Insect Immune Evasion by Dauer and Nondauer Entomopathogenic Nematodes.

The immune response of animals, including insects, is overcome by some parasites. For example, dauer larvae (DL) of the obligate entomopathogenic nematodes (EPNs) Heterorhabditis and Steinernema can invade insects, evade their defenses, and cause death. Although DL were long assumed to be the only infective stage of nematodes, recent reports suggest that L2-L3 larvae of facultative EPNs are also capable of killing insects. There are no studies, to our knowledge, about the role of nonimmunological barriers (the exoskeleton and its openings) in avoiding infection by DL and L2-L3 larvae, or whether these larval stages evade the host immune system in the same way. The objective of this study was to examine these questions by infecting Galleria mellonella with the facultative parasitic nematode Rhabditis regina. DL or L2-L3 larvae were either deposited on or near the moths or injected into their hemocoel. Once nematodes reached the hemocoel, the following host immune response parameters were quantified: prophenoloxidase, phenoloxidase, lytic activity, and the number of granular hemocytes. DL showed a greater ability to penetrate the exoskeleton than L2-L3 larvae. Once inside, however, both went unnoticed by the immune system and killed the insect. A higher number of granular hemocytes was activated by L2-L3 larvae than DL. We show for the first time that L2-L3 larvae can penetrate and evade the insect immune system. Further research is needed to compare facultative and specialized EPNs to determine which is more likely, with both DL and L2-L3 larvae, to evade insect defense barriers and produce death. The results will contribute to understanding the evolution of virulence in entomopathogenic nematodes.

Meta-analysis of cyathostomin species-specific prevalence and relative abundance in domestic horses from 1975-2020: emphasis on geographical region and specimen collection method.

BACKGROUND: Cyathostomins infect virtually all horses, and concomitant infections with 10 or more species per horse is standard. Species-specific knowledge is limited, despite potential species bias in development of disease and anthelmintic resistance. This is the first meta-analysis to examine effects of geographical region and cyathostomin collection method on reported composition of cyathostomin communities. METHODS: Thirty-seven articles published in English in 1975 or later, in which adults of individual species were systematically enumerated, were included. Seven regions; North America, South America, eastern Europe, western Europe, northern Europe, southern Africa, and Oceania, and three cyathostomin collection methods; (i) standard necropsy recovery from the large intestine, (ii) critical test collection from post-treatment feces and necropsy, and (iii) diagnostic deworming recovery solely from post-treatment feces, were considered. Generalized mixed linear models analyzed the effects of region and collection method on species-specific prevalence and relative abundance. Species richness was analyzed by mixed linear models. RESULTS: Definitively, the most prevalent and relatively abundant species were Cylicocyclus nassatus (prevalence = 93%, relative abundance = 20%), Cylicostephanus (Cys.) longibursatus (93%, 20%), and Cyathostomum catinatum (90%, 16%). A bias toward horses with high infection intensities and cyathostomin collection from feces resulted in North American critical tests and eastern European diagnostic deworming overestimating the species-specific prevalence and underestimating the relative abundance of rare/uncommon species compared to respective intra-regional standard necropsies. North American critical tests underestimated species richness due partially to identification key errors. Inter-regional standard necropsy comparisons yielded some species-specific regional differences, including a significantly higher Cys. longibursatus prevalence and relative abundance in North America (92%, 33%) than in eastern Europe (51%, 7%) (P > 0.0001). Localization of critical tests to North America and diagnostic deworming to Eastern Europe precluded expansive 'region by collection method' interaction analyses. CONCLUSION: We provide substantial data to inform study design, e.g. effect and study size, for cyathostomin research and highlight necessity for method standardization and raw data accessibility for optimal post-factum comparisons.

