Strongyloides infection screening in transplant candidates: What is the best strategy?

BACKGROUND: The potential that Strongyloides stercoralis infection has to cause major morbidity and high mortality when the disseminated form occurs in transplant patients is of particular concern. METHODS: In this study, the objective was to observe S. stercoralis infection in patients who are candidates for transplantation by using parasitological, serological, and molecular techniques and to propose an algorithm for the detection of that infection in transplant candidates. RESULTS: By parasitological techniques, 10% of fecal samples were positive. Anti-Strongyloides antibodies immunoglobulin G were detected in 19.3% and 20.7% of patients by immunofluorescence assay and enzyme-linked immunosorbent assay, respectively. S. stercoralis DNA was observed in 17.3% of samples by conventional polymerase chain reaction and 32.7% of samples by quantitative polymerase chain reaction (qPCR). CONCLUSION: The set of results allows us to reinforce that a positive result by parasitological techniques and/or qPCR indicates that the specific treatment should be applied. However, the improvement of diagnostic techniques may suggest changes in the screening for strongyloidiasis in these patients.

Immune complexes as a tool for strongyloidiasis immunodiagnosis in kidney and liver transplant candidate.

Strongyloidiasis is a chronic and asymptomatic infection in immunocompetent patients. Immunocompromised patients, such as organ transplant candidates, can develop severe forms of this disease, and the best way to prevent progression to these forms is early diagnosis. Serological techniques using specific IgG and immune complexes (IC) detection can help in the diagnosis of these patients. This study aimed to detect specific anti-Strongyloides IC and IgG antibodies in kidney transplant (KT) and liver transplant (LT) candidates. A total of 100 blood samples was collected from transplant candidates (50 blood samples each from KT and LT candidates). Serum was obtained and analysed using enzyme-linked immunosorbent assay for IC and IgG detections. The IC levels showed frequencies of 18% and 2% in the KT and LT groups, respectively, whereas anti-Strongyloides IgG was detected in 34% and 12% of KT and LT candidates, respectively. The correlation between IC and IgG detection is poor in KT candidates, while in LT candidates, there is a significant positive correlation. The detection of IC can be an additional tool for the diagnosis of strongyloidiasis, especially when associated with the detection of specific IgG anti-Strongyloides antibodies.

Experimental toxocariasis in BALB/c mice: relationship between parasite inoculum and the IgG immune response.

BALB/c mice were inoculated with 5-500 Toxocara canis infective eggs, and bled at 15-120 days post infection (dpi) to evaluate the dynamics of IgG antibody response and larvae distribution. Positive results were observed in all occasions for every inoculum, and a direct proportional relationship between antibody detection and the parasitic load was observed. In samples collected at 60 dpi, detection of IgG was more intense, especially with the 50 and 500 egg doses; also, a correlation between antibody level and egg count was observed with these two inocula. At 120 dpi, a decrease in antibody titer was observed for all groups; and at the end of the experiment, larvae were recovered from carcass, liver and brain. In the liver, larvae were only found in mice inoculated with 500 T. canis eggs. In carcasses, these were recovered in all groups, and the group inoculated with 50 eggs showed the highest percentage of larvae in the brain.

Molecular diagnosis of strongyloidiasis in tropical areas: a comparison of conventional and real-time polymerase chain reaction with parasitological methods.

This study aimed to evaluate the use of conventional polymerase chain reaction (cPCR) and real-time quantitative PCR (qPCR) in the diagnosis of human strongyloidiasis from stool samples in tropical areas. Stool samples were collected from individuals and were determined to be positive for Strongyloides stercoralis (group I), negative for S. stercoralis (group II) and positive for other enteroparasite species (group III). DNA specific to S. stercoralis was found in 76.7% of group I samples by cPCR and in 90% of group I samples by qPCR. The results show that molecular methods can be used as alternative tools for detecting S. stercoralis in human stool samples in tropical areas.

Trichuris trichiura in a post-Colonial Brazilian mummy.

