[Show abstract][Hide abstract] ABSTRACT: The potato cyst nematode Globodera pallida has biotrophic interactions with its host. The nematode induces a feeding structure - the syncytium - which it keeps alive for the duration of the life cycle and on which it depends for all nutrients required to develop to the adult stage. Interactions of G. pallida with the host are mediated by effectors, which are produced in two sets of gland cells. These effectors suppress host defences, facilitate migration and induce the formation of the syncytium.
[Show abstract][Hide abstract] ABSTRACT: SUMMARY The potato cyst nematodes (PCN) Globodera pallida and G. rostochiensis are major pests of potatoes. The G. pallida (and G. rostochiensis) life cycle includes both diapause and quiescent stages. Nematodes in dormancy (diapause or quiescent) are adapted for long-term survival and are more resistant to nematicides. This study analysed the mechanisms underlying diapause and quiescence. The effects of several compounds (8Br-cGMP, oxotremorine and atropine) on the activation of hatching were studied. The measurements of some morphometric parameters in diapaused and quiescent eggs after exposure to PRD revealed differences in dorsal gland length, subventral gland length and dorsal gland nucleolus. In addition, the expression of 2 effectors (IVg9 and cellulase) was not induced in diapaused eggs in water or PRD, while expression was slightly induced in quiescent eggs. Finally, we performed a comparative study to identify orthologues of C. elegans diapause related genes in plant-parasitic nematodes (G. pallida, Meloidogyne incognita, M. hapla and Bursaphelenchus xylophilus). This analysis suggested that it was not possible to identify G. pallida orthologues of the majority of C. elegans genes involved in the control of dauer formation. All these data suggest that G. pallida may use different mechanisms to C. elegans in regulating the survival stage.
[Show abstract][Hide abstract] ABSTRACT: Recombination is typically assumed to be absent in animal mitochondrial genomes (mtDNA). However, the maternal mode of inheritance means that recombinant products are indistinguishable from their progenitor molecules. The majority of studies of mtDNA recombination assess past recombination events, where patterns of recombination are inferred by comparing the mtDNA of different individuals. Few studies assess contemporary mtDNA recombination, where recombinant molecules are observed as direct mosaics of known progenitor molecules. Here we use the potato cyst nematode, Globodera pallida, to investigate past and contemporary recombination. Past recombination was assessed within and between populations of G. pallida, and contemporary recombination was assessed in the progeny of experimental crosses of these populations. Breeding of genetically divergent organisms may cause paternal mtDNA leakage, resulting in heteroplasmy and facilitating the detection of recombination. To assess contemporary recombination we looked for evidence of recombination between the mtDNA of the parental populations within the mtDNA of progeny. Past recombination was detected between a South American population and several UK populations of G. pallida, as well as between two South American populations. This suggests that these populations may have interbred, paternal mtDNA leakage occurred, and the mtDNA of these populations subsequently recombined. This evidence challenges two dogmas of animal mtDNA evolution; no recombination and maternal inheritance. No contemporary recombination between the parental populations was detected in the progeny of the experimental crosses. This supports current arguments that mtDNA recombination events are rare. More sensitive detection methods may be required to adequately assess contemporary mtDNA recombination in animals.
[Show abstract][Hide abstract] ABSTRACT: Animal mtDNA is typically assumed to be maternally inherited. Paternal mtDNA has been shown to be excluded from entering the egg or eliminated post-fertilization in several animals. However, in the contact zones of hybridizing species and populations, the reproductive barriers between hybridizing organisms may not be as efficient at preventing paternal mtDNA inheritance, resulting in paternal leakage. We assessed paternal mtDNA leakage in experimental crosses of populations of a cyst-forming nematode, Globodera pallida. A UK population, Lindley, was crossed with two South American populations, P5A and P4A. Hybridization of these populations was supported by evidence of nuclear DNA from both the maternal and paternal populations in the progeny. To assess paternal mtDNA leakage, a ~3.4 kb non-coding mtDNA region was analyzed in the parental populations and in the progeny. Paternal mtDNA was evident in the progeny of both crosses involving populations P5A and P4A. Further, paternal mtDNA replaced the maternal mtDNA in 22 and 40 % of the hybrid cysts from these crosses, respectively. These results indicate that under appropriate conditions, paternal leakage occurs in the mtDNA of parasitic nematodes, and supports the hypothesis that hybrid zones facilitate paternal leakage. Thus, assumptions of strictly maternal mtDNA inheritance may be frequently violated, particularly when divergent populations interbreed.