Control of Strongyloides westeri by nematophagous fungi after passage through the gastrointestinal tract of donkeys / Controle de Strongyloides westeri por fungos nematófagos após trânsito gastrintestinal em jumentas

Strongyloides westeri is the most prevalent nematode among equines aged up to four months and causes gastrointestinal disorders. The objective of this study was to observe the control of infective S. westeri larvae (L3) by the nematophagous fungi Duddingtonia flagrans (AC001) and Monacrosporium thaumasium (NF34) after passage through the gastrointestinal tract of female donkeys. Twelve dewormed female donkeys that were kept in stables were used. Two treatment groups each comprising four animals received orally 100 g of pellets made of sodium alginate matrix containing a mycelial mass of either D. flagrans (AC001) or M. thaumasium (NF34). The control group consisted of four animals that received pellets without fungus. Feces samples were then collected from the animal groups at different times (after 12, 24, 48 and 72 hours). These feces were placed in Petri dishes containing 2% water-agar medium and 1000 L3 of S. westeri. AC001 and NF34 isolates showed the ability to destroy the L3, after gastrointestinal transit, thus demonstrating their viability and predatory activity.

O Strongyloides westeri é o nematóide de maior prevalência entre equídeos com idade até quatro meses, causando distúrbios gastrintestinais. O objetivo do presente trabalho foi observar o controle de larvas infectantes (L3) de Strongyloides westeri pelos fungos nematófagos Duddingtonia flagrans (AC001) e Monacrosporium thaumasium (NF34) após trânsito gastrintestinal em jumentas. Foram utilizadas 12 jumentas, estabuladas e previamente vermifugadas. A seguir, dois grupos tratados, contendo cada um 4 animais receberam por via oral 100 g de péletes em matriz de alginato de sódio, contendo massa miceliana dos fungos D. flagrans (AC001) ou M. thaumasium (NF34). O grupo controle foi constituído de 4 animais que receberam péletes sem fungo. A seguir, amostras de fezes dos grupos de animais foram coletadas em distintos intervalos de horas (12, 24, 48 e 72). Essas fezes foram vertidas em placas de Petri contendo meio sólido ágar-água 2% e 1000 L3 de S. westeri. Os isolados AC001 e NF34 apresentaram capacidade de destruir as L3 após o trânsito, demonstrando sua viabilidade e atividade predatória.

Recent developments in research into the Cyathostominae and Anoplocephala perfoliata.

Intestinal helminths are an important cause of equine disease. Of these parasites, the Cyathostominae are the commonest group that infect horses. These nematodes consist of a complex tribe of 51 species, although individual horses tend to harbour 10 or so common species, in addition to a few rarer species. The Cyathostominae can be extremely pathogenic, and high levels of infection result in clinical symptoms ranging from chronic weight loss to colic, diarrhoea and death. As part of their life cycle, immature cyathostomins penetrate the large intestinal wall, where they can enter a state of inhibited larval development. These larvae can exist in this state for months to years, after which they subsequently re-emerge. If larvae re-emerge in large numbers (i.e. several million), severe pathological consequences ensue. The inhibited larvae are also relatively refractory to several of the currently available anthelmintics, so that horses treated previously with anthelmintics can still carry life-threatening burdens of these parasitic stages. Little is known about the cyathostomin larvae during their mucosal phase, and current research efforts are focused on investigating the biology of these stages. Much of the research described here highlights this area of research and details studies aimed at investigating the host immune responses that the mucosal larvae invoke. As part of this research effort, molecular tools have been developed to facilitate the identification of larval and egg stages of cyathostomins. These molecular tools are now proving very useful in the investigation of the relative contributions that individual, common cyathostomin species make to the pathology and epidemiology of mixed helminth infections. At the more applied level, research is also in progress to develop an immunodiagnostic test that will allow numbers of mucosal larvae to be estimated. This test utilises antigen-specific IgG(T) serum antibody responses as markers of infection. As anthelmintic resistance will be the major constraint on the future control of the Cyathostominae, researchers are now actively investigating this area and studies aimed at elucidating the molecular mechanisms of drug resistance are described. Another parasite which has assumed a clinically important role in horses is the tapeworm, Anoplocephala perfoliata. This parasite is prevalent world-wide and has been shown to be a significant cause of equine colic. Because previous methods of estimating the infection intensity of tapeworm were inaccurate, recent research has been directed at developing an immunodiagnostic ELISA for these cestodes. Specific IgG(T) responses to antigens secreted by adult tapeworms have been shown to provide a reasonable indication of infection intensity. An ELISA based on these responses is now commercially available. The steps involved in the development of this ELISA are described here. In addition to these recent advances in research, this review also outlines the principle areas for future research into these important equine parasites.

