Mitochondrial Haplotype-based Identification of Root-knot Nematodes (Meloidogyne spp.) on Cut Foliage Crops in Florida

Abstract

Florida accounts for more than 75% of the national cut foliage production. Unfortunately, root-knot nematodes (RKN) (Meloidogyne spp.) are a serious problem on these crops, rendering many farms unproductive. Currently, information on the Meloidogyne spp. occurring on most commonly cultivated cut foliage crops in Florida, and tools for their rapid identification are lacking. The objectives of this study were to (i) identify specific RKN infecting common ornamental cut foliage crops in Florida and (ii) evaluate the feasibility of using the mtDNA haplotype as a molecular diagnostic tool for rapid identification of large samples of RKN. A total of 200 Meloidogyne females were collected from cut foliage plant roots. Meloidogyne spp. were identified by PCR and RFLP of mitochondrial DNA. PCR and RFLP of mitochondrial DNA were effective in discriminating the Meloidogyne spp. present. Meloidogyne incognita is the most dominant RKN on cut foliage crops in Florida and must be a high target for making management decisions. Other Meloidogyne spp. identified include M. javanica, M. hapla, Meloidogyne sp. 1, and Meloidogyne sp. 2. The results for this study demonstrate the usefulness of the mtDNA haplotype-based designation as a valuable molecular tool for identification of Meloidogyne spp.

First Report of the Root-Knot Nematode (Meloidogyne javanica) Infecting Hops (Humulus lupulus) in Florida, USA

“…To further confirm the species identification we use the species-specific SCAR primer set Fjav (5¢-GGT GCG CGA TTG AAC TGA GC-3¢) and Rjav (5¢-CAG GCC CTT CAG TGG AAC TAT AC-3¢) (Zijlstra et al, 2000). This primer set yield a fragment of approximately 670 bp, which is identical to that previously reported for M. javanica (Zijlstra et al, 2000;Baidoo et al, 2016). Additionally, NADH dehydrogenase subunit 5 gene was amplified using NAD5F2 (5¢-TAT TTT TTG TTT GAG ATA TAT First Report of the Root-Knot Nematode (Meloidogyne javanica) Infecting Hops (Humulus lupulus) in Florida, USA TAG-3¢) and NAD5R1 (5¢-CGTGAATCTTGATTTTC-CATTTTT-3¢) primers as described by Janssen et al (2016).…”

“…For molecular analyses, DNA was extracted from individual females and mitochondrial DNA was amplified with MORF (5¢-ATC GGG GTT TAA TAA TGG G-3¢) and MTHIS (5¢-AAA TTC AAT TGA AAT TAA TAG C-3¢) primer set (Stanton et al, 1997). A fragment of approximately 740 bp was produced, which has been reported for M. incognita and M. javanica found in Florida (Baidoo et al, 2016). To further confirm the species identification we use the species-specific SCAR primer set Fjav (5¢-GGT GCG CGA TTG AAC TGA GC-3¢) and Rjav (5¢-CAG GCC CTT CAG TGG AAC TAT AC-3¢) (Zijlstra et al, 2000).…”

Meloidogyne luci, a new root‐knot nematode parasitizing potato in Portugal

“…Mitochondrial DNA COI was proposed as a molecular marker for DNA barcoding and is capable of discriminating between RKN species more than the rRNA genes (Ahmed et al ., ). Moreover, the mtDNA COII /16S rRNA region has been used in diagnosis and large‐scale regional RKN surveys through PCR and RFLP (Tigano et al ., ; Humphreys‐Pereira et al ., ; Baidoo et al ., ; Karuri et al ., ). Meloidogyne lopezi , closely related to other tropical Meloidogyne species that infect coffee, especially M. arabicida , M. izalcoensis and M. paranaensis , could be differentiated from these species by PCR‐RFLP of the mtDNA COII /16S rRNA (Humphreys‐Pereira et al ., ).…”

“…Meloidogyne lopezi , closely related to other tropical Meloidogyne species that infect coffee, especially M. arabicida , M. izalcoensis and M. paranaensis , could be differentiated from these species by PCR‐RFLP of the mtDNA COII /16S rRNA (Humphreys‐Pereira et al ., ). A nematode survey of sweet potato fields and cut foliage crops was conducted in Kenya and Florida, respectively, and the mtDNA COII /16S rRNA region was used to determine the distribution and prevalence of Meloidogyne species (Baidoo et al ., ; Karuri et al ., ). Additionally, mtDNA COII /16S rRNA data improved the discrimination sensitivity of RKN species and atypical, non‐identified isolates (Tigano et al ., ).…”

Recognition of species belonging to Meloidogyne ethiopica group and development of a diagnostic method for its detection

“…While species-specific primer pairs are developed for the more common Meloidoyne species (Zijlstra et al 2000;Adam et al 2007;Tigano et al 2010), they are not available or are not working reliably for many other 'minor' species including M. ethiopica M. luci and M. inornata (Correa et al 2014). Similar is true for PCR-RFLP approach; while it seems to work for identification of some species, the approach does not enable identification of all or at least the majority of clade I species in the genus Meloidoyne (Powers & Harris, 1993;Zijlstra et al 1995;Stanton et al 1997;Maleita et al 2012;Pagan et al 2015;Baidoo et al 2016). While we could not find any species-specific diagnostic parameters that could be used for an easy identification on morphological level, we were able to identify group-specific parameters (conserved nucleotide sites) on molecular level.…”