Sex Determination and Female Reproductive Development in the Genus Schistosoma: A Review

Parasites of the genus Schistosoma were among the first metazoans to develop separate sexes, which is chromosomally determined in the fertilized egg. Despite the occurrence of specific sex chromosomes, the females of most Schistosomatidae species do not complete their somatic development and reach no sexual maturity without the presence of males. Indeed, the most controversial and at the same time most fascinating aspect about the sexual development of Schistosoma females lies on discover the nature of the stimulus produced by males that triggers and controls this process. Although the nature of the stimulus (physical or chemical) is a source of controversy, there is agreement that mating is a necessary requirement for maturation to occur and for migration of the female to a definitive final site of residence in the vascular system of the vertebrate host. It has also been proposed that the stimulus is not species-specific and, in some cases, not even genus-specific. Despite a vast literature on the subject, the process or processes underlying the meeting of males and females in the circulatory system have not been determined and as yet no consensus exists about the nature of the stimulus that triggers and maintains female development. In the studies about their role, Schistosoma males have been considered, at times pejoratively, the brother, the muscles or even the liver of females. Indeed, it still remains to be determined whether the stimulus responsible for female maturation involves the transfer of hormones, nutrients, neuromediators, mere tactile stimulation or a combination of chemotactic and thigmotactic factors

ANTI M. leprae IgM ANTIBODY DETERMINATION BY ULTRAMICROIMMUNOENZYMATIC (UMELISA HANSEN) FOR THE DIAGNOSIS AND MONITORING LEPROSY

The relationship between the IgM antibody response, antigenic load as well as the clinical improvement after chemotherapy was studied in order to obtain useful data for the early diagnosis and monitoring leprosy. A level of 82% (94/115) agreement was obtained between IgM UMELISA HANSEN and slitskin smear examination. Discrepant results were observed in 16 patients who showed positive IgM response despite negative by the skin smear examination. In these patients, the IgM response was seen to be associated to the early signal for bacilli recurrence in the skin. In one of these patients the presence of bacilli was demonstrated in the skin, two months after IgM antibodies being detected by UMELISA HANSEN. Also in one of the treated patients positive by both diagnostic techniques, a remarkable decrease in the IgM antibody levels was seen, correlating with a significant clinical improvement. Moreover it was found a direct relationship between the IgM antibody response and bacterial antigenic load, regardless the time elapsed in the disease's evolution.

Serological diagnosis of toxoplasmosis: usefulness of IgA detection and IgG avidity determination in a patient with a persistent IgM antibody response to Toxoplasma gondii

We report the detection of specific IgA antibodies and the determination of IgG avidity in sequential serum samples from a patient exhibiting significant levels of Toxoplasma-specific IgM antibodies for seven years after the onset of the clinical symptoms of toxoplasmosis. IgM antibodies were detected by an indirect immunofluorescence test and by three commercial enzyme-linked immunosorbent assays (ELISA). Anti-T. gondii IgA was quantified by the a-capture ELISA technique using a commercial kit. As defined by the manufacturer of the IgA ELISA test used, most patients with acute toxoplasmosis have antibody levels > 40 arbitrary units per ml (AU/mL). At this cut-off level, the patient still had a positive ELISA result (45 AU/mL) in a serum sample taken one year after the beginning of clinical manifestations. The IgG avidity-ELISA test was performed with the Falcon assay screening test (F.A.S.T.®) - ELISA system. Avidity indices compatible with a recent Toxoplasma infection were found only in serum samples taken during the first 5 months after the onset of the clinical symptoms of toxoplasmosis. These results show that the interpretation of positive IgM results as indicative of recently acquired toxoplasmosis requires additional laboratory confirmation either by other tests or by the demonstration of a significant rise in the antibody titers in sequential serum samples.

