Parasitological and serological studies on Amoebiasis and other intestinal parasitic infections in Recife and its suburban area, northeast Brazil

Parasitological examinations were carried out during April to August, 1987, with 187 out-patients of the IMIP hospital, located in the center of Recife City, and 464 inhabitants of several villages around Cabo City, 50 Km southeast of Recife, Pernambuco, Brazil. Approximately 71% of the IMIP patients and 92% of the Cabo inhabitants were infected with at least one species of intestinal parasite. There was minimum difference in the prevalence rate of Trichuris trichiura between two areas, whereas the prevalence rates of Ascaris lumbricoides, hookworms, Strongyloides stercoralis, Schistosoma mansoni and Entamoeba histolytica were higher in the inhabitants of the Cabo City area. Only Giardia lamblia was more prevalent in the out-patients of IMIP hospital. Test tube cultivation revealed that the prevalence rate of Necator americanus in both areas was much higher than that of Ancylostoma duodenale , and also that the prevalence rate of S. stercoralis of the IMIP patients and Cabo inhabitants were 4.5% and 9.6%, respectively. Six hundred and fifteen sera were serologically examined for amoebiasis by the gel diffusion precipitation test (GDP) and enzyme linked immunosorbent assay (ELISA) using the antigen prepared from axenically cultured trophozoite of E. histolytica (strain HM-ITMSS). No positive reaction was observed in all of the sera as examined by GDP, while 32 out of 615 sera were positive on ELISA.

Parasitological and serological studies on amoebiasis and other intestinal parasitic infections in the rural sector around Recife, Northeast Brazil

Parasitological examinations were carried out during July to December, 1989, on 485 inhabitants of four villages in São Lourenço da Mata, 25 km northwest of Recife, Pernambuco, Brazil. Approximately 99.6% of the inhabitants were infected with at least one species of intestinal parasites. A high prevalence of Schistosoma mansoni (82.1%), hookworm (80.2%) Trichuris trichiura (69.9%), Ascaris lumbricoides (61.9%) and Entamoeba coli (36.7%) infections were demonstrated. Test tube cultivation revealed that the most common species of hookworm in this region was Necator americanus (88.4%), and also that the prevalence of Strongyloides stercoralis was 5.8%. Three hundred and thirty-four sera were serologically examined for amoebiasis by the gel diffusion precipitation test (GDP) and enzyme-linked immunosorbent assay (ELISA). No positive reaction was observed in all sera as examined by GDP, while 24 sera were positive by ELISA.

Detection of a Giardia lamblia coproantigen by using a commercially available immunoenzymatic assay, in Belo Horizonte, Brazil

It is known that fecal examination to detect Giardia lamblia cysts or trophozoites produces a high percentage of false-negative results. A commercially available immunoenzymatic assay (ProSpecT Giardia Microplate Assay, Alexon, Inc., BIOBRÁS) to detect G. lamblia specific coproantigen was evaluated for the first time in Brazil. A total of 90 specimens were tested. Each specimen was first tested as unpreserved stool, and then it was preserved in 10% Formalin to be tested 2 months later. The assay was able to identify all the 30 positive patients (sensitivity = 100.0%) by visual or spectrophotometric examination in the unpreserved specimens and was negative in 57 of the 60 patients without G. lamblia (specificity = 95.0%). The assay identified 27 of the 30 positive patients (sensitivity = 90.0%) and was negative in 59 of the 60 negatives (specificity = 98.3%) in the preserved stools according to both readings. A marked difference was observed in the optical densities in both groups, preserved and unpreserved stools, when the G. lamblia-positive specimens were compared to the negative or positive for other intestinal parasites than G. lamblia. The assay seems a good alternative for giardiasis diagnosis, especially when the fecal examination was repeatedly negative and the patient presents giardiasislike symptoms.

Diagnostic tests for amoebic liver abscess: comparison of enzyme - linked immunosorbent assay (Elisa) and counterimmunoelectrophoresis (CIE)

The liver abscess is the most frequent extraintestinal complication of intestinal amoebiasis: its diagnosis is suggested by the clinical picture but it must be confirmed by paraclinic tests. Themost stringent diagnosis requires identification of E. histolytica. But this is possible only in a few cases. Serological tests greatly improve the diagnosis of this severe complication of amoebiasis. We compared the Enzyme Linfed Immunosorbent Assay and the Counterimmunoeletrophoresis techniques. Both techniques were used to detect amoebic antibodies in 50 control patients, 30 patients with liver abscess and 30 patients with intestinal amoebiasis. All the sera from control patients gave negative results iin both techniques. When analysing the sera from patients with intestinal amoebiasis, 10% of them were positive by ELISA but non by CIE. The sera of patients with liver abscess, we found that 90% were positive by the ELISA method and 66.6% by the CIE technique. In patients with amoebic liver abscess, the results showed that the ELISA was more sensitive than the CIE, as it presented a higher sensitivity (100%) than that of the CIE technique (66%).