Trichuris trichiura is a soil-transmitted helminth which is prevalent in warm, moist, tropical and subtropical regions of the world with poor sanitation. Heavy whipworm can result either in Trichuris dysenteric syndrome - especially in children - or in a chronic colitis. In heavy infections, worms can spread proximally and may cause ileitis. Here we provide first microscopic evidence for a T. trichiura adult worm embedded in the rectum of a post-Colonial Brazilian adult mummy. During Colonial and post-Colonial times, many European chroniclers described a parasitic disease named Maculo whose symptomatology coincides with heavy helminthiasis. Based on our findings and on comparison of ancient textual evidence with modern description of heavy whipworm, we feel confident in considering that the two syndromes are expressions of the same pathological condition.

The hamster (Mesocricetus auratus) as an experimental model of toxocariasis: histopathological, immunohistochemical, and immunoelectron microscopic findings.

Toxocariasis is a globally distributed parasitic infection caused by the larval stage of Toxocara spp. The typical natural hosts of the parasite are dogs and cats, but humans can be infected by the larval stage of the parasite after ingesting embryonated eggs in soil or from contaminated hands or fomites. The migrating larvae are not adapted to complete their life cycle within accidental or paratenic hosts like humans and laboratory animals, respectively, but they are capable of invading viscera or other tissues where they may survive and induce disease. In order to characterize hamsters (Mesocricetus auratus) as a model for Toxocara canis infection, histopathological and immunohistochemistry procedures were used to detect pathological lesions and the distribution of toxocaral antigens in the liver, lungs, and kidneys of experimentally infected animals. We also attempted to characterize the immunological parameters of the inflammatory response and correlate them with the histopathological findings. In the kidney, a correlation between glomerular changes and antigen deposits was evaluated using immunoelectron microscopy. The hamster is an adequate model of experimental toxocariasis for short-term investigations and has a good immunological and pathological response to the infection. Lung and liver manifestations of toxocariasis in hamsters approximated those in humans and other experimental animal models. A mixed Th2 immunological response to T. canis infection was predominant. The hamster model displayed a progressive rise of anti-toxocaral antibodies with the formation of immune complexes. Circulating antigens, immunoglobulin, and complement deposits were detected in the kidney without the development of a definite immune complex nephropathy.

Detection of Schistosoma mansoni infection by TaqMan® Real-Time PCR in a hamster model.

An experimental study in hamsters was performed to evaluate the capability for detecting Schistosoma mansoni DNA in serum and fecal samples during the pre and post-egg-laying periods of infection using TaqMan® Real-Time PCR system (qPCR), was compared with the circumoval precipitin test (COPT) and the Kato-Katz technique, especially among individuals with low parasitic burden. Twenty-four hamsters were infected with cercariae. Three hamsters were sacrificed per week under anesthesia, from 7 days post infection (DPI) up to 56 DPI. A serum sample and a pool of feces were collected from each hamster. The presence of S. mansoni eggs in fecal samples was evaluated by Kato-Katz method and in the hamsters gutby histopathology. Detection of S. mansoni DNA was performed using qPCR and S. mansoni antibody using COPT. The first detection of eggs in feces by Kato-Katz method and S. mansoni DNA in feces by qPCR occurred 49 DPI. Nevertheless, S. mansoni DNA was detected in serum samples from 14 up to 56 DPI. COPT was positive at 35 DPI. The results not only confirm the reliability of S. mansoni DNA detection by qPCR, but also demonstrate that serum is a trustworthy source of DNA in the pre patent infection period.

Evaluation of aversive behavior in Rattus norvegicus experimentally infected by two distinct strains of Toxoplasma gondii (ME49 and VEG): study of epigenetic markers.

BACKGROUND: Behavioral changes in Rattus norvegicus infected with two strains of Toxoplasma gondii (ME49 and VEG) were investigated. METHODS: Rats were evaluated for motor activity and aversion or attraction to cat urine 60 days after infection. After euthanasia, arginine-vasopressin gene methylation in the central nervous system was evaluated. RESULTS: A significant difference was observed in the methylation of the arginine-vasopressin promoter gene between rats infected with the ME49 and VEG strains. CONCLUSIONS: Although differences were not observed in many parameters, significant differences were observed in the methylation of the arginine-vasopressin promoter gene in rats infected with the two studied strains.

Human Toxocariasis: 2010 to 2020 Contributions from Brazilian Researchers.

This is a review of the published contributions made by Brazilian researchers between 2010 and 2020 on the natural history of human toxocariasis and the effects of human toxocariasis on nonhuman paratenic hosts.

