Sedimentation in parasitological coproscopy

A sedimentation technique is described, in which a fecal suspension is placed on top of an aqueous sucrose solution of specific gravity 1.015 g/cm³. Using 100 by 15 mm test tubes, duplicate gravity sedimentation experiments were made using homogenized fecal suspensions (single-columns) and fecal suspensions placed on top of clear columns (double-columns). Egg- and cyst-counts, and turbidity determinations were made in the sediments obtained after definite time intervals. Most Ascaris lumbricoides, Trichuris trichiura and Ancylostomidae eggs sedimented within 20 minutes in single - and between 30 and 60 minutes in double-columns. Giardia duodenalis cysts required longer periods to sediment in double - than in single-columns; after 180 minutes (the maximum period of observation), double-column sediments produced 60.0% of the counts of single-columns. Double-column sediments were consistently less turbid than single-column ones

Evaluation of the double diffusion, enzyme immunoassay and immunoblotting techniques, for the diagnosis of human hydatid disease in tropical areas

Hydatid disease in tropical areas poses a serious diagnostic problem due to the high frequence of cross-reactivity with other endemic helminthic infections. The enzyme-linked-immunosorbent assay (ELISA) and the double diffusion arc 5 showed respectively a sensitivity of 73% and 57% and a specificity of 84-95% and 100%. However, the specificity of ELISA was greatly increased by using ovine serum and phosphorylcholine in the diluent buffer. The hydatic antigen obtained from ovine cyst fluid showed three main protein bands of 64,58 and 30 KDa using SDS PAGE and immunoblotting. Sera from patients with onchocerciasis, cysticercosis, toxocariasis and Strongyloides infection cross-reacted with the 64 and 58 KDa bands by immunoblotting. However, none of the analyzed sera recognized the 30 KDa band, that seems to be specific in this assay. The immunoblotting showed a sensitivity of 80% and a specificity of 100% when used to recognize the 30 KDa band.

Specific label-free and real-time detection of oxidized low density lipoprotein (oxLDL) using an immunosensor with three monoclonal antibodies

Increased levels of plasma oxLDL, which is the oxidized fraction of Low Density Lipoprotein (LDL), are associated with atherosclerosis, an inflammatory disease, and the subsequent development of severe cardiovascular diseases that are today a major cause of death in modern countries. It is therefore important to find a reliable and fast assay to determine oxLDL in serum. A new immunosensor employing three monoclonal antibodies (mAbs) against oxLDL is proposed in this work as a quick and effective way to monitor oxLDL. The oxLDL was first employed to produce anti-oxLDL monoclonal antibodies by hybridoma cells that were previously obtained. The immunosensor was set-up by selfassembling cysteamine (Cyst) on a gold (Au) layer (4 mm diameter) of a disposable screen-printed electrode. Three mAbs were allowed to react with N-hydroxysuccinimide (NHS) and ethyl(dimethylaminopropyl)carbodiimide (EDAC), and subsequently incubated in the Au/Cys. Albumin from bovine serum (BSA) was immobilized further to ensure that other molecules apart from oxLDL could not bind to the electrode surface. All steps were followed by various characterization techniques such as electrochemical impedance spectroscopy (EIS) and square wave voltammetry (SWV). The analytical operation of the immunosensor was obtained by incubating the sensing layer of the device in oxLDL for 15 minutes, prior to EIS and SWV. This was done by using standard oxLDL solutions prepared in foetal calf serum, in order to simulate patient's plasma with circulating oxLDL. A sensitive response was observed from 0.5 to 18.0 mg mL 1 . The device was successfully applied to determine the oxLDL fraction in real serum, without prior dilution or necessary chemical treatment. The use of multiple monoclonal antibodies on a biosensing platform seemed to be a successful approach to produce a specific response towards a complex multi-analyte target, correlating well with the level of oxLDL within atherosclerosis disease, in a simple, fast and cheap way.

