAR marker for its diagnosis. Plant Pathology, v.63, n.2, p.476-483, 2014. Available from: http://onlinelibrary.wiley.com/doi/10.1111/ppa.12108/full>. Accessed: Jan. 17, 2017. doi: 10.1111/ppa.12108. http://onlinelibrary.wiley.com/doi/10.11... )
	meth-R	TGTTGTTTCATGTGCTTCGGCATC
M. exígua (SCAR)	exD15-F	CATCCGTGCTGTAGCTGCGAG	562	RANDIG et al. (2002RANDIG, O. et al. A species-specific satellite DNA family in the genome of the coffee root nematode Meloidogyne exigua: application to molecular diagnosis of the parasite. MolecularPlant Pathology , v.3, p.431-437, 2002. Available from: http://onlinelibrary.wiley.com/doi/10.1046/j.1364-3703.2002.00134.x/full>. Accessed: Dec. 12, 2016. doi: 10.1046/j.1364-3703.2002.00134.x http://onlinelibrary.wiley.com/doi/10.10... )
	exD15-R	CTCCGTGGGAAGAAAGACTG
M. hapla (SCAR)	-	CAGGCCCTTCCAGCTAAAGA	960	WILLIAMSON et al. (1997WILLIAMSON, V.M. et al. A PCR assay to identify and distinguish single juveniles of Meloidogyne hapla and M. chitwoodi. Journal of Nematology , v.29, n.1, p.9-15, 1997. Available from: http://journals.fcla.edu/jon/article/view/66873/64541>. Accessed: Dec. 03, 2016. http://journals.fcla.edu/jon/article/vie... )
	-	CTTCGTTGGGGAACTGAAGA
M. hapla (SCAR)	-	TGACGGCGGTGAGTGCGA	610	ZIJLSTRA (2000bZIJLSTRA, C. Identification of Meloidogyne chitwoodi, M. fallax and M. hapla based on SCAR-PCR: a powerful way of enabling reliable identification of populations or individuals that share common traits. European Journal ofPlant Pathology , v.106, n.3, p.283-290, 2000b. Available from: https://link.springer.com/content/pdf/10.1023%2FA%3A1008765303364.pdf>. Accessed: Dec. 03, 2017. doi: 10.1023/A:1008765303364. https://link.springer.com/content/pdf/10... )
	-	TGACGGCGGTACCTCATAG
M. hapla (SCAR)	-	GGCTGAGCATAGTAGATGATGTT	1500	DONG et al. (2001DONG, K. et al. Development of PCR primers to identify species of root-knot nematodes: Meloidogyne arenaria, M. hapla, M. incognita and M. javanica. Nematropica, v.31, n.2, p.271-280, 2001. Available from: http://journals.fcla.edu/nematropica/article/viewFile/69633/67293>. Accessed: Mar. 03, 2017. http://journals.fcla.edu/nematropica/art... )
	-	ACCCATTAAAGAGGAGTTTTGC
M. incognita (SCAR)	Finc	CTCTGCCCAATGAGCTGTCC	1200	ZIJLSTRA (2000aZIJLSTRA, C. et al. Identification of Meloidogyne incognita, M. javanica and M. arenaria using sequence characterized amplified regions (SCAR) based PCR assays. Nematology , v.2, n.8, p.847-853, 2000a. Accessed: Dec. 03, 2017. doi: 10.1163/156854100750112798. https://doi.org/10.1163/1568541007501127... )
	Rinc	CTCTGCCCTCACATTAAG
M. incognita (SCAR)	-	TAGGCAGTAGGTTGTCGGG	1350	DONG et al. (2001DONG, K. et al. Development of PCR primers to identify species of root-knot nematodes: Meloidogyne arenaria, M. hapla, M. incognita and M. javanica. Nematropica, v.31, n.2, p.271-280, 2001. Available from: http://journals.fcla.edu/nematropica/article/viewFile/69633/67293>. Accessed: Mar. 03, 2017. http://journals.fcla.edu/nematropica/art... )
	-	CAGATATCTCTGCATTGGTGC
M. incognita (SCAR)	Inc-K14-F	GGGATGTGTAAATGCTCCTG	399	RANDIG et al. (2002RANDIG, O. et al. A species-specific satellite DNA family in the genome of the coffee root nematode Meloidogyne exigua: application to molecular diagnosis of the parasite. MolecularPlant Pathology , v.3, p.431-437, 2002. Available from: http://onlinelibrary.wiley.com/doi/10.1046/j.1364-3703.2002.00134.x/full>. Accessed: Dec. 12, 2016. doi: 10.1046/j.1364-3703.2002.00134.x http://onlinelibrary.wiley.com/doi/10.10... )
	Inc-K14-R	CCCGCTACACCCTCAACTTC
M. incognita (SCAR)	MI-F	GTGAGGATTCAGCTCCCCAG	955	MENG et al. (2004MENG, Q.P. et al. PCR assays for rapid and sensitive identification of three major root-knot nematodes, Meloidogyne incognita, M. javanica and M. arenaria. Acta Phytopathologica Sinica , v.34, p.204-210, 2004. Accessed: Mar. 11, 2017.)
