Antibodies to variable surface antigens induce antigenic variation in the intestinal parasite Giardia lamblia.

The genomes of most protozoa encode families of variant surface antigens. In some parasitic microorganisms, it has been demonstrated that mutually exclusive changes in the expression of these antigens allow parasites to evade the host's immune response. It is widely assumed that antigenic variation in protozoan parasites is accomplished by the spontaneous appearance within the population of cells expressing antigenic variants that escape antibody-mediated cytotoxicity. Here we show, both in vitro and in animal infections, that antibodies to Variant-specific Surface Proteins (VSPs) of the intestinal parasite Giardia lamblia are not cytotoxic, inducing instead VSP clustering into liquid-ordered phase membrane microdomains that trigger a massive release of microvesicles carrying the original VSP and switch in expression to different VSPs by a calcium-dependent mechanism. This novel mechanism of surface antigen clearance throughout its release into microvesicles coupled to the stochastic induction of new phenotypic variants not only changes current paradigms of antigenic switching but also provides a new framework for understanding the course of protozoan infections as a host/parasite adaptive process.

Efficient oral vaccination by bioengineering virus-like particles with protozoan surface proteins.

Intestinal and free-living protozoa, such as Giardia lamblia, express a dense coat of variant-specific surface proteins (VSPs) on trophozoites that protects the parasite inside the host's intestine. Here we show that VSPs not only are resistant to proteolytic digestion and extreme pH and temperatures but also stimulate host innate immune responses in a TLR-4 dependent manner. We show that these properties can be exploited to both protect and adjuvant vaccine antigens for oral administration. Chimeric Virus-like Particles (VLPs) decorated with VSPs and expressing model surface antigens, such as influenza virus hemagglutinin (HA) and neuraminidase (NA), are protected from degradation and activate antigen presenting cells in vitro. Orally administered VSP-pseudotyped VLPs, but not plain VLPs, generate robust immune responses that protect mice from influenza infection and HA-expressing tumors. This versatile vaccine platform has the attributes to meet the ultimate challenge of generating safe, stable and efficient oral vaccines.

Case report of clinical salmonellosis by Salmonella Typhimurium that occurred in Portuguese children.

AIMS: Aim of this study is to characterize clinical isolates of Salmonella Typhimurium that occurred in Portuguese children on the basis of their virulence and antimicrobial resistance profiles and pulsed-field gel electrophoresis typing and to analyse possible strain relatedness. METHODS AND RESULTS: Different Salmonella serotypes were isolated from clinical cases of salmonellosis that had occurred in two Portuguese hospitals (a total of 259 isolates). All Salm. Typhimurium strains, with the age of the patients known, (total of 26 isolates) were selected for this study. These isolates were characterized for their virulence gene profiles (agfA, iroB, slyA, hin/H2, spv), antimicrobial resistance profiles and investigated for the occurrence of multidrug-resistant Salm. Typhimurium DT 104 by PCR. Salmonella isolates showed high rates of resistance to four or more antibiotics, 100% resistance to sulfadiazine and a high percentage of strains with the resistance profile of Salm. Typhimurium DT 104, two of them with this phage type (determined by PCR). A relationship between some clusters and their resistance and virulence profiles was detected, each cluster having the same profile. CONCLUSIONS: This study showed high-antibiotic resistance of the Salmonella strains investigated, and the presence of multidrug-resistant Salm. Typhimurium DT104 in infections of Portuguese children. SIGNIFICANCE AND IMPACT OF THE STUDY: Study is based on regarding the increase in antibiotic resistance by Salmonella strains isolated from infections in Portuguese children and on the presence of Salm. Typhimurium DT 104 circulating in Portugal.

Characterization of microbial population of 'Alheira' (a traditional Portuguese fermented sausage) by PCR-DGGE and traditional cultural microbiological methods.

AIMS: This study evaluates the microbial ecology of 'Alheira' by traditional microbiological analysis and a PCR-denaturing gradient gel electrophoresis (DGGE) protocol. METHODS AND RESULTS: Total microbial DNA from 'Alheiras' was extracted directly from the products and subjected to PCR using Eubacterial primers for 16S rDNA. The amplicons were separated by DGGE. The results demonstrated that different products of the same batch display identical profiles, whereas products from different batches of the same producer could display different DGGE profiles. 'Alheiras' from different producers were distinguishable based on the respective DGGE profiles. The obtained sequences from prevalent phylotypes affiliated with order Lactobacillales and order Bacillales and class Gammaproteobacteria. The same samples were subjected to traditional microbiological analysis. In both methods, lactic acid bacteria were dominant and were present together with other organisms, mainly members of the family Micrococcaceae. CONCLUSIONS: The approach explored in this study allowed the description of the microbial community present in 'Alheira' in particular the diversity of lactic acid bacteria. SIGNIFICANCE AND IMPACT OF THE STUDY: This can be useful for the microbiological characterization of traditional products in order to develop new methods of quality control capable of supporting a standardization of the processes, while preserving their typical traits.

