Parascaris spp. eggs in horses of Italy: a large-scale epidemiological analysis of the egg excretion and conditioning factors.

BACKGROUND: Equine ascariosis, caused by Parascaris spp., is a worldwide endoparasitic disease affecting young horses in particular. Despite the great number of horses reared in Italy, large-scale epidemiological surveys dealing with ascariosis prevalence in the country are not reported in the current literature. For this reason, the present survey aims to describe, for the first time, the spread and infestation of Parascaris spp. in a large population of Italian horses (6896 animals) using faecal egg counts, and further to identify risk factors associated with ascarid egg shedding. METHODS: Individual rectal faecal samples collected during routine veterinary examinations were used and Parascaris spp. prevalence was tested against the animal's age, sex, housing conditions, geographic provenance as well as the respective sampling season. RESULTS: Among the examined stables, 35.8% showed at least one horse to be positive for Parascaris spp. eggs and an overall prevalence of 6.3% was found. Ascariosis rates tended to decrease significantly with age and, proportionally, 80.0% of the recorded Parascaris spp. eggs were found in 0.7% of the examined animals. Statistically significant differences among prevalence rates were found between the different geographic areas of provenance and prevalence was found to be higher in horses reared outdoors compared to those raised indoors. Analysis of data based on sex and season did not show any significant differences. Despite the lower prevalence found compared to other European countries, ascariosis was concluded to represent a significant health challenge for horses reared in Italy, especially foals. Age (foals and yearlings) and outdoor rearing were identified to be significant risk factors for Parascaris spp. egg shedding. Furthermore, the relevance of the infected horses over 6 years of age should not be underestimated as these represent a significant source of contamination for younger animals. CONCLUSIONS: The development of improved treatment protocols based on regular faecal examination combined with follow-up assessment of the efficacy of integrated action plans would prove beneficial in regard to animal health and anthelmintic resistance reduction in the field.

Functional Ultrastructure of the Excretory Gland Cell in Zoonotic Anisakids (Anisakidae, Nematoda).

Excretory and secretory products are crucial for parasite infectivity and host immunomodulation, but the functioning and ultrastructure of the excretory gland cell (EC) that produces these products are still scarcely understood and described. In light of growing reports on anisakiasis cases in Europe, we aimed to characterise the EC of larval Anisakispegreffii and adult Pseudoterranovaazarasi. In the latter, EC starts 0.85 mm from the head tip, measuring 1.936 × 0.564 mm. Larval EC shows a long nucleus with thorn-like extravaginations toward the cytoplasm, numerous electron-dense and -lucent secretory granules spanning from the perinuclear to subplasmalemmal space, an elevated number of free ribosomes, small, spherical mitochondria with few cristae and a laminated matrix, small and few Golgi apparatuses, and few endoplasmic reticula, with wide cisternae complexes. Ultrastructure suggests that anaerobic glycolysis is the main metabolic pathway, obtained through nutrient endocytosis across the pseudocoelomic surface of the EC plasmalemma and its endocytic canaliculi. Thorn-like extravaginations of EC karyotheca likely mediate specific processes (Ca2+ signaling, gene expression, transport, nuclear lipid metabolism) into the extremely wide EC cytosol, enabling focal delivery of a signal to specific sites in a short time. These functional annotations of parasitic EC should help to clarify anisakiasis pathogenesis.

Pixel by pixel: real-time observation and quantification of passive flotation speeds of three common equine endoparasite egg types.

The efficacy of anthelmintic treatments against populations of endoparasites infecting livestock throughout the world is decreasing. To mitigate this, the use of fecal egg counts is recommended to determine both the necessity, and to ensure the appropriate choice, of anthelmintic treatment. Traditionally, and in order to facilitate easier identification and/or enumeration, samples are analysed after separating eggs from other fecal particulates by exposing them to a solution with a density higher than that of the eggs, but lower than the remaining fecal contents. While many parasite egg flotation protocols exist, little is known about the characteristics of these eggs with respect to their movement through a flotation solution. In this study, we have demonstrated a novel method for the observation and quantification of microscopic (65-100 µm) objects as they experience unassisted flotation. This also represents, to our knowledge for the first time, that the flotation of parasite eggs has been observed and their movement characteristics quantified as they float through solution. Particle tracking and video analysis software were utilised to automatically detect and track the movement of individual eggs as they floated. Three 30 s videos and one 2 min video of each egg type were analysed. If the first 30 s of video were discounted, the differences in mean flotation speed among all videos was statistically significant between egg types (P = 0.0004). Strongyle type eggs (n = 201) moved the fastest with a mean 51.08 µm/s (95% confidence interval: 47.54-54.62). This was followed by Parascaris spp. (n = 131) and Anoplocephala perfoliata eggs (n = 322), with mean speeds of 44.43 µm/s (95% confidence interval: 39.47-49.4) and 31.11 µm/s (95% confidence interval: 29.6-32.61), respectively. This method for evaluating the mean speed of passive flotation may represent a first step towards further optimizing fecal egg flotation and be of interest to parasitologists and veterinary practitioners.

