Organ-specific Toxocara canis larvae migration and host immune response in experimentally infected mice.

We investigated organ specific Toxocara canis larval migration in mice infected with T. canis larvae. We observed the worm burden and systemic immune responses. Three groups of BALB/c mice (n=5 each) were orally administered 1,000 T. canis 2nd stage larvae to induce larva migrans. Mice were sacrificed at 1, 3, and 5 weeks post-infection. Liver, lung, brain, and eye tissues were collected. Tissue from 2 mice per group was digested for larval count, while the remaining 3 mice underwent histological analysis. Blood hematology and serology were evaluated and compared to that in a control uninfected group (n=5) to assess the immune response. Cytokine levels in bronchoalveolar lavage (BAL) fluid were also analyzed. We found that, 1 week post-infection, the mean parasite load in the liver (72±7.1), brain (31±4.2), lungs (20±5.7), and eyes (2±0) peaked and stayed constant until the 3 weeks. By 5-week post-infection, the worm burden in the liver and lungs significantly decreased to 10±4.2 and 9±5.7, respectively, while they remained relatively stable in the brain and eyes (18±4.2 and 1±0, respectively). Interestingly, ocular larvae resided in all retinal layers, without notable inflammation in outer retina. Mice infected with T. canis exhibited elevated levels of neutrophils, monocytes, eosinophils, and immunoglobulin E. At 5 weeks post-infection, interleukin (IL)-5 and IL-13 levels were elevated in BAL fluid. Whereas IL-4, IL-10, IL-17, and interferon-γ levels in BAL fluid were similar to that in controls. Our findings demonstrate that a small portion of T. canis larvae migrate to the eyes and brain within the first week of infection. Minimal tissue inflammation was observed, probably due to increase of anti-inflammatory cytokines. This study contributes to our understanding of the histological and immunological responses to T. canis infection in mice, which may have implications to further understand human toxocariasis.

A Reevaluation of the Tolerability and Effects of Single-Dose Ivermectin Treatment on Onchocerca volvulus Microfilariae in the Skin and Eyes in Eastern Ghana.

Mass administration of ivermectin (IVM) has significantly reduced onchocerciasis prevalence, intensity, and morbidity in most endemic areas. Most IVM clinical trials were performed long ago in persons with high-intensity infections that are uncommon in West Africa today. This cohort treatment study recruited participants from a hypoendemic area in eastern Ghana to reevaluate the efficacy and tolerability of IVM with a special focus on the kinetics of microfilaria (Mf) clearance. Mf in the skin and anterior chambers (AC) were assessed by skin snip and slit lamp examinations at baseline and at 3 and 6 months after treatment with IVM 150 µg/kg. Most participants (184-231, 79.7%) enrolled were treatment-naïve. The baseline geometric mean skin Mf count was 12.67/mg (range 3-86). Although persons with MfAC at baseline (64/231, 27%) had significantly higher skin Mf counts than people without MfAC, 7 of 39 (15%) of persons with skin Mf counts in the range of 3-5 Mf/mg had MfAC. Skin Mf were detected in 14% (31/218) and 45% (96/216) of participants 3 and 6 months after IVM treatment, respectively. MfAC were detected in 12 of 212 (5.7%) study participants at 6 months. 81% (187 of 231) of participants experienced 439 adverse events within 7 days after treatment; all adverse events were mild (96.1%) or moderate. This study has provided new data on the kinetics of Mf in the skin and eyes after IVM treatment of persons with light to moderate intensity Onchocerca volvulus infections that are common in Africa at this time.

SAMPLE COMPOSITION: PARASITE ECOLOGY'S DIRTY LITTLE SECRET.

In comparative studies, the advantage of increased sample sizes might be outweighed by detrimental effects on sample homogeneity and comparability when small numbers of hosts from a different demographic of the same species are included in samples. A mixed sample of sunfishes (Lepomis spp.) was subdivided in different ways and examined using cumulative performance curves to determine whether the exclusion of larger hosts from a single-species sample and/or the inclusion of hosts of the same size demographic from closely related host species would produce more homogeneous samples. The exclusion of larger hosts from the single-species samples tended to reduce the aggregation of the infrapopulation samples, and mixed-species samples of smaller fishes tended to have lower degrees of aggregation for a given sample size relative to the single-species sample. Cumulative performance curves for diversity and richness, in concert with nonmetric multidimensional scaling of the infracommunities, demonstrated sunfish size to be a more reliable determinant of infracommunity similarity than sunfish species in this particular sample. The results demonstrate that cumulative aggregation curves can be an effective tool for delineating homogeneous and comparable subsamples and that, under some circumstances, it is possible to offset the smaller sample sizes that result from the exclusion of older/larger hosts by the addition of congeneric or confamilial hosts within the same size/age classes as the stratified sample.

