Nematode Growth Research Articles

Identification and preliminary characterization of Hc-clec-160, a novel C-type lectin domain-containing gene of the strongylid nematode Haemonchus contortus

BackgroundThe strongylid parasite Haemonchus contortus causes severe anemia in domestic animals worldwide. Effective preventive and therapeutical agents are lacking, because of drug resistance and that little is known about the molecular mechanism of the interaction between H. contortus and host cells.MethodsA new gene, Hc-clec-160, was discovered with RT-PCR. Transcriptional levels of Hc-clec-160 and Ce-clec-160 throughout different growth phases of corresponding nematodes were assayed by qPCR. Immunofluorescence staining of paraffin section were performed to determine the protein localization in adult worms of H. contortus. To monitor the promoter capacity of the 5'-flanking region of Ce-clec-160, micro-injection was used. Overexpression and RNAi constructs was carried out in the N2 strain of Caenorhabditis elegans to find out the gene function of Hc-clec-160.ResultsThe full-length cDNA of 1224 bp of Hc-clec-160 was cloned by RT-PCR. The corresponding gene contained twelve exons. Its transcripts peaked in male adult worms. Hc-CLEC-160 was predicted to have a Willebrand factor type A (vWA) domain and a C-type lectin domain. The proteins were not detected by expression in C. elegans or paraffin section experiments in adult of H. contortus. Knockdown of Ce-clec-160 expression in C. elegans by RNAi resulted in shortened body length and decreased brood size.ConclusionsIn this experiment, a new gene Hc-clec-160 was obtained in H. contortus and its function was addressed using a model organism: C. elegans. Our study showed that Hc-clec-160 possesses characteristics similar to those of Ce-clec-160 and plays an important role in the growth and reproduction of this parasitic nematode.

Evaluation of four rhizobacteria on tomato growth and suppression of root-knot nematode, Meloidogyne javanica under greenhouse conditions, a pilot study

Growth-promoting rhizobacteria are free-living bacteria that colonize the roots and stimulate the plant growth. Many of these bacteria secrete a range of extracellular metabolites that can be involved in the biological control of plant pathogens. In this study, the effect of four plant growth-promoting rhizobacteria (PGPR) was evaluated on tomato growth parameters as well as biological control of the root-knot nematode (RKN) (Meloidogyne javanica). Tomato seedlings were inoculated with four isolates of PGPR including Pseudomonas fluorescens, Pseudomonas striata, Bacillus subtilis, and Paenibacillus polymyxa and cultivated in the presence or absence of RKN. The results showed that the PGPR significantly increased the plant growth parameters. Bacterial strains also significantly affected the reproductive factor of Meloidogyne javanica significantly so that P. fluorescens and B. subtilis reduced the reproductive factor from 112.15 to 24.94 and 24.96, respectively. Based on these results, among applied rhizobacteria in this study, P. polymyxa can be regarded as the best candidate for promoting the growth and biological control of M. javanica in tomato crop under greenhouse conditions.

Differential Selection by Nematodes of an Introduced Biocontrol Fungus vs. Indigenous Fungi in Nonsterile Soil.

Trophic interactions of introduced biocontrol fungi with soil animals can be a key determinant in the fungal proliferation and activity. This study investigated the trophic interaction of an introduced biocontrol fungus with soil nematodes. The biocontrol fungus Trichoderma harzianum ThzID1-M3 and the fungivorous nematode Aphelenchoides sp. (10 per gram of soil) were added to nonsterile soil, and microbial populations were monitored for 40 days. Similar results were obtained when the experiment was duplicated. ThzID1-M3 stimulated the population growth of indigenous nematodes (p < 0.05), regardless of whether Aphelenchoides sp. was added. Without ThzID1-M3, indigenous nematodes did not increase in number and the added Aphelenchoides sp. nematodes almost disappeared by day 10. With ThzID1-M3, population growth of nematodes was rapid between 5 and 10 days after treatment. ThzID1-M3 biomass peaked on day 5, dropped at day 10, and then almost disappeared at day 20, which was not influenced by the addition of nematodes. In contrast, a large quantity of ThzID1-M3 hyphae were present in a heat-treated soil in which nematodes were eliminated. Total fungal biomass in all treatments peaked on day 5 and subsequently decreased. Addition of nematodes increased the total fungal biomass (p < 0.05), but ThzID1-M3 addition did not affect the fungal biomass. Hyphae of total fungi when homogenously distributed did not support the nematode population growth; however, hyphae of the introduced fungus did when densely localized. The results suggest that soil fungivorous nematodes are an important constraint on the hyphal proliferation of fungal agents introduced into natural soils.