Quinacrine sterilization (QS) experience in the Philippines: a preliminary report.

OBJECTIVE: The first clinical trial of Quinacrine Sterilization (QS) in the Philippines was undertaken in Ceby City on January 10, 2000, to evaluate the accetability, safety, effectiveness and side effects of this technology. We intend to recruit 500 patients to utilize this technique for limiting family size. For the purposes of this report, our cut-off date is April 11, 2003. METHODS: Over more than two years, QS was performed on 36 volunteer patients. After careful explanation of the procedure and given the opportunity to ask questions, they had signed an informed consent. The trial involved transcervical insertion of 252 mg quinacrine in the form of pellets, and placed at the tip of the uterine fundus on two occasions, a month apart. Condoms were routinely provided to all patients except those on oral contraceptive pills and DMPA after the first insertion to be used for six weeks after the second one. As the numbers are small, no statistical evaluation was called for. RESULTS: The accumulated experience was 515 woman-months. There were no pregnancies, neither ectopic nor intrauterine. Adverse events (AE) were mild. Some patients complained of a yellow discharge and itching. Fifty percent experienced midl abdominal discomfort which was easily managed with mefenamic acid. CONCLUSIONS: Although this is a small study, we believe that QS is both safe and effective and we are strongly encouraged to continue to offer this nonsurgical sterilization method to our patients.

Quinacrine sterilization (QS) experience in the Philippines: a preliminary report.

OBJECTIVE: The first clinical trial of Quinacrine Sterilization (QS) in the Philippines was undertaken in Cebu City on January 10, 2000, to evaluate the acceptability, safety, effectiveness and side effects of this technology. We intend to recruit 500 patients to utilize this technique for limiting family size. For the purposes of this report, our cut-off date is April 11, 2003. METHODS: Over more than two years, QS was performed on 36 volunteer patients. After careful explanation of the procedure and given the opportunity to ask questions, they had signed an informed consent. The trial involved transcervical insertion of 252 mg quinacrine in the form of pellets, and placed at the tip of the uterine fundus on two occasions, a month apart. Condoms were routinely provided to all patients except those on oral contraceptive pills and DMPA after the first insertion to be used for six weeks after the second one. As the numbers are small, no statistical evaluation was called for. RESULTS: The accumulated experience was 515 woman-months. There were no pregnancies, neither ectopic nor intrauterine: Adverse events (AE) were mild. Some patients complained of a yellow discharge and itching. Fifty percent experienced mild abdominal discomfort which was easily managed with mefenamic acid. CONCLUSIONS: Although this is a small study, we believe that QS is both safe and effective and we are strongly encouraged to continue to offer this nonsurgical sterilization method to our patients.

Gellan gum biosynthesis in Sphingomonas paucimobilis ATCC 31461: genes, enzymes and exopolysaccharide production engineering.

The commercial gelling agent, gellan, is an extracellular polysaccharide (EPS) produced by Sphingomonas paucimobilis ATCC 31461. In recent years, significant progress in understanding the relationship between gellan structure and properties and elucidation of the biosynthesis and engineering of this recent product of biotechnology has been made. This review focuses on recent advances in this field. Emphasis is given to identification and characterization of genes and enzymes involved, or predicted to be involved, in the gellan biosynthetic pathway, at the level of synthesis of sugar-activated precursors, of the repeat unit assembly and of gellan polymerization and export. Identification of several genes, biochemical characterization of the encoded enzymes and elucidation of crucial steps of the gellan pathway indicate that possibilities now exist for exerting control over gellan production at any of the three levels of its biosynthesis. However, a better knowledge of the poorly understood steps and of the bottlenecks and regulation of the pathway, the characterization of the composition, structure and functional properties of gellan-like polymers produced either by the industrial strain under different culture conditions or by mutants are still required for eventual success of the metabolic engineering of gellan production.