Epidemic of unilateral panuveitis in children from Brazilian Amazonia: clinical and etiological aspects in seven patients.

OBJECTIVES: To describe clinical presentation and results of diagnostic and therapeutic procedures in seven children from an epidemic of panuveitis in the Brazilian Amazonia, as well as environmental analysis and etiological aspects involved. METHODS: Patients underwent full pediatric and ophthalmic examinations, B-scan, ultrasound biomicroscopy, and serological tests. Ocular samples were thoroughly analyzed, including two enucleation specimens. Environmental investigation encompassed water, soil, and river fauna. RESULTS: All patients had bathed in the waters of a regional river, the Araguaia. Six of them presented with intermediate uveitis, with snowbanking. Five had cataract and four showed inferior endothelial opacity, with localized anterior synechiae. One showed total leukoma, with flat anterior chamber. Only two had active uveitis, one of them with anterior chamber nodule. Serology revealed high prevalence of anti-Toxocara canis immunoglobulin G (IgG) antibodies. In three cases, vitreous and lens samples disclosed spicules of freshwater sponges Drulia uruguayensis and D. ctenosclera, also detected in the waters of the river. CONCLUSION: Freshwater sponge spicules could be potential new etiological agents of ocular pathology, but further studies are needed, considering the heterogeneity of the ocular lesions and results of serological and environmental studies.

IgG reactivity with 40-35 kDa soluble and membrane antigen of Strongyloides venezuelensis in immunocompromised patients.

Immunocompromised patients constitute a risk group for the development of severe clinical forms of human strongyloidiasis. The diagnosis of this infection is primarily performed by parasitological techniques, but with low sensitivity. Serological techniques appear as an alternative, especially with heterologous antigens use. The aim of this study was to perform the Western blot technique by using S. venezuelensis infective third stage larva (iL3) soluble (TS) and membrane (TM) saline antigens to reveal immunoreactive bands in immunocompromised patients with strongyloidiasis. Serum samples from 117 parasitologically well-characterized patients were divided into four groups: S. stercoralis positive and immunocompetent (S + IC); S. stercoralis positive and immunocompromised (S + IP); negative and immunocompetent (S-IC); negative and immunocompromised (S-IP). A 40-35 kDa band was recognized by 100% of patients in the S + IC group in both antigenic fractions, and by 62.5% and 50% in the S + IP group using the TS and TM fractions, respectively. A 29 kDa band was recognized by 86.3% and 72.7% (for TS and TM, respectively) of patients in the S + IC group, and only by 12.5% of patients in the S + IP group on the TM antigen. Regardless of the patients' immunological condition, the 40-35 kDa band from S. venezuelensis was detected more frequently and can be used as an important marker to the immunodiagnosis of human strongyloidiasis.

Evaluation of aversive behavior in Rattus norvegicus experimentally infected by two distinct strains of Toxoplasma gondii (ME49 and VEG): study of epigenetic markers

ABSTRACT Background: Behavioral changes in Rattus norvegicus infected with two strains of Toxoplasma gondii (ME49 and VEG) were investigated. Methods: Rats were evaluated for motor activity and aversion or attraction to cat urine 60 days after infection. After euthanasia, arginine-vasopressin gene methylation in the central nervous system was evaluated. Results: A significant difference was observed in the methylation of the arginine-vasopressin promoter gene between rats infected with the ME49 and VEG strains. Conclusions: Although differences were not observed in many parameters, significant differences were observed in the methylation of the arginine-vasopressin promoter gene in rats infected with the two studied strains.

Frequency of soil contamination by Toxocara canis eggs in the south region of São Paulo municipality (SP, Brazil) in a 18 month period.

Soil contamination by embryonic eggs of Toxocara canis is the main source of human infection by this ascarid larvae resulting, sometimes, in the occurrence of visceral larva migrans syndrome. The objective of the present research is to determine the frequency of T. canis eggs in soil samples monthly collected in nine public places, located at the South Region of São Paulo municipality in a 18-month period, from February 2004 to July 2005. The soil samples collected were treated with a 30% antiformine solution and with a sodium dichromate solution (d = 1.40) and microscopic slides were prepared and examined under light microscopy for searching T. canis eggs. Two peaks of higher frequency had been found, one in February-May 2004 and the other in April-July 2005.