Chronic toxicity of hexabromocyclododecane(HBCD) induced by oxidative stress and cell apoptosis on nematode Caenorhabditis elegans

In order to gain insights into the chronic effects and mechanisms of hexabromocyclododecane (HBCD), the animal model Caenorhabditis elegans (C. elegans) was chosen for toxicity study. Multiple endpoints, including the physiological (growth and locomotion behaviors), biochemical (reactive oxygen species (ROS) production, lipofuscin accumulation, and cell apoptosis), and molecular (stress-related gene expressions) levels, were tested by chronic exposure for 10 d to low concentrations of HBCD (0.2 nM-200 nM). The results revealed that chronic exposure to HBCD at concentrations more than 20 nM would significantly influence the growth, locomotion behaviors, ROS formation, lipofuscin accumulation, and cell apoptosis of nematodes. Treatment with antioxidants of ascorbate and N-acetyl-l-cysteine (NAC) suppressed the toxicity induced by HBCD. The integrated gene expression profiles showed that the chronic exposure to 200 nM of HBCD significantly increased the expression levels of stress-related genes (e.g., hsp-16.2, hsp-16.48, sod-1, sod-3, and cep-1 genes). Among these genes, the sod-1, sod-3, and cep-1 gene expressions were significantly correlated with HBCD-induced physiological effects by the Pearson correlation test. The mutations of sod-3 and cep-1 induced more severe toxicity compared to wild-type nematodes. Therefore, HBCD exposure induced oxidative stress by ROS accumulation and cell apoptosis, which resulted in HBCD-induced toxicity on nematodes, and sod-3 and cep-1 played important roles in protecting nematodes against HBCD-induced toxicity.

Development of Phasmarhabditis hermaphrodita (and members of the Phasmarhabditis genus) as new genetic model nematodes to study the genetic basis of parasitism.

The genetic mechanisms of how free-living nematodes evolved into parasites are unknown. Current genetic model nematodes (e.g. Caenorhabditis elegans) are not well suited to provide the answer, and mammalian parasites are expensive and logistically difficult to maintain. Here we propose the terrestrial gastropod parasite Phasmarhabditis hermaphrodita as a new alternative to study the evolution of parasitism, and outline the methodology of how to keep P. hermaphrodita in the lab for genetic experiments. We show that P. hermaphrodita (and several other Phasmarhabditis species) are easy to isolate and identify from slugs and snails from around the UK. We outline how to make isogenic lines using 'semi-natural' conditions to reduce in-lab evolution, and how to optimize growth using nematode growth media (NGM) agar and naturally isolated bacteria. We show that P. hermaphrodita is amenable to forward genetics and that unc and sma mutants can be generated using formaldehyde mutagenesis. We also detail the procedures needed to carry out genetic crosses. Furthermore, we show natural variation within our Phasmarhabditis collection, with isolates displaying differences in survival when exposed to high temperatures and pH, which facilitates micro and macro evolutionary studies. In summary, we believe that this genetically amenable parasite that shares many attributes with C. elegans as well as being in Clade 5, which contains many animal, plant and arthropod parasites, could be an excellent model to understand the genetic basis of parasitism in the Nematoda.