Comparison between immunomagnetic separation, coupled with immunofluorescence, and the techniques of Faust et al. and of Lutz for the diagnosis of Giardia lamblia cysts in human feces

In the present study, the performance of Immunomagnetic Separation technique, coupled with Immunofluorescence (IMS-IFA), was compared with the FAUST et al. and Lutz parasitological techniques for the detection of Giardia lamblia cysts in human feces. One hundred and twenty-seven samples were evaluated by the three techniques at the same time showing a rate of cyst detection of 27.5% by IMS-IFA and 15.7% by both Faust et al. and Lutz techniques. Data analysis showed a higher sensitivity of IMS-IFA for the detection of G. lamblia cysts in comparison with the techniques of FAUST et al. and Lutz. The use of this methodology as a routine procedure enables the processing of many samples simultaneously, in order to increase recovery rate of G. lamblia cysts and reduce the time of sample storage.

Dermoid Tumor of the Lateral Wall of the Cavernous Sinus

BACKGROUND: Congenital intracranial dermoid tumors are very rare. The location of these dermoid lesions in the cavernous sinus and the complexity of the operative procedure for these lesions have been noted by several authors. Dermoid tumors originating in the cavernous sinus are usually interdural, and thus blurred vision is an uncommon presentation. CASE DESCRIPTION: Herein we report the first incidental case of a cavernous sinus dermoid cyst in a 21-year-old woman. CONCLUSIONS: A literature review was done and the possible treatments and approaches for this lesion are discussed. We consider that surgical treatment is indicated in most incidental cavernous sinus dermoid lesions due to the possible symptoms related to compression or rupture leading to chemical meningitis.

Preliminary report of HIV and Toxoplasma gondii occurrence in pregnant women from Mozambique

Toxoplasmosis, a protozoan disease, causes severe disease in fetuses during pregnancy and deadly encephalitis in HIV patients. There are several studies on its seroprevalence around the world, but studies focusing on African countries are limited in number and mostly anecdotal. We studied two groups of samples from Mozambique by ELISA, using serum samples from 150 pregnant women and six Cerebrospinal fluid (CSF) samples from AIDS patients with encephalitis. HIV status was confirmed, and CD4 blood counts were obtained from HIV-positive pregnant women. IgG seroprevalence of the group as a whole was 18.7% (28/150), with a higher prevalence in HIV-positive individuals compared to those who were HIV-negative (31.3%, [18/58] vs. 10.9%, [10/92]) patients. These data may be biased due to cumulative effects of exposition affecting disease prevalence. If corrected, this data may indicate an interaction of HIV and T. gondii. Prevalence of both diseases increases with age, but this is more clearly seen for toxoplasmosis (p < 0.005) than HIV infection, possibly explained by higher transmission of HIV after childhood. In HIV patients suffering from encephalitis, CSF serology showed that 33% of specific IgG CSF had a high avidity, which was in accordance with the data from the group of pregnant women. Lower prevalence rates of both infections in older groups could be explained by more deaths in the infected groups, resulting in an artificially lower prevalence. Using CD4 counts as a marker of time of HIV infection, and correcting for age, patients with contact with T. gondii had fewer CD4 cells, suggesting prolonged HIV disease or other causes. Toxoplasma IgG prevalence is higher in HIV+ groups, which could be ascribed to HIV- and T. gondii-associated risk factors, such as exposure to higher and more diverse social contacts. The low incidence of Toxoplasma IgG in younger age groups shows that transmission could be related to better access to cyst-containing meat in adulthood, as environmental transmission due to oocysts is usually blamed for higher incidence in children. Taken together, these data support the urgent need of research in toxoplasmosis in Africa, especially in the presence of HIV epidemics.

Oral cavity eumycetoma

Mycetoma is a pathological process in which eumycotic (fungal) or actinomycotic causative agents from exogenous source produce grains. It is a localized chronic and deforming infectious disease of subcutaneous tissue, skin and bones. We report the first case of eumycetoma of the oral cavity in world literature. CASE REPORT: A 43-year-old male patient, complaining of swelling and fistula in the hard palate. On examination, swelling of the anterior and middle hard palate, with fistula draining a dark liquid was observed. The panoramic radiograph showed extensive radiolucent area involving the region of teeth 21-26 and the computerized tomography showed communication with the nasal cavity, suggesting the diagnosis of periapical cyst. Surgery was performed to remove the lesion. Histopathological examination revealed purulent material with characteristic grain. Gram staining for bacteria was negative and Grocott-Gomori staining for the detection of fungi was positive, concluding the diagnosis of eumycetoma. The patient was treated with ketoconazole for nine months, and was considered cured at the end of treatment. CONCLUSION: Histopathological examination, using histochemical staining, and direct microscopic grains examination can provide the distinction between eumycetoma and actinomycetoma accurately.

