RNA interference in parasitic helminths: current situation, potential pitfalls and future prospects

Parasitology ◽  
2006 ◽  
Vol 134 (5) ◽  
pp. 609-619 ◽  
Author(s):  
P. GELDHOF ◽  
A. VISSER ◽  
D. CLARK ◽  
G. SAUNDERS ◽  
C. BRITTON ◽  
...  
Keyword(s):  
Rna Interference ◽  
Culture Conditions ◽  
Gene Knockdown ◽  
Future Application ◽  
Parasitic Nematodes ◽  
Parasitic Helminths ◽  
Rnai Treatment ◽  
Free Living Nematode ◽  
Invaluable Tool

SUMMARYRNA interference (RNAi) has become an invaluable tool for the functional analysis of genes in a wide variety of organisms including the free-living nematode Caenorhabditis elegans. Recently, attempts have been made to apply this technology to parasitic helminths of animals and plants with variable success. Gene knockdown has been reported for Schistosoma mansoni by soaking or electroporating different life-stages in dsRNA. Similar approaches have been tested on parasitic nematodes which clearly showed that, under certain conditions, it was possible to interfere with gene expression. However, despite these successes, the current utility of this technology in parasite research is questionable. First, problems have arisen with the specificity of RNAi. Treatment of the parasites with dsRNA resulted, in many cases, in non-specific effects. Second, the current RNAi methods have a limited efficiency and effects are sometimes difficult to reproduce. This was especially the case in strongylid parasites where only a small number of genes were susceptible to RNAi-mediated gene knockdown. The future application of RNAi in parasite functional genomics will greatly depend on how we can overcome these difficulties. Optimization of the dsRNA delivery methods and in vitro culture conditions will be the major challenges.

Pharmacology of anthelmintic resistance

Parasitology ◽  
1996 ◽  
Vol 113 (S1) ◽  
pp. S201-S216 ◽  
Author(s):  
N. Sangster
Keyword(s):  
Acetylcholine Receptors ◽  
Anthelmintic Resistance ◽  
Cross Resistance ◽  
Parasitic Nematodes ◽  
Parasitic Helminths ◽  
Nematode Parasites ◽  
Neuromuscular Systems ◽  
Free Living Nematode ◽  
Molecular Aspects

SUMMARYAnthelmintic resistance has compromised the control of nematode parasites in several animal-based industries. Studies of resistance have not only improved our understanding of this phenomenon but also shed light on physiological systems of parasitic helminths. In addition, research on molecular aspects of anthelmintic resistance may provide selectable markers for use in future transfection studies with helminths. Several anthelmintics act on helminth neuromuscular systems. Drugs such as levamisole are cholinergic agonists and, based on pharmacological studies, levamisole-resistant nematodes appear to have altered acetylcholine receptors. It is likely that anticholinesterase anthelmintics share cross resistance with levamisole. Ivermectin appears to be a glutamate agonist.In vitrostudies of ivermectin-resistant nematodes suggest that IVM receptors are located on pharyngeal and somatic muscle. The free-living nematodeCaenorhabditis elegansmay provide a model for anthelmintic resistance. It has been useful in cloning drug receptors from parasites but differences between its life history and habitat compared with parasitic nematodes may limit its usefulness for studying resistance in these parasites.

scholarly journals Comparison of two in vitro methods for the detection of ivermectin resistance in Haemonchus contortus in sheep

2016 ◽  
Vol 53 (2) ◽  
pp. 120-125 ◽  
Author(s):  
M. Urda Dolinská ◽  
A. Königová ◽  
M. Babják ◽  
M. Várady
Keyword(s):  
Drug Resistance ◽  
Haemonchus Contortus ◽  
Anthelmintic Resistance ◽  
Economic Losses ◽  
Parasitic Nematodes ◽  
Migration Inhibition ◽  
In Vitro Methods ◽  
Parasitic Helminths

SummaryGastrointestinal parasitic nematodes in sheep cause severe economic losses. Anthelmintics are the most commonly used drugs for prophylaxis and therapy against parasitic helminths. The problem of drug resistance has developed for all commercially available anthelmintics in several genera and classes of helminths. In vitro and in vivo tests are used to detect anthelmintic resistance. Two in vitro methods (larval migration inhibition test and micromotility test) for the detection of ivermectin (IVM) resistance were compared using IVM-resistant and IVM-susceptible isolates of Haemonchus contortus. The degree of resistance for each test was expressed as a resistance factor (RF). The micromotility test was more sensitive for quantitatively measuring the degree of resistance between susceptible and resistant isolates. The RFs for this test for IVM and eprinomectin ranged from 1.00 to 108.05 and from 3.87 to 32.32, respectively.