[Show abstract][Hide abstract] ABSTRACT: Potato cyst nematodes cost the potato industry substantial financial losses annually. Through the use of molecular markers, the distribution and infestation routes of these nematodes can be better elucidated, permitting the development of more effective preventative methods. Here we assess the ability of three molecular markers to resolve multiple representatives of five Globodera pallida populations as monophyletic groups. Molecular markers included a region of the rbp-1 gene (an effector), a non-coding nuclear DNA region (the ITS region), and a novel marker for G. pallida, a ∼3.4 kb non-coding mitochondrial DNA (mtDNA) region. Multiple phylogenetic analysis methods were performed on the three DNA regions separately, and on a data set of these three regions combined. The analyses of the combined data set were similar to that of the sole mtDNA marker; resolving more populations as monophyletic groups, relative to that of the ITS region and rbp-1 gene region. This suggests that individual markers may be inadequate for distinguishing populations of G. pallida. The use of this new non-coding mtDNA marker may provide further insights into the historical distribution of G. pallida, as well as enable the development of more sensitive diagnostic methods.
Journal of nematology 03/2012; 44(1):7-17. · 0.52 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: In this article, we describe the analysis of over 9000 expressed sequence tags (ESTs) from cDNA libraries obtained from various life cycle stages of Globodera pallida. We have identified over 50 G. pallida effectors from this dataset using bioinformatics analysis, by screening clones in order to identify secreted proteins up-regulated after the onset of parasitism and using in situ hybridization to confirm the expression in pharyngeal gland cells. A substantial gene family encoding G. pallida SPRYSEC proteins has been identified. The expression of these genes is restricted to the dorsal pharyngeal gland cell. Different members of the SPRYSEC family of proteins from G. pallida show different subcellular localization patterns in plants, with some localized to the cytoplasm and others to the nucleus and nucleolus. Differences in subcellular localization may reflect diverse functional roles for each individual protein or, more likely, variety in the compartmentalization of plant proteins targeted by the nematode. Our data are therefore consistent with the suggestion that the SPRYSEC proteins suppress host defences, as suggested previously, and that they achieve this through interaction with a range of host targets.
[Show abstract][Hide abstract] ABSTRACT: Two homologues of map-1, which encodes a putative avirulence factor, were found in a single egg mass line of the root-knot nematode Meloidogyne javanica that was virulent to the Mi-resistant gene. The main difference between the two encoded proteins of these homologues, MJ-MAP-1 and MJ-MAP-2, was the position of a 13 amino acid repeat region. Genes encoding two related but novel proteins, MJAP-1 and MJAP-2 (290 and 283 amino acids, respectively, including a potential signal secretion peptide), were also isolated from M. javanica. They have high similarity to MAP at the C-terminus. MJAP-1 and MJAP-2 differ from each other in the number and position of a seven-amino-acid repeat and in five other amino acids. The mjap genes are expressed in the subventral pharyngeal glands of second-stage juveniles of M. javanica, and transcription analysis in different developmental stages showed expression in the juvenile stage but not in eggs or adult females. Both mjap-1 and mjap-2 were expressed in both Mi-virulent and avirulent lines of M. javanica.
[Show abstract][Hide abstract] ABSTRACT: Following the discovery of Globodera pallida in the Uzhhorod region (Zakarpats'ka oblast') of Ukraine, two populations from this region were further examined. Firstly, their virulence was assessed in relation to two sources of resistance to G. pallida and to other G. pallida populations from Europe and South America. The results showed that the Ukrainian populations were very similar in their virulence to the other European populations. One of the test host genotypes was a new cultivar derived from Solanum tuberosum spp. andigena CPC2802, which proved to be more highly resistant than previously available partially resistant cultivars. The second experiment was a comparison of the mitochondrial Cytochrome B gene from the Ukrainian populations with other European and South American populations. The data indicated that the Ukrainian populations were similar to other European populations.
