Detection of Anti-Strongyloides Antibodies in the Serum of Liver Transplant Recipients: Need of Screening for This Neglected Helminthiasis.

Immunosuppressed patients, particularly transplant recipients, can develop severe strongyloidiasis. This study aimed to detect anti-Strongyloides IgG antibodies in a panel of sera from liver transplant patients. Two techniques were used: ELISA as the initial screening test and Western blotting as a confirmatory test. ELISA reactivity of 10.9% (32/294) was observed. The 40-30 kDa fraction was recognised in 93.7% (30/32) of the patients, resulting in a positivity rate of 10.2%. These data highlight the importance of serological screening for Strongyloides stercoralis infection in liver transplant recipients.

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.

Proteomic analysis of the excretory-secretory products from Strongyloides venezuelensis infective larvae: new insights for the immunodiagnosis of human strongyloidiasis.

Serodiagnosis of human strongyloidiasis is a practical alternative to parasitological methods due to its high sensitivity. However, cross-reactivity with other helminth infections limits its utility, and this problem is due to the use of homologous or heterologous somatic extracts of the parasite as an antigen source. Excretory-secretory (E/S) products from Strongyloides infective larvae can be used to improve the serodiagnosis. The combined use of western blot and proteomics became an interesting strategy to identify immunological markers for the serodiagnosis of strongyloidiasis. The present study describes the proteomic analysis of the antigenic components from E/S products of S. venezuelensis infective larvae that were recognized by IgG antibodies from patients with strongyloidiasis. Our results showed that IgG antibodies from patients with strongyloidiasis recognized between 15 and 16 antigenic bands in the E/S products from S. venezuelensis that were incubated in PBS or in RPMI culture medium, respectively. Overall, antigenic bands of low and high molecular weight were more specific than those of intermediate molecular weight, which were cross-reactive. A 36-kDa antigenic band was 93% sensitive and 100% specific (a probably arginine kinase of 37 kDa), while other antigenic bands were highly sensitive but low specific. Proteomic analysis revealed differences between the protein profiles from E/S-RPMI and E/S-PBS since only one-third of all proteins identified were common in both types of E/S products. Bioinformatic analysis showed that more than 50% of the proteins from E/S products are secreted within extracellular vesicles and only a small percentage of them are actually released by the classical secretory pathway. Several components from the E/S products were identified as plasminogen-binding proteins, probably used as an immune evasion mechanism. The data provided here provide valuable information to increase understanding of E/S products from S. venezuelensis infective larvae. This may help us to find new targets for the immunodiagnosis of human strongyloidiasis.

A case report of COVID-19 in a patient with non-Hodgkin's lymphoma.

BACKGROUND: The current literature is scarce as to the outcomes of COVID-19 infection in non-Hodgkin's lymphoma patients and whether immunosuppressive or chemotherapeutic agents can cause worsening of the patients' condition during COVID-19 infection. CASE PRESENTATION: Our case is a 59-year-old gentleman who presented to the Emergency Department of the Cancer Institute of Hospital das Clínicas da Universidade de São Paulo, São Paulo, Brazil on 10th May 2020 with a worsening dyspnea and chest pain which had started 3 days prior to presentation to the Emergency Department. He had a past history of non-Hodgkin's lymphoma for which he was receiving chemotherapy. Subsequent PCR testing demonstrated that our patient was SARS-CoV-2 positive. CONCLUSION: In this report, we show a patient with non-Hodgkin lymphoma in the middle of chemotherapy, presented a mild clinical course of COVID-19 infection.

Diagnostic accuracy of somatic and excretory-secretory antigens from Strongyloides venezuelensis infective larvae for the immunodiagnosis of human strongyloidiasis.