	MI-R	ACGAGGAACATACTTCTCCGTCC
M. incognita (SCAR)	Mi2F4	ATGAAGCTAAGACTTTGGGCT	300	KIEWNICK et al. (2013KIEWNICK, S. et al. Identiﬁcation of the tropical root-knot nematode species Meloidogyne incognita, M. javanica and M. arenaria using a multiplex PCR assay. Nematology , v.15, p.891-894, 2013. Accessed: Feb. 17, 2017. doi: 10.1163/15685411-00002751. https://doi.org/10.1163/15685411-0000275... )
	Mi1R1	TCCCGCTACACCCTCAACTTC
M. javanica (SCAR)	Fjav	GGTGCGCGATTGAACTGAGC	670	ZIJLSTRA (2000aZIJLSTRA, C. et al. Identification of Meloidogyne incognita, M. javanica and M. arenaria using sequence characterized amplified regions (SCAR) based PCR assays. Nematology , v.2, n.8, p.847-853, 2000a. Accessed: Dec. 03, 2017. doi: 10.1163/156854100750112798. https://doi.org/10.1163/1568541007501127... )
	Rjav	CAGGCCCTTCAGTGGAACTATAC
M. javanica (SCAR)	-	CCTTAATGTCAACACTAGAGCC	1650	DONG et al. (2001DONG, K. et al. Development of PCR primers to identify species of root-knot nematodes: Meloidogyne arenaria, M. hapla, M. incognita and M. javanica. Nematropica, v.31, n.2, p.271-280, 2001. Available from: http://journals.fcla.edu/nematropica/article/viewFile/69633/67293>. Accessed: Mar. 03, 2017. http://journals.fcla.edu/nematropica/art... )
	-	GGCCTTAACCGACAATTAGA
M. javanica (SCAR)	-	ACGCTAGAATTCGACCCTGG	517	MENG et al. (2004MENG, Q.P. et al. PCR assays for rapid and sensitive identification of three major root-knot nematodes, Meloidogyne incognita, M. javanica and M. arenaria. Acta Phytopathologica Sinica , v.34, p.204-210, 2004. Accessed: Mar. 11, 2017.)
	-	GGTACCAGAAGCAGCCATGC
M. paranaensis (SCAR)	Par-F	GCCCGACTCCATTTGACGGA	208	RANDIG et al. (2002RANDIG, O. et al. A species-specific satellite DNA family in the genome of the coffee root nematode Meloidogyne exigua: application to molecular diagnosis of the parasite. MolecularPlant Pathology , v.3, p.431-437, 2002. Available from: http://onlinelibrary.wiley.com/doi/10.1046/j.1364-3703.2002.00134.x/full>. Accessed: Dec. 12, 2016. doi: 10.1046/j.1364-3703.2002.00134.x http://onlinelibrary.wiley.com/doi/10.10... )
	Par-R	CCGTCCAGATCCATCGAAGTC
	JMV1	GGATGGCGTGCTTTCAAC
M. hapla, M. chitwoodie M. fallax (IGS - SCAR)	JMV2	TTTCCCCTTATGATGTTTACCC	440, 540, 670	WISHART et al. (2002WISHART, J. et al. Ribosomal intergenic spacer: a PCR diagnostic for Meloidogyne chitwoodi, M. fallax and M. hapla. Phytopathology , v.92, n.8, p.884-892, 2002. Available from: http://apsjournals.apsnet.org/doi/pdf/10.1094/PHYTO.2002.92.8.884>. Accessed: Feb. 17, 2017. doi: 10.1094/PHYTO.2002.92.8.884. http://apsjournals.apsnet.org/doi/pdf/10... )
	JMV	AAAAATCCCCTCGAAAAATCCACC

Table 2 Primers for the identification of Meloidogyne species by real-time PCR (qPCR) and loop-mediated isothermal amplification (LAMP), including probes for qPCR.

