Proteomic Profiling Reveals New Insights into the Allergomes of Anisakis simplex, Pseudoterranova decipiens, and Contracaecum osculatum.

Anisakis simplex, Pseudoterranova decipiens, and Contracaecum osculatum third-stage larvae (L3) are fish-borne nematodes that can cause human anisakidosis. Although A. simplex is a known source of allergens, knowledge about the allergic potential of P. decipiens and C. osculatum is limited. Therefore, we performed comparative proteomic profiling of A. simplex, P. decipiens, and C. osculatum L3 larvae using liquid chromatography-tandem mass spectrometry. In total, 645, 397, and 261 proteins were detected in A. simplex, P. decipiens, and C. osculatum L3 larvae, respectively. Western blot analysis confirmed the cross-reactivity of anti-A. simplex immunoglobulin (Ig)G antibodies with protein extracts from P. decipiens and C. osculatum L3 larvae. The identified proteins of the Anisakidae proteomes were characterized by label-free quantification and functional analysis, and proteins involved in many essential biological mechanisms, such as parasite survival, were identified. In the proteome of A. simplex 14, the following allergens were identified: Ani s 1, Ani s 2 (2 isomers), Ani s 3 (2 isomers), Ani s 4, Ani s 8, Ani s 9, Ani s 10, Ani s 11-like, Ani s 13, Ani s fructose 1,6-bisphosphatase, Ani s phosphatidylethanolamine-binding protein (PEPB), and Thu a 3.0101. The following 8 allergens were detected in P. decipiens: Ani s 2, Ani s 3 (2 isomers), Ani s 5, Ani s 8, Ani s 9, Ani s PEPB, and Ani s troponin. In C. osculatum 4, the following allergens were identified: Ani s 2, Ani s 5, Ani s 13, and Asc l 3. Furthermore, 28 probable allergens were predicted in A. simplex and P. decipiens, whereas in C. osculatum, 25 possible allergens were identified. Among the putative allergens, heat shock proteins were most frequently detected, followed by paramyosin, peptidyl-prolyl cis-trans isomerase, enolase, and tropomyosin. We provide a new proteomic data set that could be beneficial for the discovery of biomarkers or drug target candidates. Furthermore, our findings showed that in addition to A. simplex, P. decipiens and C. osculatum should also be considered as potential sources of allergens that could lead to IgE-mediated hypersensitivity.

Antigen detection: Insights into Toxocara and other ascarid infections in dogs and cats.

Detection of ascarid excreted or secreted (E/S) molecules is an alternative approach to the identification of infection by egg flotation. E/S molecules serve as direct markers for the ascarid nematode commonly found in cats and dogs (Toxocara spp., Toxascaris leonina and Baylisascaris procyonis). The nematode derived E/S material mixes with the intestinal contents of the host animal and is available for detection as a coproantigen in the host's faeces. Antigen capture immunological techniques allow sensitive coproantigen detection. Different patterns of antigen to egg agreement are demonstrated in an experimental Toxocara canis infection throughout the prepatent, patent, and post-treatment phases. Examination of faecal samples from a large field population of dogs and cats tested for both egg shedding and antigen indicates that more infections were identified by antigen. Host age influences the agreement of antigen and Toxocara egg results. Older dogs and cats were less likely to have a patent infection (egg positive and antigen positive) result pattern. An egg observation in the absence of antigen detection may indicate a spurious egg. The impact of spurious eggs was further examined by comparisons of cohorts of dogs separated by presence or absence of a pseudoparasite observation or by egg semi-quantification bin. Lastly, the antigen to egg agreement was calculated for other ascarid species.

Baylisascariasis: A young boy with neural larva migrans due to the emerging raccoon round worm.

A 17-month-old boy from Vancouver, Canada, presented with a 5-day history of progressive somnolence, ataxia, and torticollis. Additional investigations revealed eosinophilic encephalitis with deep white matter changes on MR imaging. On day 13, serology came back positive for Baylisascaris procyonis antibodies. While prophylaxis after ingestion of soil or materials potentially contaminated with raccoon feces can prevent baylisascariasis, timely treatment can sometimes alter a disastrous outcome. Populations of infected raccoons are propagating globally, but cases of Baylisascaris neural larva migrans have so far only been reported from North America.

