Behaviour of Heterodera glycines and Meloidogyne incognita infective juveniles exposed to nematode FMRFamide-like peptides in vitro

Abstract

The effects of 1 mM solutions of FMRFamide-like peptides (FLPs) on the behaviours of Heterodera glycines and Meloidogyne incognita infective juveniles (J2) were examined in vitro. Seven FLPs, representing products of five flp genes and comprising a variety of amino acid sequences, were tested for their effects on J2 head movement frequency. Distinct differences in species responses were observed. KHEYLRFa and KSAYMRFa caused increased head movement frequencies in each species. In H. glycines KHEYLRFa was 2.9-fold more potent than KSAYMRFa. In M. incognita the potencies were equal. KHEYLRFa and KSAYMRFa each stimulated H. glycines J2 more effectively than M. incognita J2. However, two additional FLPs, AQTFVRFa and SAPYDPNFLRFa, were stimulatory in M. incognita but not in H. glycines. KPNFIRFa, KPNFLRFa and RNSSPLGTMRFa had no effect on either species. Substitution of d-amino acids at any position in KHEYLRFa resulted in decreased stimulation of head movement relative to the native peptide in each species, but all of the d-amino acid KHEYLRFa analogues were stimulatory relative to untreated controls. d-amino acid substitutions in KSAYMRFa eliminated stimulatory activity in M. incognita by all analogues except dKSAYMRFa. In H. glycines, only KSdAYMRFa and KSAYMdRFa were not stimulatory, and KSAYdMRFa stimulated equal to the native peptide. The remaining four analogues each stimulated relative to controls but below the native peptide level. Analysis of the head movement behaviour of large numbers of J2 of each species demonstrated that behaviours are quite stable and responses to FLP treatment are highly predictable.

FMRFamide-Like Peptide 22 Influences the Head Movement, Host Finding, and Infection of Heterodera glycines

“…Hg-FLP-22b and Hg-FLP-22c displayed significant stimulatory effects on head movement and stylet penetration ability compared with Hg-FLP-22a, suggesting the two peptides may be significantly more active isoforms of Hg-FLP-22 for activation of putative cognate receptors. Similar observations were also reported by previous studies indicating obvious differences in physiological function and potency between FLP peptides encoded on the same gene in H. glycines , Panagrellus redivivus , or C. elegans ( Masler et al, 2012 ; Atkinson et al, 2016 ). It has been documented that a single FLP receptor can be activated by multiple FLPs, whereas the potency of receptor activation by structurally similar FLPs is different ( Rogers et al, 2003 ; Mertens et al, 2005 ; Cohen et al, 2009 ).…”

“…Applications of synthetic Hg-FLP-22 peptides (FLP-22b and FLP-22c) significantly increased the head movement frequency of H. glycines . This is similar to the observation which showed that FLP-6 and FLP-14, both highly conserved single-copy peptides in nematode species ( Peymen et al, 2014 ), also have significant stimulatory effects on the anterior movement frequency of H. glycines and M. incognita ( Masler et al, 2012 ). Once hatched from eggs, the infective J2s of PPN need to quickly move toward host roots and penetrate the cortex layer cells via thrusting their stylet before their energy reserves are depleted ( Perry, 1996 ).…”

“…Then, J2s were transferred to a cell counting chamber (75 mm × 25 mm × 1.8 mm; chamber depth = 100 μm) with four rooms (10 μl loading volume), and head movements were observed for a randomly selected individual nematode under a stereomicroscope (OLYMPUS SZX-16, Tokyo, Japan). Head movement frequencies of J2s were determined following a definition previously described by Masler et al (2012) . Head movements of each J2 were counted for 1 min.…”

“…The neuromuscular system is considered as the potential target of the effective chemotherapeutic treatments for animalparasitic nematode management because a series of normal parasite biological functions is governed through the nervemuscle coordinated function (Maule et al, 2002;McVeigh et al, 2012). The role of the neuromuscular system in PPN has also been demonstrated in various facets of parasitic activities, such as host location, penetration, and migration inside and outside of host tissues, and even slight head motion at the feeding site (Perry, 1996;Maule et al, 2002;Curtis, 2008;Reynolds et al, 2011;Han et al, 2018).…”

“…To date, studies of FLP functions in PPN have primarily focused on root-knot nematodes, M. incognita and M. graminicola ; only in vitro physiological experiments have tested the proteolysis of two synthetic peptides (FLP-6 and FLP-14) and their effects on the behavior of H. glycines J2s ( Masler, 2010 ; Masler et al, 2012 ). Chromatographic analysis of the FaRP immunoactivity from H. glycines further indicated that FLPs are present in all development stages and the highest FLP immunoactivity occurs in the J2 stage ( Masler et al, 1999 ).…”