The diagnostic significance of the induced acrosome reaction

1994 ◽  
Vol 3 (3) ◽  
pp. 159-178 ◽  
Author(s):  
Christopher J De Jonge
Keyword(s):  
Male Infertility ◽  
Acrosome Reaction ◽  
Culture Conditions ◽  
Human Spermatozoa ◽  
Future Application ◽  
Clinical Implications ◽  
General Description ◽  
Diagnostic Significance ◽  
Comprehensive Literature Review

In trying to discern potential causes of male infertility, little investigation has been done to determine if there is predictive value in assessing acrosome reaction inducibility of human spermatozoa for clinical interpretation and application. The purpose of this review is to outline what is currently known in terms of the diagnostic significance of the induced acrosome reaction and to discuss what might be anticipated for the future. In constructing this review, every effort has been made to provide a comprehensive literature review for the reader and a number of review articles have been cited to serve as sources for additional related material. Regarding the aforementioned, it is essential to qualify that, with the exception of the Section entitled: ‘General characteristics of capacitation and the acrosome reaction’, only characteristics, properties and areas relevant to human spermatozoa will be addressed. Thus, in total, this review will: (1) provide a general description of capacitation and the acrosome reaction; (2) briefly characterize the acrosome and its properties; (3) address the signalling mechanism(s) by which the acrosome reaction occurs; (4) outline various methods for detecting the acrosome reaction; (5) define in vitro culture conditions that facilitate the acrosome reaction; (6) define agents, i.e., biological and chemical, that induce the reaction; (7) discuss the association of the acrosome reaction with fertilization; (8) discuss the clinical implications of the induced acrosome reaction; (9) discuss the relevance of sperm function assays in general; and lastly, (10) discuss future application of acrosome reaction assessment for the clinical diagnosis of male infertility.

Morphological observations and the effects of artificial digestive fluids on the survival of Diploscapter coronata from a Japanese patient

2006 ◽  
Vol 80 (4) ◽  
pp. 341-348 ◽  
Author(s):  
N. Morimoto ◽  
M. Korenaga ◽  
K. Yagyu ◽  
N. Kagei ◽  
M. Fujieda ◽  
...  
Keyword(s):  
Intestinal Tract ◽  
Gastric Fluid ◽  
Culture Technique ◽  
Parasitic Nematodes ◽  
Japanese Female ◽  
Old Japanese ◽  
Free Living Nematode ◽  
Gastro Intestinal Tract ◽  
Schonlein Purpura

AbstractUnusual non-human parasitic nematodes and eggs were detected in the faeces of an 8-year-old Japanese female suffering from Henoch-Schönlein purpura. The worms were adult female rhabditiform nematodes measuring 325.6–441.2 μm in length and 18.3–26.5 μm in width. One pair of the labia oris was notched with many spiny projections, while the other pair was strongly curved outwards. The worms were identified using light and scanning electron microscopy as the free-living nematode Diploscapter coronata (Cobb) based on their characteristic morphology. The patient's faeces containing worms and eggs were cultured using a filter-paper culture technique and after 7 days of culture, male as well as female worms were recovered. Worm survival time and hatchability of the eggs were examined in vitro after treatment with an artificial gastric or intestinal fluid. Although adult worms survived for less than one minute, eggs hatched after treatment with artificial gastric fluid. This suggests that eggs accidentally ingested or produced by adult D. coronata could develop in the human gastro-intestinal tract. Some morphological features of male D. coronata are also described.

The Use of RNA Interference in Enhancing Plant Resistance against Nematodes

2020 ◽  
Vol 2 (1) ◽  
Author(s):  
Siye Chen
Keyword(s):  
Rna Interference ◽  
Plant Resistance ◽  
Parasitic Nematodes ◽  
Great Success ◽  
Plant Parasitic Nematodes ◽  
In Planta ◽  
Rnai Technology ◽  
Wide Range ◽  
Plant Parasitic

Plant-parasitic nematodes caused severe yield loss in major crops all over the world. The most wild-used strategies to combat the nematodes is the chamical nematicides, but the overuse of synthetic nematicides threaten sustainable agriculture development. Other strategies, like resistance cultivars and crop rotation, have limited efficiency. Thus, the utilization of molecular biotechnology like RNA interference (RNAi) would be one of the alternative ways to enhance plant resistance against nematodes. RNAi has already used as a tool for gene functional analysis in a wide range of species, especially in the non-parasitic nematode, Caenorhabtidis elegans. In plant-parasitic nematodes, RNAi is induced by soaking nematodes with double-strand RNA(dsRNA) solution mixed with neurostimulants, which is called in vitro RNAi delivery method. In another way around, in planta RNAi method, which is Host-mediated RNAi approach also showed a great success in conferring the resistance against root-knock nematodes. Two main advantages of RNAi-based transgenics are RNAi technology do not produce any functional foreign proteins and it target organisms in a sequence-specific way. Even though the development of RNAi-based transgenics against plant-parasitic nematodes is still in the initial phase, it offers the prospect into a novel nematode control strategy in the future.