Treatment of larva migrans syndrome with long-term administration of albendazole.

BACKGROUND: Larva migrans syndrome is a food-borne parasitic disease in humans, caused by accidental ingestion of eggs or larvae of ascarid nematodes, namely, Toxocara canis, Toxocara cati, or Ascaris suum, the roundworms commonly found in the intestines of dogs, cats and pigs respectively. When a patient is diagnosed as having larva migrans syndrome, oral-administration of albendazole is recommended, however, the regimen remains controversial worldwide. In Japan, the duration of albendazole administration is longer than those of European and North American countries. The purpose of this study was to assess the efficacy and safety of long-term administration treatment of albendazole for larva migrans syndrome. METHODS: From 2004 to 2014, our laboratory was involved in the diagnosis of 758 larva migrans syndrome cases, of which 299 cases could be followed up after the treatment. We analyzed these 299 follow-up cases on the ELISA results before and after the treatment as well as on anthelmintic used, dose and duration of medication, clinical findings, and side effects, recorded on a consultation sheet provided by the attending physicians. We have 288 cases as the subjects of this study. RESULTS: Albendazole represented a 78.0% efficacy rate. The side effects represented 15.0% in using albendazole alone cases; however, the side effects were mild to moderate and there were no severe cases reported. CONCLUSIONS: The long-term administration treatment of albendazole is safe and effective for larva migrans syndrome.

Comparative analysis of excretory-secretory antigens of Anisakis simplex, Pseudoterranova decipiens and Contracaecum osculatum regarding their applicability for specific serodiagnosis of human anisakidosis based on IgG-ELISA.

Serodiagnosis of human anisakidosis is presently hampered by the current lack of standardised serological assays that allow sensitive and specific detection of Anisakidae-specific antibodies in human patients. In the present study, we comparatively evaluated the diagnostic value (by IgG-ELISA) of excretory-secretory antigens (ESAgs) of Anisakis simplex, Pseudoterranova decipiens and Contracaecum osculatum, representing the most frequently found genera responsible for human infection. In addition, we tested also a mix of the three ES preparations (Mix-ESAgs) as well as two recombinant allergens of A. simplex, rAni s 1 and rAni s 7. ES antigen from C. osculatum yielded the best diagnostic performance in IgG-ELISA-based serodiagnosis of the Spanish anisakidosis patients investigated in this study (relative serodiagnostic sensitivity 100%; specificity 89%) as compared to A. simplex ES-antigen (93% versus 57%) and P. decipiens (67% versus 93%) or a mix of the three ES antigens (100% versus 44%), respectively. Cross-reactions of C. osculatum ES antigen with serum-antibodies from patients suffering from other helminth infections were rare and were exclusively found with few sera from toxocariasis, ascariasis, and filariasis patients. The two recombinant allergens rAni s 1 and rAni s 7 did not prove sufficiently sensitive and specific in order to justify a further evaluation of these antigens regarding their suitability in IgG-ELISA-based serodiagnosis of human anisakidosis. In conclusion, the C. osculatum-ESAg-ELISA remains as key candidate to be further assessed for the serodiagnosis of symptomatic anisakidosis in different endemic regions.

Detection and Evaluation of Antibody Response to a Baylisascaris-Specific Antigen in Rodent Hosts with the Use of Western Blotting and Elisa.