Rapid environmental effects on gut nematode susceptibility in rewilded mice.

Genetic and environmental factors shape host susceptibility to infection, but how and how rapidly environmental variation might alter the susceptibility of mammalian genotypes remains unknown. Here, we investigate the impacts of seminatural environments upon the nematode susceptibility profiles of inbred C57BL/6 mice. We hypothesized that natural exposure to microbes might directly (e.g., via trophic interactions) or indirectly (e.g., via microbe-induced immune responses) alter the hatching, growth, and survival of nematodes in mice housed outdoors. We found that while C57BL/6 mice are resistant to high doses of nematode (Trichuris muris) eggs under clean laboratory conditions, exposure to outdoor environments significantly increased their susceptibility to infection, as evidenced by increased worm burdens and worm biomass. Indeed, mice kept outdoors harbored as many worms as signal transducer and activator of transcription 6 (STAT6) knockout mice, which are genetically deficient in the type 2 immune response essential for clearing nematodes. Using 16S ribosomal RNA sequencing of fecal samples, we discovered enhanced microbial diversity and specific bacterial taxa predictive of nematode burden in outdoor mice. We also observed decreased type 2 and increased type 1 immune responses in lamina propria and mesenteric lymph node (MLN) cells from infected mice residing outdoors. Importantly, in our experimental design, different groups of mice received nematode eggs either before or after moving outdoors. This contrasting timing of rewilding revealed that enhanced hatching of worms was not sufficient to explain the increased worm burdens; instead, microbial enhancement and type 1 immune facilitation of worm growth and survival, as hypothesized, were also necessary to explain our results. These findings demonstrate that environment can rapidly and significantly shape gut microbial communities and mucosal responses to nematode infections, leading to variation in parasite expulsion rates among genetically similar hosts.

Ratiometric Calcium Imaging of Individual Neurons in Behaving &lt;em&gt;Caenorhabditis Elegans&lt;/em&gt;

It has become increasingly clear that neural circuit activity in behaving animals differs substantially from that seen in anesthetized or immobilized animals. Highly sensitive, genetically encoded fluorescent reporters of Ca2+ have revolutionized the recording of cell and synaptic activity using non-invasive optical approaches in behaving animals. When combined with genetic and optogenetic techniques, the molecular mechanisms that modulate cell and circuit activity during different behavior states can be identified. Here we describe methods for ratiometric Ca2+ imaging of single neurons in freely behaving Caenorhabditis elegans worms. We demonstrate a simple mounting technique that gently overlays worms growing on a standard Nematode Growth Media (NGM) agar block with a glass coverslip, permitting animals to be recorded at high-resolution during unrestricted movement and behavior. With this technique, we use the sensitive Ca2+ reporter GCaMP5 to record changes in intracellular Ca2+ in the serotonergic Hermaphrodite Specific Neurons (HSNs) as they drive egg-laying behavior. By co-expressing mCherry, a Ca2+-insensitive fluorescent protein, we can track the position of the HSN within ~ 1 µm and correct for fluctuations in fluorescence caused by changes in focus or movement. Simultaneous, infrared brightfield imaging allows for behavior recording and animal tracking using a motorized stage. By integrating these microscopic techniques and data streams, we can record Ca2+ activity in the C. elegans egg-laying circuit as it progresses between inactive and active behavior states over tens of minutes.

