RÉSUMÉ
Rat lungworm disease or neuroangiostrongyliasis is a cerebral parasitic infection that affects humans and animals alike. Its clinical signs and symptoms can range from mild self-resolving to serious life-threatening conditions. Studies suggest therapeutic interventions during the early stages of infection to be more effective than in later stages. However, early diagnosis of infection is usually problematic without the knowledge of exposure and/or detection of the parasite's DNA or antibody against the parasite in the cerebrospinal fluid. This requires a lumbar puncture, which is an invasive procedure that generally requires hospitalization. This study evaluates an affordable and less invasive alternative to detect parasitic DNA by PCR from the peripheral blood of potentially infected animals. Blood samples from 58 animals (55 dogs and 3 cats) with clinical suspicion of infection were submitted to our lab between February 2019 and August 2022 by local, licensed veterinarians. DNA was extracted from whole blood, plasma, serum, and/or packed cells using the Qiagen DNeasy Blood & Tissue Kit as per the manufacturer's protocol. All 58 animals were tested by real-time PCR using the AcanITS1 assay and 32 of these animals (31dogs; 1 cat) were also tested using the AcanR3990 assay. The PCR results for both assays were classified into strongly positive > positive > weakly positive > negative, and equivocal for ambiguous results, based on the strength of the signal. The percent infection detected using the AcanITS1 and AcanR3990 assays was 12.72% (7/55) and 20.68% (6/29), respectively. The overall percent infection detected was 34.37% (11/32), with only two animals testing positive by both assays. The three cats involved in this study tested negative by both assays. These results are promising and warrant further investigations to increase sensitivity including variables that might affect detection in the blood, such as parasite load, and laboratory methodologies.
Sujet(s)
Angiostrongylus cantonensis , Maladies des chats , Réaction de polymérisation en chaine en temps réel , Infections à Strongylida , Animaux , Angiostrongylus cantonensis/isolement et purification , Angiostrongylus cantonensis/génétique , Infections à Strongylida/médecine vétérinaire , Infections à Strongylida/parasitologie , Infections à Strongylida/diagnostic , Infections à Strongylida/sang , Réaction de polymérisation en chaine en temps réel/méthodes , Réaction de polymérisation en chaine en temps réel/médecine vétérinaire , Chats , Maladies des chats/parasitologie , Maladies des chats/diagnostic , Maladies des chats/sang , Chiens , Maladies des chiens/parasitologie , Maladies des chiens/diagnostic , Maladies des chiens/sang , Sensibilité et spécificité , ADN des helminthes/génétique , ADN des helminthes/sangRÉSUMÉ
Angiostrongylus cantonensis is a globally distributed nematode and the leading cause of eosinophilic meningitis in humans. As a global hotspot for this disease, Hawaii's agricultural exports may be contributing to the spread of A. cantonensis. Phytosanitary irradiation doses of 150 or 400 Gy provide quarantine security against multiple insect pests. We evaluated the in vitro and in vivo effects of phytosanitary irradiation on infectious, third-stage, A. cantonensis larvae. In vitro experiments directly exposed larvae to irradiation doses ranging from 200 to 1,000 Gy. Results showed low mortality and no dose response across all treatments 27 days post-irradiation. In vivo studies isolated larvae from wild-caught Parmarion martensi after exposure to x-ray irradiation at doses of 0, 150, and 400 Gy and infected them into laboratory rats. Fourteen rats were assigned to each treatment and infected with 50 larvae from their assigned irradiation dose. Results at 3 and 6 weeks post-infection demonstrated a significant negative dose response in regard to the number of larvae that migrated to the brain and adults found in the pulmonary artery. No irradiated larvae that grew into adults were able to produce eggs. These findings indicate that x-ray irradiation does not result in the direct mortality of A. cantonensis larvae; however, it does affect the infectivity and reproduction of A. cantonensis within its definitive host, the rat. Phytosanitary irradiation at doses ≥150 Gy appears to be an effective means of preventing the establishment of viable populations of A. cantonensis, thus reducing the potential for global spread due to agricultural exports from Hawaii.