Caucasian Familial Moyamoya Syndrome With Rare Multisystemic Malformations

Moyamoya disease is an idiopathic progressive steno-occlusive disorder of the intracranial arteries located at the base of the brain. It is associated with the development of compensatory extensive network of fine collaterals. Moyamoya disease is considered syndromic when certain genetic or acquired disorders such as polycystic kidney disease, neurofibromatosis, or meningitis are also present. Although the genetic contribution in moyamoya is indisputable, its cause and pathogenesis remain under discussion. Herein, we report a rare occurrence of moyamoya syndrome in two European Caucasian siblings in association with unusual multisystemic malformations (polycystic kidney disease in one, and intestinal duplication cyst in the other). The karyotype was normal. No mutation in the RFN213 gene was found, and none of the HLA types linked to moyamoya disease or described in similar familial cases were identified. By describing these multisystemic associations, polycystic kidney disease for the second time, and intestinal malformation for the first time in the literature, our report expands the phenotypic variability of moyamoya syndrome. The coexistence of disparate malformations among close relatives suggests an underlying common genetic background predisposing to structural or physiological abnormalities in different tissues and organs.

HUMAN TOXOPLASMOSIS OUTBREAKS AND THE AGENT INFECTING FORM. FINDINGS FROM A SYSTEMATIC REVIEW

SUMMARY Toxoplasmosis, a worldwide highly prevalent zoonotic infection, is transmitted either by the oocysts, from water and soil, or the tissue cysts, in raw or undercooked infected meat, of Toxoplasma gondii. An ongoing debate is whether there are differences between the clinical and epidemiological characteristics of the outbreaks due to one or the other infective form of the agent. We performed a systematic review, recovering 437 reported outbreaks of which 38 were selected. They were complete reports containing ascribed Toxoplasma infecting form, and clinical and demographic data. There was no gender or age group selection in the outbreaks, which were described more often in the Americas. A large number of individuals were affected when oocysts, associated with soil and water contaminated with cat feces, were considered the transmission source. Onset of symptoms occurred early when the infection was ascribed to meat tissue cysts (11.4 ± 6.7 days) with sharpened temporal distribution of cases, while a broader and prolonged appearance of new cases was observed when oocysts in water were the source of the infection (20 ± 7 days, p < 0.001). Such information may be useful in the design and implementation of control strategies.

Toxoplasma-SPECIFIC IgG SUBCLASS ANTIBODY RESPONSE IN CEREBROSPINAL FLUID SAMPLES FROM PATIENTS WITH CEREBRAL TOXOPLASMOSIS

SUMMARY Cerebral toxoplasmosis can be highly debilitating and occasionally fatal in persons with immune system deficiencies. In this study, we evaluated the Toxoplasma gondii-specific IgG subclass antibody response in 19 cerebrospinal fluid (CSF) samples from patients with cerebral toxoplasmosis who had a positive IgG anti-T. gondii ELISA standardized with a cyst antigen preparation. There were no significant differences between the rates of positivity and the antibody concentrations (arithmetic means of the ELISA absorbances, MEA) for IgG1 and IgG2, but the rates of positivity and MEA values for these two IgG subclasses were significantly higher than those for IgG3 and IgG4. The marked IgG2 response in CSF from patients with cerebral toxoplasmosis merits further investigation.

Cutaneous Leiomyomatosis in a Mother and Daughter

A 34-year-old woman with no known medical history was evaluated for multiple painful brown nodules and papules on the anterior aspect of the trunk. She mentioned a history of similar cutaneous findings on her mother. Biopsies of three lesions revealed piloleiomyomata. Renal and adrenal ultrasound revealed an isolated simple cortical cyst, and pelvic and endovaginal ultrasound revealed two uterine myomata. The clinical diagnosis of hereditary leiomyomatosis and renal cell cancer was corroborated by the identification of a heterozygous variant on exon 5 of the fumarate hydratase gene (c.578C>T p.T193I). Identification of the tumor piloleiomyoma should alert the dermatologist to this rare genodermatosis, which is associated with an increased risk of renal cell tumors, demanding multidisciplinary follow-up, and personal and family counseling.