Ratiometric Calcium Imaging of Individual Neurons in Behaving &lt;em&gt;Caenorhabditis Elegans&lt;/em&gt;

It has become increasingly clear that neural circuit activity in behaving animals differs substantially from that seen in anesthetized or immobilized animals. Highly sensitive, genetically encoded fluorescent reporters of Ca2+ have revolutionized the recording of cell and synaptic activity using non-invasive optical approaches in behaving animals. When combined with genetic and optogenetic techniques, the molecular mechanisms that modulate cell and circuit activity during different behavior states can be identified.Here we describe methods for ratiometric Ca2+ imaging of single neurons in freely behaving Caenorhabditis elegans worms. We demonstrate a simple mounting technique that gently overlays worms growing on a standard Nematode Growth Media (NGM) agar block with a glass coverslip, permitting animals to be recorded at high-resolution during unrestricted movement and behavior. With this technique, we use the sensitive Ca2+ reporter GCaMP5 to record changes in intracellular Ca2+ in the serotonergic Hermaphrodite Specific Neurons (HSNs) as they drive egg-laying behavior. By co-expressing mCherry, a Ca2+-insensitive fluorescent protein, we can track the position of the HSN within ~ 1 µm and correct for fluctuations in fluorescence caused by changes in focus or movement. Simultaneous, infrared brightfield imaging allows for behavior recording and animal tracking using a motorized stage. By integrating these microscopic techniques and data streams, we can record Ca2+ activity in the C. elegans egg-laying circuit as it progresses between inactive and active behavior states over tens of minutes.

5,7-Dihydroxy-4-Methoxyflavone a bioactive flavonoid delays amyloid beta-induced paralysis and attenuates oxidative stress in transgenic Caenorhabditis elegans

Background: The various herbal remedies have been used in Ayurveda which symbolizes traditional medicine system since ancient times. The increasing popularity of herbal medicines has prompted us toward the development of natural therapeutics for preventing neurodegenerative disease such as Alzheimer's disease (AD) in living organisms. This study focused on a flavonoid compound 5,7-dihydroxy-4-methoxyflavone (DMF) also known as acacetin which is a major constituent of Premna odorata (L.) plant. This bioactive flavonoid exhibits several medicinal properties such as antimicrobial, anti-inflammatory, antioxidant, and anti-carcinogenic. Objective: The aim is to determine the protective effects of DMF against amyloid beta (Aβ)-induced toxicity and oxidative stress in transgenic Caenorhabditis elegans model of AD. Materials and Methods: The therapeutic potential of DMF treatments (5, 25, and 50 μM) was investigated to counteract Aβ paralysis and oxidative stress through paralysis assay, reactive oxygen species (ROS) detection, protein carbonylation, aldicarb assay, and mRNA quantification using transgenic C. elegans model of AD. Results: The present study reports that DMF effectively delayed Aβ-induced paralysis, attenuated ROS level reduced protein carbonylation and conferred aldicarb resistance. In addition, DMF was also found to up-regulate the expression of stress modulating (sod-1, sod-2, sod-3, ctl-1, hsp-16.2, and gst-4), acetylcholine transporter(unc-17), regulator of nicotinic acetylcholine receptor (unc-50), and choline acetyltransferase (cha-1) related genes. Conclusion: These findings suggest DMF may provide protection against Aβ toxicity and oxidative stress due to its antioxidant activity. Therefore, the bioactive flavonoid DMF may provide invaluable medicinal and health benefits which can delay the onset of age-related diseases. Abbreviations used: DMF: 5, 7-dihydroxy-4-methoxyflavone; ROS: Reactive oxygen species; C. elegans: Caenorhabditis elegans; AD: Alzheimer's disease; Aβ: Amyloid beta; DMSO: Dimethyl sulphoxide; NGM: Nematode growth media; ACh: Acetylcholine; Ald: Aldicarb; APP: Amyloid precursor protein.

Molecular genetic and biochemical characterization of a putative family of zinc metalloproteins in Caenorhabditis elegans.