To evaluate the diagnostic accuracy of three types of antigenic preparations from Strongyloides venezuelensis infective larvae for detection of serum IgG anti-Strongyloides antibodies by enzyme-linked immunosorbent assay (ELISA). Soluble somatic fractions (SSF) and membrane somatic fractions (MSF) and excretory−secretory (E/S) products from S. venezuelensis infective larvae were evaluated against 71 sera from individuals with strongyloidiasis, 105 sera from healthy individuals, and 84 sera from individuals with other helminth infections. Using an ELISA cut-off for 100% sensitivity, E/S products were 97.88% specific followed by MSF (93.12%) and then by SSF (85.2%). The occurrence of cross-reactivity with other helminths was 4.76% (4/84) with E/S products, 8.33% (7/84) with MSF, and 17.86% (15/84) with SSF. For a cut-off for 100% specificity, E/S products showed a sensitivity of 88.73% whereas MSF and SSF showed sensitivities of 59.15% and 53.52%, respectively. In conclusion, E/S products were the best antigenic option for the serodiagnosis of human strongyloidiasis.

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.

Evaluation of real-time PCR assay to detect Schistosoma mansoni infections in a low endemic setting.

BACKGROUND: Schistosomiasis constitutes a major public health problem, and 200 million people are estimated to be infected with schistosomiasis worldwide. In Brazil, schistosomiasis has been reported in 19 states, showing areas of high and medium endemicity and a wide range of areas of low endemicity (ALE). Barra Mansa in Rio de Janeiro state has an estimated prevalence of 1%. ALE represent a new challenge for the helminth control because about 75% of infected individuals are asymptomatic and infections occur with a low parasite load (<100 eggs per gram of feces), causing a decrease in sensitivity of stool parasitological techniques, which are a reference for the laboratory diagnosis of this helminth. The objective of this study was to evaluate the performance of a TaqMan quantitative polymerase chain reaction (qPCR) technique in serum and feces DNA samples using the techniques of Kato-Katz (KK), Hoffman, Pons and Janer (HH) as references, during an epidemiological survey using fecal samples and sera from randomized residents from an ALE. METHODS: A cross-sectional study conducted from April to December 2011 using a probabilistic sampling that collected 572 fecal and serum samples. The laboratory diagnostic techniques used were: KK, HH and qPCR (feces and serum). RESULTS: We obtained the following results using the different diagnostic techniques: KK and HH, 0.9% (n =5); qPCR-feces, 9.6% (n =55); and qPCR-serum, 1.4% (n =8). The qPCR-feces presented the highest positivity, whereas the techniques of HH and KK were the least sensitive to detect infections (0.8%). Compared to HH and KK, qPCR-feces showed a statistically significant difference in positivity (p <0.05), although with poor agreement. CONCLUSION: The positivity rate presented by the qPCR approach was far higher than that obtained by parasitological techniques. The lack of adequate surveillance in ALE of schistosomiasis indicates a high possibility of these areas being actually of medium and high endemicity. This study presents a control perspective, pointing to the possibility of using combined laboratory tools in the diagnosis of schistosomiasis in ALE.

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.

A silent threat behind bars: Serosurvey and associated risk factors for strongyloidiasis in female inmates and correctional officers.

Strongyloidiasis has been a neglected parasitic infection caused by Strongyloides genus parasites. Despite assessment of S. stercoralis exposure in different vulnerable populations, seroprevalence in inmates worldwide remains to be fully established. Due to poor sanitation and lack of personal hygienic practices, incarcerated individuals have been considered prone to spread infectious illnesses. Accordingly, the present study has assessed exposure and associated risk factors for strongyloidiasis in women inmates and correctional officers at the Women's State Penitentiary of Parana, part of the third largest incarceration complex in Brazil at the time. Blood samplings were performed in 2020 and 2021from a total of 503 women inmates and 92 correctional officers. Participants voluntarily responded to an epidemiological questionnaire to assess associated risk factors to strongyloidiasis. Serological analysis was performed by ELISA for anti-S. stercoralis IgG detection. Statistical analysis was performed using R software, adopting a 5% level of significance. The data were submitted to univariate analysis by chi-square or Fisher´s Exact test for assessing the association among seropositivity and the variables. The variables with p-value < 0.2 in the univariate analysis were considered fit to be included in the logistic regression. In overall, 356/503 (70.8%; 95% CI: 66.7-74.6) inmates were seropositive for anti-S. stercoralis antibodies, with no statistically associated risk factor to seropositivity. A total of 57/92 (62.0%; 95% CI: 51.8-71.2) correctional officers were seropositive, and logistic regression revealed that individuals older than 50 years were more likely seropositive. In conclusion, the high endemicity observed herein has indicated a history of previous exposure to S. stercoralis and warned for a systematic strongyloidiasis screening for inmates, to prevent long term morbidity and disseminated infection during incarceration.