How effective is freezing at killing Anisakis simplex, Pseudoterranova krabbei, and P. decipiens larvae? An experimental evaluation of time-temperature conditions.

The consumption of raw or inadequately cooked marine fish can lead to several disorders caused by the ingestion of viable anisakid nematodes. Although anisakid larvae can be killed by subzero temperatures, making freezing an important control measure for this potential health hazard, these parasites can survive freezing under some conditions. Therefore, the aim of the present study was to experimentally evaluate the time-temperature conditions needed to kill Anisakis simplex and Pseudoterranova spp. The effectiveness of freezing was tested on two species of fish: cod, Gadus morhua from the North Atlantic, and herring, Clupea harengus membras from the southern Baltic Sea. Samples, which comprised skinless fillets of cod (n = 40) with visible parasites and whole herring (n = 240), were separately frozen at - 15, - 18, or - 20 °C for 24 h, or at - 20 °C for 48 h in the single-compressor freezer and at - 20, - 25, or - 35 °C for 24 h in the double-compressor freezer. After thawing, parasites were stained with malachite green and examined under the microscope for viability. All A. simplex and Pseudoterranova spp. larvae in cod fillets died at a temperature of - 15 °C or lower. However, freezing did not kill all the A. simplex larvae in whole herring: spontaneous movement of these parasites was observed in samples stored in the single-compressor freezer at - 15, - 18, and - 20 °C over 24 h. Our results demonstrate that the freezing procedure must consider both the capability of the freezing device and the nature of the fish product to ensure consumer safety.

Lensless phase microscopy and diffraction tomography with multi-angle and multi-wavelength illuminations using a LED matrix.

We demonstrate lensless quantitative phase microscopy and diffraction tomography based on a compact on-chip platform, using only a CMOS image sensor and a programmable color LED matrix. Based on the multi-wavelength phase retrieval and multi-angle illumination diffraction tomography, this platform offers high quality, depth resolved images with a lateral resolution of 3.72µm and an axial resolution of 5µm, across a wide field-of-view of 24mm2. We experimentally demonstrate the success of our method by imaging cheek cells, micro-beads, and fertilized eggs of Parascaris equorum. Such high-throughput and miniaturized imaging device can provide a cost-effective tool for telemedicine applications and point-of-care diagnostics in resource-limited environments.

Larva migrans in squirrel monkeys experimentally infected with Baylisascaris potosis.

Roundworms of the genus Baylisascaris are natural parasites primarily of wild carnivores, and they can occasionally cause infection in humans and animals. Infection results in visceral larva migrans and/or neural larva migrans, which can be severe or fatal in some animals. Recently, Baylisascaris nematodes isolated from kinkajous (Potos flavus) and previously referred to as Baylisascaris procyonis were renamed as Baylisascaris potosis; however, data regarding the pathogenicity of B. potosis towards animals and humans are lacking. In the present study, we experimentally infected squirrel monkeys (Saimiri sciureus) with B. potosis to determine the suitability of the monkey as a primate model. We used embryonated eggs of B. potosis at two different doses (10,000 eggs and 100,000 eggs) and examined the animals at 30 days post-infection. Histopathological examination showed the presence of B. potosis larvae and infiltration of inflammatory cells around a central B. potosis larvae in the brain, intestines, and liver. Nevertheless, the monkeys showed no clinical signs associated with infection. Parasitological examination revealed the presence of B. potosis larvae in the intestines, liver, lung, muscles, brain, kidney, and diaphragm. Our findings extend the range of species that are susceptible to B. potosis and provide evidence for the zoonotic potential of larva migrans in high dose infections.

New host records of three juvenile nematodes in Egypt: Anisakis sp. (Type II), Hysterothylacium patagonense (Anisakidae), and Echinocephalus overstreeti (Gnathostomatidae) from the greater lizard fish Saurida undosquamis of the Red Sea.