Sujet(s)
Angiostrongylus cantonensis , Gastropoda , Infections à Strongylida , Humains , Rats , Animaux , Rayons X , Larve/physiologie , ReproductionRÉSUMÉ
Angiostrongylus cantonensis is the leading cause of neuroangiostrongyliasis worldwide, and east Hawaii Island is a hotspot for the disease in the United States. A combination of glycoproteins with molecular weight of 31 kDa has been used as antigen to evaluate antibody response in human serum samples in Thailand with high specificity and sensitivity. In a previous pilot study, the Thailand-isolated 31-kDa proteins showed efficacy in dot-blot tests using serum samples from 435 human volunteers on Hawaii Island. However, we hypothesized that native antigen isolated from Hawaii A. cantonensis may exhibit higher specificity than the Thailand-isolated 31-kDa antigen due to potential minor variation in epitopes between isolates. In this study, 31-kDa glycoproteins were isolated by sodium dodecyl-sulfate polyacrylamide gel electrophoresis from adult A. cantonensis nematodes collected from rats captured on east Hawaii Island. The resultant proteins were purified by electroelution, pooled, bioanalyzed, and quantified. A subset of 148 samples from human participants of the original cohort of 435 was consented for this study, including 12 of the original 15 clinically diagnosed participants. Results of ELISA using the Hawaii-isolated 31-kDa antigen were compared with results of the same serum samples previously tested with both crude Hawaii antigen ELISA and Thailand 31-kDa antigen dot blot. This study shows a seroprevalence in the general population of East Hawaii Island of 25.0%, similar to previous findings of 23.8% seroprevalence in this cohort using crude antigen from Hawaii A. cantonensis and 26.5% using Thailand 31-kDa antigen.
Sujet(s)
Angiostrongylus cantonensis , Infections à Strongylida , Adulte , Humains , Animaux , Rats , Hawaï/épidémiologie , Projets pilotes , Études séroépidémiologiques , Glycoprotéines , Infections à Strongylida/diagnostic , Infections à Strongylida/épidémiologieRÉSUMÉ
Angiostrongylus cantonensis is the main causative agent for eosinophilic meningoencephalitis in humans. Larvae are rarely found in the cerebral spinal fluid (CSF). Consequently, serology and DNA detection represent important diagnostic tools. However, interpretation of the results obtained from these tools requires that more extensive accuracy studies be conducted. The aim of the present study is to update guidelines for diagnosis and case definitions of neuroangiostrongyliasis (NA) as provided by a working group of a recently established International Network on Angiostrongyliasis. A literature review, a discussion regarding criteria and diagnostic categories, recommendations issued by health authorities in China and an expert panel in Hawaii (USA), and the experience of Thailand were considered. Classification of NA cases and corresponding criteria are proposed as follows: minor (exposure history, positive serology, and blood eosinophilia); major (headache or other neurological signs or symptoms, CSF eosinophilia); and confirmatory (parasite detection in tissues, ocular chambers, or CSF, or DNA detection by PCR and sequencing). In addition, diagnostic categories or suspected, probable, and confirmatory are proposed. Updated guidelines should improve clinical study design, epidemiological surveillance, and the proper characterization of biological samples. Moreover, the latter will further facilitate accuracy studies of diagnostic tools for NA to provide better detection and treatment.
RÉSUMÉ
Neuroangiostrongyliasis (NAS) is an emerging tropical disease in humans and some animals which is caused by infection with the parasitic nematode Angiostrongylus cantonensis. It is the leading cause of eosinophilic meningitis worldwide. Diagnoses in humans and susceptible animals are generally presumptive and easily confused with other central nervous system disorders. The 31 kDa antigen is currently the only NAS immunodiagnostic assay that has achieved 100% sensitivity. However, little is known about the humoral immune response against the 31 kDa antigen in NAS infections, which would be critical for widespread adoption of this assay. We used the Hawai'i 31 kDa isolate in an indirect ELISA assay to confirm the presence of immunoglobulin IgG, IgM, IgA, and IgE isotypes in six-week post-infection plasma from lab-reared rats infected with 50 live, third-stage, A. cantonensis larvae isolated from a wild Parmarion martensi semi-slug. Our results confirmed the presence of all four isotypes against the Hawaii 31 kDa isolate, with sensitivity ranging from 22-100%. The IgG isotype showed 100% sensitivity in detecting A. cantonensis infection, which validates the use of IgG indirect ELISA with 31 kDa antigen as an effective immunodiagnostic assay for rats six weeks post-infection. Given each isotype may be present at different times during NAS infections, our data provides preliminary information on the humoral immune response to A. cantonensis infection in lab-reared rats and serves as a baseline for future studies.