Diagnosis of parasitic diseases that involve tissue-stage larvae is challenging, and serology remains the most effective antemortem test for detecting these infections. Baylisascaris procyonis, the raccoon roundworm, is a zoonotic ascarid. Raccoons are the usual definitive host, and humans may be infected as accidental hosts. More than 150 species of birds and mammals may act as paratenic hosts, and rodents play an important role in the transmission and maintenance of this parasite in nature. Migratory larvae in paratenic host tissues can produce ocular disease and severe to fatal neurologic disease, but not all infected hosts develop signs. A sensitive and specific Western blot (WB) assay based on a recombinant Baylisascaris-specific antigen (rBpRAG-1) has been developed for use in humans. We evaluated the use of this antigen to detect Baylisascaris spp. infections in rodent paratenic hosts. With the use of 4 species of Peromyscus mice ( Peromyscus californicus, Peromyscus leucopus, Peromyscus maniculatus, Peromyscus polionotus) from a previous infection trial, we developed species-adapted WB and ELISA assays and evaluated performance compared to detection of larvae in tissue samples. These assays revealed species-level differences in seroconversion and terminal antibody concentrations, with P. leucopus developing significantly greater antibody concentrations than P. californicus and P. polionotus at all dose levels, and P. maniculatus at the low dose. Some P. californicus and P. polionotus failed to seroconvert despite the recovery of larvae from their tissues. WB and ELISA results were correlated; however, the WB demonstrated higher sensitivity than the ELISA overall (72.2% versus 63.9%, respectively). With the use of experimental samples, specificity was 100% for WB and 94.1% for ELISA. A WB was also used to test Mus and Rattus samples, and although numbers were too limited to evaluate sensitivity and specificity, all animals known to be infected by tissue digestion were WB positive, and all uninfected animals were negative. Finally, the Peromyscus-adapted WB and ELISA were used to test a set of serum samples from wild-trapped P. maniculatus and Rattus rattus. Both assays were generally sensitive, but specificity was equivocal. This emphasizes the challenge of using serology for investigation of wildlife diseases, in which hosts have unknown exposure histories. Nevertheless, serologic methods have utility in the study of Baylisascaris spp. in paratenic hosts, either wild or captive, and have advantageous attributes (non-lethal, high-throughput), but results should be interpreted carefully.

Immunomodulatory effects of excretory/secretory compounds from Contracaecum osculatum larvae in a zebrafish inflammation model.

Excretory/secretory (ES) compounds isolated from third-stage larvae of the anisakid nematode Contracaecum osculatum parasitizing liver of Baltic cod were investigated for effects on immune gene expression in a zebrafish LPS-induced inflammation model. ES products containing a series of proteins, of which some had enzymatic activity, were injected solely or with LPS. ES proteins alone induced up-regulation of a number of immune-related genes, but generally to a lower degree compared to LPS. When co-injected with LPS, the worm products exacerbated merely expression of five genes affecting Th1, Th2, Th17 and innate responses compared to the LPS-injected group. However, the level of overexpression decreased in an inverse dose-dependent manner. The immune regulating action of C. osculatum ES products is interpreted as an important evolutionary ability of larval parasites in the transport host which makes it less susceptible to host immune responses whereby the probability of reaching the final host is increased.

Allergenic activity of Pseudoterranova decipiens (Nematoda: Anisakidae) in BALB/c mice.

BACKGROUND: Anisakis simplex is the only fishery-product associated parasite causing clinical allergic responses in humans so far. However, other anisakids, due to the presence of shared or own allergens, could also lead to allergic reactions after sensitization. The aim of this study was to determine if Pseudoterranova decipiens belonging to the family Anisakidae has allergenic activity and is able to induce sensitization after oral administration in a murine (BALB/c mice) model. RESULTS: The ingestion of A. pegreffii proteins by BALB/c mice, which had been previously sensitized by intraperitoneal inoculation with the corresponding live L3 larvae, triggers signs of allergy within 60 min, whereas P. decipiens did to a lesser extent. Beside symptoms, allergic reactions were furtherly supported by the presence of histamine in sera of sensitized mice. Specific IgG1 and IgE responses were detected in sera of all sensitized mice from week four. Specific IgG2a response was detected in sera from mice sensitized to P. decipiens. After polyclonal or specific activation with anti-CD3/anti-CD28 or antigens, respectively, splenocytes from mice infected i.p. with A. pegreffii or P. decipiens larvae showed significantly higher production of IL-10 than naïve mice. After stimulation with specific antigens, significantly higher IL-5 and IL-13 amounts were produced by specific antigen stimulated splenocytes than by the naïve cells; only P. decipiens proteins induced IFN-É£. CONCLUSIONS: The overall results suggest that infection with P. decipiens can sensitize mice to react to subsequent oral challenge with anisakid proteins, as described for A. simplex (sensu stricto) and A. pegreffii infections. The results show that anisakid proteins induce a dominant Th2 response, although P. decipiens could also induce a mixed type 1/type 2 pattern.