Frequency and risk factors for toxocariasis in children from a pediatric outpatient center in southeastern Brazil.

To estimate the frequency of anti-Toxocara sp. antibodies, and evaluate factors associated with this infection, sera from 242 male and female children, aged from one to fifteen years old, attended at the Hospital of the Federal University of Uberlândia, Minas Gerais State, Brazil, were analyzed by ELISA. Information on the patients was collected and registered using an investigative questionnaire, and details on possible clinical alterations were obtained from the medical charts of 187 patients. Of a total of 242 samples, 21 (8.7%) were positive for anti-Toxocara sp. antibodies. The presence of dogs and cats and the school variable (place of contact), appeared to be significantly associated (p<0.05) with a positive serology. Respiratory symptoms and eosinophil counts greater than 20% also showed a positive statistical correlation with a positive serology for Toxocara sp.. Factors such as sex and age, and symptoms like headache, stomach ache, convulsive crises and anemia were not associated with toxocariasis.

DIAGNOSIS OF Strongyloides stercoralis INFECTION IN IMMUNOCOMPROMISED PATIENTS BY SEROLOGICAL AND MOLECULAR METHODS.

Strongyloidiasis is a potentially serious infection in immunocompromised patients. Thus, the availability of sensitive and specific diagnostic methods is desirable, especially in the context of immunosuppressed patients in whom the diagnosis and treatment of strongyloidiasis is of utmost importance. In this study, serological and molecular tools were used to diagnose Strongyloides stercoralis infections in immunosuppressed patients. Serum and stool samples were obtained from 52 patients. Stool samples were first analyzed by Lutz, Rugai, and Agar plate culture methods, and then by a quantitative real time polymerase chain reaction (qPCR). Serum samples were evaluated by an enzyme-linked immunosorbent assay (ELISA) using a soluble (AS) or a membrane fractions antigen (AM) obtained from alkaline solutions of the filariform larvae of Strongyloides venezuelensis. Of the 52 immunosuppressed patients, three (5.8%) were positive for S. stercoralis by parasitological methods, compared to two patients (3.8%) and one patient (1.9%) who were detected by ELISA using the AS and the AM antigens, respectively. S. stercoralis DNA was amplified in seven (13.5%) stool samples by qPCR. These results suggest the utility of qPCR as an alternative diagnostic tool for the diagnosis of S. stercoralis infection in immunocompromised patients, considering the possible severity of this helminthiasis in this group of patients.

Infection by Cryptosporidium sp. in immunocompromised haematological patients.

The frequency of intestinal infection by Cryptosporidium sp. was determined in 60 patients, attended at the Haematological and Haemotherapeutical Service of "Santa Casa de Misericórdia" of São Paulo, suffering lymphoproliferative diseases (Group 1). As control group (Group 2) 59 persons without haematological diseases, but with the same life time and living at the same place of that of haematological patients, had been examined. The stool parasitological tests performed disclosed Cryptosporidium sp. oocysts in six (10%) individuals belonging to Group 1, whereas, in Group 2, nobody showed infection by this coccidian. Among the patients infected by Cryptosporidium sp. only one showed diarrhoeal faeces.

EFFICACY OF NITAZOXANIDE AGAINST Toxocara canis: LARVAL RECOVERY AND HUMORAL IMMUNE RESPONSE IN EXPERIMENTALLY INFECTED MICE.

The efficacy of nitazoxanide (NTZ) against toxocariasis was investigated in an experimental murine model and results were compared to those obtained using mebendazole. Sixty male BALB/c mice, aged six to eight weeks-old, were divided into groups of 10 each; fifty were orally infected with 300 larvaed eggs of T. canis and grouped as follows, G I: infected untreated mice; G II: infected mice treated with MBZ (15 mg/kg/day) 10 days postinfection (dpi); G III: infected mice treated with NTZ (20 mg/kg/day) 10 dpi; G IV: infected mice treated with MBZ 60 dpi; G V: infected mice treated with NTZ 60 dpi; GVI: control group comprising uninfected mice. Mice were bled via retro-orbital plexus on four occasions between 30 and 120 dpi. Sera were processed using the ELISA technique to detect IgG anti- Toxocara antibodies. At 120 dpi, mice were sacrificed for larval recovery in the CNS, liver, lungs, kidneys, eyes and carcass. Results showed similar levels of anti- Toxocara IgG antibodies among mice infected but not submitted to treatment and groups treated with MBZ or NTZ, 10 and 60 dpi. Larval recovery showed similar values in groups treated with NTZ and MBZ 10 dpi. MBZ showed better efficacy 60 dpi, with a 72.6% reduction in the parasite load compared with NTZ, which showed only 46.5% reduction. We conclude that administration of these anthelmintics did not modify the humoral response in experimental infection by T. canis. No parasitological cure was observed with either drug; however, a greater reduction in parasite load was achieved following treatment with MBZ.

