Molecular imprinted nanoelectrodes for ultra sensitive detection of ovarian cancer marker

The relentless discovery of cancer biomarkers demands improved methods for their detection. In this work, we developed protein imprinted polymer on three-dimensional gold nanoelectrode ensemble (GNEE) to detect epithelial ovarian cancer antigen-125 (CA 125), a protein biomarker associated with ovarian cancer. CA 125 is the standard tumor marker used to follow women during or after treatment for epithelial ovarian cancer. The template protein CA 125 was initially incorporated into the thin-film coating and, upon extraction of protein from the accessible surfaces on the thin film, imprints for CA 125 were formed. The fabrication and analysis of the CA 125 imprinted GNEE was done by using cyclic voltammetry (CV), differential pulse voltammetry (DPV) and electrochemical impedance spectroscopy (EIS) techniques. The surfaces of the very thin, protein imprinted sites on GNEE are utilized for immunospecific capture of CA 125 molecules, and the mass of bound on the electrode surface can be detected as a reduction in the faradic current from the redox marker. Under optimal conditions, the developed sensor showed good increments at the studied concentration range of 0.5–400 U mL−1. The lowest detection limit was found to be 0.5 U mL−1. Spiked human blood serum and unknown real serum samples were analyzed. The presence of non-specific proteins in the serum did not significantly affect the sensitivity of our assay. Molecular imprinting using synthetic polymers and nanomaterials provides an alternative approach to the trace detection of biomarker proteins.

Molecular screening of Plasmodium sp. asymptomatic carriers among transfusion centers from Brazilian Amazon region

The transmission of malaria in Brazil is heterogeneous throughout endemic areas and the presence of asymptomatic Plasmodium sp. carriers (APCs) in the Brazilian Amazon has already been demonstrated. Malaria screening in blood banks is based on the selection of donors in respect to possible risks associated with travel or residence, clinical evidence and/or inaccurate diagnostic methods thereby increasing the probability of transfusion-transmitted infection. We evaluated the frequency of APCs in four blood services in distinct areas of the Brazilian Amazon region. DNA was obtained from 400 human blood samples for testing using the phenol-chloroform method followed by a nested-PCR protocol with species-specific primers. The positivity rate varied from 1 to 3% of blood donors from the four areas with an average of 2.3%. All positive individuals had mixed infections for Plasmodium vivax and Plasmodium falciparum. No significant differences in the results were detected among these areas; the majority of cases originated from the transfusion centres of Porto Velho, Rondônia State and Macapá, Amapá State. Although it is still unclear whether APC individuals may act as reservoirs of the parasite, efficient screening of APCs and malaria patients in Brazilian blood services from endemic areas needs to be improved.

Neutralization of bitis parviocula (Ethiopian mountain adder) venom by the south african institute of medical research (SAIMR) antivenom

BACKGROUND: The Ethiopian mountain adder (Bitis parviocula) is a viperid known only from a few locations in southwestern Ethiopia. METHODS: a total of 30 µg of B. arietans and B. parviocula venoms were run on a 10-20% Tricine gel. To assay lethality dose fifty (LD50), five groups of eight mice for each venom were used. Hemorrhagic activity for crude venom was tested. Fibrinogenolytic activity of crude venom was measured using (2.5 mg/mL) of fibrinogen solution and (0.03 mg/mL) of crude venom. Gelatinase activity of the venom was tested on a Kodak X-OMAT TM film. Crude venoms of B. parviocula and B. arietans were tested for their abilities to affect clotting time, clotting rate and platelet function on whole human blood. RESULTS: The (SAIMR) antivenom was confirmed in this study to neutralize the lethal activity of venom from Bitis parviocula. The ED50s of SAIMR antivenom on B. parviocula and B. arietans neutralized half of 18.2 and 66.7 mg of venom, respectively. The hemorrhagic activities (MHDs) of B. parviocula and B. arietans were 0.88 and 1.7 µg, respectively. Bitis arietans and B. parviocula venoms degradated α and β chains at different times. The γ chains remained unaffected. Bitis parviocula venom did not exhibit gelatinase activity, while B. arietans had a MGD of 6.9 µg. At 3 mg/mL, the crude venoms of B. parviocula and B. arietans did not significantly affect clotting time or clotting rate. CONCLUSIONS: The SAIMR antivenom is very effective in neutralizing the venom of B. parviocula and should be considered in treating envenomations by these snakes.