Parenterally administered gastrointestinal nematode infective larvae viable after more than 15 years in liquid nitrogen.

After cryopreservation for 13.3-15.8 years, the viability of the infective larvae (L3) of Trichostrongylus axei, T. colubriformis, Oesophagostomum columbianum, Haemonchus contortus, Ostertagia circumcincta, T. falculatus, Nematodirus spathiger, Chabertia ovina and Dictyocaulus filaria was assessed in sheep, by being deposited at their predilection sites. D. filaria was, however, an exception, in that the L3 were injected into the jugular vein. The mean development of all the species was 22.8%, but if three species (O. columbianum, C. ovina and D. filaria), that developed poorly are disregarded, then the mean development was 33.4%, similar to previous tests after shorter periods of cryopreservation. The L3 of some of the species appeared sluggish when examined 10-15 min after being thawed, and in the case of H. contortus practically all the larvae of the original batch tested in the previous trials of the series appeared dead when thawed for use in the present trial, and were replaced by another batch of L3 of the same species. When re-examined after about 8 h, however, a high percentage of the L3 of the original batch appeared to have become revitalised, and their viability was tested in a trial reported elsewhere. The intestinal cells of the majority of the L3 of N. spathiger, O. circumcincta and C. ovina were vesiculated when they were thawed. Nevertheless, the degree of development of the former two species was of the highest in the trial, and it can be concluded that this phenomenon does not necessarily impede the viability of larvae.

Pathogenicity of cyathostome infection.

Cyathostomes are now the principle parasitic pathogen of the horse: a remarkable transformation during the last 25 years from virtual obscurity to focus of attention in equine parasitology. This rise to prominence coincides with the marked decrease in prevalence of large strongyle infections as a result of widespread use of modern anthelmintic compounds. On the basis that strongyle-associated diseases continue to commonly occur in the absence of these large strongyle species, clinical attention has turned to the pathogenicity of cyathostomes. Although many horses harbour burdens of tens of thousands of cyathostomes without developing detectable illness, these parasites can result in an inflammatory enteropathy affecting the caecum and colon. Although the principle clinical effect of cyathostomosis is weight loss, affected individuals may exhibit other signs including diarrhoea and/or subcutaneous oedema and/or pyrexia. Clinical cyathostomosis occurs more commonly in young horses in late winter/early spring but there is lifelong susceptibility to cyathostomes and they can cause clinical disease in any age of horse during any season. Animals with cyathostomosis often develop hypoalbuminaemia and/or neutrophilia but there are no clinicopathological features specific for the disease. Experimental infections with cyathostomes have resulted in both clinical and pathological features similar to those of naturally-occuring cyathostomosis cases. From the experimental infection studies, it is evident that cyathostomes are pathogenic at times of both penetration into and emergence from the large intestinal mucosa. An unusual feature of cyathostome biology is the propensity for arrested larval development within the large intestinal mucosa for more than 2 years. From limited studies it appears that this arrested larval development is favoured by: feedback from luminal to mucosal worms; larger size of challenge dose of larvae and trickle (versus single bolus) infection. During arrested larval development cyathostomes have minimal susceptibility to all anthelmintic compounds, thus, limiting the effectiveness of therapeutic and/or control strategies. Although, the relative importance of individual cyathostomes is not known, the development of species-specific DNA methods for identification of cyathostomes provides a means by which the pathogenicity of different species might be established.

The pathogenic effects of experimental cyathostome infections in ponies.

Nine pony breed foals were reared indoors, then allocated to one of three groups infected with either 3.9 million (Group A) or 3.15 million (Group B) cold-conditioned third stage cyathostome larvae or kept as uninfected controls (Group C). The larvae were administered as a 'trickle' infection of 150000 larvae per dose, three times weekly. Blood biochemical and haematological analyses were performed weekly and faecal worm egg counts bi-weekly. Complete parasitological examinations were performed on all ponies at various times post-initial infection (PI): one infected animal at 9 weeks PI, four animals (three infected, one control) at 20 weeks PI and four animals (two infected, two controls) at 60-62 weeks PI. All ponies in the infected groups experienced a marked reduction in weight gain and two animals developed clinical disease: one pony developed intermittent diarrhoea and colic 8 weeks PI; another pony developed intermittent diarrhoea between 30 and 52 weeks PI. All infected ponies had decreased serum fructosamine concentrations and five had decreased serum albumin, which were first apparent 4-6 weeks PI. Alterations in the composition of serum globulins were detected in all ponies. Transient neutrophilia was observed in five animals from the infected groups 3-9 weeks PI. Serum alkaline phosphatase concentrations were increased in one pony between 30 and 60 weeks PI. During the course of the experiment, faecal samples from all infected animals were negative for worm eggs. At necropsy, cyathostome larvae were present within the mucosa of the large intestine of all infected ponies, however the mucosal larval counts varied considerably between individuals.