Anti-phenolic glycolipid-I (PGL-I) determination using blood collection on filter paper in leprosy patients

The authors studied 70 leprosy patients and 20 normal individuals, comparing the traditional sera collection method and the finger prick blood with the conservation on filter paper for specific antibodies against the native phenolic glycolipid-I (PGL-I) from Mycobacterium leprae. The finger prick blood dried on filter paper was eluated in phosphate buffer saline (PBS) containing 0.5% gelatin. The classical method for native PGL-I was performed for these eluates, and compared with the antibody determination for sera. It was observed that there is a straight correlation comparing these two methods; although the titles found for the eluates were lower than those obtained for serology. This blood collection method could be useful for investigation of new leprosy cases in field, specially in contacts individuals.

Seroprevalence of hepatitis B virus infection in individuals with clinical evidence of hepatitis in Goiânia, Goiás: detection of viral DNA and determination of subtypes

The presence of serological markers for hepatitis B virus (HBsAg, anti-HBc IgM and Anti-HBc total) was investigated in the serum of 1,396 individuals who had clinical suspect of hepatitis. It was observed that 50.7% of the individuals were positive and, from the total of the studied individuals, 14.5% were positive for HBsAg. From these, 8.5% were also positive for anti-HBc IgM. The analysis in relation to gender showed a higher seroprevalence index among male individuals (p < 0.0001). It was observed the occurrence of subtypes adw2 (62.7%), ayw3 (23.5%), ayw2 (9.8%) and adw4 (3.9%). The viral DNA was detected in 61 (33.9%) HBsAg positive samples and in one sample positive only for anti-HBc total. These results indicate an important incidence of the HBV infection in this population, and reinforce previous studies regarding this virus in the central west region of Brazil.

Determination of human cytomegalovirus genetic diversity in different patient populations in Costa Rica

Seroprevalence of HCMV in Costa Rica is greater than 95% in adults; primary infections occur early in life and is the most frequent congenital infection in newborns. The objectives of this study were to determine the genetic variability and genotypes of HCMV gB gene in Costa Rica. Samples were collected from alcoholics, pregnant women, blood donors, AIDS patients, hematology-oncology (HO) children and HCMV isolates from neonates with cytomegalic inclusion disease. A semi-nested PCR system was used to obtain a product of 293-296 bp of the gB gene to be analyzed by Single Stranded Conformational Polymorphism (SSCP) and sequencing to determine the genetic polymorphic pattern and genotypes, respectively. AIDS patients showed the highest polymorphic diversity with 14 different patterns while fifty-six percent of HO children samples showed the same polymorphic pattern, suggesting in this group a possible nosocomial infection. In neonates three genotypes (gB1, gB2 and gB3), were determined while AIDS patients and blood donors only showed one (gB2). Of all samples analyzed only genotypes gB1, 2 and 3 were determined, genotype gB2 was the most frequent (73%) and mixed infections were not detected. The results of the study indicate that SSCP could be an important tool to detect HCMV intra-hospital infections and suggests a need to include additional study populations to better determine the genotype diversity and prevalence.

Genotyping, serotyping and determination of mating-type of Cryptococcus neoformans clinical isolates from São Paulo State, Brazil

The basidiomycetous yeast Cryptococcus neoformans is an important fungal pathogen mainly in immunocompromised patients. In this study, 47 clinical isolates of C. neoformans from regions of São Paulo State were studied serologically by using the Crypto Check Iatron RM 304-K kit, their genetic diversity was estimated by PCR-fingerprinting with a microsatellite-specific sequence (GACA)4, RAPD with primer 6 (Amersham Pharmacia Biotech), PCR-restriction fragment length polymorphism (RFLP) analysis of the phospholipase B gene (PLB1) digested with AvaI and mating type analysis by PCR. All 47 strains isolated from HIV positive patients included in this study were serotype A and MATalpha. The majority of the isolates (45/47) were VNI and only two were VNII by PCR-fingerprinting and PCR-RFLP analysis. High degree of homogeneity was observed when (GACA)4 was used, being highly correlated (> 0.9). In contrast, the RAPD analysis was more heterogeneous with higher number of molecular profiles. By PCR-RFLP, no new molecular type was found, enhancing the suggestion that the differences based on conserved gene as PLB1, can be resultant of ongoing divergent evolution within the C. neoformans complex, into the current eight subtypes. Our results furnish new information on the molecular epidemiology of C. neoformans in the southeast region of Brazil.