RÉSUMÉ
Albendazole is considered the anthelmintic of choice for the management of rat lungworm disease (neuroangiostrongyliasis), due to its broad spectrum of nematocidal activity and its ability to cross the blood-brain barrier. Albendazole binds to ß-tubulins, preventing their polymerization into microtubules, thereby corrupting the cascade of cell division at metaphase, which ultimately leads to the death of individual cells and eventually the death of the parasite. Inhibition of microtubule formation will also hinder the axoplasmic transport system, affecting the neuronal activities of the parasite. While this mechanism has been explicated in other parasitic and non-parasitic nematodes, it has never been evaluated in Angiostrongylus cantonensis. This study evaluates the antimitotic effects of albendazole sulphoxide (active metabolite) on the microtubules of adult A. cantonensis using the tubulin polymerization assay and measures its effects on worm viability using the colorimetric MTT assay. Three different concentrations of albendazole (62.5 µM, 250 µΜ, and 1 mM) were evaluated. We saw a statistically significant dose-dependent reduction in the band intensity of polymerized tubulins (or microtubules) (P = 0.019), suggesting that albendazole imparts its antimitotic effect in a dose-dependent manner. Similarly, our MTT assay showed a dose-dependent decrease in formazan intensity (proportional to cell viability), suggesting that the rate of nematocidal activity of albendazole is also proportional to its concentration. In compiling the results from both these experiments, a correlation between the microtubule assembly and worm viability is evident.
Sujet(s)
Angiostrongylus cantonensis , Anthelminthiques , Antimitotiques , Infections à Strongylida , Animaux , Rats , Angiostrongylus cantonensis/physiologie , Albendazole/pharmacologie , Albendazole/usage thérapeutique , Tubuline , Antimitotiques/pharmacologie , Antimitotiques/usage thérapeutique , Formazanes , Anthelminthiques/pharmacologie , Anthelminthiques/usage thérapeutique , Antihelminthiques antinématodes/pharmacologie , Infections à Strongylida/traitement médicamenteux , Infections à Strongylida/parasitologieRÉSUMÉ
The parasitic nematode Angiostrongylus cantonensis (rat lungworm) is the leading cause of human eosinophilic meningitis worldwide. Most human infections occur through the accidental consumption of A. cantonensis hidden within produce as infectious third-stage larvae (L3), yet little research has been published addressing possible methods to mitigate this means of transmission. Here, we describe our tests of ozone gas-an oxidizing agent-and ultrasound, both used for disinfection of food and municipal water supplies and in industrial cleaning. We found that exposure to ozone, produced using two different commercially available ozone generators over varying durations of time and concentrations, was capable of achieving 100% larval mortality. In addition, we evaluated the impact of different sound frequencies on A. cantonensis L3 survival using two different commercially available ultrasonic cleaners, and found that 60 s of 40 kHz produced 46% mortality within 2 h. The combined use of ultrasound and ozone gas simultaneously resulted in a minimum of 89% normalized mean percent mortality within 2 h of treatment. Our study suggests that both ozone and ultrasound show high larvicidal efficacy, both independently and together, and thus show promise as methods for reducing the risk of rat lungworm infection via accidental consumption.
RÉSUMÉ
Rat lungworm (Angiostrongylus cantonensis) is a neurotropic nematode, and the leading cause of eosinophilic meningitis worldwide. The parasite is usually contracted through ingestion of infected gastropods, often hidden in raw or partially cooked produce. Pharmaceutical grade pyrantel pamoate was evaluated as a post-exposure prophylactic against A. cantonensis. Pyrantel pamoate is readily available over-the-counter in most pharmacies in the USA and possesses anthelmintic activity exclusive to the gastrointestinal tract (GIT). Administering pyrantel pamoate immediately after exposure should theoretically paralyze the larvae in the GIT, causing the larvae to be expelled via peristalsis without entering the systemic circulation. In this study, pyrantel pamoate (11 mg/kg) was orally administered to experimentally infected rats at 0, 2-, 4-, 6-, or 8-h post-infection. The rats were euthanized six weeks post-infection, and worm burden was evaluated from the heart-lung complex. This is the first in vivo study to evaluate its efficacy against A. cantonensis. This study demonstrates that pyrantel pamoate can significantly reduce worm burden by 53-72% (P = 0.004), and thus likely reduce the severity of infection that is known to be associated with worm burden. This paralyzing effect of pyrantel pamoate on the parasite may also be beneficial for delaying the establishment of infection until a more suitable anthelmintic such as albendazole is made available to the patient.