Four highly similar genes (W08E12.2, W08E12.3, W08E12.4 and W08E12.5) which are consecutively aligned on chromosome IV of the C. elegans genome are predicted to code for small (120-141aa) yet cysteine rich (18-19 cysteines) proteins. Cloning and sequencing of the genomic regions of the isoforms confirmed the presence and order of all genes. The generation of transgenic worms strains with an integrated single copy or extrachromosomal multi-copy PW08E12.3;W08E12.4::GFP uncovered that W08E12.3 and W08E12.4 are constitutively expressed in the pharynx and significantly induced in worms exposed to 100 μM Zn. Knockdown by RNAi did not have a marked consequence on reproductive performance nor was a Zn-dependent effect on nematode growth observed. However, RNAi of these genes led to an accumulation of Zn in the intestinal cells. W08E12.3 was recombinantly expressed in E. coli and the purified protein was shown to be able to bind up to 6.5 Zn molecules at neutral pH. Zn-binding was acid-labile and the apo protein was observed at pH < 4.3. This characterization suggests W08E12.2, W08E12.3, W08E12.4 and W08E12.5 belong to a family of putative Metalloproteins which, akin to metallothioneins, may play an important role in Zn-sensing, homeostasis and/or detoxification.

Antioxidant Activity of Water Extract from Fermented Mycelia of Cordyceps sobolifera (Ascomycetes) in Caenorhabditis elegans.

This research aimed to evaluate the potential of Cordyceps sobolifera in mycelial biomass production via liquid culture and to assay the safety and determine the antioxidative and antiaging activities of Caenorhabditis elegans. A C. sobolifera isolate was cultured using the one-factor-at-a-time method to illustrate its carbon and nitrogen requirements. To assess safety, we determined the lethality, locomotion behavior, and reproduction of C. elegans cultured on a mycelial water extract (MWE) containing nematode growth medium (NGM). To investigate antiaging activity, C. elegans treated with MWE was incubated on NGM plates. The lethality was recorded throughout the whole life cycle. To identify antioxidant activity, C. elegans treated with MWE was exposed to paraquat, causing superoxide conditions. The results showed that C. sobolifera was favored by glucose and peptone as carbon and nitrogen sources, respectively. MWE was considered to be safe, as no abnormal behaviors were observed in C. elegans. Compared with nematodes pretreated with no MWE but with water instead, MWE at 1.0 mg/mL significantly prolonged the mean lifespan of C. elegans by 24%. We observed an obvious dose-effect relation between concentration and mean lifespan. The effective antioxidant activity was recorded at the high concentration of MWE. These findings demonstrate the potential antiaging and antioxidant properties of C. sobolifera as functional food and dietary supplement.

Effect of soil texture on infectivity of root knot nematode (Meloidogyne incognita) in chillies

The study was conducted to examine infectivity (penetration, gall & egg-mass formations) of the root-knot nematode, Meloidogyne incognita on chilli grown in 6 different soil textures viz sandy, red sodic, clay, sandy loam soils and pot mixture. The nematode penetrations in sand soil were significantly higher than other. However, gall and eggmass formations were remarkably increased in the pot mixture followed by sandy loam and sandy loam soils which was coincided with the general aspects of severe nematode infestations in sandy soil. This suggested the increased gall and egg-mass formations of M. incognita derived from the increased penetration in the sandy soil conditions, which provided sufficient aeration due to coarse soil nature for the nematodes, leading to their mobility increased for the enhanced root penetration. It is suggested that the sandy soil conditions affected positively on the healthy plant growth with little accumulation of the inhibitory materials and sufficient aeration, enhancing the nematode growth and feeding activities.

Role of Indigenous Rhizosphere Bacteria in Suppressing Root-knot Nematode and Improve Plant Growth Tomato