DETERMINATION OF VIRAL TROPISM BY GENOTYPING AND PHENOTYPING ASSAYS IN BRAZILIAN HIV-1-INFECTED PATIENTS

The clinical application of CCR5 antagonists involves first determining the coreceptor usage by the infecting viral strain. Bioinformatics programs that predict coreceptor usage could provide an alternative method to screen candidates for treatment with CCR5 antagonists, particularly in countries with limited financial resources. Thus, the present study aims to identify the best approach using bioinformatics tools for determining HIV-1 coreceptor usage in clinical practice. Proviral DNA sequences and Trofile results from 99 HIV-1-infected subjects under clinical monitoring were analyzed in this study. Based on the Trofile results, the viral variants present were 81.1% R5, 21.4% R5X4 and 1.8% X4. Determination of tropism using a Geno2pheno[coreceptor] analysis with a false positive rate of 10% gave the most suitable performance in this sampling: the R5 and X4 strains were found at frequencies of 78.5% and 28.4%, respectively, and there was 78.6% concordance between the phenotypic and genotypic results. Further studies are needed to clarify how genetic diversity amongst virus strains affects bioinformatics-driven approaches for determining tropism. Although this strategy could be useful for screening patients in developing countries, some limitations remain that restrict the wider application of coreceptor usage tests in clinical practice.

Determination of germ tube, phospholipase, and proteinase production by bloodstream isolates of Candida albicans

Introduction Candida albicans is a commensal and opportunistic agent that causes infection in immunocompromised individuals. Several attributes contribute to the virulence and pathogenicity of this yeast, including the production of germ tubes (GTs) and extracellular hydrolytic enzymes, particularly phospholipase and proteinase. This study aimed to investigate GT production and phospholipase and proteinase activities in bloodstream isolates of C. albicans. Methods One hundred fifty-three C. albicans isolates were obtained from blood samples and analyzed for GT, phospholipase, and proteinase production. The assays were performed in duplicate in egg yolk medium containing bovine serum albumin and human serum. Results Detectable amounts of proteinase were produced by 97% of the isolates, and 78% of the isolates produced phospholipase. GTs were produced by 95% of the isolates. A majority of the isolates exhibited low levels of phospholipase production and high levels of proteinase production. Conclusions Bloodstream isolates of C. albicans produce virulence factors such as GT and hydrolytic enzymes that enable them to cause infection under favorable conditions.

Experimental determination of bone cortex holding power of orthopedic screw

Cylindrical specimens of bone measuring 15 mm in diameter were obtained from the lateral cortical layer of 10 pairs of femurs and tibias. A central hole 3.2 mm in diameter was drilled in each specimen. The hole was tapped, and a 4.5 mm cortical bone screw was inserted from the outer surface. The montage was submitted to push-out testing up to a complete strip of the bone threads. The cortical thickness and rupture load were measured, and the shear stress was calculated. The results were grouped according to the bone segment from which the specimen was obtained. The results showed that bone cortex screw holding power is dependent on the bone site. Additionally, the diaphyseal cortical bone tissue is both quantitatively and qualitatively more resistant to screw extraction than the metaphyseal tissue.

Determination of fat soluble vitamins from amazonian fresh-water fishes. I. HPLC analysis of tambaqui, pirarucu and cuiu-cuiu livers .

It was found that fish livers from the Amazon have considerable amounts of vitamins A, D and E compared with the values of the standartized cod-liver oil. Tambaqui liver oil has high concentration of vitamin A1(retinol) and vitamin A2 (degidroretinol) whereas the liver oils of pirarucu and cuiu-cuiu have preferently the vitamin A2. The contents of the vitamins D and E observed in the liver oils of tambaqui and cuiu-cuiu was extremely high.