Target	Code	Sequence (5’→3’)	Citation
-----------------------------------------------------------------------------------qPCR-----------------------------------------------------------------------------------			Citation
Ma^*	QareF	TCCATTTCTCCTTGGGTTTG	1
	QareR	GCCATCCCTTCACACGTTAT
Mi^*	RKNf	GCTGGTGTCTAAGTGTTGCTGATAC	2
	RKNr	GAGCCTAGTGATCCACCGATAAG
	Forward	TGGTTCAGGGTCATTTTTCTATAAAGT
Mh	Reverse	CAAATCGCTGCGTACCAACA	3
	Probe	FAM-CCATTGGCACTATAAC-MGB
	Ment17F	TGTGGTGGCTCATTTTCATTA
Me	Ment17R	AAAAACCCTAAAAATACCCCAAA	4
	Probe	AGGAGCTG
	Mminor_f299	CCGTGACTGAATATGAGGTGA
Mm+	Mminor_r362	GAGGCTCATTAAGTCTTACGATTAT	5
	Probe	FAM-ATGTTAGGATTATCG-MGBNFQ
RKN^*	Melo-Rlong	GGCCTCACTTAAGAGGCTCA	6
	Melo-R short	TATACAGCCACGGACGTTCA
-----------------------------------------------------------------------------------LAMP-----------------------------------------------------------------------------------
	RKN-F3	CTGCCCTTTGTACACACC
	RKN-B3	GACACCAGCGACAGCCGTT
RKN	RKN-FIP	CTGCGATTAAATTGGTTTCCATCAACGGGACTGAGCCATTTCG	7
	RKN-BIP	GCTTGAACCGGGCAAAAGTCCATAAAGTAATGATCCAGCAGC
	RKN-LB	GTAACAAGGTAGCTGTAGGTGAAC
	Mi-F3	TATGTCAGCCCCCGGTTC
Mi	Mi-B3	GAGAAGGAAAAGAGTGCCAA	7
	Mi-FIP	CTTTCCTTGGAATTGGAACAGGGTCAATTGCTTTATATCAAACACC
	Mi-BIP	GGACGGAGAAGTATGTTCCTCTCTGGAAAAGAAAAATCAGTCTT
	Me-F3	CCAAGTACTAAGGAAGCCC
	Me-B3	ATCCTAATTTTYCTCCCACACA
Me	Me-FIP	ACAGTGATTACGACCATACCGCGTTCGTTGCTTAACTTGCCAGA	8
	Me-BIP	TCTAAGGCAAAGTGGGCGGAGCTCTYTTTGCCTTAAACCATTCCC
	Me-LF	AAGCACGCCATCCCGTC
	Me-LB	TGTTGTTCGCTGTTCGC
	Mh-F3	GAATATGAGGTGACATGTTAGG
	Mh-B3	TCAATGTTTCTGCAGTTCG
Mh	Mh-FIP	TGAAAAAAATATTGCTGGCGTCCACCTTAATCGGGTTTAAGACT	9
	Mh-BIP	TCTATCCTTATCGGTGGATCACTCCACAAATTATCGCAGTTAGCT
	Mh-LB	GGCTCGTGGATCCATGAAGAACG

^*
SYBR Green system. ⁺Quarantine pest for Brazil. RKN: Universal sets for root-knot nematodes. Targets: Ma - M. arenaria; Me - M. enterolobii; Mi - M. incognita; Mh - M. hapla; Mm - M. minor. Citations: 1 - AGUDELO et al. (2011AGUDELO, P. et al. Validation of a real-time polymerase chain reaction assay for the identification of Meloidogyne arenaria. Plant Disease, v.95, p.835-838, 2011. Available from: https://doi.org/10.1094/PDIS-09-10-0668>. Accessed: Dec. 03, 2016. doi: 10.1094/ PDIS-09-10-0668.