COLONIZATION OF PALM TREES BY Rhodnius neglectus AND HOUSEHOLD AND INVASION IN AN URBAN AREA, ARAÇATUBA, SÃO PAULO STATE, BRAZIL

The objective of this study is to report on the colonization of palm trees by Rhodnius neglectus, its invasion in an urban area, in Araçatuba - São Paulo, and the control and surveillance measures that have been put in place. Domiciliary triatomine searches occurred in apartments upon the inhabitants' notification. The collected insects were identified and examined for natural infection and food sources with a precipitin test. To search the palm trees, tarps were used to cover the floor, and a “Munck” truck equipped with a tree-pruning device was utilized. Chemical control was performed with the utilization of a manual compression. In 2009, 81 specimens of Rhodnius neglectus were collected from the domiciles by the population. The precipitin test revealed a presence of human blood in 2.7% of the samples. Entomological studies were carried out in these domiciles and in those located within a radius of 200 meters. The search performed in the palm trees resulted in the capture of 882 specimens of triatomines, negative for tripanosomatids. Mechanical and chemical controls were carried out. New searches conducted in the palm trees in the same year resulted in the capture of six specimens. The mechanical and chemical controls of the palm trees, together with the population's work, proved to be effective, therefore preventing these insects' colonization of the city's domiciles.

Transimpedance amplifier for integrated SpO2 optic sensor

Dissertação para obtenção do Grau de Mestre em Engenharia Electrotécnica e de Computadores

HDL in HIV-1 infection: a quality perspective through paraoxonase-1 activities

Dissertação para obtenção do Grau de Mestre em Genética Molecular e Biomedicina

Study on Chagas disease occurrence in the municipality of Monte Negro, State of Rondônia, Brazilian Amazon

INTRODUCTION: Studies on Chagas disease deal with the perspective of its occurrence in the Amazon region, which is directly correlated to the population growth and the spread of the bug biotope. The state of Rondônia has an immense source of vectors (Triatomine) and reservoirs of Trypanosoma cruzi. Environmental changes brought forth by the deforestation in the region may cause vector behavior changes and bring these vectors to a closer contact with humans, increasing the probability of vector infection. METHODS: This study was carried out to check the occurrence of Chagas disease in the municipality of Monte Negro, Rondônia, Brazil, based on a random sampling of the farms and people wherein blood collection from the population and capturing triatomines were done. The blood samples were submitted to serologic tests to detect antibodies of the IgG class against T. cruzi. The triatomines that were collected had their digestive tract checked for the presence of trypanosomatidae with morphology resembling that of the T. cruzi. RESULTS: The population examined was mostly from other states. From the 322 bugs examined on the microscope, 50% showed parasites with morphology compatible with T. cruzi. From the serology of 344 random samples of human blood, 1.2% was found positive, 6% showed inconclusive results, and 92.8% were negative. CONCLUSIONS: Monte Negro shows low prevalence of human infection by T. cruzi and none active vector transmission; however, preventive and surveying measures, which are not performed until now, shall be taken due to the abundance of vectors infected by trypanosomatidae.

Risk factors for hepatitis C virus transmission in the municipality of Catanduva, State of São Paulo: a case-control study

Introduction Hepatitis C virus (HCV) is primarily transmitted via contact with the blood of infected patients, although the form of contact has not been identified for a significant percentage of carriers. The present study evaluated possible risk factors for HCV transmission in a medium-sized town located in the northwest region of the State of São Paulo. Methods This was a case-control study, with the case group consisting of 190 chronic HCV carriers older than 18 years residing in the municipality of Catanduva. The control group also consisted of 190 individuals with HCV-negative serology. The groups were paired (1:1) for gender, age range (± five years), and place of residence. The same structured questionnaire was applied to all subjects, who gave written informed consent to participate in the study. The data were statistically analyzed using crude and adjusted logistic regression, and the results were expressed as odds ratios with a 95% confidence interval. Results The demographic profiles of the groups indicated a predominance of males (68.9%) and mean ages of 47.1 years (case group) and 47.3 years (control group). After adjusting for conditional regression, the following factors were found to represent risks for HCV: history of sexually transmitted disease (STD) and blood transfusion; accidents with syringes and/or needles; tattoos; and the use of non-injectable drugs and injectable medications. Conclusions The transmission of HCV via the blood route has been well characterized. Other forms of contact with human blood and/or secretions are likely to transmit the virus, although with a lower frequency of occurrence.