Detection of the acute phase of abdominal angiostrongyliasis with a parasite-specific IgG enzyme linked immunosorbent assay

Angiostrongylus costaricensis may cause intestinal lesions of varied severity when it accidentally infects man in Central and South America. First-stage larvae have never been detected in stools. Therefore, a parasite-specific IgG ELISA was evaluated for the determination of the acute phase of infection. The specificity and the sensitivity of the immunoassay was shown to be 76.2% and 91.1%, respectively. Eight serum samples taken from patients with histopathological diagnosis, at different time points (3 to 15 months) after surgical treatment, showed a sharp and early decline in antibody reactivity. The titration of anti-A. costaricensis antibodies has proved to be a useful method for the diagnosis of acute abdominal angiostrongyliasis.

Antimicrobial resistance of Enterococcus sp. isolated from the intestinal tract of patients from a university hospital in Brazil

This study reports the results about antimicrobial resistance of Enterococcus spp. isolated from intestinal tract of patients from a university hospital in Brazil. The identification of strains at species level was performed by conventional biochemical tests, API 20 Strep (bioMérieux), and polymerase chain reaction assay. The specie distribution was E. faecium (34%), followed by E. faecalis (33%), E. gallinarum (23.7%), E. casseliflavus (5.2%), E. avium (1%), and E. hirae (1%). Intrinsic resistance to vancomycin characterized by presence of vanC genes was found in E. gallinarum and E. casseliflavus. The high prevalence of VanC phenotype enterococci is very important because these species have been reported as causing a wide variety of infections. Vancomycin-resistant E. faecium or E. faecalis were not found and no one isolate of these species was a beta-lactamase producer. Thirteen clinical isolates of enterococci (13.4%) showed multiresistance patterns, which were defined by resistance to three classes of antibiotics plus resistance to at least one aminoglycoside (gentamicin and/or streptomycin). The resistance to several antimicrobials shown by enterococcal strains obtained in this study is of concern because of the decrease in the therapeutic options for treatment of infections caused by enterococci.

Detection of Entamoeba histolytica/Entamoeba dispar in stool specimens by using enzyme-linked immunosorbent assay

Entamoeba histolytica actually comprises two genetically distinct but morphologically indistinguishable species. E. histolytica can cause invasive intestinal and extra intestinal disease, while E. dispar cannot. Identification and differentiation of E. dispar and E. histolytica in stool sample by microscopy is imprecise. Several weeks of culture and isoenzyme analysis are required to differentiate E. histolytica from E. dispar. In this study, we have used an enzyme-linked immunosorbent assay (ELISA) for detection of E. histolytica/E.dispar and compared it with microscopy. Eighty-eight samples were evaluated, trichrome staining was positive in 20.4% (18) and ELISA was positive in 29.5% (26). Both tests were positive in 14 (15.9%) samples, 4 (4.5%) only with direct microscopy, and 12 (13.6%) only with ELISA. Both tests were negative in 58 (65.9%) samples. Microscopy has low sensitivity and high specificity, low negative predictive value and high positive predictive value in comparison with ELISA. E. histolytica/E. dispar antigen detection ELISA tests are inexpensive compared to the specific tests, yield objective results and do not require experienced microscopists and can therefore be recommended for screening of stools worldwide and the results can be taken for treatment that are fitting with its clinic.