Efficacy of fipronil/(S)-methoprene/eprinomectin/praziquantel (Broadline®) against Thelazia callipaeda in naturally infected cats.

BACKGROUND: The present clinical field trial was conducted to assess the efficacy of a broad-spectrum parasiticide spot-on formulation containing eprinomectin (Broadline®) against Thelazia callipaeda eyeworm in naturally infected cats. METHODS: Fifteen privately owned cats harboring at least one live adult T. callipaeda were included in the study. Cats were randomly allocated to an untreated control group of seven cats or to a Broadline®-treated group of eight cats. Cats were treated on Day 0; ocular examinations were performed at inclusion and on Days 7 and 14; eyeworms were recovered and counted on Day 14. The primary efficacy assessment was based on group comparison of number of T. callipaeda on Day 14. RESULTS: Seven days after treatment, six of eight treated cats were negative for eyeworm infection per visual examination, and on Day 14 no eyeworms were found in the treated cats while the seven untreated cats were still infected (geometric mean: 1.97). All cats had inflammatory ocular signs at inclusion; on Day 14, five of eight treated cats had recovered while all untreated control cats were still symptomatic. All collected parasites were confirmed to be T. callipaeda by morphology and molecular characterization. CONCLUSIONS: A single treatment with Broadline® provided 100% efficacy against feline thelaziosis and improved related ocular inflammation signs.

Confirmed cases of human Onchocerca lupi infection: a systematic review of an emerging threat.

Diverse Onchocerca species are present mostly parasitizing ungulates, with the exception of Onchocerca volvulus (O. volvulus) in humans and O. lupi in canids and cats. The human cases due to the O. lupi have been more highlighted during last years. So, the present review was performed to determine the detailed characteristics of confirmed human O. lupi case reports documented worldwide. Hence, a systematic search was done using English international databases (Scopus, PubMed, Web of Science, Embase, ProQuest, and Google Scholar). Totally, 14 confirmed human cases were documented during the last decade, mostly from the USA and Turkey with 7 and 3 cases, respectively. Most cases (7 individuals) were male with the age range of 22-month-old to 54-year-old. The parasite was frequently isolated from the right eye (5 cases), followed by the left eye (4 cases), cervical spinal canal (3 cases), scalp, and right forearm (one case each). Molecular identification of the isolated agent was the preferred way of diagnosis in most cases (9 records). In conclusion, human O. lupi cases have been more highlighted in recent years, whether due to the improved diagnostics and/or host-switching phenomenon, and both veterinarians and healthcare authorities should be alerted.

Towards an odour-baited trap to control Musca sorbens, the putative vector of trachoma.

Musca sorbens is a synanthropic filth fly that aggressively attacks people to feed from mucous membranes of the eyes, nose or mouth, from open sores, or from sweat. It has long been suspected that this fly contributes to the transmission of eye infections, particularly trachoma, and recent work has added to the evidence base that M. sorbens is a trachoma vector in Ethiopia. There are few options to control M. sorbens, largely due to a lack of evidence. Space spraying with insecticides is effective, but an environmentally sound and long-term sustainable solution would be better, for example, mass trapping. We tested commercially available and homemade trap types in a pilot (laboratory) study and three field studies. A homemade design, built from a bucket and two empty water bottles, baited with a commercially available lure, The Buzz, was found to be most effective. This trap caught 3848 M. sorbens over 26 trap 'events' (3- or 4-day periods); mean/median per 24 h 43.6 (standard deviation 137.10)/2.25 (IQR 0.25-12.67). The Buzz lure is cheap and effective for 4 weeks, and trap components cheap and locally available. Further studies are needed to understand the impact of this trap on local fly populations and the local transmission of trachoma.