Sujet(s)
Angiostrongylus cantonensis , Anthelminthiques , Albendazole , Animaux , Anthelminthiques/usage thérapeutique , Pamoate de pyrantel/usage thérapeutique , RatsRÉSUMÉ
While there have been legitimate concerns in the past regarding the use of anthelmintics for the management of neuroangiostrongyliasis (rat lungworm disease), recent studies demonstrate that they can be considered safe and efficacious, particularly albendazole, which is regarded as the choice anthelmintic for its management. However, physician hesitancy to prescribe, as well as problems of availability persist, at least in Hawaii, which is considered the epicenter of this disease in the US. As a result, many patients suffer a diminished quality of life or even death. Here, we discuss recent studies that provide insights into new treatments and preventative interventions, which can be more rigorously used for the management of neuroangiostrongyliasis. In summary, results from recent studies suggest that albendazole and avermectins are beneficial for post-exposure management, pyrantel pamoate is beneficial as a post-exposure prophylactic, and levamisole is deserving of further study for the treatment of neuroangiostrongyliasis.
RÉSUMÉ
The safety and efficacy of benzimidazole anthelmintics for the treatment of rat lungworm disease (neuroangiostrongyliasis) have been questioned regardless of numerous experimental animal studies and clinical reports. In this review, 40 of these experimental animal studies and 104 clinical reports are compiled with a focus on albendazole. Among the 144 articles involving an estimated 1034 patients and 2561 animals, 4.1% were inconclusive or vague regarding the use of benzimidazoles. Of the remaining 138 articles, 90.5% found benzimidazoles to be safe and effective (885 patients, 2530 animals), 4.3% as safe but ineffective (73 patients, 3 animals), and 5.0% caused adverse reactions (7 patients, 28 animals). Among those clinical reports that described a confirmed diagnosis of neuroangiostrongyliasis in which albendazole monotherapy was used, 100% reported high efficacy (743 patients, 479 animals). In those where albendazole-corticosteroid co-therapy was used, 97.87% reported it to be effective (323 patients, 130 animals).
Sujet(s)
Angiostrongylus cantonensis , Anthelminthiques , Albendazole/effets indésirables , Animaux , Anthelminthiques/effets indésirables , Benzimidazoles/effets indésirables , Humains , Rats , Résultat thérapeutiqueRÉSUMÉ
BACKGROUND: Phytosanitary irradiation is used to control insect pests of quarantine concern on exported fresh horticultural products. Generic irradiation doses of 150 and 400 Gy are approved for tephritid fruit flies and all other insects, respectively. Other invertebrates such as gastropods (snails and slugs) may be classified as quarantine pests and require a disinfestation treatment. Parmarion martensi Simroth (Stylommatophora: Ariophantidae) is a semi-slug quarantine pest sometimes found on fresh sweet potatoes and other fruits and vegetables exported from Hawai'i to the continental USA. Also, P. martensi is a host of the parasitic nematode Angiostrongylus cantonensis (Rhabditida: Angiostrongylidae), the causative agent of neuroangiostrongyliasis or rat lungworm disease in humans. We conducted a study to determine if phytosanitary irradiation could control P. martensi and thereby reduce the risk of transmitting A. cantonensis in the USA. RESULTS: Two-, 12-, and 21-week-old P. martensi were treated with X-ray radiation at a dose of 150 or 400 Gy or left untreated as controls then held in the laboratory for up to 250 days. Survivorship and reproduction were recorded every 2-3 days and individual weights were measured biweekly. Irradiation at 150 and 400 Gy reduced growth and increased the mortality rate compared to untreated controls and prevented reproduction. CONCLUSION: Phytosanitary irradiation treatment at doses ≥150 Gy will prevent the establishment of viable populations of P. martensi. The literature on radiation tolerance in gastropods suggests that the internationally approved generic dose for tephritid fruit flies of 150 Gy may be effective against many slug and snail pest species.