Use of heterologous antigens for the immunodiagnosis of abdominal angiostrongyliasis by an enzyme-linked immunosorbent assay

Angiostrongylus costaricensis has a broad geographic distribution spanning from North to South America and the infections of vertebrates with this nematode can result in abdominal complications. Human infections are diagnosed by histological or serological methods because the isolation of larvae from feces is not feasible, as most parasites become trapped in intestinal tissues due to intense eosinophilic inflammation. Because A. costaricensis is difficult to maintain in the laboratory, an immunodiagnostic IgG enzyme-linked immunosorbent assay (ELISA) using antigens from the congeneric Angiostrongylus cantonensis species was evaluated against a panel of serum samples from patients who were histologically diagnosed with A. costaricensis infections. Sera from uninfected individuals and individuals infected with other parasites were used as controls. The sensitivity and specificity of the assay were estimated at 88.4% and 78.7%, respectively. Because the use of purified or cloned antigens has not been established as a reliable diagnostic tool, the use of heterologous antigens may provide a viable alternative for the development of an ELISA-based immunodetection system for the diagnosis of abdominal angiostrongyliasis.

An HPLC-UV method for the measurement of permeability of marker drugs in the Caco-2 cell assay

The Caco-2 cell line has been used as a model to predict the in vitro permeability of the human intestinal barrier. The predictive potential of the assay relies on an appropriate in-house validation of the method. The objective of the present study was to develop a single HPLC-UV method for the identification and quantitation of marker drugs and to determine the suitability of the Caco-2 cell permeability assay. A simple chromatographic method was developed for the simultaneous determination of both passively (propranolol, carbamazepine, acyclovir, and hydrochlorothiazide) and actively transported drugs (vinblastine and verapamil). Separation was achieved on a C18 column with step-gradient elution (acetonitrile and aqueous solution of ammonium acetate, pH 3.0) at a flow rate of 1.0 mL/min and UV detection at 275 nm during the total run time of 35 min. The method was validated and found to be specific, linear, precise, and accurate. This chromatographic system can be readily used on a routine basis and its utilization can be extended to other permeability models. The results obtained in the Caco-2 bi-directional transport experiments confirmed the validity of the assay, given that high and low permeability profiles were identified, and P-glycoprotein functionality was established.

Fluorescent quantitative PCR of Mycobacterium tuberculosis for differentiating intestinal tuberculosis from Crohn's disease

Intestinal tuberculosis (ITB) and Crohn's disease (CD) are granulomatous disorders with similar clinical manifestations and pathological features that are often difficult to differentiate. This study evaluated the value of fluorescent quantitative polymerase chain reaction (FQ-PCR) for Mycobacterium tuberculosis (MTB) in fecal samples and biopsy specimens to differentiate ITB from CD. From June 2010 to March 2013, 86 consecutive patients (38 females and 48 males, median age 31.3 years) with provisional diagnoses of ITB and CD were recruited for the study. The patients' clinical, endoscopic, and histological features were monitored until the final definite diagnoses were made. DNA was extracted from 250 mg fecal samples and biopsy tissues from each patient. The extracted DNA was amplified using FQ-PCR for the specific MTB sequence. A total of 29 ITB cases and 36 CD cases were included in the analysis. Perianal disease and longitudinal ulcers were significantly more common in the CD patients (P<0.05), whereas night sweats, ascites, and circumferential ulcers were significantly more common in the ITB patients (P<0.05). Fecal FQ-PCR for MTB was positive in 24 (82.8%) ITB patients and 3 (8.3%) CD patients. Tissue PCR was positive for MTB in 16 (55.2%) ITB patients and 2 (5.6%) CD patients. Compared with tissue FQ-PCR, fecal FQ-PCR was more sensitive (X2=5.16, P=0.02). We conclude that FQ-PCR for MTB on fecal and tissue samples is a valuable assay for differentiating ITB from CD, and fecal FQ-PCR has greater sensitivity for ITB than tissue FQ-PCR.

Imunofluorescência para neuropeptídeos na mucosa nasal humana: avaliação de técnica para peptídeo intestinal vasoativo (VIP)