Resposta imunológica a antígenos de Hysterothylacium deardorffoverstreetorum de peixes teleósteos / Imunne response against Hysterothylacium deardorffoverestreetorum from teleost fish

Anisaquidose é uma doença provocada por parasitos da família Anisakidae e se caracteriza por manifestações gastrointestinais e alérgicas. O Anisakis simplex é o parasito mais patogênico ao homem e altamente alergênico. Porém, outros anisaquídeos também são danosos aos humanos, mas é desconhecida a imunogenicidade dessas larvas. O objetivo deste trabalho foi avaliar o potencial imunogênico do parasito Hysterothylacium deardorffoverestreetorum (HD) em modelo murino. Camundongos da linhagem BALB/c foram divididos em três grupos experimentais e receberam as preparações antigênicas obtidas de larvas de HD: extrato bruto de larvas (EBH), extrato secretado/ excretado de larvas (ESH) e extrato bruto de larvas após excreção/secreção (EEH). Amostras séricas foram obtidas em diferentes dias após imunização para determinação dos níveis de anticorpos específicos pelo ensaio imunoenzimático (ELISA). Os resultados demonstram aumento na produção de imunoglobulina (Ig) G após a segunda imunização, com aumento progressivo após a terceira imunização. Já em relação à IgE, a reatividade foi mais tardia, demonstrando aumento progressivo após a terceira imunização. Foi avaliada a imunidade celular por meio da intradermorreação, como resultado estatisticamente significativo em relação ao controle utilizado. Este experimento é a primeira descrição da potencialidade patogênica desse parasito em mamíferos e representa um avanço no diagnóstico da anisaquidose humana.(AU)

Anisaquidosis is a disease caused by parasites of Anisakidae family and is characterized by gastrointestinal and allergic reactions. The Anisakis simplex is a more pathogenic Anisakidae to humans and is highly allergenic. However, other species of this family also have characteristics that are harmful to humans, but little is known about the immunogenicity this parasites. The objective of this study was to experimentally assess the immunogenic potential of the parasite Hysterothylacium deardorffoverestreetorum (H.D) in mice. Mice of inbred BALB/c strain were divided into three groups and received three immunizations of the following antigenic preparations obtained from L3 larvae H.D: Crude larval extract of H.D (CEH) Extract secreted / excreted larvae H.D. (ESH) and crude extract of larvae after excretion / secretion (EEH). Serum samples were obtained on different days after immunization to determine the levels of circulating specific antibodies by enzyme-linked immunosorbent assay (ELISA). The results show increased production of immunoglobulin (Ig) G after the second immunization with a gradual increase after the third immunization. Regarding IgE reactivity, this occurred later, demonstrating a progressive increase only after the third immunization. Cellular immunity was evaluated by intradermal, and showed statistically significant result compared to the control used. This experiment is the first description of the pathogenic potential of this parasite in mammals and represents a breakthrough in the diagnosis of human Anisakidosis.(AU)

Baylisascaris procyonis Roundworm Seroprevalence among Wildlife Rehabilitators, United States and Canada, 2012-2015.

Baylisascaris procyonis roundworms can cause potentially fatal neural larva migrans in many species, including humans. However, the clinical spectrum of baylisascariasis is not completely understood. We tested 347 asymptomatic adult wildlife rehabilitators for B. procyonis antibodies; 24 were positive, suggesting that subclinical baylisascariasis is occurring among this population.

Equine antibody response to larval Parascaris equorum excretory-secretory products.

Parascaris equorum is an intestinal nematode of foals and young horses that can produce mild to severe pathology. Current diagnosis is limited to detection of patent infections, when parasite eggs are identified during fecal examinations. This study examined the use of larval P. equorum excretory-secretory (ES) products in a western blot test for diagnosis of prepatent equine P. equorum infection. Sera from adult mares negative for patent P. equorum infections, foals prior to consuming colostrum, and P. equorum infected foals were used as controls in this study. Study samples included sera from 18 broodmares prior to parturition and sera from their foals throughout the process of natural infection. Sera from study horses were examined for IgG(T) antibody recognition of ES products. Foals naturally infected with P. equorum possessed IgG(T) antibodies against 19kDa, 22kDa, 26kDa, and 34kDa ES products. However, passive transfer of colostral antibodies from mares was shown to preclude the use of the crude larval ES product-based western blot test for diagnosis of prepatent P. equorum infections in foals.

Potential of recombinant inorganic pyrophosphatase antigen as a new vaccine candidate against Baylisascaris schroederi in mice.