Sujet(s)
Angiostrongylus cantonensis , Escargots/parasitologie , Angiostrongylus cantonensis/effets des radiations , Animaux , Reproduction , Rayons XSujet(s)
Angiostrongylus cantonensis/physiologie , Méningoencéphalite , Infections à Strongylida , Animaux , Humains , Méningoencéphalite/diagnostic , Méningoencéphalite/parasitologie , Méningoencéphalite/thérapie , Méningoencéphalite/transmission , Infections à Strongylida/diagnostic , Infections à Strongylida/thérapie , Infections à Strongylida/transmission , Infections à Strongylida/médecine vétérinaireRÉSUMÉ
The semi-slug, Parmarion martensi, is an intermediate host of the zoonotic nematode, Angiostrongylus cantonensis, the aetiological agent of neuroangiostrongyliasis or rat lungworm disease in humans. Rearing methods were developed for P. martensi to facilitate studies on nematode transmission and control. Parmarion martensi exhibited high survivorship when reared on a diet of dog food and fresh fruits and vegetables in temperature-controlled cabinets at 21.4°C, 98% relative humidity and 12:12 L:D cycle. Rearing containers were lined with moist paper towels for substrate and plastic pots were provided for hiding/resting and egg-laying. Under these conditions, time to first reproduction was 165.3 ± 12.3 days, fecundity was approximately 34.5 ± 7.8 eggs per adult, and hatch rate was 52.7 ± 3.2%. Survivorship post egg hatch was 86.2 ± 2.9% at 30 days (neonates had a mortality rate of about 14%) and 99% thereafter for up to a year. The demographics of laboratory-reared and wild-caught P. martensi were similar except for the weight of reproductive adults, which was significantly higher in laboratory-reared adults (4.0 ± 0.2 g) than in field-collected adults (1.5 ± 0.1 g).
Sujet(s)
Angiostrongylus cantonensis/parasitologie , Gastropoda/physiologie , Interactions hôte-parasite , Animaux , Démographie , Hawaï , Laboratoires , Dynamique des populationsRÉSUMÉ
Angiostrongylus cantonensis (rat lungworm) is a tropical and subtropical parasitic nematode, with infections in humans causing angiostrongyliasis (rat lungworm disease), characterized by eosinophilic meningitis. Hawaii has been identified as a global hotspot of infection, with recent reports of high infection rates in humans, as well as rat definitive and snail intermediate hosts. This study investigated variation in A. cantonensis infection, both prevalence and intensity, in wild populations of two species of rats (Rattus exulans and R. rattus) and one species of snail (Parmarion martensi). An overall infection prevalence of 86.2% was observed in P. martensi and 63.8% in rats, with R. exulans (77.4%) greater than R. rattus (47.6%). We found infections to vary with environmental and host-related factors. Body mass was a strong predictor of infection in all three species, with different patterns seen between sexes and species of rats. Infection prevalence and intensity for R. exulans were high in May 2018 and again in February 2019, but generally lower and more variable during the intervening months. Information on sources of variability of infection in wild host populations will be a crucial component in predicting the effectiveness of future disease surveillance or targeted management strategies.
Sujet(s)
Angiostrongylus cantonensis/physiologie , Gastropoda/parasitologie , Maladies des rongeurs/épidémiologie , Infections à Strongylida/médecine vétérinaire , Animaux , Femelle , Hawaï/épidémiologie , Mâle , Prévalence , Rats , Maladies des rongeurs/parasitologie , Spécificité d'espèce , Infections à Strongylida/épidémiologie , Infections à Strongylida/parasitologieRÉSUMÉ
Angiostrongylus cantonensis is the leading cause of eosinophilic meningitis worldwide, with life-threatening complications if not managed correctly. Previous in vitro studies have utilized change in motility patterns of adult female worms to assess the efficacy of anthelmintics qualitatively. However, it is the third stage larvae (L3) that are infectious to humans. With differential staining using propidium iodide penetration as the indicator of death, we can distinguish between dead and live larvae. This assay has enabled us to quantify the in vitro efficacy of nine clinically established anthelmintics on A. cantonensis L3. All drugs were tested at a 1 mm concentration. Piperazine and niclosamide were ineffective in inducing larval death; however, albendazole sulfoxide, pyrantel pamoate, diethylcarbamazine, levamisole and praziquantel were effective as compared to unexposed controls (P < 0.05). Ivermectin and moxidectin did not induce significant levels of mortality, but they considerably reduced larval motility almost immediately. This study indicates the need for further in vivo studies to determine the optimal dose and time frame for post-infection treatment with anthelmintics that demonstrated efficacy.
