Biomedical subjects

Fernando Ferreira

Publications and source records attributed to Fernando Ferreira.

17 recordsLinked to original sources

PubMed2006

Risk factors associated with sero-positivity to Toxoplasma gondii in captive neotropical felids from Brazil.

From September 1995 to February 2001, blood samples were collected from 865 neotropical felids belonging to 8 different species. These animals were housed in 86 institutions located in 78 cities of 20 Brazilian states. Our goal was to identify the risk factors associated with sero-positivity to Toxoplasma gondii in captive neotropical felids from Brazil. All serum samples were tested by the modified agglutination test (MAT), using formalin-fixed whole tachyzoites and mercaptoethanol. For each animal an individual questionnaire was filled with questions about tattoo number, felid species, age, sex, origin, number of animals in the group, introduction of new animals in the group, time in the institution, eating meat previously frozen for a period <7 days in the last 6 months, eating meat of run-over or euthanized animals in the last 6 months, predation of rodents or birds in the last 6 months and presence of domestic cats near the enclosures in the last 6 months. The total sero-prevalence was 55% (95% CI: 52%, 57%). We estimated a prevalence of 46% (95% CI: 40%, 54%) for jaguarundi (Puma yagouaroundi); 58% (95% CI: 53%, 63%) for ocelot (Leopardus pardalis); 50% (95% CI: 45%, 56%) for oncilla (L. tigrinus); 54% (95% CI: 46%, 62%) for margay (L. wiedii); 12% (95% CI: 4%, 31%) for Pampas-cat (L. colocolo); 83% (95% CI: 65%, 93%) for Geoffroy's-cat (L. geoffroyi); 64% (95% CI: 50%, 68%) for jaguar (Panthera onca) and 48% (95% CI: 42%, 54%) for puma (Puma concolor). Multiple logistic regression was used to evaluate the association between the variables in the questionnaire and sero-positivity to T. gondii. We concluded that the independent risk factors for toxoplasmosis were: age >3 years (OR=4.75 [2.75; 8.2]), eating meat previously frozen for a period <7 days (OR=2.23 [1.24; 4.01]), and consumption of animals that were run-over or euthanized (OR=1.64; [1.14; 2.37]).

Agglutination Tests↗

PubMed2006

Characterization of myo-inositol hexakisphosphate deposits from larval Echinococcus granulosus.

The abundant metabolite myo-inositol hexakisphosphate (InsP6) can form vesicular deposits with cations, a widespread phenomenon in plants also found in the cestode parasite, Echinococcus granulosus. In this organism, the deposits are exocytosed, accumulating in a host-exposed sheath of extracellular matrix termed the laminated layer. The formation and mobilization of InsP6 deposits, which involve precipitation and solubilization reactions, respectively, cannot yet be rationalized in quantitative chemical terms, as the solids involved have not been formally described. We report such a description for the InsP6 deposits from E. granulosus, purified as the solid residue left by mild alkaline digestion of the principal mucin component of the laminated layer. The deposits are largely composed of the compound Ca5H2L.16H2O (L representing fully deprotonated InsP6), and additionally contain Mg2+ (6-9% molar ratio with respect to Ca2+), but not K+. Calculations employing recently available chemical constants show that the precipitation of Ca5H2L.16H2O is predicted by thermodynamics in secretory vesicle-like conditions. The deposits appear to be similar to microcrystalline solids when analysed under the electron microscope; we estimate that each crystal comprises around 200 InsP6 molecules. We calculate that the deposits increase, by three orders of magnitude, the surface area available for adsorption of host proteins, a salient ability of the laminated layer. The major inositol phosphate in the deposits, other than InsP6, is myo-inositol (1,2,4,5,6) pentakisphosphate, or its enantiomer, inositol (2,3,4,5,6) pentakisphosphate. The compound appears to be a subproduct of the intracellular pathways leading to the synthesis and vesicular accumulation of InsP6, rather than arising from extracellular hydrolysis of InsP6.

Animals↗

PubMed2006

Prevalence of Rickettsia felis in the fleas Ctenocephalides felis felis and Ctenocephalides canis from two Indian villages in Sao Paulo Municipality, Brazil.

We evaluated the presence of Rickettsia infection among fleas collected on domestic dogs in two Guarani Indian communities in the suburban area of São Paulo Municipality, Brazil. A total of 114 Ctenocephalides felis felis and 47 Ctenocephalides canis were collected from 40 dogs. A total of 41 C. felis felis (36.0%) and 9 C. canis (19.1%) fleas yielded expected bands by PCR, which were all shown by DNA sequencing to be indentical to the corresponding sequence of a fragment of the Rickettsia felis gltA gene deposited in GenBank. The overall prevalence of R. felis was 31.0% (49/161).

Animals↗

PubMed2005

Universal solid supports for the synthesis of oligonucleotides via a transesterification of H-phosphonate diester linkage.