Quantitative determination by HPLC of iridoids in the bark and latex of himatanthus sucuuba

The main iridoids from the bark and latex of Himatanthus sucuuba were isolated and characterised by spectroscopic methods. HPLC was used for the quantitative analyses of these iridoids and the chromatograms of bark and latex showed a similar iridoid composition. Both parts of the plant are used in folk medicine for the treatment of various ailments.

On line pre-concentration for simultaneous determination of low molecular weight organic acids and inorganic anions in Amazonian river water samples employing ion chromatography with conductivity detection

An ion chromatography procedure, employing an IonPac AC15 concentrator column was used to investigate on line preconcentration for the simultaneous determination of inorganic anions and organic acids in river water. Twelve organic acids and nine inorganic anions were separated without any interference from other compounds and carry-over problems between samples. The injection loop was replaced by a Dionex AC15 concentrator column. The proposed procedure employed an auto-sampler that injected 1.5 ml of sample into a KOH mobile phase, generated by an Eluent Generator, at 1.5 mL min-1, which carried the sample to the chromatographic columns (one guard column, model AG-15, and one analytical column, model AS15, with 250 x 4mm i.d.). The gradient elution concentrations consisted of a 10.0 mmol l-1 KOH solution from 0 to 6.5 min, gradually increased to 45.0 mmol l-1 KOH at 21 min., and immediatelly returned and maintained at the initial concentrations until 24 min. of total run. The compounds were eluted and transported to an electro-conductivity detection cell that was attached to an electrochemical detector. The advantage of using concentrator column was the capability of performing routine simultaneous determinations for ions from 0.01 to 1.0 mg l-1 organic acids (acetate, propionic acid, formic acid, butyric acid, glycolic acid, pyruvate, tartaric acid, phthalic acid, methanesulfonic acid, valeric acid, maleic acid, oxalic acid, chlorate and citric acid) and 0.01 to 5.0 mg l-1 inorganic anions (fluoride, chloride, nitrite, nitrate, bromide, sulfate and phosphate), without extensive sample pretreatment and with an analysis time of only 24 minutes.

Sex determination in the Giant fish of Amazon Basin, Arapaima gigas (Osteoglossiformes, Arapaimatidae), using laparoscopy

The Giant of Amazon basin, pirarucu, Arapaima gigas, is the largest scaled freshwater fish in the world. pirarucu cultivation has recently started, driven by the decline in natural populations and high market value. Currently, there are no reliable methods for sexual differentiation in this species other than direct examination of gonads, which requires dissection of specimens. A non-lethal and less invasive method for sexual identification is highly desirable in order to properly group broodstock for mating and offspring production. We utilized laparoscopic examination in anesthetized pirarucu to differentiate between male and female individuals. This method allowed for the observation and differentiation of the reproductive organs within an individual. Our results suggest that laparoscopy is an efficient method for sex differentiation in pirarucu causing minimal stress to the fish.

Brazil nuts: determination of natural elements and aflatoxin

A study was carried out to evaluate the association of levels of radioactivity, selenium and aflatoxin in shelled Brazil nuts, which were classified in different sizes, for export. The selenium determinations were performed by inductively coupled plasma optical emission spectrometry (LOQ = 3.0 µg g-1), and aflatoxins were detected by Liquid chromatography-mass spectrometry (LOQ = 0.85 µg kg-1), recovery rates were between 92 and 100%. Radioactivity was measured by high-resolution gamma spectrometry. The selenium mean concentration was (22.7 ± 7.4) µg g-1. (n = 30). Mean activities determined for the following radium isotopes were: 15.77 Bq kg-1 for 224Ra, 104.8 Bq kg-1 for 226Ra and 99.48 Bq kg-1 for 228Ra. For 226Ra, the levels did not vary significantly with nut sizes, although such differences were observed for 224Ra and 228Ra. There was no statistically significant association between the level of selenium and the activity of radionuclides, however, there was correlation between the radionuclides. Aflatoxins above the quantification limit were not found.