Lactacidemia: métodos de dosagem do ácido lático

A critical study of three methods for the determination of lactic acid (EDWARDS, MENDEL & GOLDSCHEIDER, MILLER & MUNTZ) is presented and some modifications are proposed. It was shown t hat more accurate results could be obtained with Edward's technic when an Iena glass filter is connected with the absorption tube. Before the dropping of the permanganate solution it is necessary to pass a current of air through the reaction flask to avoid the oxidation of the non-lactic acid substances which interfere with the reaction. The absorption tube must be maintained at 18°C during the destillation and the titration of the bisulphite binding aldehyde at 4°C. When the sample contains more than 5 mg it is useful to work with greater quantities of the bisulphite. More permanganate is consumed when the lactic acid concentration is higher. The sensivity of the method permits the titration of 0.04 mg to 5 mg of lactic acid in the sample. The calculated error of the method gave 0.018 % and the normal values for blood determined in 20 human cases averaged 10.30 mg per 100 ml (Table VI). MENDEL and GOLDSCHEIDER'S method was modified in the following details: Somogyis deproteinization was performed instead metaphosphoric acid as in the original method; to avoid the evaporation of the acetic aldehyde during the heating time with sulfuric acid a special glass stopped tube is proposed (Fig. 2). The reaction with sulfuric acid and veratrol is performed in an ice bath. Blood proteins precipitants were tried and Somogyi's lattest tecnic showed better results (Table V). Colorimetric readings were done in the PULFRICH photometer using filter S 53 and a 10 mm cup. The method is accurate within an error of 0.23 % and samples of 5 to 70 microg. could be easily determined. Normal values for human blood averaged 10.78 mg per 100 ml. More accurate results were obtained with the technic of MILLER & MUNTZ. Slight modifications were introduced: deproteinization with copper sulfate and sodium tungstate; satured p-hydroxydiphenyl solution according to KOENEMANN which is stable for 5 months when stored in the ice-box. Using the PULFRICH step-photometer the error is 0.17% with samples varying from 0.1 to 10 microg. of lactic acid. The filter employed was S 57 with the 5 mm cup. The method was adapted to 0.1 ml of blood. Normal values for human blood gave an average of 10.58 mg per 100 ml.

Endogenous and exogenously-induced immunomodulation of tumour-host responsiveness

In spite of the availability of multiple effector mechanisms of the immune system to combat tumour growth and metastases, their impairment frequently accompanies the appearance of cancer. Factors contributing to this impairment may be related to properties of the host and/or the tumour itself and may be with respect to their origin -endogenous or exogenour. Based on the unique biological behavior of prostate cancer (PCa), and its apparent escape from immune surveillance in the presence of tumour immuno genicity, continuing investigation of endogenous and exogenous factors thought to be relevant to its pathogenesis have been made. For this purpose further studies of the suggested role of human seminal plasma (SePl) and the synthetic oestrogen, diethylstiboestrol (DES), as representative endogenous and exogenous immunomodulatory factors (IMF) of tumour-host responsiveness, together with evaluation of human prostatic tissue extracts and leuprolide (the luteinizing-hormone-releasing-hormone proposed as an alternate to DES therapy) have been made by evaluating their effect on the lytic activity of natural killer (NK) cells. SePl and prostate extracts significantly suppressed NK cell lysis. Physicochemical studies suggest SePl and prostate IMF to be associated with high and low molecular weight macromolecules; and implicate the participation of transglutaminase and prostaglandins. Comparative study of therapeutic levels of DES vs. leuprolide on NK cell lysis demonstrated significant suppression by DES vs. a negligible effect of leuprolide. Metastases are highly prevalent in PCa, and contribute significantly to its morbidity and mortality. Further knowledge of the range of effects of endogenous and exogenous IMF on effector mechanisms of tumour-host responsiveness, to include suppression of NK cells, and elucidation of their nature, may contribute toward our understanding of the unique biological behavior of tumours of the prostate, in addition to improvement in their clinical management.

Hemolytic activity of Trichomonas vaginalis and Tritrichomonas foetus

The hemolytic activity of live isolates and clones of Trichomonas vaginalis and Tritrichomonas foetus was investigated. The isolates were tested against human erythrocytes. No hemolytic activity was detected by the isolates of T. foetus. Whereas the isolates of T. vaginalis lysed erythrocytes from all human blood groups. No hemolysin released by the parasites could be detected. Our preliminary results suggest that hemolysis depend on the susceptibility of red cell membranes to destabilization and the intervention of cell surface receptors as a mechanism of the hemolytic activity. The mechanism could be subject to strain-species-genera specific variation of trichomonads. The hemolytic activity of T. vaginalis is not due to a hemolysin or to a product of its metabolism. Pretreatment of trichomonads with concanavalin A reduced levels of hemolysis by 40%.

Bloodmeal microfilariae density and the uptake and establishment of Wuchereria bancrofti infections in Culex quinquefasciatus and Aedes aegypti

The relationship between ingestion of microfilariae (mf), production of infective larvae (L3) and mf density in human blood has been suggested as an important determinant in the transmission dynamics of lymphatic filariasis. Here we assess the role of these factors in determining the competence of a natural vector Culex quinquefasciatus and a non vector Aedes aegypti to transmit Wuchereria bancrofti. Mosquitoes were infected via a membrane feeding procedure. Both mosquito species ingested more than the expected number of microfilariae (concentrating factor was 1.28 and 1.81 for Cx. quinquefasciatus and Ae. aegypti, respectively) but Cx. quinquefasciatus ingested around twice as many mf as Ae. aegypti because its larger blood meal size. Ae. aegypti showed a faster mf migration capacity compared to Cx. quinquefasciatus but did not allow parasite maturation under our experimental conditions. Similar proportions of melanized parasites were observed in Ae. aegypti (2.4%) and Cx. quinquefasciatus (2.1%). However, no relationship between rate of infection and melanization was observed. We conclude that in these conditions physiological factors governing parasite development in the thorax may be more important in limiting vectorial competence than the density of mf ingested.