Autochthonous, zoonotic Onchocerca lupi in a South Texas dog, United States.

BACKGROUND: Onchocerca lupi is an emerging, zoonotic filarioid nematode associated with ocular disease in companion animals in North America and the Old World. The areas where this parasite is assumed to be endemic in the USA comprise southwestern states. Thus far, all cases reported outside of the southwest are associated with travel or animal movement. METHODS: An 11-year-old, castrated male Pitbull dog from McAllen, Hidalgo County, southern Texas, with no travel history, was diagnosed with a perforating corneal ulceration of the right eye. Enucleation was performed and tissues submitted for histopathology. RESULTS: Histologically, sections of two filarioid nematodes were observed. DNA was extracted from formalin-fixed paraffin-embedded tissue using a commercial kit. We performed PCR targeting the cox1 gene of the mitochondrial DNA, followed by sequencing and phylogenetic analysis. Altogether, these results confirmed the identification of the nematode specimens as O. lupi, phylogenetically belonging to haplotype 1. CONCLUSION: We report the first autochthonous case of O. lupi in a dog from Hidalgo County, southern Texas, USA. Our finding suggests Texas as an additional state where this zoonotic nematode is endemic. Further investigations are required to understand the epidemiology of this parasite along the USA/Mexico border.

Altered neuronal activity in the visual processing region of eye-fluke-infected fish.

Fish, like most vertebrates, are dependent on vision to varying degrees for a variety of behaviours such as predator avoidance and foraging. Disruption of this key sensory system therefore should have some impact on the ability of fish to execute these tasks. Eye-flukes, such as Tylodelphys darbyi, often infect fish where they are known to inflict varying degrees of visual impairment. In New Zealand, T. darbyi infects the eyes of Gobiomorphus cotidianus, a freshwater fish, where it resides in the vitreous chamber between the lens and retina. Here, we investigate whether the presence of the parasite in the eye has an impact on neuronal information transfer using the c-Fos gene as a proxy for neuron activation. We hypothesized that the parasite would reduce visual information entering the eye and therefore result in lower c-Fos expression. Interestingly, however, c-Fos expression increased with T. darbyi intensity when fish were exposed to flashes of light. Our results suggest a mechanism for parasite-induced visual disruption when no obvious pathology is caused by infection. The more T. darbyi present the more visual stimuli the fish is presented with, and as such may experience difficulties in distinguishing various features of its external environment.

Pathophysiology of ocular toxoplasmosis: Facts and open questions.

Infections with the protozoan parasite Toxoplasma gondii are frequent, but one of its main consequences, ocular toxoplasmosis (OT), remains poorly understood. While its clinical description has recently attracted more attention and publications, the underlying pathophysiological mechanisms are only sparsely elucidated, which is partly due to the inherent difficulties to establish relevant animal models. Furthermore, the particularities of the ocular environment explain why the abundant knowledge on systemic toxoplasmosis cannot be just transferred to the ocular situation. However, studies undertaken in mouse models have revealed a central role of interferon gamma (IFNγ) and, more surprisingly, interleukin 17 (IL17), in ocular pathology and parasite control. These studies also show the importance of the genetic background of the infective Toxoplasma strain. Indeed, infections due to exotic strains show a completely different pathophysiology, which translates in a different clinical outcome. These elements should lead to more individualized therapy. Furthermore, the recent advance in understanding the immune response during OT paved the way to new research leads, involving immune pathways poorly studied in this particular setting, such as type I and type III interferons. In any case, deeper knowledge of the mechanisms of this pathology is needed to establish new, more targeted treatment schemes.

Ocular Sparganosis: The First Report of Spirometra ranarum in Thailand.

A 22-year-old Thai man from the Northeast region presented with acute eye swelling, itching, and discharge on his left eye. He was suspected of having gnathostomiasis and treated with albendazole and prednisolone for 3 weeks. Nine months later, he was treated with high-dose oral prednisolone for the preliminary and differential diagnoses with thyroid-associated orbitopathy and lymphoma. He had been administered prednisolone intermittently over a few years. Then he developed a painless movable mass at the left upper eyelid and recurrent pseudotumor oculi was suspected. The surgical removal of the mass was performed. A white pseudosegmented worm revealed a definite diagnosis of ocular sparganosis by a plerocercoid larva. Molecular diagnosis of the causative species was made based on the mitochondrial cytochrome c oxidase subunit 1 (cox1) gene. Proper technique of extraction and amplification of short fragments DNA from formalin-fixed paraffin-embedded tissue successfully identified parasite species. The result from the sequencing of the PCR-amplified cox1 fragments in this study showed 99.0% sequence homology to Spirometra ranarum. This is the first report of S. ranarum in Thailand.

