Lagochilascaris minor: antibody production in experimentally infected mice

Lagochilascaris minor is the causative agent of lagochilascariosis, a disease that affects the neck region and causes festering abscesses, with eggs, adult parasites and L3/L4 larvae within the purulent exudates. Today, mice are considered to be intermediate hosts for the parasite. C57BL/6 mice produce immunoglobulin IgM, IgA and IgG against the crude extract of the parasite; on the other hand, antibodies produced against the secreted/excreted antigens of Lagochilascaris minor present lower levels of IgM, IgA and IgG. This is the first description of antibody detection against different antigens of Lagochilascaris minor.

Functional genomic analysis of heat stress in Vitis vinifera

Grapevine (Vitis vinifera) is one of most agro-economically important fruit crops worldwide, with a special relevance in Portugal where over 300 varieties are used for wine production. Due to global warming, temperature stress is currently a serious issue affecting crop production especially in temperate climates. Mobile genetic elements such as retrotransposons have been shown to be involved in environmental stress induced genetic and epigenetic modifications. In this study, sequences related to Grapevine Retrotransposon 1 (Gret1) were utilized to determine heat induced genomic and transcriptomic modifications in Touriga Nacional, a traditional Portuguese grapevine variety. For this purpose, growing canes were treated to 42 oC for four hours and leaf genomic DNA and RNA was utilized for various techniques to observe possible genomic alterations and variation in transcription levels of coding and non-coding sequences between non-treated plants and treated plants immediately after heat stress (HS-0 h) or after a 24 hour recovery period (HS-24 h). Heat stress was found to induce a significant decrease in Gret1 related sequences in HS-24 h leaves, indicating an effect of heat stress on genomic structure. In order to identify putative heat induced DNA modifications, genome wide approaches such as Amplified Fragment Length Polymorphism were utilized. This resulted in the identification of a polymorphic DNA fragment in HS-0 h and HS-24 h leaves whose sequence mapped to a genomic region flanking a house keeping gene (NADH) that is represented in multiple copies in the Vitis vinifera genome. Heat stress was also found to affect the transcript levels of various non-coding and gene coding sequences. Accordingly, quantitative real time PCR results established that Gret1 related sequences are up regulated immediately after heat stress whereas the level of transcript of genes involved in identification and repair of double strand breaks are significantly down regulated in HS-0 h plants. Taken together, the results of this work demonstrated heat stress affects both genomic integrity and transcription levels.

Production of anti-Cryptosporidium polyclonal antibodies and standardization of direct immunofluorescence for detecting oocysts in water

INTRODUCTION: The production of anti-Cryptosporidium polyclonal antibodies and its use in direct immunofluorescence assays to determine the presence of Cryptosporidium in water are described in the present work. METHODS: Two rabbits were immunized with soluble and particulate antigens from purified Cryptosporidium oocysts. The sera produced were prepared for immunoglobulin G extraction, which were then purified and conjugated with fluorescein isothiocyanate (FITC). Slides containing known amounts of oocysts were prepared to determine the sensitivity of the technique. To test the specificity, slides containing Giardia duodenalis cysts were prepared. RESULTS: The conjugate was successfully used in water samples experimentally contaminated with Cryptosporidium oocysts, and it was possible to detect up to five oocysts/spot, corresponding to contamination of 250 oocysts/mL. CONCLUSIONS: The three immunizations performed in the rabbits were enough to produce antibodies against Cryptosporidium, the standard direct immunofluorescence assay permitted the detection of five oocysts in 20% of the samples, and no cross-reaction with Giardia duodenalis cysts occurred.