https://doi.org/10.1094/PDIS-09-10-0668... ); 2 - TOYOTA et al. (2008TOYOTA, K. et al. Development of a real-time PCR method for the potato-cyst nematode Globodera rostochiensis and the root-knot nematode Meloidogyne incognita. Soil Science and Plant Nutrition, v.54, n.1, p.72-76, 2008. Available from: http://onlinelibrary.wiley.com/doi/10.1111/j.1747-0765.2007.00212.x/epdf>. Accessed: Jan. 01, 2017. doi: 10.1111/j.1747-0765.2007.00212.x.
http://onlinelibrary.wiley.com/doi/10.11... ); 3 - SAPKOTA et al. (2016SAPKOTA, R. et al. A TaqMan real-time PCR assay for detection of Meloidogyne hapla in root galls and in soil. Nematology , v.18, p.147-154, 2016. Available from: https://www.researchgate.net/publication/284712140_A_TaqMan_real-time_PCR_assay_for_detection_of_Meloidogyne_hapla_in_root_galls_and_in_soil>. Accessed: Jan. 01, 2017. doi: 10.1163/15685411-00002950.
https://www.researchgate.net/publication... ); 4 - KIEWNICK et al. (2015KIEWNICK, S. et al. Development and validation of LNA-based quantitative real-time PCR assays for detection and identification of the root-knot nematode Meloidogyne enterolobii in complex DNA backgrounds. Phytopathology , v.105, n.9, p.1245-1249, 2015. Available from: http://dx.doi.org/10.1094/PHYTO-12-14-0364-R>. Accessed: Feb. 17, 2017.doi: 10.1094/PHYTO-12-14-0364-R.
http://dx.doi.org/10.1094/PHYTO-12-14-03... ); 5 - DE WEERDT et al. (2011DE WEERDT, M. et al. A real-time PCR assay to identify Meloidogyne minor. Journal of Phytopathology, v.159, p.80-84, 2011. Available from: https://doi.org/10.1111/j.1439-0434.2010.01717.x>. Accessed: Dec. 03, 2017. doi: 10.1111/j.1439-0434.2010.01717.x.
https://doi.org/10.1111/j.1439-0434.2010... ); 6 - BERRY et al. (2008BERRY, S.D. et al. Detection and quantification of root-knot nematode (Meloidogyne javanica), lesion nematode (Pratylenchus zeae) and dagger nematode (Xiphinema elongatum) parasites of sugarcane using real-time PCR. Molecular and Cellular Probes, v.22, n.3, p.168-176, 2008. Available from: https://doi.org/10.1016/j.mcp.2008.01.003>. Accessed: Jan. 01, 2017. doi: 10.1016/j.mcp.2008.01.003.
https://doi.org/10.1016/j.mcp.2008.01.00... ); 7 - NIU et al. (2011NIU, J.H. et al. Rapid detection of Meloidogyne spp. by LAMP assay in soil and roots. Crop Protection, v.30, n.8, p.1063-1069, 2011. Available from: https://doi.org/10.1016/j.cropro.2011.03.028>. Accessed: Feb. 17, 2017. doi: 10.1016/j.cropro.2011.03.028.
https://doi.org/10.1016/j.cropro.2011.03... ); 8 - NIU et al. (2012)NIU, J.H. et al. Evaluation of loop-mediated isothermal amplification (LAMP) assays based on 5S rDNA-IGS2 regions for detecting Meloidogyne enterolobii. Plant Pathology , v.61, p.809-819, 2012. Available from: https://www.researchgate.net/publication/262890929>. Accessed: Jan. 15, 2017. doi: 10.1111/j.1365-3059.2011.02562.x.
https://www.researchgate.net/publication... ; 9 - PENG et al. (2017PENG, H. et al. Rapid, simple and direct detection of Meloidogyne hapla from infected root galls using loop-mediated isothermal amplification combined with FTA technology. Scientific Reports, v.7, Article number 44853, 2017. Available from: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5377304/>. Accessed: May 24, 2017. doi: 10.1038/srep44853.
https://www.ncbi.nlm.nih.gov/pmc/article... ).

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Diagnostic methods for identification of root-knot nematodes species from Brazil

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[1] E-mail: lvisotto@yahoo.com.br. ^*Corresponding author