Membrane fractions from Strongyloides venezuelensis in the immunodiagnosis of human strongyloidiasis.

Strongyloides venezuelensis is a parasitic nematode of rodents frequently used to obtain heterologous antigens for the immunological diagnosis of human strongyloidiasis. The aim of this study was to evaluate membrane fractions from S. venezuelensis for human strongyloidiasis immunodiagnosis. Soluble and membrane fractions were obtained in phosphate saline (SS and SM) and Tris-HCl (TS and TM) from filariform larvae of S. venezuelensis. Ninety-two serum samples (n = 92) were obtained from 20 strongyloidiasis patients (Group I), 32 from patients with other parasitic diseases (Group II), and 40 from healthy individuals (Group III), and were analyzed by enzyme-linked immunosorbent assay (ELISA). Soluble fractions (SS and TS) showed 90.0% sensitivity and 88.9% specificity, whereas the membrane fractions (SM and TM) showed 95.0% sensitivity and 94.4% specificity. The present results suggest the possible use of membrane fractions of S. venezuelensis as an alternative antigen for human strongyloidiasis immunodiagnosis.

IMMUNODIAGNOSIS OF HUMAN STRONGYLOIDIASIS: USE OF SIX DIFFERENT ANTIGENIC FRACTIONS FROM Strongyloides venezuelensis PARASITIC FEMALES.

The aim of this study was to evaluate six different antigenic fractions from Strongyloides venezuelensis parasitic females for the immunodiagnosis of human strongyloidiasis. Soluble and membrane fractions from S. venezuelensis parasitic females were prepared in phosphate-buffered saline (SSF and SMF, respectively), Tris-HCl (TSF and TMF, respectively), and an alkaline buffer (ASF and AMF, respectively). Serum samples obtained from patients with strongyloidiasis or, other parasitic diseases, and healthy individuals were analyzed by enzyme-linked immunosorbent assay (ELISA). Soluble fractions SSF, TSF, and ASF showed 85.0%, 75.0%, and 80.0% sensitivity and 93.1%, 93.1%, and 87.5% specificity, respectively. Membrane fractions SMF, TMF, and AMF showed 80.0%, 75.0%, and 85.0% sensitivity, and 95.8%, 90.3%, and 91.7% specificity, respectively. In conclusion, the present results suggest that the fractions obtained from parasitic females, especially the SSF and SMF, could be used as alternative antigen sources in the serodiagnosis of human strongyloidiasis.

Immunofluorescence assay for diagnosis of strongyloidiasis in immunocompromised patients.

BACKGROUND: Strongyloides stercoralis is a parasite that causes human strongyloidiasis. The disease ranges from asymptomatic to severe forms, which are often fatal in immunocompromised individuals. Laboratory diagnosis is challenging owing to limitations in the use of conventional parasitological techniques. The present study aimed to evaluate the indirect immunofluorescence assay (IFA) using infective larvae of S. venezuelensis as an antigen for the immunodiagnosis of human strongyloidiasis in immunocompromised patients. METHODS: Serum and stool samples from 200 immunocompromised patients (HIV-positive, HTLV-1-positive, and renal, liver, and/or bone marrow transplantation candidates) were used. Stool samples were examined using three parasitological methods: Lutz, Rugai, and culture agar plate. IFA was performed using sections of infective larvae of S. venezuelensis as antigens, and showed 95.4% sensitivity and 95.8% and specificity. RESULTS: Among the 200 patients, 17 (8.5%) were positive for S. stercoralis by at least one parasitological method, and 43 (21.5%) were positive by IFA. CONCLUSIONS: IFA can be used as a screening method for the detection of S. stercoralis in immunocompromised patients.