[Disseminated cysticercosis affecting the brain, the eye and the skin on a Senegalese patient]. / Cysticercose disséminée à localisation neurologique, oculaire et cutanée chez une patiente sénégalaise.

Cysticercosis is a neglected tropical disease set as health priority by WHO. Most of the reported cases included isolated types of cysticercosis affecting the skin, the eyes or the brain . Disseminated types, however, are rare. We here report a case of disseminated cysticercosis affecting the brain, the eyes and the skin in a Senegalese female patient aged 66 years admitted with headaches and chronic seizures. Clinical examination showed cerebellar syndrome associated with generalized and painless nodular subcutaneous lesions. Diagnosis was confirmed based on histopathological examination of skin biopsy which showed cysticerci. Patient's outcome was good under albendazole therapy.

Suspected autochthonous Thelazia callipaeda infection in a dog in northern Germany.

A 12-year old Elo dog was presented with recurring symptoms of conjunctivitis in November 2019. A single whitish nematode was found upon inspection of the eye and identified as a Thelazia callipaeda male. The morphological identification of the eye worm was supported by analysis of a partial cytochrome c oxidase I (cox1) gene sequence. The dog lived in Lower Saxony, northwestern Germany, and had not visited regions known to be endemic for T. callipaeda. This suggests that a local transmission cycle of this zoonotic nematode may exist in Germany.

Humic-acid-driven escape from eye parasites revealed by RNA-seq and target-specific metabarcoding.

BACKGROUND: Next generation sequencing (NGS) technologies are extensively used to dissect the molecular mechanisms of host-parasite interactions in human pathogens. However, ecological studies have yet to fully exploit the power of NGS as a rich source for formulating and testing new hypotheses. METHODS: We studied Eurasian perch (Perca fluviatilis) and its eye parasite (Trematoda, Diplostomidae) communities in 14 lakes that differed in humic content in order to explore host-parasite-environment interactions. We hypothesised that high humic content along with low pH would decrease the abundance of the intermediate hosts (gastropods), thus limiting the occurrence of diplostomid parasites in humic lakes. This hypothesis was initially invoked by whole eye RNA-seq data analysis and subsequently tested using PCR-based detection and a novel targeted metabarcoding approach. RESULTS: Whole eye transcriptome results revealed overexpression of immune-related genes and the presence of eye parasite sequences in RNA-seq data obtained from perch living in clear-water lakes. Both PCR-based and targeted-metabarcoding approach showed that perch from humic lakes were completely free from diplostomid parasites, while the prevalence of eye flukes in clear-water lakes that contain low amounts of humic substances was close to 100%, with the majority of NGS reads assigned to Tylodelphys clavata. CONCLUSIONS: High intraspecific diversity of T. clavata indicates that massively parallel sequencing of naturally pooled samples represents an efficient and powerful strategy for shedding light on cryptic diversity of eye parasites. Our results demonstrate that perch populations in clear-water lakes experience contrasting eye parasite pressure compared to those from humic lakes, which is reflected by prevalent differences in the expression of immune-related genes in the eye. This study highlights the utility of NGS to discover novel host-parasite-environment interactions and provide unprecedented power to characterize the molecular diversity of cryptic parasites.

First report of Thelazia callipaeda in a free-ranging Iberian wolf (Canis lupus signatus) from Spain.

Thelazia callipaeda (Spirurida, Thelaziidae) is a vector-borne zoonotic eyeworm able to infect a broad spectrum of carnivores. Here, we describe the first case of bilateral infection by T. callipaeda in the eyes of an adult female Iberian wolf (Canis lupus signatus) in central Spain. Nematodes collected were morphologically identified (n = 42), and two specimens were molecularly characterized. At the sequence analysis of the partial mitochondrial cytochrome c oxidase subunit 1 gene, T. callipaeda haplotype 1 (the only haplotype circulating in Europe) was detected. The role of the Iberian wolf as a natural reservoir for T. callipaeda in the life cycle of this emerging zoonosis and the implications in conservation are discussed.