The effect of ionic osmotic potential (Ψ), and matric potential (Ψm) in the range of -0.2 to -4.0 Mpa on mycelial growth of three species of Pleurotus (P.florida, P.ostrenatus and P.safor-caju) were determined over a range of temperature (15-3) on a 2% malt extract agar medium and compared with the Ψ effect on growth of two strains of Trichoderma green mould. With the ionic solute KCl, optimun Ψfor growth was -0.2 MPa for P.floreda and in the range of -0.2 to -0.5 MPa, with slight growth at -3.0 MPa and with nogrowth at -4.0 MPa. Of the species of Pleurotus, P.florida grew signigicantly slower than the other two species. Growt of the species of Pleurocus was significantly slower when water potential (Ψ) was modified matrically with polyethylene glycol (PEG) 8000 then osmotically with KCl. They were also more sensitive to changes in Ψm than ΨThe optimum Ψm of the Pleurotus was -0.5 Ψm, with no growth below -3.0 MPa. Of the species of Pleurotus, P.florida was most sensitive and P.sajor-caju was more tolerent to lowered Ψ,but P.sajor-caju was most sensitive to lowered Ψm. The growth rate of the Trichoderma green mould strains was much faster than that observed for the Pleurotus spp. Optimum growth for bot strains of Trichoderma was in the range of -0.2 to -0.5 MPa. Strain CNU 503 was more tolerant to water stress than strain CNU 501. Both strains were able to grow up to 30% of optimum growth at -4.0 MPa at 25-3.

Changes emerging with calcinerin inhibition in Caenorhabditis elegans individuals as an aging model

Objective: It is known that calcineurin inhibition prolongs life in individuals with Caenorhabditis elegans ( C. elegans ). An important part of C. elegans genes shows great similarity to human genes. This study aims to evaluate the effects of a calcineurin inhibitor on vital functions in C. elegans individuals. Method: In this study, solutions of cyclosporine A which is a calcineurin inhibitör 1 μM (1.dose), 0.1 μM (2.dose), 0.01 μM (3.dose) and 0.001 μM (4.dose) were prepared and 1 ml from each solution was taken and added to petri dishes, containing 10 ml Nematode Growth Media (NGM). The control group was fed the medium which was not added cyclosporine. The indicated doses of cyclosporine whose synchronization has been completed were transferred to petri dishes in order that each would have 20 pieces. C. elegans were observed using bottom light stereo microscop that provided 6X-50X magnification. Live animals at the same time every day were counted and compared with the control group until all C. elegans in all petri dishes died. Results: The results have showed that there was a statistically significant increase in life span periods of C. elegans exposed to cyclosporin depending on dose increase (p<0,05). It has been found out that physical development of C. elegans exposed to increased doses of cyclosporin decreased significantly (p<0,05). The increase in doses of cyclosporin have not caused statitically significant change in the number of pharynx pumping (p>0,05). Conclusions: Calcineurin inhibition slows down the developmental process in C. elegans and prolongs survival, and may produce positive effects on aging.

Dynamics of plant parasitic and free living nematode populations in sugarcane &lt;i&gt;Saccharum officinarum&lt;/i&gt; in Fiji

&#x0D; &#x0D; &#x0D; Parasitic nematodes are one of the main constraints in the production of sugarcane, Saccharum officinarum, in Fiji and other sugarcane-growing countries. Both plant parasitic and free-living nematodes are associated with roots of the plant, and the level of infestation affects both soil health and cane yield. This study examined the dynamics of plant parasitic and free-living nematode populations in sugarcane crops in Drasa Estate, Lautoka, Fiji. Soil samples to quantify nematode populations were collected ve times during the crop cycle. There was an exponential increase in the plant parasitic nematode population with time after planting (nematode population=64.316e0.0112x; R2=0.56, x=days after planting). However, the increase was not consistent among the different nematode groups. Significant increases in Lesion (P&lt;0.05), Rootknot (P&lt;0.05), Spiral (P&lt;0.01), and Ring (P&lt;0.03) nematode populations were found but not for Reniforms (P&gt;0.05). Populations of free-living nematodes built up 90 days after planting and remained elevated in the ratoon crop. Populations of parasitic nematodes increased with the growth of cane (as measured by cane height) with increases occurring faster in the ratoon than in the plant crop (P=0.001). The number of free-living nematodes may have affected population growth of parasitic nematodes.&#x0D; &#x0D; &#x0D;

Effects of simulated microgravity on gene expression and biological phenotypes of a single generation Caenorhabditis elegans cultured on 2 different media