RNA interference in plant parasitic nematodes: a summary of the current status

Parasitology ◽  
2012 ◽  
Vol 139 (5) ◽  
pp. 630-640 ◽  
Author(s):  
C. J. LILLEY ◽  
L. J. DAVIES ◽  
P. E. URWIN
Keyword(s):  
Rna Interference ◽  
Gene Silencing ◽  
Cell Types ◽  
Current Status ◽  
Parasitic Nematodes ◽  
Rnai Phenotype ◽  
Plant Parasitic Nematodes ◽  
Systemic Rnai ◽  
Plant Parasitic

SUMMARYRNA interference (RNAi) has emerged as an invaluable gene-silencing tool for functional analysis in a wide variety of organisms, particularly the free-living model nematode Caenorhabditis elegans. An increasing number of studies have now described its application to plant parasitic nematodes. Genes expressed in a range of cell types are silenced when nematodes take up double stranded RNA (dsRNA) or short interfering RNAs (siRNAs) that elicit a systemic RNAi response. Despite many successful reports, there is still poor understanding of the range of factors that influence optimal gene silencing. Recent in vitro studies have highlighted significant variations in the RNAi phenotype that can occur with different dsRNA concentrations, construct size and duration of soaking. Discrepancies in methodology thwart efforts to reliably compare the efficacy of RNAi between different nematodes or target tissues. Nevertheless, RNAi has become an established experimental tool for plant parasitic nematodes and also offers the prospect of being developed into a novel control strategy when delivered from transgenic plants.

scholarly journals A Transcription Factor DAF5 Functions in Haemonchus Contortus Development

2021 ◽  
Author(s):  
Wenda Di ◽  
Fangfang Li ◽  
Li He ◽  
Chunqun Wang ◽  
Caixian Zhou ◽  
...  
Keyword(s):  
Haemonchus Contortus ◽  
Developmental Stages ◽  
Reproductive Organs ◽  
Bimolecular Fluorescence Complementation ◽  
Parasitic Nematodes ◽  
C Elegans ◽  
Free Living Nematode ◽  
And Function ◽  
Reproductive Processes

Abstract Background: Daf5 (Dauer abnormal formation gene), located in the downstream of DAF-7 signalling pathway, mainly functions in dauer formation and reproductive processes in the free-living nematode Caenorhabditis elegans. Although its structure and function have been studied clearly in C. elegans, it was totally unknown in Haemonchus contortus, a socio-economically important parasitic nematode of gastric ruminants.Methods: Here, we identified and characterized a homologue of Daf5, Hcdaf5 and its inferred product (HcDAF5) in H. contortus. Using an integrated molecular approach, we studied the transcriptional profiles of Hcdaf5 and the anatomical expression of HcDAF5 in H. contortus. RNA interference (RNAi) was performed to explore its function in transition from the exsheathed third-stage larvae (xL3) to the fourth-stage larvae (L4) in vitro. Interaction of HcDAF5 and HcDAF3 (a co-SMAD) was also detected by bimolecular fluorescence complementation system (BiFc) in vitro.Results: Here, we showed that HcDAF5 is a member of the Sno/Ski superfamily. Hcdaf5 was transcribed in all developmental stages of H. contortus, with a significant up-regulation in L3. Immunohistochemistry localized native HcDAF5 to the reproductive organs, cuticle and intestine. RNAi revealed specific siRNAs (small interfering RNA) could retard the xL3 development. In addition, the interaction between HcDAF5 and HcDAF3 indicated the SDS box region of HcDAF5 is dispensable for the binding of HcDAF5 to HcDAF3 and the region in HcDAF3 that binds to HcDAF5 is MH2 domain.Conclusion: In summary, these findings show that Hcdaf5 functions in developmental processes of H. contortus, and this is the first characterization of daf-5 gene in parasitic nematodes.