Higher memory responses in HIV-infected and kidney transplanted patients than in healthy subjects following priming with the pandemic vaccine.

BACKGROUND: Memory responses require immune competence. We assessed the influence of priming with AS03-adjuvanted pandemic vaccine (Pandemrix®) on memory responses of HIV patients, kidney recipients (SOT) and healthy controls (HC). METHOD: Participants (HIV: 197, SOT: 53; HC: 156) were enrolled in a prospective study and 390/406 (96%) completed it. All had been primed in 2009/2010 with 1 (HC) or 2 (patients) doses of Pandemrix®, and were boosted with the 2010/2011 seasonal influenza vaccine. Geometric mean titres and seroprotection rates were measured 12 months after priming and 4 weeks after boosting. Primary and memory responses were directly compared in 191 participants (HCW: 69, HIV: 71, SOT: 51) followed during 2 consecutive seasons. RESULTS: Most participants (HC: 77.8%, HIV: 77.6%, SOT: 66%) remained seroprotected at 12 months post-priming. Persisting A/09/H1N1 titers were high in HIV (100.2) and HC (120.1), but lower in SOT (61.4) patients. Memory responses reached higher titers in HIV (507.8) than in HC (253.5) and SOT (136.9) patients. Increasing age and lack of HAART reduced persisting and memory responses, mainly influenced by residual antibody titers. Comparing 2009/2010 and 2010/2011 titers in 191 participants followed for 2 seasons indicated lower post-2010/2011 titers in HC (240.2 vs 313.9), but higher titers in HIV (435.7 vs 338.0) and SOT (136 vs 90.3) patients. CONCLUSIONS: Priming with 2 doses of Pandemrix® elicited persistent antibody responses and even stronger memory responses to non-adjuvanted seasonal vaccine in HIV patients than 1 dose in healthy subjects. Adjuvanted influenza vaccines may improve memory responses of immunocompromised patients. TRIAL REGISTRATION: ClinicalTrials.gov NCT01022905.

Malaria vectors in the municipality of Serra do Navio, State of Amapá, Amazon Region, Brazil

We conducted a survey to determine the vectors of malaria in six localities of Serra do Navio municipality, State of Amapá, from 1990 to 1991. Malaria infection rates of 29.3%, 6.2% and 20.4% were detected by human blood smears in Colônia Água Branca, Porto Terezinha and Arrependido, respectively. There was no malaria infection detected in Serra do Navio. Fifteen species were identified among 3,053 anopheline mosquitoes collected by human bait and 64.4% were identified as Anopheles albitarsis s.l., 16.7% An. braziliensis, 9.5% An. nuneztovari and 5.8% An. triannulatus. An. darlingi, the main vector of malaria in the Amazon region of Brazil, was scare. Using enzyme-linked immunosorbent assay (ELISA), a total positive rate of 0.8% (23/2876) was found for six species: fifteen An. albitarsis s.l., four An. nuneztovari, and one of each: An. braziliensis, An. triannulatus, An. oswaldoi and An. rangeli. Nine of 23 positive mosquitoes were infected with Plasmodium malariae, eight with P. vivax VK210, three with P. vivax VK247 and three with P. falciparum. Since An. albitarsis s.l. was collected feeding on humans, was present in the highest density and was positive by ELISA for malaria sporozoites, it probably plays an important role in malaria transmission in this area.

Environmental characteristics of the cemeteries of Buenos Aires City (Argentina) and infestation levels of Aedes aegypti (Diptera: Culicidae)

Cemeteries with many water-filled containers, flowers, sources of human blood, and shade are favorable urban habitats for the proliferation of Aedes aegypti, a vector of yellow fever and dengue. A total of 22,956 containers was examined in the five cemeteries of the city of Buenos Aires, Argentina. The vector was found in four cemeteries that showed an average infestation level of 5.5% (617 positive out of 11,196 water-filled containers). The four cemeteries positive for Ae. aegypti showed significantly different (p<0.01) infestation levels. Vegetation cover and percentage of infestation were significantly correlated (p<0.01), but neither cemetery area nor number of available containers were significantly related to the proportion of positive vases. Our results suggest that the cemeteries of Buenos Aires represent a gradient of habitat favorableness for this vector species, some of which may act as foci for its proliferation and dispersal.