Sujet(s)
Angiostrongylus cantonensis/effets des médicaments et des substances chimiques , Anthelminthiques/pharmacologie , Infections à Strongylida/traitement médicamenteux , Angiostrongylus cantonensis/croissance et développement , Animaux , Femelle , Larve/effets des médicaments et des substances chimiques , Larve/croissance et développementRÉSUMÉ
Education is essential for lowering cases of Angiostrongyliasis in Hawaii. A professional development course on rat lungworm disease (RLWD) prevention was offered to Hawaii K-12 teachers during the 2018-2019 school year. Fourteen teachers, 11 from Hawaii Island and three from Maui, representing grades K, 2, 6-8, and 10-12, completed workshops and activities and received course credit. Teachers reached 652 students and educated 86 teachers and staff and approximately 900 community members including parents. A pre-course survey showed 55% of teachers agreed to a basic understanding of rat lungworm and its impact on Hawaii; this increased to 100% post-course. A sixth-grade class was the first to document the arrival of a highly competent intermediate host of the rat lungworm in a new location, an important discovery that initiated community awareness and control efforts. Six, 1-day workshops were held in 2018-2019, each on one of the main Hawaiian Islands. These were attended by 106 participants including teachers, community educators, and interested individuals from agencies, non-profits, businesses and the private sector. Of participants surveyed, 100% responded the workshop improved their overall understanding of RLWD. Efforts are being made to continue these programmes.
Sujet(s)
Santé publique/enseignement et éducation , Établissements scolaires , Infections à Strongylida/parasitologie , Angiostrongylus cantonensis , Animaux , HawaïRÉSUMÉ
BACKGROUND: Angiostrongylus cantonensis (Ac), or the rat lungworm, is a major cause of eosinophilic meningitis. Humans are infected by ingesting the 3rd stage larvae from primary hosts, snails, and slugs, or paratenic hosts. The currently used molecular test is a qPCR assay targeting the ITS1 rDNA region (ITS1) of Ac. METHODS: In silico design of a more sensitive qPCR assay was performed based on tandem repeats predicted to be the most abundant by the RepeatExplorer algorithm. Genomic DNA (gDNA) of Ac were used to determine the analytical sensitivity and specificity of the best primer/probe combination. This assay was then applied to clinical and environmental samples. RESULTS: The limit of detection of the best performing assay, AcanR3990, was 1 fg (the DNA equivalent of 1/100 000 dilution of a single 3rd stage larvae). Out of 127 CDC archived CSF samples from varied geographic locations, the AcanR3990 qPCR detected the presence of Ac in 49/49 ITS1 confirmed angiostrongyliasis patients, along with 15/73 samples previously negative by ITS1 qPCR despite strong clinical suspicion for angiostrongyliasis. Intermediate hosts (gastropods) and an accidental host, a symptomatic horse, were also tested with similar improvement in detection observed. AcanR3990 qPCR did not cross-react in 5 CSF from patients with proven neurocysticercosis, toxocariasis, gnathostomiasis, and baylisascariasis. AcanR3990 qPCR failed to amplify genomic DNA from the other related Angiostrongylus species tested except for Angiostrongylus mackerrasae (Am), a neurotropic species limited to Australia that would be expected to present with a clinical syndrome indistinguishable from Ac. CONCLUSION: These results suggest AcanR3990 qPCR assay is highly sensitive and specific with potential wide applicability as a One Health detection method for Ac and Am.