Quantification of Toxocara canis DNA by qPCR in mice inoculated with different infective doses.

The nematode Toxocara canis is of public health importance and is the main causative agent of toxocariasis in humans. This disease is difficult to diagnose due to several factors, including the possibility of cross-reactions with other nematodes in the ELISA. To overcome this problem, molecular tests have been recommended as an alternative to identify the parasite. The quantitative real-time polymerase chain reaction (qPCR) technique was used in this study to identify and quantify the parasite load of T. canis in the mouse brain. To this end, 24 mice were divided into six groups, five of which were challenged with different infective doses of T. canis larvae (L3) (1000, 500, 250, 100 and 50 larvae), while the sixth group, uninfected, acted as negative control. Forty-five days after infection, the animals were euthanized to collect the brain, from which two portions of 20 mg of tissue were taken for DNA extraction, while the rest of the brain tissue was digested to quantify the number of larvae by microscopy. The number of DNA copies was calculated from the standard DNA quantification curve, showing values of E = 93.4%, R2 = 0.9655 and Y = -3.415. A strong positive correlation (R = 0, 81; p < .001) was found between the number of copies and the recovery of larvae from brain. However, the parasite's DNA was also identified even in animals from whose brain no larvae were recovered after tissue digestion. The results of this study therefore confirm that the qPCR technique can be a valuable tool for the detection and quantification of T. canis DNA in murine hosts, even in animals whose with tissues contain very few parasites.

Canine trypanosomosis in Sri Lanka: An emerging problem reported from three distinct geographic locations.

Here we report three cases of canine trypanosomosis presented to the Veterinary Teaching Hospital in the Faculty of Veterinary Medicine and Animal Sciences at the University of Peradeniya, Sri Lanka during 2018. The cases were presented to the hospital when the dogs were already in critical condition. Confirmation of the cases was done by microscopic examination of Giemsa-stained thin blood smears. All three dogs had bilateral keratitis and anterior chamber cloudiness in eyes. Despite the intramuscular administration of diminazine aceturate, all of them subsequently died. Amplification and sequencing of a fragment of the internal transcribed spacer 1 (ITS1) of the nuclear ribosomal DNA confirmed the parasite as Trypanosoma. evansi. This is the first record of clinical cases of canine trypanosomosis in Sri Lanka. The three cases reported here came from widely separated geographical locations within the country: Balangoda, Mullaitivu and Kadawatha.

Temperature is a common climatic descriptor of lachryphagous activity period in Phortica variegata (Diptera: Drosophilidae) from multiple geographical locations.

BACKGROUND: The drosophilid Phortica variegata is known as vector of Thelazia callipaeda, the oriental eyeworm native to Asia that has become an emergent zoonotic agent in several European regions. Unlike almost all other arthropod vectors of pathogens, only P. variegata males feed of lachrymal secretions of animals, ingesting first-stage larvae (L1) of the worm living in the orbital cavities of the host, and allowing with the same behaviour the introduction of infective L3. Despite the increased detection of T. callipaeda in many European countries, information about the length of the lachryphagous activity period of P. variegata and a deep knowledge of the environmental and climatic variables involved are still limited. METHODS: We herein present the results of a multicentre study involving five sites from four different countries (Italy, Spain, UK and USA) where canine thelaziosis is endemic and/or where it has already been ascertained the presence of P. variegata. Field data have been obtained on a fortnightly basis from mid-April to the end of November 2018 from a contemporary standardized sampling (same sampling effort and time of collection in all sites) of lachryphagous flies collected around the eyes of a human bait using an entomological net. These data have been associated to data collection of local climatic variables (day length, temperature, wind speed, barometric pressure and relative humidity). RESULTS: Overall, a total of 4862 P. variegata flies (4637 males and 224 females) were collected, with high differences in densities among the different sampling sites. Significant positive correlations were found between P. variegata male density and temperature and wind speed, while negative correlations were observed for barometric pressure and relative humidity. However, the above significant differences are confirmed in each sampling site separately only for the temperature. CONCLUSIONS: This multicentre study highlights that temperature is the major common environmental driver in describing the lachryphagous activity of P. variegata in Europe and USA and, therefore, the transmission risk of thelaziosis.