Evaluation of larval surface antigens from infective larvae of Strongyloides venezuelensis for the serodiagnosis of human strongyloidiasis.

Serodiagnosis of strongyloidiasis is usually performed by ELISA for the detection of IgG antibodies due to its high sensitivity and practicality, but its main limitation is a constant source of S. stercoralis antigens. The use of S. venezuelensis as a heterologous source of antigens has facilitated several published studies on the serodiagnosis and epidemiology of human strongyloidiasis. The main objective of this study was to evaluate the diagnostic accuracy of surface cuticle antigens of infective larvae of S. venezuelensis extracted with CTAB detergent (L3-CTAB) in comparison with soluble somatic extracts (L3-SSE) using a panel of sera from immunocompetent and immunocompromised individuals, at three different cut-offs. ROC curve analysis showed that L3-CTAB had an AUC of 0.9926. At the first cut-off value (OD 450 nm = 0.214), sensitivity and specificity were 100% and 90.11%, respectively, with a diagnostic accuracy of 0.93. At a second cut-off value (OD 450 nm = 0.286), sensitivity and specificity were 70% and 100%, respectively, with a diagnostic accuracy of 0.91. However, at an alternative third cut-off value (OD 450 nm = 0.589), sensitivity and specificity were 95% and 97.8%, respectively, with a diagnostic accuracy of 0.97. Using L3-CTAB as an antigenic source, the seropositivity rate in immunocompromised patients was 28.13% (9/32) whereas a seropositivity rate of 34.38% (11/32) was found when L3-SSE was used in ELISA. Therefore, the L3-CTAB is simple and practical to obtain and was found to be highly sensitive and specific.

Seroprevalence and associated risk factors of strongyloidiasis in indigenous communities and healthcare professionals from Brazil.

Strongyloides stercoralis, a pathogenic roundworm, is considered endemic in several tropical and subtropical areas worldwide. Indigenous populations have the highest soil-transmitted helminthiases-related mortality rates, but the prevalence and risk factors associated with S. stercoralis in Brazilian indigenous populations have not been established. Thus, the present study aimed to assess the seroprevalence and associated risk factors for S. stercoralis in indigenous communities and the healthcare professionals serving them in Brazil. Indigenous populations living in nine communities and healthcare professionals were tested for anti- S. stercoralis antibodies by ELISA. A questionnaire was used to assess socio-epidemiological information. Associated risk factors for seropositivity were tested by chi-square or Fisher's exact tests, using univariate analyses and multivariate logistic regression. Overall, 174/463 (37.6%; CI 95%: 33.3-42.1) indigenous persons and 77/147 (52.4%; 95% CI: 44.3-60.3) healthcare professionals were seropositive for anti- S. stercoralis antibodies. Seropositivity among the two groups was statistically significant (p = 0.0016; OR = 0.547; 95% CI: 0.376-0.796) and revealed that healthcare professionals were 1.83 times more likely to be seropositive. The multivariate analysis showed that being male or being adult were also risk factors, while having a septic tank as a sanitary facility represented a protective factor for S. stercoralis exposure in indigenous persons. None of the variables evaluated were associated with S. stercoralis exposure in the professional group. The study herein has reported a high seroprevalence to Strongyloides stercoralis in indigenous communities of Brazil and healthcare professionals, warning for potential public health concerns of strongyloidiasis in such populations.