Investigation of the chemosensory function of amphids of Syngamus trachea using electrophysiological techniques.

Syngamus trachea, the gape nematode, has been used as a model to study the chemosensory function of amphids. Extracellular electrophysiological recordings were performed directly on amphids. The amphids were stimulated by blood serum from a host bird and by D-tryptophan. The spike frequency produced by the amphid increased significantly after the application of the serum and the application of D-tryptophan. Two types of amphidial neurones responded to the presence of the stimuli. These extracellular recordings have verified the chemosensory function of the amphids.

Case report of and description of parasite in Mammomonogamus laryngeus (human syngamosis) infection.

About 100 cases of human infections with Mammomonogamus laryngeus (Syngamidae, Syngaminae) have been reported, with virtually all cases originating in the Caribbean Islands and Brazil. This report describes the fifth patient in North America infected with M. laryngeus and the first case documented to originate in Jamaica. The patient complained of a characteristic persistent nonproductive cough and a lump in her throat. M. laryngeus is a nematode in which the male and female are permanently joined in copula, producing a distinctive Y shape. Since there is scant clinical information about this parasite, this report includes a description of the adult worms and eggs, a summary of the epidemiology, and the clinical manifestations in humans. Illustrations are presented to facilitate future identification of this parasite and to expedite treatment.

Case report of a description of parasite in mommomonogamus laryngeus (human syngamosis) infection

About 100 cases of human infections with Mammomonogamus laryngeus (Syngamidae, Syngaminae) have been reported, with virtually all cases originating in the Caribbean Islands and Brazil. This report describes the fifth patient in north America infected with M. laryngeus and the first case documented to originate in Jamaica. The patient complained of a characteristic persistent nonproductive cough and a lump in her throat. M. laryngeus is a nematode in which the male and female are permanently joined in copula, producing a distinctive Y shape. Since there is scant clinical information about this parasite, this report includes a description of the adult worms and eggs, a summary of the epidemiology, and the clinical manifestations in humans. Illustrations are presented to facilitate furture identification of this parasite and to expedite treatment (AU)

Control of strongylosis in horses by alternate grazing of horses and sheep and some other aspects of the epidemiology of Strongylidae infections.

Alternate grazing of horses and sheep as a control measure for gastrointestinal helminthiasis was studied in three grazing experiments in 1981, 1982 and 1983. Each year a group of three mare yearling Shetland ponies, which were kept on a small pasture from spring to autumn, were compared with a similar group which grazed a similar or the same pasture until July and were subsequently removed to a similar pasture which had been grazed by sheep from April to July. In addition both groups were treated with an anthelmintic when the latter group was removed to the sheep pasture. Pasture larval counts and worm counts and, in 1982 and 1983, faecal egg counts, clinical condition, total protein, albumin and beta-globulin levels demonstrated that the groups removed to sheep pasture acquired considerably lower burdens of nematodes of the subfamilies Cyathostominae and Strongylinae, but considerably higher burdens of Trichostrongylus axei than the groups which were not moved. These T. axei infections resulted in higher serum pepsinogen levels in the former groups compared to the latter in 1981 and 1982. At necropsy an important part of the T. axei burdens and, in 1982 and 1983, the Cyathostominae burdens consisted of inhibited early third stage larvae. A total of 20 species of the subfamily Cyathostominae and 7 species of the Strongylinae were found. Generally the composition of species was in agreement with other observations in western Europe, the most common species being: Cylicostephanus longibursatus, Cylicostephanus minutus, Cylicostephanus calicatus, Cylicostephanus goldi, Cylicostephanus poculatus, Cyathostomum labratum, Cyathostomum coronatum, Cyathostomum catinatum, Cylicocyclus leptostomus, Cylicocyclus nassatus, Cylicocyclus insigne, Strongylus edentatus and Strongylus vulgaris.