Studies of multigenerational Caenorhabditis elegans exposed to long-term spaceflight have revealed expression changes of genes involved in longevity, DNA repair, and locomotion. However, results from spaceflight experiments are difficult to reproduce as space missions are costly and opportunities are rather limited for researchers. In addition, multigenerational cultures of C. elegans used in previous studies contribute to mixture of gene expression profiles from both larvae and adult worms, which were recently reported to be different. Usage of different culture media during microgravity simulation experiments might also give rise to differences in the gene expression and biological phenotypes of the worms. In this study, we investigated the effects of simulated microgravity on the gene expression and biological phenotype profiles of a single generation of C. elegans worms cultured on 2 different culture media. A desktop Random Positioning Machine (RPM) was used to simulate microgravity on the worms for approximately 52 to 54 h. Gene expression profile was analysed using the Affymetrix GeneChip® C. elegans 1.0 ST Array. Only one gene (R01H2.2) was found to be downregulated in nematode growth medium (NGM)-cultured worms exposed to simulated microgravity. On the other hand, eight genes were differentially expressed for C. elegans Maintenance Medium (CeMM)-cultured worms in microgravity; six were upregulated, while two were downregulated. Five of the upregulated genes (C07E3.15, C34H3.21, C32D5.16, F35H8.9 and C34F11.17) encode non-coding RNAs. In terms of biological phenotype, we observed that microgravity-simulated worms experienced minimal changes in terms of lifespan, locomotion and reproductive capabilities in comparison with the ground controls. Taking it all together, simulated microgravity on a single generation of C. elegans did not confer major changes to their gene expression and biological phenotype. Nevertheless, exposure of the worms to microgravity lead to higher expression of non-coding RNA genes, which may play an epigenetic role in the worms during longer terms of microgravity exposure.

HexA is a versatile regulator involved in the control of phenotypic heterogeneity of Photorhabdus luminescens.

Phenotypic heterogeneity in microbial communities enables genetically identical organisms to behave differently even under the same environmental conditions. Photorhabdus luminescens, a bioluminescent Gram-negative bacterium, contains a complex life cycle, which involves a symbiotic interaction with nematodes as well as a pathogenic association with insect larvae. P. luminescens exists in two distinct phenotypic cell types, designated as the primary (1°) and secondary (2°) cells. The 1° cells are bioluminescent, pigmented and can support nematode growth and development. Individual 1° cells undergo phenotypic switching after prolonged cultivation and convert to 2° cells, which lack the 1° specific phenotypes. The LysR-type regulator HexA has been described as major regulator of this switching process. Here we show that HexA controls phenotypic heterogeneity in a versatile way, directly and indirectly. Expression of hexA is enhanced in 2° cells, and the corresponding regulator inhibits 1° specific traits in 2° cells. HexA does not directly affect bioluminescence, a predominant 1° specific phenotype. Since the respective luxCDABE operon is repressed at the post-transcriptional level and transcriptional levels of the RNA chaperone gene hfq are also enhanced in 2° cells, small regulatory RNAs are presumably involved that are under control of HexA. Another phenotypic trait that is specific for 1° cells is quorum sensing mediated cell clumping. The corresponding pcfABCDEF operon could be identified as the first direct target of HexA, since the regulator binds to the pcfA promoter region and thereby blocks expression of the target operon. In summary, our data show that HexA fulfills the task as repressor of 1° specific features in 2° cells in a versatile way and gives first insights into the complexity of regulating phenotypic heterogeneity in Photorhabdus bacteria.

Climate change is predicted to alter the current pest status of Globodera pallida and G.rostochiensis in the United Kingdom.