The intestinal nematode Baylisascaris schroederi is an important cause of death for wild and captive giant pandas. Inorganic pyrophosphatases (PPases) are critical for development and molting in nematode parasites and represent potential targets for vaccination. Here, a new PPase homologue, Bsc-PYP-1, from B. schroederi was identified and characterized, and its potential as a vaccine candidate was evaluated in a mouse challenge model. Sequence alignment of PPases from nematode parasites and other organisms show that Bsc-PYP-1 is a nematode-specific member of the family I soluble PPases. Immunohistochemistry revealed strong localization of native Bsc-PYP-1 to the body wall, gut epithelium, ovary and uterus of adult female worms. Additionally, Bsc-PYP-1 homologues were found in roundworms infecting humans (Ascaris lumbricoides), swine (Ascaris suum) and dogs (Toxocara canis). In two vaccine trials, recombinant Bsc-PYP-1 (rBsc-PYP-1) formulated with Freund complete adjuvant induced significantly high antigen-specific immunoglobulin (Ig)G but no IgE or IgM responses. Analysis of IgG-subclass profiles revealed a greater increase of IgG1 than IgG2a. Splenocytes from rBsc-PYP-1/FCA-immunized mice secreted low levels of T helper (Th)1-type cytokines, interferon-γ and interleukin (IL)-2, while producing significantly high levels of IL-10 and significantly elevated levels of IL-4 (Th2 cytokines) after stimulation with rBsc-PYP-1 in vitro. Finally, vaccinated mice had 69.02-71.15% reductions (in 2 experiments) in larval recovery 7 days post-challenge (dpc) and 80% survival at 80 dpc. These results suggest that Th2-mediated immunity elicited by rBsc-PYP-1 provides protection against B. schroederi, and the findings should contribute to further development of Bsc-PYP-1 as a candidate vaccine against baylisascariasis.

Interlaboratory optimization and evaluation of a serological assay for diagnosis of human baylisascariasis.

A Western blot assay using a recombinant protein, recombinant Baylisascaris procyonis RAG1 protein (rBpRAG1), was developed for the diagnosis of human baylisascariasis concurrently by the Centers for Disease Control and Prevention (CDC) in Atlanta, Georgia, and the National Reference Centre for Parasitology (NRCP) in Montreal, Canada. Assay performance was assessed by testing 275 specimens at the CDC and 405 specimens at the NRCP. Twenty specimens from 16 cases of baylisascariasis were evaluated. Eighteen were positive, with the assay correctly identifying 14 of 16 patients. The rBpRAG1 Western blot assay showed no cross-reactivity with Toxocara-positive serum and had an overall sensitivity of 88% and a specificity of 98%.

Recent advances in diagnosing pathogenic equine gastrointestinal helminths: the challenge of prepatent detection.

Parasites infecting horses are ubiquitous and clinically important across the world. The major parasitic threats to equine health are cyathostomins, Parascaris equorum, Anoplocephala perfoliata, and Strongylus vulgaris. Increasing levels of anthelmintic resistance reported world wide in equine parasites have led to recommendations of constructing sustainable parasite control programmes based on systematic surveillance of parasite levels. Regulations at the European Union level now make anthelmintics available on prescription-only basis and disallow prophylactic treatment. This emphasizes the needs for reliable and practical diagnostic tools for detection of major parasites infecting equines. The current, widely used coprological techniques are important and useful, but they do have considerable limitations as they are incapable of diagnosing the pathogenic migrating stages. Species-specific molecular assays have been developed for diagnosing patent infections with 21 cyathostomin species, A. perfoliata, and S. vulgaris, but none of these have found use in practice. An antibody-directed enzyme-linked immunosorbent assay (ELISA) has been developed, validated and made commercially available for diagnosing A. perfoliata infection, but interpretation is complicated by the fact that horses not harbouring tapeworms can maintain elevated antibody titres. Recent work with a coproantigen ELISA has shown promise for reliable detection of current A. perfoliata infection. Perhaps most remarkable is the fact that the pathogenic larval stages of cyathostomins and large strongyles cannot be detected by any of the available diagnostics. With the lengthy prepatency periods characterizing these parasites, there is a huge need for developing such assays. The recent identification of a possible diagnostic marker for encysted cyathostomins holds great promise, and could become very useful in clinical practice. Several attempts have been made to construct assays for diagnosing the highly pathogenic migrating larvae of S. vulgaris, but none of these have performed sufficiently to make a useful test. The present review illustrates that classical coprological techniques remain the cornerstone of equine parasitology diagnosis and surveillance, and will remain so in a foreseeable future. However, promising progress has been made for developing assays capable of diagnosing prepatent stages of strongyle infection, and there is reason to hope for validated and useful assays in the relative near future.