Toxocara canis 30-35 kDa excretory-secretory antigen is an important marker in mice challenged by inocula containing different parasite load levels.

The Western-blotting technique was applied to identify antigenic fractions of excretory-secretory Toxocara canis antigen recognized by IgG antibodies throughout an experimental infection in mice challenged by different inocula. Mice were inoculated with 5, 50 and 500 embryonated eggs and serum samples were collected 15, 30, 60, 90 and 120 days post-infection. Serum samples were analyzed using an excretory-secretory Toxocara antigen. Antibodies recognized antigenic fractions from 30 to 90 kDa. The protein fraction of 30-35 kDa was the most frequently recognized regardless of the size of inoculum and the stage of infection represented by the different collection times, but the antigenic recognition was more evident in groups infected with 50 and 500 eggs. This study presents an antigenic panel of the excretory-secretory antigen of T. canis and suggests that the 30-35 kDa antigenic fraction is a promising marker of the infection and should be further explored in future studies on experimental toxocariasis.

Importance of detection of Strongyloides stercoralis DNA in fecal samples from patients with type 2 diabetes mellitus.

OBJECTIVE: The association between diabetes and Strongyloides infection remains controversial. This study aimed to detect Strongyloides stercoralis DNA in the feces of patients with Diabetes Mellitus type 2 (DM2). METHODS: Fecal samples were analyzed via the Lutz, Rugai, and agar plate culture methods. PCR amplification was performed using two targets (PCR-genus and PCR-species) located on the S. stercoralis 18S ribosomal. RESULTS: The positivity for S. stercoralis using parasitological methods was 1.1%. PCR-genus (14.13%) demonstrated a higher positivity than PCR-species (9.78%). CONCLUSION: The results confirm the greater positivity of the molecular diagnosis in relation to parasitological methods, reinforcing its use as an additional tool for the diagnosis of S. stercoralis infection in patients with DM2 living in endemic areas for this helminthiasis.

Management and treatment of decompensated hepatic fibrosis and severe refractory Schistosoma mansoni ascites with transjugular intrahepatic portosystemic shunt.

This study aimed to report the first case of a patient with hepatosplenic schistosomiasis mansoni, refractory ascites and portal vein thrombosis treated with a transjugular intrahepatic portosystemic shunt (TIPS), at the Instituto de Radiologia, Hospital das Clinicas, Faculdade de Medicina, Universidade de Sao Paulo, Brazil. After the procedure, the patient recovered favorably and progressed with portal pressure reduction and no deterioration of the liver function. Endovascular shunt modification is a conservative medical approach that often helps in reducing symptoms significantly, making it a less invasive and a safer alternative to liver transplantation for the treatment of schistosomiasis with portal hypertension.

Current status of research regarding Blastocystis sp., an enigmatic protist, in Brazil.

The present study aimed to evaluate the occurrence of Blastocystis sp. in Brazilian studies over a period of years (2000-2020), as well as point out relevant aspects of this enigmatic organism. We performed a literature search using six sources of international databases. The data were divided into diagnostic by parasitological and molecular techniques, and relevant aspects. After applying the inclusion and exclusion criteria, 52 studies were included in the final analysis. The occurrence of Blastocystis sp. in Brazil ranged from 0.5% to 86.6%, as determined using parasitological techniques. The highest occurrence was in the North (27.3%) and the lowest, in the Midwest region (13.4%). In Brazil, most studies have employed molecular techniques and are concentrated in the Southeast region. The Blastocystis sp. subtype ST3 had the highest average positivity, followed by ST1 and ST2. These findings represent a panorama that reflects the reality of Brazil; thus, we believe that the effectiveness of parasitological diagnosis should be considered with regard to making an appropriate choice of technique for detecting Blastocystis sp. Additionally, we emphasize the importance of further studies in the context of molecular epidemiology with regard to this genus. Blastocystis sp. is not well understood yet, and very little information regarding this genus is available; hence, further research regarding this genus is urgently needed.