Getting to the root of neuronal signalling in plant-parasitic nematodes using RNA interference

Nematology ◽  
2007 ◽  
Vol 9 (3) ◽  
pp. 301-315 ◽  
Author(s):  
Steven McMaster ◽  
Susan McKinney ◽  
Aaron Maule ◽  
Michael Kimber ◽  
Colin Fleming ◽  
...  
Keyword(s):  
Rna Interference ◽  
Control Strategies ◽  
Large Family ◽  
Parasitic Nematodes ◽  
Plant Parasitic Nematodes ◽  
In Planta ◽  
Nematode Parasites ◽  
Neuronal Signalling ◽  
Plant Parasitic

AbstractA variety of genes expressed in preparasitic second-stage juveniles (J2) of plant-parasitic nematodes appear to be vulnerable to RNA interference (RNAi) in vitro by coupling double-stranded (ds)RNA soaking with the artificial stimulation of pharyngeal pumping. Also, there is mounting evidence that the in planta generation of nematode-specific double-stranded RNAs (dsRNAs) has real utility in the control of these pests. Although neuronally-expressed genes in Caenorhabditis elegans are commonly refractory to RNAi, we have discovered that neuronally-expressed genes in plant-parasitic nematodes are highly susceptible to RNAi and that silencing can be induced by simple soaking procedures without the need for pharyngeal stimulation. Since most front-line anthelmintics that are used for the control of nematode parasites of animals and humans act to disrupt neuromuscular coordination, we argue that intercellular signalling processes associated with neurons have much appeal as targets for transgenic plant-based control strategies for plant-parasitic nematodes. FMRFamide-like peptides (FLPs) are a large family of neuropeptides which are intimately associated with neuromuscular regulation, and our studies on flp gene function in plant-parasitic nematodes have revealed that their expression is central to coordinated locomotory activities. We propose that the high level of conservation in nervous systems across nematodes coupled with the RNAi-susceptibility of neuronally-expressed genes in plant-parasitic nematodes provides a valuable research tool which could be used to interrogate neuronal signalling processes in nematodes.

scholarly journals RNA Interference of Dual Oxidase in the Plant Nematode Meloidogyne incognita

2005 ◽  
Vol 18 (10) ◽  
pp. 1099-1106 ◽  
Author(s):  
Manjula Bakhetia ◽  
Wayne Charlton ◽  
Howard J. Atkinson ◽  
Michael J. McPherson
Keyword(s):  
Rna Interference ◽  
Gene Function ◽  
Meloidogyne Incognita ◽  
Feeding Strategy ◽  
Model Organisms ◽  
Parasitic Nematodes ◽  
In Planta ◽  
Oxidase Gene ◽  
Dual Oxidase

RNA interference (RNAi) is a powerful tool for the analysis of gene function in model organisms such as the nematode Caenorhabditis elegans. Recent demonstrations of RNAi in plant parasitic nematodes provide a stimulus to explore the potential of using RNAi to investigate disruption of gene function in Meloidogyne incognita, one of the most important nematode pests of global agriculture. We have used RNAi to examine the importance of dual oxidases (peroxidase and NADPH oxidase), a class of enzyme associated with extracellular matrix cross-linking in C. elegans. RNAi uptake by M. incognita juveniles is highly efficient. In planta infection data show that a single 4-h preinfection treatment with double-stranded RNA derived from the peroxidase region of a dual oxidase gene has effects on gene expression that are phenotypically observable 35 days postinfection. This RNAi effect results in a reduction in egg numbers at 35 days of up to 70%. The in vitro feeding strategy provides a powerful tool for identifying functionally important genes, including those that are potential targets for the development of new agrochemicals or transgenic resistance strategies.

Neutrophil migration through in vitro interstitial matrix

1992 ◽  
Vol 50 (1) ◽  
pp. 894-895
Author(s):  
J. Roemer ◽  
S.R. Simon
Keyword(s):  
Extracellular Matrix ◽  
Smooth Muscle ◽  
Cell Migration ◽  
Smooth Muscle Cells ◽  
Inflammatory Cell ◽  
Neutrophil Migration ◽  
Culture Conditions ◽  
Aortic Smooth Muscle ◽  
Specific Culture

We are developing an in vitro interstitial extracellular matrix (ECM) system for study of inflammatory cell migration. Falcon brand Cyclopore membrane inserts of various pore sizes are used as a support substrate for production of ECM by R22 rat aortic smooth muscle cells. Under specific culture conditions these cells produce a highly insoluble matrix consisting of typical interstitial ECM components, i.e.: types I and III collagen, elastin, proteoglycans and fibronectin.

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