Different culture media containing methyldopa for melanin production by Cryptococcus species

INTRODUCTION: Melanin production by species of Cryptococcus is widely used to characterize C. neoformans complex in mycology laboratories. This study aims to test the efficacy of methyldopa from pharmaceutical tablet as a substrate for melanin production, to compare the production of melanin using different agar base added with methyldopa, and to compare the melanin produced in those media with that produced in Niger seed agar and sunflower seed agar by C. neoformans, C. laurentii, and C. albidus. Two isolates of each species, C. neoformans, C. laurentii, and C. albidus, and one of Candida albicans were used to experimentally detect conditions for melanin production. METHODS: The following media were tested: Mueller-Hinton agar (MHA), brain and heart infusion agar (BHIA), blood agar base (BAB), and minimal medium agar (MMA), all added with methyldopa, and the media Niger seed agar (NSA) and sunflower seed agar (SSA). RESULTS: All isolates grew in most of the culture media after 24h. Strains planted on media BAB and BHIA showed growth only after 48h. All isolates produced melanin in MMA, MHA, SSA, and NSA media. CONCLUSIONS: Methyldopa in the form pharmaceutical tablet can be used as a substrate for melanin production by Cryptococcus species; minimal medium plus methyldopa was more efficient than the BAB, MHA, and BHIA in the melanin production; and NSA and SSA, followed by MMA added with methyldopa, were more efficient than other media studied for melanin production by all strains studied.

Production of cytokine and chemokines by human mononuclear cells and whole blood cells after infection with Trypanosoma cruzi

INTRODUCTION: The innate immune response is the first mechanism of protection against Trypanosoma cruzi, and the interaction of inflammatory cells with parasite molecules may activate this response and modulate the adaptive immune system. This study aimed to analyze the levels of cytokines and chemokines synthesized by the whole blood cells (WBC) and peripheral blood mononuclear cells (PBMC) of individuals seronegative for Chagas disease after interaction with live T. cruzi trypomastigotes. METHODS: IL-12, IL-10, TNF-α, TGF-β, CCL-5, CCL-2, CCL-3, and CXCL-9 were measured by ELISA. Nitrite was determined by the Griess method. RESULTS: IL-10 was produced at high levels by WBC compared with PBMC, even after incubation with live trypomastigotes. Production of TNF-α by both PBMC and WBC was significantly higher after stimulation with trypomastigotes. Only PBMC produced significantly higher levels of IL-12 after parasite stimulation. Stimulation of cultures with trypomastigotes induced an increase of CXCL-9 levels produced by WBC. Nitrite levels produced by PBMC increased after the addition of parasites to the culture. CONCLUSIONS: Surface molecules of T. cruzi may induce the production of cytokines and chemokines by cells of the innate immune system through the activation of specific receptors not evaluated in this experiment. The ability to induce IL-12 and TNF-α contributes to shift the adaptive response towards a Th1 profile.

Establishment and evaluation of flexible insect cell lines for rapid production of recombinant proteins

Fundação para a Ciência e a Tecnologia (FCT) - (PTDC/EBB-EBI/102266/2008 and SFRH/BD/43830/2008, respectively) and by European Community’s FP7/2007-2013 (grant agreement nº 270089)

Production and diagnostic application of recombinant domain III of West Nile envelope protein in Brazil

INTRODUCTION: West Nile virus (WNV) is a flavivirus with a natural cycle involving mosquitoes and birds. Over the last 11 years, WNV has spread throughout the Americas with the imminent risk of its introduction in Brazil. METHODS: Envelope protein domain III of WNV (rDIII) was bacterially expressed and purified. An enzyme-linked immunosorbent assay with WNV rDIII antigen was standardized against mouse immune fluids (MIAFs) of different flavivirus. RESULTS: WNV rDIII reacted strongly with St. Louis encephalitis virus (SLEV) MIAF but not with other flaviviruses. CONCLUSIONS: This antigen may be a potentially useful tool for serologic diagnosis and may contribute in future epidemiological surveillance of WNV infections in Brazil.

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.