[Show abstract][Hide abstract] ABSTRACT: This essay considers biotrophic cyst and root-knot nematodes in relation to their biology, host-parasite interactions and molecular genetics. These nematodes have to face the biological consequences of the physical constraints imposed by the soil environment in which they live while their hosts inhabit both above and below ground environments. The two groups of nematodes appear to have adopted radically different solutions to these problems with the result that one group is a host specialist and reproduces sexually while the other has an enormous host range and reproduces by mitotic parthenogenesis. We consider what is known about the modes of parasitism used by these nematodes and how it relates to their host range, including the surprising finding that parasitism genes in both nematode groups have been recruited from bacteria. The nuclear and mitochondrial genomes of these two nematode groups are very different and we consider how these findings relate to the biology of the organisms.
[Show abstract][Hide abstract] ABSTRACT: We sequenced a mitochondrial subgenome from the nematode Globodera rostochiensis, in two overlapping pieces. The subgenome was 9210 bp and contained four protein-coding genes (ND4, COIII, ND3, Cytb) and two tRNA genes (tRNA(Thr), tRNA(Gln)). Genome organization was similar to that of Globodera pallida, which is multipartite. Together with the small number of genes on this subgenome, this suggests that the mitochondrial genome of G. rostochiensis is also multipartite. In the initial clones sequenced, COIII and ND3 were full-length, while ND4 and Cytb were interrupted by premature stop codons and contained point indels that disrupted the reading frame. However, sequencing of multiple clones, from DNA extracted both from multiple individuals and from single cysts, revealed a predominant source of variation-in the length of polythymidine tracts. Comparison of our genomic sequences with ESTs similarly revealed variation in the length of polythymidine tracts. We subsequently sequenced both genomic DNA and mRNA from populations of G. pallida. In each case, variation in the length of polythymidine tracts was observed. The levels of expression of mitochondrial genes in G. pallida were representative of the subgenomes present: little evidence of differential expression was observed. These observations are consistent with the operation of posttranscriptional editing in Globodera mitochondria, although this is difficult to show conclusively in the presence of intraindividual gene sequence variation. Further, alternative explanations cannot be discounted; these include the operation of slippage during translation or that genomic copies of most genes are pseudogenes with a small proportion of full-length sequences able to maintain mitochondrial function.
Journal of Molecular Evolution 04/2008; 66(3):197-209. · 2.15 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: SummaryA cloned cDNA copy of the satellite RNA of groundnut rosette virus (GRV), labelled with either 32P or digoxigenin, was used as a probe to detect the satellite RNA in infected leaves. The test was successfully applied to N. benthamiana and to groundnuts, infected with isolates of GRV from East and West Africa and with isolates which cause different types of symptom in groundnuts, including one which is almost symptomless. Although the probe did not react with extracts from plants infected with GRV isolates from which the satellite RNA had been artificially eliminated, all naturally occurring GRV isolates contain the satellite RNA. The test therefore provides a reliable indicator of infection by GRV.
[Show abstract][Hide abstract] ABSTRACT: The cellulose-binding protein gene, cbp, of Meloidogyne javanica encodes a protein that contains a signal secretion peptide and is expressed in the eggs and pre-invasive stage J2s but not from adult females. Several highly similar forms were found in the three root knot nematode species, M. javanica, Meloidogyne incognita and Meloidogyne arenaria indicating very limited divergence of the orthologues and paralogues. The genomic sequences included three introns which differed in size between species and between isolates of M. incognita and M. arenaria, however, the coding regions of the various forms were highly similar. In M. javanica only one form of cbp was found. Treatment of pre-invasive stage J2 nematodes with cbp-dsRNA resulted in a reduction in the ability of J2s to penetrate tomato roots and consequently in a reduction in the number of egg masses produced by the cpb-dsRNA treated nematodes compared to the control nematodes, consistent with an important role of CBP in the early stages of penetration and invasion.