Diagnosis of human strongyloidiasis: Application in clinical practice.

This review considers the advantages and disadvantages of parasitological techniques, methods of detecting antibodies and antigens, as well as molecular biology techniques in the diagnosis of human strongyloidiasis. In addition, it elucidates the potential of different techniques for rapid and effective detection of clinical cases, thus enabling early treatment and preventing fatal consequences of this helminthiasis.

Blastocystis subtypes in patients with diabetes mellitus from the Midwest region of Brazil.

Blastocystis sp. is an enteric protist commonly found in human fecal samples. In Brazil, few studies have been developed, but none of them has explored the presence of Blastocystis in patients with diabetes mellitus. We evaluated the occurrence and molecular identification of Blastocystis sp. among patients with diabetes mellitus in the Midwest region, Goias State, Brazil. Genomic DNA was obtained from 175 fecal samples (99 from the diabetic group and 76 from the control group). PCR was performed using pan-Blastocystis primers from the SSU-rDNA gene. Microscopic examination revealed positivity of 12.1% and 7.9% for Blastocystis in diabetics and in controls, respectively. Amplification of Blastocystis DNA was observed in 34.4% (34 of 99) and 30.3% (23 of 76) from the diabetic and control groups, respectively. Phylogenetic analyses and BLAST searches revealed six subtypes among Blastocystis isolates in the diabetic group, represented by ST1 (38.2%), ST2 (11.8%), ST3 (35.3%), ST6 (2.9%), ST7 (2.9%) and ST8 (8.8%). In the control group, ST1 (21.8%), ST2 (21.8%), ST3 (43.5%), ST6 (4.4%) and ST8 (8.7%) were identified. This study is the first report regarding the occurrence and subtypes distribution of Blastocystis in patients with diabetes mellitus in Brazil. The results reinforce the potential risk of Blastocystis infection in patients with diabetes, in addition, it contributes to the understanding of the genetic diversity of this enigmatic organism.

Culture isolation and molecular identification of Blastocystis sp. in Brazilian human isolates: preliminary results.

Blastocystis sp. is a protist commonly found in stool samples of humans and animals. Biological and genetic factors of this organism remain controversial. The present study aimed to develop and implement the Blastocystis in vitro culture of Brazilian human isolates for routine use. The fecal isolates (n = 20) were maintained in our laboratory by several passages in Pavlova's medium. Cultures were monitored every 72 h by light microscopy. Genomic DNA was extracted to identify the subtypes (STs). In most isolates, the vacuolar form was prevalent. The amoeboid, granular and cystic forms were observed during in vitro cultivation. STs 1, 2, 3, 4 and 7 were identified. Our preliminary results show the generation time and forms present in the in vitro culture of Blastocystis subtypes isolated from Brazilian human isolates. Therefore, we emphasize the use of in vitro culture as a tool in future studies for the better understanding of the biological aspects of Blastocystis sp.

Molecular detection of prepatent Schistosoma mansoni infection in Biomphalaria glabrata snail vectors.

Approximately 240 million people worldwide are infected by Schistosoma. In Brazil, one of the main intermediate hosts of this parasite is Biomphalaria glabrata snails. The early detection of larval stages in intermediate hosts is an important challenge to public health, but it also represents an opportunity as a new alternative to indicate earlier natural infections before cercariae differentiation and emergence. In this context, we demonstrated that PCR amplification of a 28S gene fragment from the parasite does demonstrate S. mansoni infection in snails 14 days post infection. This conventional polymerase chain reaction amplified clear bands and was able to detect parasitic infection in the intermediate host B. glabrata under experimental conditions. However, we reinforce that this approach requires deeper investigations and further comparisons to confirm its specificity and sensitivity in earlier time points after miracidia infection. This approach has relevant potential as an effective molecular-based strategy for the monitoring of schistosomiasis transmission.