Pathogenesis of helminths in equines.

This review summarizes information on the clinical signs, gross and microscopic lesions associated with nematode and cestode infections and discusses the development of these conditions in the equine host.

Demonstration of collagenase activity in adult Strongyloides ratti (Nematoda:Strongyloididae) and its absence in the infective larvae.

Larvae and adults of Strongyloides ratti were examined for collagenolytic activity on 14C proline-labelled, native, guinea-pig skin collagen substrate. The activity was measured by determining either the amount of hydroxyproline released or the amount of radioactivity in the solubilized fraction of the collagen substrate. Bacterial collagenase was used for enzyme control and trypsin served as substrate control. No collagenolytic activity was found in living larvae, their extracts or metabolites. The collagenolytic activity of the metabolites of adult worms appeared weak, whereas that of the extracts of the adults was pronounced. It is suggested that collagenase is active in the adult females at the time of migration in the intestinal mucosa during oviposition.

Studies on Strongylus asini. I. Experimental infestation of equines.

Infective larvae were harvested from a culture of eggs collected from adult Strongylus asini recovered from a free-ranging Burchell's zebra, Equus burchelli, in the Kruger National Park. Worm-free zebra, horse and donkey foals were successfully infested, but infestation failed in a mule foal. At slaughter, 117-125 days post-infestation, S. asini in their 4th moult were recovered from the liver and portal veins. This is the first report of successful experimental infestation of these hosts with S. asini.

[Motility and infestivity of first-stage Protostrongylid larvae. Factors of variation (author's transl)]. / Motilité et infestivité des larves L1 de protostrongylides: facteurs de variation.

The infection of land-snails was realised in order to relate the motility of Protostrongylid first-stage larvae to their infectivity. Various factors such as the age of larvae, their density, the composition of the conservation liquid and the temperature were tried on the motility of larvae. A clear relation appeared between motility and infestivity of larvae. The age, density of larvae, and cold did influence negatively their motility. On the contrary increases in temperature and optimal concentrations of magnesium, calcium and sodium salts did influence positively the motility. Induced variations of motility did provoke related fluctuations of infestivity.

Studies in vitro on the reaction of peritoneal exudate cells from mice immune to infection with Nematospiroides dubius with the infective third stage larvae of this parasite.

The results of the present study show that peritoneal exudate cells from mice immune to infection with Nematospiroides dubius are able in vitro to damage the third stage infective larvae as measured by a loss in infectivity. The ability of these cells to function in the absence of specific antibody seems to be related to the presence of trypsin labile factors on their surfaces. Lymphocytes from immune mice are also able to damage the larvae. The suggestion is made that 'activated' macrophages may play an important role in immunity to this infection.

Angiostrongylus cantonensis: phospholipase in nonsensitized and sensitized rats after challenge.

Rats given an initial infection with Angiostrongylus cantonensis had moderately elevated phospholipase B activity in the lungs at 8 and 15 days after challenge, and greatly elevated levels were evident at 35, 43, and 49 days. In the brain, the values were elevated at 15 through 35 days. These periods of increased activity in the lungs and brain coincided with the migration patterns of the third stage larvae and the adult worms in this host. The elevated enzyme levels also were were correlated with increased numbers of eosinophils in the bone marrow at 8 and 15 days and again at 36, 43, and 49 days after infection. Similarly infected rats exhibited leukocytosis at 1 through 10 weeks of observation after challenge, and striking eosinophilia at 1, 7, 8, and 9 weeks. Rats reinfected after removal of the worms of the initial infection by thiabendazole treatment showed an anamnestic response characterized by (i) elevated enzyme values in both the lungs and brain at 1 day after reinfection and (ii) eosinophilia in the bone marrow by day 4. These accelerated responses were accompanied by a significant reduction in the worm burden of the rats. The results, which support our hypothesis that inflammation, elevated phospholipase B activity, and reduction in worm burden are causally related, are discussed in light of similar findings reported earlier from our studies with Trichinella spiralis and Nippostrongylus brasiliensis.