[Structure: see text]. Three universal solid supports exhibiting an hydroxyl function were prepared. The introduction of a first H-phosphonate diester linkage which was kept throughout the elongation allowed the release of 3'-hydroxyl oligonucleotides by a transesterification mechanism. The transesterification was performed in a few minutes with either amino alcohols or K2CO3/methanol. Starting from a hydroxyl solid support, tandem oligonucleotides were synthesized and the solid support was easily recyclable. This strategy was extended to the release of an oligonucleotide from the solid support by a nonbasic treatment opening the way to the synthesis of base-sensitive oligonucleotides thanks to the selective deprotection of a hydroxyl in beta of the H-phosphonate diester linkage.

Esterification↗

PubMed2005

Silyl protecting groups for oligonucleotide synthesis removed by a ZnBr2 treatment.

An oligonucleotide protected with N-(trimethylsilyloxycarbonyl) (Teoc) and P-(trimethylsilylethanol) (Tse) groups was synthesized and deprotected by a single ZnBr2 treatment. Finally it was released from the solid support by cleavage of a disulfide linkage with TCEP. The olgonucleotide was obtained without any basic treatment.

Bromides↗

PubMed2005

Adoption of shelter dogs in a Brazilian community: assessing the caretaker profile.

A survey in Ibiúna, Sao Paulo, Brazil, of caregivers (owners) who adopted shelter dogs assessed length of ownership, proportion of male and female dogs adopted, and owners' characteristics. It addressed breeding, neutering, vaccination, and veterinary care. It used no testing to provide a good "match" between dog and future owner. Of adopted dogs, 58% were male. Only 36% of owners were located. Mean ownership length was 14.8 months (95% confidence interval = 12.4 to 17.2 months), estimated through a survival analysis method. Of adopted dogs, 40.9% lived with their owners; 34.9% had died (some had lived on the streets); 15.0% were donated; 4.3% ran away; 3.2% were returned to the city shelter. Of interviewees, 57% reported no difficulties with the adoption; 23.1% cited the animal's illness and death as the main difficulty. For contraception, 87 owners (46.7%) chained dogs to prevent contact with other animals; 56.5% were against neutering. Reasons given were compassion (58.1%), unnecessary procedure (11.4%), cost (9.5%), and behavior change (4.8%). This research motivated a design for Ibiúna shelter dog adoption to improve the proportion of successful adoptions.

Animal Welfare↗

PubMed2005

Rickettsial infection in animals and Brazilian spotted fever endemicity.

We compared the rickettsial infection status of Amblyomma cajennense ticks, humans, dogs, and horses in both Brazilian spotted fever (BSF)-endemic and -nonendemic areas in the state of Sao Paulo, Brazil. Most of the horses and few dogs from BSF-endemic areas had serologic titers against Rickettsia rickettsii antigens. In contrast, no dogs or horses from BSF-nonendemic areas had serologic titers against R. rickettsii antigens, although they were continually exposed to A. cajennense ticks. All human serum samples and ticks from both areas were negative by serologic assay and polymerase chain reaction, respectively. Our results indicate that surveys of horse serum are a useful method of BSF surveillance in areas where humans are exposed to A. cajennense ticks. In addition, we successfully performed experimental infection of A. cajennense ticks with R. parkeri.

Animals↗

PubMed2004

Unique precipitation and exocytosis of a calcium salt of myo-inositol hexakisphosphate in larval Echinococcus granulosus.

The ubiquitous intracellular molecule myo-inositol hexakisphosphate (IP6) is present extracellularly in the hydatid cyst wall (HCW) of the parasitic cestode Echinococcus granulosus. This study shows that extracellular IP6 is present as its solid calcium salt, in the form of deposits that are observed, at the ultrastructural level, as naturally electron dense granules some tens of nanometers in diameter. The presence of a calcium salt of IP6 in these structures was determined by two different electron microscopy techniques: (i) the analysis of the spatial distribution of phosphorus and calcium in the outer, acellular layer of the HCW (the laminated layer, LL) through electron energy loss spectroscopy, and (ii) the observation, by transmission electron microscopy, of HCW that were selectively depleted of IP6 by treatment with EGTA or phytase, an enzyme that catalyses the dephosphorylation of IP6. The deposits of the IP6-Ca(II) salt are also observed inside membrane vesicles in cells of the germinal layer (the inner, cellular layer of the HCW), indicating that IP6 precipitates with calcium within a cellular vesicular compartment and is then secreted to the LL. Thus, much as in plants (that produce vesicular IP6 deposits), the existence of transporters for IP6 or its precursors in internal membranes is needed to explain the compound's cellular localisation in E. granulosus.

6-Phytase↗

PubMed2004

The Williams syndrome transcription factor interacts with PCNA to target chromatin remodelling by ISWI to replication foci.