Sequence analysis of the Bs-Ag1 gene of Baylisascaris schroederi from the giant panda and an evaluation of the efficacy of a recombinant Baylisascaris schroederi Bs-Ag1 antigen in mice.

The Baylisascaris schroederi infection rate among wild giant pandas may reach over 50% or even 100%, making it one of the leading causes of death from primary or secondary infection in wild populations. Until now, little was known about how protective immunity to B. schroederi infection could be achieved. The present study was conducted to evaluate the immunogenicity and protective efficacy of recombinant Bs-Ag1 from B. schroederi, by cloning the full-length Bs-Ag1 gene of B. schroederi and expressing it in a heterologous host, Escherichia coli BL21. In mice vaccinated with rBs-Ag1 coupled with Freund's complete adjuvant (FCA), there was a significant reduction (69.2%) in the recovery of challenged B. schroederi L3 compared with either nonvaccinated controls or mice vaccinated with FCA alone. Our study supports the use of Bs-Ag1 as a potential candidate for vaccination against B. schroederi infection and provides basic data for further vaccination trials with mixtures of antigens (with Bs-Ag2 and Bs-Ag3) to B. schroederi.

Recombinant antigen-based enzyme-linked immunosorbent assay for diagnosis of Baylisascaris procyonis larva migrans.

Baylisascaris larva migrans is an important zoonotic disease caused by Baylisascaris procyonis, the raccoon roundworm, and is being increasingly considered in the differential diagnosis of eosinophilic meningoencephalitis in children and young adults. Although a B. procyonis excretory-secretory (BPES) antigen-based enzyme-linked immunosorbent assay (ELISA) and a Western blot assay are useful in the immunodiagnosis of this infection, cross-reactivity remains a major problem. Recently, a recombinant B. procyonis antigen, BpRAG1, was reported for use in the development of improved serological assays for the diagnosis of Baylisascaris larva migrans. In this study, we tested a total of 384 human patient serum samples in a BpRAG1 ELISA, including samples from 20 patients with clinical Baylisascaris larva migrans, 137 patients with other parasitic infections (8 helminth and 4 protozoan), and 227 individuals with unknown/suspected parasitic infections. A sensitivity of 85% and a specificity of 86.9% were observed with the BpRAG1 ELISA, compared to only 39.4% specificity with the BPES ELISA. In addition, the BpRAG1 ELISA had a low degree of cross-reactivity with antibodies to Toxocara infection (25%), while the BPES antigen showed 90.6% cross-reactivity. Based on these results, the BpRAG1 antigen has a high degree of sensitivity and specificity and should be very useful and reliable in the diagnosis and seroepidemiology of Baylisascaris larva migrans by ELISA.

Molecular cloning of an immunogenic protein of Baylisascaris procyonis and expression in Escherichia coli for use in developing improved serodiagnostic assays.

Larva migrans caused by Baylisascaris procyonis is an important zoonotic disease. Current serological diagnostic assays for this disease depend on the use of the parasite's larval excretory-secretory (ES) antigens. In order to identify genes encoding ES antigens and to generate recombinant antigens for use in diagnostic assays, construction and immunoscreening of a B. procyonis third-stage larva cDNA expression library was performed and resulted in identification of a partial-length cDNA clone encoding an ES antigen, designated repeat antigen 1 (RAG1). The full-length rag1 cDNA contained a 753-bp open reading frame that encoded a protein of 250 amino acids with 12 tandem repeats of a 12-amino-acid long sequence. The rag1 genomic DNA revealed a single intron of 837 bp that separated the 753-bp coding sequence into two exons delimited by canonical splice sites. No nucleotide or amino acid sequences present in the GenBank databases had significant similarity with those of RAG1. We have cloned, expressed, and purified the recombinant RAG1 (rRAG1) and analyzed its diagnostic potential by enzyme-linked immunosorbent assay. Anti-Baylisascaris species-specific rabbit serum showed strong reactivity to rRAG1, while only minimal to no reactivity was observed with sera against the related ascarids Toxocara canis and Ascaris suum, strongly suggesting the specificity of rRAG1. On the basis of these results, the identified RAG1 appears to be a promising diagnostic antigen for the development of serological assays for specific detection of B. procyonis larva migrans.