Fruta com cheiro: The implementation of a new brand of fruit in the Portuguese market

‘’Fruta com Cheiro’’ is an idea for a new brand of fruit that would be introduced in the Portuguese market that would differentiate itself from the brands already in the market and from other non-branded fruit producers. In order to prove that the idea was valid and would have a place in the market, two methods of exploratory research were used – in-depth interviews and focus groups – to understand attitudes and behaviors regarding fruit selection and purchase and also people’s perceptions to ‘’Fruta com Cheiro’’. After these two steps, several considerations were made in relation to preliminary marketing aspects such as brand creation and positioning. There was also a final remark on the fact that this thesis is not a business plan and its purpose was to show how viable would the project be.

Evolution of coauthorship networks: worldwide scientific production on leishmaniasis

Introduction Collaboration is one of the defining features of contemporary scientific research, and it is particularly important with regard to neglected diseases that primarily affect developing countries. Methods The present study has identified publications on leishmaniasis in the Medline database from 1945 to 2010, analyzing them according to bibliometric indicators and statistics from social network analysis. Examining aspects such as scientific production, diachronic evolution, and collaboration and configuration of the research groups in the field, we have considered the different types of Leishmania studied and the institutional affiliation and nationality of the authors. Results Seven-hundred and thirty-five authors participate in 154 prominent research clusters or groups. Although the most predominant and consolidated collaborations are characterized by members from the same country studying the same type of Leishmania, there are also notable links between authors from different countries or who study different clinical strains of the disease. Brazil took the lead in this research, with numerous Brazilian researchers heading different clusters in the center of the collaboration network. Investigators from the USA, India, and European countries, such as France, Spain, the United Kingdom, and Italy, also stand out within the network. Conclusions Research should be fostered in countries such as Bangladesh, Nepal, Sudan, and Ethiopia, where there is a high prevalence of different forms of the disease but limited research development with reference authors integrated into the collaboration networks.

Shrinking shrinkage:How and why is fruit shrinkage being caused in Sonaemc's convenience stores? Finding opportunities of improvement envisioning its optimization

This work project regards a challenge presented by a Portuguese organization on the retail sector, SONAEMC, which is a case study of how and why fruit shrinkage occurs in the fruit supply chain within their convenience stores. A qualitative research methodology enabled to infer in which stages throughout the chain shrinkage’s causes occur and, to conclude that internal rules for procedures and processes are not always followed and whose compliance would be enough to reduce fruit shrinkage. The key conclusion is that if fruit stock loss is reduced by as much as 15% the category’s profitability could increase about 8%.

Lucuma as an exotic high quality fruit imported into Portugal and the UE

This project has two different goals, one of them is to promote the consumption of exotic fruits with high quality. The other goal of the project is to look for the viability of turning this work project into a real business, focusing in two different channels to diversify its revenues: B2B and B2C. In order to achieve this second goal, this project aims to see the best way to commercialize this product (Lucuma powder and Pulp of Lucuma) and how to make it in an efficient way with right companies. Therefore, the project aims to create a company to commercialize the product between the producers in Peru and possibly small businesses interested in acquire the processed fruit and also individuals interested in own consumption in small quantities. This project, if successful, tries to diversify the consumption into other good organic healthy products in the long-term.

Production of metallo-β-lactamase among Pseudomonas aeruginosa strains isolated in the State of Sergipe, Brazil

INTRODUCTION: Acquired production of metallo-β-lactamases is an important mechanism of resistance in Pseudomonas aeruginosa. The objective of this study was to investigate the production of metallo-β-lactamase and the genetic diversity among ceftazidime-resistant P. aeruginosa isolates from State of Sergipe, Brazil. METHODS: Metallo-β-lactamase was investigated using the disk approximation test and polymerase chain reaction (PCR). Genetic diversity was evaluated by pulsed-field gel electrophoresis (PFGE). RESULTS: A total of 48 (51.6%) isolates were resistant to ceftazidime. Six (12.2%) of these were positive for metallo-β-lactamase production. Only two (4.1%) of the ceftazidime-resistant isolates carried the bla SPM-1 gene. CONCLUSIONS: Production of metallo-β-lactamases was not the main mechanism of resistance to ceftazidime and carbapenems among P. aeruginosa strains in Sergipe, Brazil.