Sujet(s)
Angiostrongylus cantonensis , Angiostrongylus , Méningite , Infections à Strongylida , Angiostrongylus cantonensis/génétique , Animaux , Equus caballus , Humains , Rats , Infections à Strongylida/diagnosticRÉSUMÉ
On Hawai'i Island, an increase in human neuroangiostrongyliasis cases has been primarily associated with the accidental ingestion of Angiostrongylus cantonensis L3 in snails or slugs, or potentially, from larvae left behind in the slug's slime or feces. We evaluated more than 40 different treatments in vitro for their ability to kill A. cantonensis larvae with the goal of identifying a safe and effective fruit and vegetable wash in order to reduce the risk of exposure. Our evaluation of treatment lethality was carried out in two phases; initially using motility as an indicator of larval survival after treatment, followed by the development and application of a propidium iodide staining assay to document larval mortality. Treatments tested included common household products, consumer vegetable washes and agricultural crop washes. We found minimal larvicidal efficacy among consumer-grade fruit and vegetable washes, nor among botanical extracts such as those from ginger or garlic, nor acid solutions such as vinegar. Alkaline solutions, on the other hand, as well as oxidizers such as bleach and chlorine dioxide, did show larvicidal potential. Surfactants, a frequent ingredient in detergents that lowers surface tension, had variable results, but dodecylbenzene sulfonic acid as a 70% w/w solution in 2-propanol was very effective, both in terms of the speed and the thoroughness with which it killed A. cantonensis L3 nematodes. Thus, our results suggest promising directions for future investigation.
Sujet(s)
Angiostrongylus cantonensis/effets des médicaments et des substances chimiques , Anthelminthiques/pharmacologie , Angiostrongylus cantonensis/croissance et développement , Animaux , Larve/effets des médicaments et des substances chimiques , Larve/croissance et développementRÉSUMÉ
Angiostrongylus cantonensis is a parasitic nematode known to infect humans through the ingestion of third stage larvae which can cause inflammation and damage to the central nervous system. Currently, polymerase chain reaction (PCR) is one of the most reliable diagnostic methods for detecting A. cantonensis in humans as well as in gastropod hosts, but requires expensive and specialized equipment. Here, we compare the sensitivity and accuracy of a recombinase polymerase amplification Exo (RPA-EXO) assay, and a recombinase polymerase amplification lateral flow assay (RPA-LFA) with a traditional quantitative PCR (qPCR) assay currently available. The three assays were used to test 35 slugs from Hawai'i for the presence of A. cantonensis DNA. Consistent results among the three tests were shown in 23/35 samples (65.7%), while 7/35 (20%) were discordant in low infection level samples (<0.01 larvae per mg tissue), and 5/35 (14.3%) were equivocal. To evaluate sensitivity, a partial ITS1 gene was cloned, and serial plasmid dilutions were created ranging from 100 copies µL-1 to ~1 copy µL-1. All three assays consistently detected 50-100 copies µL-1 in triplicate and qPCR was able to detect ~13 copies µL-1 in triplicate. RPA-EXO was able to detect 25 copies µL-1 in triplicate and RPA-LFA was not able to amplify consistently below 50 copies µL-1. Thus, our RPA-EXO and RPA-LFA assays do not appear as sensitive as the current qPCR assay at low DNA concentrations; however, these tests have numerous advantages that may make them useful alternatives to qPCR.
Sujet(s)
Angiostrongylus cantonensis/isolement et purification , ADN des helminthes/analyse , Tests diagnostiques courants/méthodes , Gastropoda/parasitologie , Angiostrongylus cantonensis/enzymologie , AnimauxRÉSUMÉ
The rat lungworm (Angiostrongylus cantonensis) has emerged as an important human and animal health concern in Hawaii, US. Although the life cycle of the parasite requires both rat and gastropod hosts, other animals acting as paratenic hosts, such as frogs and centipedes, have been identified as sources of infection. We investigated the occurrence of rat lungworm infections in potential paratenic hosts in Hawaii to provide information on how they might be involved in transmission of angiostrongyliasis. We confirmed the presence of rat lungworm in 87% (21/24) of introduced Puerto Rican coqui frogs (Eleutherodactylus coqui) in Hilo, Hawaii, by real-time PCR. Additionally, four Cuban greenhouse frogs (Eleutherodactylus planirostris), two cane toads (Rhinella marina), and three centipedes (Scolopendra subspinipes) were found to be infected. In the frogs and toads, multiple tissue types were positive, including stomach and intestine, muscle, liver, heart, and brain, indicating larval migration. We identified rat lungworm infections in frogs, toads, and centipedes in Hawaii and highlighted the lack of knowledge of the role paratenic hosts may be playing in the transmission and life cycle maintenance of rat lungworm in Hawaii.