Experimental lagochilascariosis in X-chromosome-linked immunodeficient mice.

Lagochilascaris minor is the etiological agent of lagochilascariosis, a disease that affects the neck region and causes exudative abscesses, with eggs, adult parasites and L3/L4 larvae in the purulent exudates. Mice are now considered to be intermediate hosts for the parasite. To determine the pattern of infection in B1 cell-deficient mice, experimental lagochilascariosis was studied in BALB/c and X-chromosome-linked immunodeficient (xid) mice. BALB.xid-infected mice showed lower numbers of larvae. Third-stage larvae, fourth-stage larvae and adult parasites were found in both strains. BALB/c mice produced IgM, IgG, IgA and IgE against the crude extract and secreted/excreted antigens of the parasite. On the other hand, BALB.xid mice did not produce IgM and produced lower levels of IgG and IgA, and similar quantities of IgE.

Experimental lagochilascariosis in X-chromosome-linked immunodeficient mice / Lagochilascariose experimental em camundongos com imunodeficiência ligada ao cromossomo X

Lagochilascaris minor is the etiological agent of lagochilascariosis, a disease that affects the neck region and causes exudative abscesses, with eggs, adult parasites and L3/L4 larvae in the purulent exudates. Mice are now considered to be intermediate hosts for the parasite. To determine the pattern of infection in B1 cell-deficient mice, experimental lagochilascariosis was studied in BALB/c and X-chromosome-linked immunodeficient (xid) mice. BALB.xid-infected mice showed lower numbers of larvae. Third-stage larvae, fourth-stage larvae and adult parasites were found in both strains. BALB/c mice produced IgM, IgG, IgA and IgE against the crude extract and secreted/excreted antigens of the parasite. On the other hand, BALB.xid mice did not produce IgM and produced lower levels of IgG and IgA, and similar quantities of IgE.

Lagochilascaris minor é o agente etiológico da lagochilascariose, uma doença que afeta a região de pescoço provocando abscessos exudativos contendo ovos, parasitas adultos e larvas L3/L4 nos exudates purulentos. Atualmente, camundongos são considerados hospedeiros intermediários do parasita. Para determinar o padrão de infecção em camundongos deficientes de células B1, a lagochilascariose experimental foi estudada em camundongos BALB/c e em camundongos com imunodeficiência ligada ao cromossomo X (xid). Camundongos BALB.xid infectados mostraram menor número de larvas. Larvas L3, L4 e parasitas adultos foram encontrados em ambas as linhagens. Camundongos BALB/c produziram IgM, IgG, IgA e IgE contra o extrato bruto e antígenos secretados/excretados do parasita; por outro lado, camundongos BALB.xid não produziram IgM, produziram baixos níveis de IgG e IgA, e quantidades semelhantes de IgE.

Sequence analysis of Bs-Ag2 gene from Baylisascaris schroederi of giant panda and evaluation of the efficacy of a recombinant Bs-Ag2 antigen in mice.

The gene of Bs-Ag2 from Baylisascaris schroederi was amplified, cloned and sequenced. Sequence analysis indicated that the nucleotide sequences of the Bs-Ag2 from adult, L2 and L3 of B. schroederi were completely identical. A homology search performed by BLAST revealed that Bs-Ag2 shared the highest amino acid sequence identity with As16 protein from Ascaris suum (94%). The recombinant Bs-Ag2 proteins can be successfully expressed in Escherichia coli BL21 (DE3). The rBs-Ag2 was used to evaluate their ability to induce immune protective responses in BALB/c mice against L3-challenge infection in a mouse-B. schroederi model. There was a 63.66% reduction (P<0.001) of recovery of larvae compared with that in the control group. Specific anti-Bs-Ag2 antibodies from immune protected mice had significantly higher levels of immunoglobulin G (IgG) (P<0.0001). Our data supported the use of Bs-Ag2 as a potential candidate for vaccination against B. schroederi infection.