Risky business: influence of eye flukes on use of risky microhabitats and conspicuousness of a fish host.

A prerequisite for a parasitic manipulation to be considered adaptive is that it confers a fitness benefit to the parasite, such as increased transmission to another host. These manipulations can involve alterations to a wide range of host phenotypic traits, including microhabitat choice. Eye flukes of the trematode family Diplostomidae use fish as intermediate hosts and must be transmitted by predation to a piscivorous bird. In New Zealand, the diplostomid Tylodelphys darbyi infects the eyes of a widespread endemic freshwater fish, the common bully Gobiomorphus cotidianus. Within the eye, T. darbyi metacercariae achieve large sizes and move freely about the aqueous and vitreous humors of the eye. We hypothesized that higher intensities of T. darbyi would (i) cause bullies to show increased activity and spend more time moving about in open space (i.e., more conspicuous, risky microhabitat) and (ii) reduce their ability to compete for shelter with fish harboring lower infection levels. Our experiments showed that heavily infected fish were more active and spent more time in the open, although the effect was age-dependent, with immature fish displaying decreases in activity and time spent in the open with increasing intensities of infection. We also demonstrated that heavily infected female bullies have a lower probability of using shelter, but males show the opposite pattern. It is possible that using more risky microhabitats increases the likelihood of the fish being eaten by the parasite's predatory avian definitive hosts. However, our findings indicate that age- and sex-dependent effects call for a more nuanced interpretation.

The First Acanthamoeba keratitis Case of Non-Contact Lens Wearer with HIV Infection in Thailand.

Acanthamoeba keratitis (AK) is a rare sight-threatening corneal infection, often reporting from contact lens wearers. An asymptomatic human immunodeficiency virus (HIV)-infected Thai male without history of contact lens use complained foreign body sensation at his left eye during motorbike riding. He had neither specific keratitis symptoms nor common drugs responding, which contributed to delayed diagnosis. By corneal re-scraping, Acanthamoeba-like cysts were detected by calcofluor white staining and agar culture. The etiological agent obtained from the culture was molecularly confirmed by Acanthamoeba spp.-specific PCR, followed by DNA sequencing. The results from BLAST and phylogenetic analysis based on the DNA sequences, revealed that the pathogen was Acanthamoeba T4, the major genotype most frequently reported from clinical isolates. The infection was successfully treated with polyhexamethylene biguanide resulting in corneal scar. This appears the first reported AK case from a non-contact lens wearer with HIV infection in Thailand. Although AK is sporadic in developing countries, a role of free-living Acanthamoeba as an opportunistic pathogen should not be neglected. The report would increase awareness of AK, especially in the case presenting unspecific keratitis symptoms without clinical response to empirical antimicrobial therapy.

First report of eyeworm infection by Thelazia callipaeda in gray wolf (Canis lupus) from Serbia.

Thelazia callipaeda, originally known as an "Oriental eyeworm," is a small nematode parasitizing the conjunctival sacs of domestic and wild animals and humans. Previous studies conducted in Serbia have reported the eyeworm infections in dogs, cats, and foxes, as well as in a human patient. As the data regarding thelaziosis from wildlife is still scarce, the aim of this study was to investigate the presence of T. callipaeda in gray wolf (Canis lupus) from Serbia. All collected nematodes were morphologically identified as T. callipaeda males (n = 64) or females (n = 225). Molecular characterization, conducted by PCR amplification followed by sequence analysis of partial cytochrome c oxidase subunit 1 gene (cox1), revealed only haplotype 1 of T. callipaeda. The overall prevalence of thelaziosis was 38.1% (8/21). In all positive animals, both eyes were affected, with a total parasitic load ranging from four to 132 worms per animal. Our results indicate the important epidemiological role of wolves as wildlife reservoirs of T. callipaeda, expanding geographic range of infection, as well as intra- and interspecies contact rates, although the role of other wild carnivore species (i.e., foxes and jackals) should be investigated in future studies.