Tyrosinase-based phenol remediation

Dissertation for the obtention of the Master Degree in Biotechnology

Parasitological stool sample exam by spontaneous sedimentation method using conical tubes: effectiveness, practice, and biosafety

INTRODUCTION: Spontaneous sedimentation is an important procedure for stool examination. A modification of this technique using conical tubes was performed and evaluated. METHODS: Fifty fecal samples were processed in sedimentation glass and in polypropylene conical tubes. Another 50 samples were used for quantitative evaluation of protozoan cysts. RESULTS: Although no significant differences occurred in the frequency of protozoa and helminths detected, significant differences in protozoan cyst counts did occur. CONCLUSIONS: The use of tube predicts a shorter path in the sedimentation of the sample, increases concentration of parasites for microscopy analysis, minimizes the risks of contamination, reduces the odor, and optimizes the workspace.

Analysis of the chemical components of hydatid fluid from Echinococcus granulosus

Introduction The aim of this study was to explore the environment of Echinococcus granulosus (E. granulosus) protoscolices and their relationship with their host. Methods Proteins from the hydatid-cyst fluid (HCF) from E. granulosus were identified by proteomics. An inductively coupled plasma atomic emission spectrometer (ICP-AES) was used to determine the elements, an automatic biochemical analyzer was used to detect the types and levels of biochemical indices, and an automatic amino acid analyzer was used to detect the types and levels of amino acids in the E. granulosus HCF. Results I) Approximately 30 protein spots and 21 peptide mass fingerprints (PMF) were acquired in the two-dimensional gel electrophoresis (2-DE) pattern of hydatid fluid; II) We detected 10 chemical elements in the cyst fluid, including sodium, potassium, calcium, magnesium, copper, and zinc; III) We measured 19 biochemical metabolites in the cyst fluid, and the amount of most of these metabolites was lower than that in normal human serum; IV) We detected 17 free amino acids and measured some of these, including alanine, glycine, and valine. Conclusions We identified and measured many chemical components of the cyst fluid, providing a theoretical basis for developing new drugs to prevent and treat hydatid disease by inhibiting or blocking nutrition, metabolism, and other functions of the pathogen.

Clinical and serological outcomes with different surgical approaches for human hepatic hydatidosis

ABSTRACTINTRODUCTION:Hydatidosis is the result of infection with the larval stages of some species of the genus Echinococcus. Treatment approaches for hydatid cysts include the use of albendazole, surgery, and/or medico-surgical procedures. The choice of the therapeutic surgical approach depends on the cyst number and localization, surgeon expertise, and presence of complications. The present study aimed to compare the outcomes of the following therapeutic approaches for the treatment of hepatic hydatid cysts: pericystectomy; the puncture, aspiration, injection, and reaspiration (PAIR) technique; and the PAIR technique followed by deroofing, evacuation of cysts, and omentoplasty.METHODS:The 54 patients were divided into 3 groups: Group I (14 patients) who underwent pericystectomy, Group II (23 patients) who underwent the PAIR technique, and Group III (17 patients) who underwent the PAIR technique followed by deroofing and omentoplasty. The diagnosis of hydatid cysts was based on serological testing using enzyme-linked immunosorbent assay, abdominal ultrasound, and parasitological examination of the cyst contents. Morbidity, mortality, length of hospital stay, recurrence, and postoperative complications were evaluated.RESULTS:Postoperative bleeding, infection, and recurrence were reported in Groups I and II; Group III did not experience postoperative infection and had shorter hospital stays. Recurrence and postoperative complications did not occur in Group III.CONCLUSIONS:The partial surgical procedure with deroofing, evacuation of the cysts, and omentoplasty, as performed in the present study, is recommended as a safe and effective method for elimination of the entire parasite with minimal possibility for intra-peritoneal spillage.