Three juvenile nematode parasites were collected naturally from 90 (75 %) out of 120 specimens of the marine greater lizard fish Saurida undosquamis captured from water coasts at Hurghada City along the Red Sea in Egypt during the period from September 2013 to April 2014. Worms were identified on the basis of light and scanning electron microscopy. Two of the recovered worms were isolated from the peritoneal cavity of the infected fish around the wall of the stomach as encapsulated larvae. The anisakid juvenile Anisakis sp. (Type II) was characterized by an anteroventrally triangular mouth, with a boring tooth; its postanal tail was rounded, without a terminal mucron or spine. The gnathostomatid Echinocephalus overstreeti was characterized by the presence of a cephalic bulb armed with six transverse rows of spines which were slightly more compact near the anterior end of bulb with maximal separation near the midbulb; the cephalic bulb terminated at a pseudolabia which situated dorsoventrally and reached its greatest width at the posterior one third of the body, The postanal tail terminated at a pointed mucron. The third juvenile species, Hysterothylacium patagonense (Anisakidae), was isolated from the intestine of the infected fish; they are characterized by a small-sized body with a conical tail provided by a nodulose apex, and the anterior end was equipped with three lips. A dorsal lip slightly smaller than the two subventrals left a deep postlabial groove and prominent lateral flanges in between, and the proximal part of each lip was smooth. The three described species were compared morphologically and morphometrically with some of the previously recorded species of the same genus. From this comparison, the similarity and variations between these species were described and concluded that the present study should be considered as a new host record in Egypt.

Dujardinascaris mormyropsis n. sp. (Nematoda: Anisakidae) from the osteoglossiform fish Mormyrops anguilloides (Linnaeus) (Mormyridae) in Central Africa.

A new nematode species, Dujardinascaris mormyropsis n. sp. (Anisakidae), is described from specimens found in the stomach and intestine of the cornish jack Mormyrops anguilloides (Linnaeus) (Osteoglossiformes; Mormyridae) in the Sangha River (Congo River basin) in Dzanga-Sangha Protected Areas, Central African Republic. Based on light and scanning electron microscopical examination, the new species differs from the only other congeneric species parasitising fishes in Africa, D. malapteruri (Baylis, 1923), mainly in the presence of dentigerous ridges on lips, absence of lateral caudal alae in the cloacal region, in the anteriorly curved, non-bifid distal tip of the gubernaculum and larger eggs (60-90 × 52-78 vs 45 × 30 µm); males of D. mormyropsis are characterised by the presence of a ventral precloacal cuticular ornamentation not observed in other Dujardinascaris spp. Dujardinascaris mormyropsis and D. malapteruri also differ in the order of their fish hosts (Osteoglossiformes vs Siluriformes). Specimens previously reported as Dujardinascaris graberi Troncy, 1969, a junior synonym of D. malapteruri, from Mormyrops engystoma Boulenger in Chad belong to the new species D. mormyropsis.

Water compartments and osmoregulation in the parasitic nematode Pseudoterranova decipiens.

A study of the time course of penetration of 3H2O into whole worms suggests that worms immersed in a hypo-osmotic environment (15% artificial sea water) reach full exchange equilibrium more slowly than worms in an iso-osmotic environment (40% artificial sea water). The apparent water content, determined by dry mass, matches that determined by 3H2O exchange when worms are immersed for 24 h in 40% artificial sea water (ASW), but the water content measured by 3H2O exchange is lower when worms are kept in a hypo-osmotic environment for 24 h. These differences disappear after 48 h. No such differences are apparent when sacs, consisting of cylinders of body wall lacking their intestines and pseudocoelomic fluid and closed at both ends by ligatures, are immersed in either 40% or 15% ASW for 24 h. The placing of ligatures at the head, but not at the tail, results in a failure of worms immersed in 40% ASW or 15% ASW containing 3H2O to achieve full exchange equilibrium within 24 h. These results suggest that although worms immersed in an iso-osmotic environment drink, those immersed in a hypo-osmotic environment do not, a conclusion supported by studies involving the addition of [14C]inulin to the medium. The application of ligatures to the head and tail of worms immersed in 40% ASW results in a slower penetration of 3H2O into the pseudocoelomic fluid, whereas similar ligatures do not further retard the penetration in worms exposed to 15% ASW. The results are consistent with a model which sees the pseudocoelomic fluid as consisting of two compartments containing water, one of which exchanges more slowly than the other.