Os neuropeptídeos são neurotransmissores relevantes na fisiologia nasal e o conhecimento crescente acerca de seu papel na fisiopatologia de doenças nasais abre novas perspectivas. A sua investigação na mucosa nasal humana baseia-se em grande parte em marcação imunológica, método complexo e sujeito a inúmeros fatores de erro. Com o intuito de viabilizar este tipo de pesquisa em nosso meio, um método de imunofluorescência para peptídeo intestinal vasoativo (VIP) na mucosa nasal humana é proposto e avaliado. FORMA DE ESTUDO: Coorte transversal. MATERIAL E MÉTODO: Oito pacientes submetidos a cirurgia funcional do nariz têm um fragmento de mucosa coletado da concha inferior. O tecido foi fixado em solução de Zamboni (paraformaldeído 4% tamponado e ácido pícrico 0,4%), congelado em nitrogênio líquido e armazenado. Cortes de 14 µm foram realizados e submetidos à reação de imunofluorescência para VIP (Península Laboratories). As imagens microscópicas foram documentadas em fotografia convencional. A especificidade, sensibilidade e reprodutibilidade de execução foram avaliadas. A reprodutibilidade de interpretação de resultados foi avaliada através da comparação de graus de marcação (0 a 4) atribuídos às fotos por seis observadores. RESULTADOS: Os resultados mostraram ser o método suficientemente específico, sensível, além de reprodutível em sua execução. A interpretação de resultados mostrou depender do perfeito esclarecimento do observador no julgamento das imagens de imunofluorescência, mas mostrou uniformidade. CONCLUSÃO: O método proposto foi considerado útil na pesquisa de neuropeptídeos na mucosa nasal humana.

Importância da reabsorção do ferro da hemorragia intestinal provocada pela ação dos vermes na progressão da anemia

Foram estudados 10 doentes, portadores de anemia ancilostomótica e de grande parasitose. Foram determinados alguns parâmetros hematológicos como dosagem de hemoglobina, contagem de hemácias, hematócrito, volume corpuscular médio, hemoglobina corpuscular média, ferro sérico e siderofilina. Foram estudados o volume da perda de sangue intestinal, o ferro perdido nesta hemorragia e eliminado nas fezes, e o ferro reaborvido da hemorragia intestinal. Conclui-se que a reabsorção do ferro da hemoglobina por dia no tubo digestivo é de suma importância no retardamento da instalação da anemia ferropriva, causada pela parasitose ancilostomótica.

The impact of improvement of water supply and sanitation facilities on diarrhea and intestinal parasites: a Brazilian experience with children in two low-income urban communities

During the second half of 1986 the impact of the improvement of water supply and excreta disposal facilities on diarrheal diseases and intestinal parasitosis was studied in 254 children up to six years of age from two favelas (shanty towns) of Belo Horizonte, Brazil. The estimated incidence of diarrhea was 6.2 episodes/child year and the estimated period prevalence reached 31.0 episode days/ child/ year. The point prevalence of parasitosis was 70.7% (Ascaris lumbricoides: 55.4%, Trichuris trichiura: 19.6%, Giardia lamblia: 17.9%). The estimated prevalence of diarrhea decreased with improvement of water supply and sanitation facilities to 45% and 44% respectively, but no statistically significant impact was observed in the case of parasitosis. School education and weaning practice were found to be other important determinants of diarrhea.

Campylobacter intestinal carriage among stray and pet dogs

The natural distribution of thermotolerant Campylobacter sp. in dogs (150 stray animals and 64 pets) was studied. Campylobacters were more frequently isolated (p<0.01) from stray dogs (51.3%) rather than from pet dogs (21.9%). All the biotypes described by Lior for C. jejuni and C. coli were found among stray animals, whereas only C. jejuni biotypes I and II and C. coli biotype II were found among pet dogs. The need for more studies related to the role of environmental sanitary conditions in the spreading of Campylobacter species is noted.

Human cryptosporidiosis: detection of specific antibodies in the serum by an indirect immunofluorescence

Cryptosporidium sp., a coccidian parasite usually found in the faeces of cattle, has been recently implicated as an agent of human intestinal disease, mainly in immunocompromised patients. In the study realized, by an indirect immunofluorescence technique, specific immunoglobulins (IgG and IgM) have been demonstrated in human serum against Cryptosporidium oocysts. Purified oocysts were used as antigens in the indirect immunofluorecence assay. After analyzing this test in sera from selected groups of patients, the frequency of both specific IgG and IgM of immunocompetent children who were excreting oocysts in their faeces was 62% and in children with negative excretion of oocysts was 20% and 40%, respectively. In adults infected with the human immunodeficiency virus (HIV) and who were excreting Cryptosporidium in their stools, the frequency was 57% for IgG but only 2% for IgM. Twenty three percent of immunocompromised adults with not determined excretion of oocysts in their stools had anti-Cryptosporidium IgG in their sera. Children infected with human immunodeficiency virus had no IgM and only 14% had IgG detectable in their sera. The indirect immunoflorescence assay, when used with other parasitological techniques appears to be useful for retrospective population studies and for diagnosis of acute infection. The humoral immune response of HIV positive patients to this protozoan agent needs clarification.