Physiological and Molecular Plant Pathology. 01/2008;
[Show abstract][Hide abstract] ABSTRACT: Four field populations of the nematode Globodera pallida were subjected to selection pressure for increased reproductive ability by rearing sub-populations continuously on four partially resistant potato genotypes for 12 generations. The resistance was derived from either Solanum vernei or from S. tuberosum spp. andigena CPC2802. After the 12th generation the original and sub-populations of nematodes were assessed for their reproductive ability on a susceptible genotype and on each of the partially resistant genotypes. Selection pressure was shown to have increased reproductive ability but the increases were specific to the source of resistance used. The average increase on the ex S. vernei clones was from 11% reproduction by the unselected populations to 35·5% reproduction after selection. On the clones derived from CPC2802, which had higher levels of resistance, the increases were larger with an average of 6·6% reproduction for the unselected but 47·4% reproduction after selection. The response to selection differed amongst the initial field populations with some rates of reproduction increased to as much as 79%. A RAPD based analysis of the original and sub-populations after selection indicated small but consistent changes in the genetic structure, which could have been a result of the selection pressure per se and/or the bottlenecks that the populations had gone though.
[Show abstract][Hide abstract] ABSTRACT: Recently, a multipartite mitochondrial genome was characterized in the potato cyst nematode, Globodera pallida. Six subgenomic circles were detectable by PCR, while full-length genomes were not. We investigate here whether this subgenomic organization occurs in a close relative of G. pallida. We amplified and sequenced one entire mitochondrial subgenome from the cyst-forming nematode, Globodera rostochiensis. Comparison of the noncoding region of this subgenome with those reported previously for G. pallida facilitated the design of amplification primers for a range of subgenomes from G. rostochiensis. We then randomly sequenced five subgenomic fragments, each representative of a unique subgenome. This study indicates that the multipartite structure reported for G. pallida is conserved in G. rostochiensis. A comparison of subgenomic organization between these two Globodera species indicates a considerable degree of overlap between them. Indeed, we identify two subgenomes with an organization identical with that reported for G. pallida. However, other subgenomes are unique to G. rostochiensis, although some of these have blocks of genes comparable to those in G. pallida. Dot-plot comparisons of pairs of subgenomes from G. rostochiensis indicate that the different subgenomes share fragments with high sequence identity. We interpret this as evidence that recombination is operating in the mitochondria of G. rostochiensis.
Journal of Molecular Evolution 10/2007; 65(3):308-15. · 2.15 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The discovery that the potato cyst nematode Globodera pallida has a multipartite mitochondrial DNA (mtDNA) composed, at least in part, of six small circular mtDNAs (scmtDNAs) raised a number of questions concerning the population-level processes that might act on such a complex genome. Here we report our observations on the distribution of some scmtDNAs among a sample of European and South American G. pallida populations. The occurrence of sequence variants of scmtDNA IV in population P4A from South America, and that particular sequence variants are common to the individuals within a single cyst, is described. Evidence for recombination of sequence variants of scmtDNA IV in P4A is also reported. The mosaic structure of P4A scmtDNA IV sequences was revealed using several detection methods and recombination breakpoints were independently detected by maximum likelihood and Bayesian MCMC methods.
Journal of Molecular Evolution 07/2007; 64(6):689-701. · 2.15 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: We sequenced four mitochondrial subgenomes from the potato cyst nematode Globodera pallida, previously characterized as one of the few animals to have a multipartite mitochondrial genome. The sequence data indicate that three of these subgenomic mitochondrial circles are mosaics, comprising long, multigenic fragments derived from fragments of the other circles. This pattern is consistent with the operation of intermitochondrial recombination, a process generally considered absent in animal mitochondria. We also report that many of the duplicated genes contain deleterious mutations, ones likely to render the gene nonfunctional; gene conversion does not appear to be homogenizing the different gene copies. The proposed nonfunctional copies are clustered on particular circles, whereas copies that are likely to code functional gene products are clustered on others.
Journal of Molecular Evolution 05/2007; 64(4):463-71. · 2.15 Impact Factor