The potato cyst nematodes Globodera pallida and G. rostochiensis are economically important plant pathogens causing losses to UK potato harvests estimated at £50 m/ year. Implications of climate change on their future pest status have not been fully considered. Here, we report growth of female G. pallida and G. rostochiensis over the range 15 to 25°C. Females per plant and their fecundity declined progressively with temperatures above 17.5°C for G. pallida, whilst females per plant were optimal between 17.5 and 22.5°C for G. rostochiensis. Relative reproductive success with temperature was confirmed on two potato cultivars infected with either species at 15, 22.5 and 25°C. The reduced reproductive success of G. pallida at 22.5°C relative to 15°C was also recorded for a further seven host cultivars studied. The differences in optimal temperatures for reproductive success may relate to known differences in the altitude of their regions of origin in the Andes. Exposure of G. pallida to a diurnal temperature stress for one week during female growth significantly suppressed subsequent growth for one week at 17.5°C but had no effect on G. rostochiensis. However, after two weeks of recovery, female size was not significantly different from that for the control treatment. Future soil temperatures were simulated for medium‐ and high‐emission scenarios and combined with nematode growth data to project future implications of climate change for the two species. Increased soil temperatures associated with climate change may reduce the pest status of G. pallida but benefit G. rostochiensis especially in the southern United Kingdom. We conclude that plant breeders may be able to exploit the thermal limits of G. pallida by developing potato cultivars able to grow under future warm summer conditions. Existing widely deployed resistance to G. rostochiensis is an important characteristic to retain for new potato cultivars.

Effects of Soil Textures on Infectivity of Root-Knot Nematodes on Carrot.

This study was conducted to examine infectivity (penetration and gall and egg-mass formations) of the root-knot nematodes, Meloidogyne incognita and M. hapla, on carrots grown in soil conditions of 5 different soil textures consisting of bed-soil (b) and sand (s) mixtures (b-s mixtures) at the ratios of 10:0, 7:3, 5:5, 3:7, and 0:10. For M. incognita, the nematode penetration rates in b-s of 0:10 (100% sand) were significantly higher than in the other b-s mixtures, more greatly at 2 and 5 days after inoculation than at 10 DAI, while no significant differences in the penetration rates were mostly shown for M. hapla at the above DAI. However, for both nematodes, gall and egg-mass formations were remarkably increased in the b-s mixture of 0:10, compared to the other b-s mixtures, which is coincided with the general aspects of severe nematode infestations in sandy soils. This suggests the increased gall and egg-mass formations of M. incognita should be derived from the increased penetration rates in the sandy soil conditions, which provide a sufficient aeration due to coarse soil nature for the nematodes, leading to their mobility increased for the enhanced root penetration. For M. hapla, it is suggested that the sandy soil conditions affect positively on the healthy plant growth with little accumulation of the inhibitory materials and sufficient aeration, enhancing the nematode growth and feeding activities. All of these aspects provide information reliable for the development screening techniques efficient for the evaluation of the nematode resistance in the breeding programs.

Screening of lentil (Lens culinaris Medik.) germplasm for resistance to root-knot nematode, Meloidogyne incognita

The root-knot nematode is one of the major limiting factors affecting lentil growth and yield. In the present study, 300 accessions of lentil (Lens culinaris Medik) were screened to find source of resistance to root-knot nematode, Meloidogyne incognita. Among them, nine accessions: EC223269, EC076551-C, EC267577-D, EC267555, EC255504, EC267690, IC282663, IC559890 and IC559673 were resistant, while 291 were susceptible to M. incognita. Root gall formation was reduced by 94–98% in the resistant accessions as compared to susceptible (IC560206). The nematode penetration into roots and egg-mass formation in all resistant accessions of lentil were significantly lowered compared to the susceptible accession (IC560206). Results suggest that the disease resistance in lentil accessions may be related to both post-infectional (nematode growth and development) as well as pre-infectional (penetration and establishment) defense mechanisms. Variations in nematode penetration into roots, root gall formation and egg mass formation in the resistant accessions may reflect their genetic differences related to the nematode resistance and their planting could provide a useful tool to manage root-knot nematodes in lentil crop.