Chromatin states have to be faithfully duplicated during DNA replication to maintain cell identity. It is unclear whether or how ATP-dependent chromatin-remodelling factors are involved in this process. Here we provide evidence that the Williams syndrome transcription factor (WSTF) is targeted to replication foci through direct interaction with the DNA clamp PCNA, an important coordinator of DNA and chromatin replication. WSTF, in turn, recruits imitation switch (ISWI)-type nucleosome-remodelling factor SNF2H to replication sites. These findings reveal a novel recruitment mechanism for ATP-dependent chromatin-remodelling factors that is fundamentally different from the previously documented targeting by sequence-specific transcriptional regulators. RNA-interference-mediated depletion of WSTF or SNF2H causes a compaction of newly replicated chromatin and increases the amount of heterochromatin markers, including HP1beta. This increase in the amount of HP1beta protein is mediated by progression through S phase and is not the result of an increase in HP1beta mRNA levels. We propose that the WSTF-ISWI complex has a role in the maintenance of chromatin structures during DNA replication.

Adenosine Triphosphatases↗

PubMed2004

[Estimate of the owned canine and feline populations in urban area in Brazil].

OBJECTIVE: Given the importance of assessing owned dog and cat populations to adequate planning and evaluation of rabies control measures in urban areas, it is proposed the use of an estimate of these populations based on human population parameters. METHODS: The ratio between human population and owned animal (dogs and cats) population was calculated in the municipality of Taboão da Serra, state of São Paulo, Brazil. This municipality was divided into two distinct social and economic homogeneous areas through k-mean algorithm, allowing for comparison between the ratios of the two homogeneous areas. RESULTS: A 5.14 ratio was calculated for human and dog populations and a 30.57 ratio for human and feline populations. A significant difference was not observed when comparing the ratios for human and animal populations of the two homogeneous areas. CONCLUSIONS: An estimate of owned dog and cat populations based on the human population is the better choice for assessing the animal population instead of an animal census because it can be easily implemented.

Animals↗

PubMed2004

Human topoisomerase IIalpha: targeting to subchromosomal sites of activity during interphase and mitosis.

Mammalian topoisomerase IIalpha (topo IIalpha) plays a vital role in the removal of topological complexities left on DNA during S phase. Here, we developed a new assay to selectively identify sites of catalytic activity of topo IIalpha with subcellular resolution. We show that topo IIalpha activity concentrates at replicating heterochromatin in late S in a replication-dependent manner and at centric heterochromatin during G2 and M phases. Inhibitor studies indicate that this cell cycle-dependent concentration over heterochromatin is sensitive to chromatin structure. We further show that catalytically active topo IIalpha concentrates along the longitudinal axis of mitotic chromosomes. Finally, we found that catalytically inert forms of the enzyme localize predominantly to splicing speckles in a dynamic manner and that this pool is differentially sensitive to changes in the activities of topo IIalpha itself and RNA polymerase II. Together, our data implicate several previously unsuspected activities in the partitioning of the enzyme between sites of activity and putative depots.

Antigens, Neoplasm↗

PubMed2003

Sialyl-Tn antigen expression and O-linked GalNAc-Thr synthesis by Trypanosoma cruzi.

Most Trypanosoma cruzi O-glycans are linked to Thr/Ser residues via N-acetylglucosamine. We report that the mucin-type carcinoma-associated sialyl-Tn antigen (NeuAc-GalNAc-O-Ser/Thr) is expressed by T. cruzi. A specific MAb allowed us to localize the antigen on the surface of epimastigotes and to identify reactive components in parasite lysates (32, 60, and 94kDa). In addition, ppGalNAc-T activity was characterized in epimastigotes, and direct evidence was obtained for the in vitro incorporation of GalNAc to a synthetic peptide derived from a T. cruzi mucin. These results add an as yet unknown complexity to the pathways of O-glycan biosynthesis in this protozoan parasite.

Acetylgalactosamine↗

PubMed2003

Preparative purification of soybean agglutinin by affinity chromatography and its immobilization for polysaccharide isolation.

Optimized procedures for the affinity purification of soybean agglutinin (SBA) from soybean flour, and its further immobilization, were developed. Lectin purification on galactosyl-Sepharose yielded 44.5+/-3.5 mg of pure SBA/50 g of flour. To prepare SBA adsorbents, the lectin was immobilized onto 1-cyano-4-(dimethylamino)pyridinium tetrafluoroborate (CDAP) activated Sepharose with high yields (77%). Feasibility of the use of this improved SBA adsorbent for affinity purification of Streptococcus pneumoniae capsular polysaccharides from strain 14 (CPS-14) at laboratory scale was demonstrated. Using SBA-Sepharose adsorbent (7.0 mg lectin per ml), amounts of 6.3 mg of pure CPS-14 per cycle were produced, the adsorbent being reused up to four times without loss of capacity.

Chromatography, Affinity↗

PubMed2003

Quantitative determination of pneumococcal capsular polysaccharide serotype 14 using a modification of phenol-sulfuric acid method.

The capsular polysaccharide of Streptococcus pneumoniae, serotype 14, is part of every pneumococcal vaccine presently in the market or under development. A strategy for the quantitative determination of this polysaccharide by the phenol-sulfuric acid method is described. The modality of acid addition is shown to be the critical step for obtaining reproducible test results between different technicians. Raising the incubation temperature above 80 degrees C increased the consistency of the method by more than 60% regardless of the acid addition modality, but at the expense of some loss of sensitivity. Incubation at 110 degrees C was found necessary to obtain reproducible results within 3% for this technique, which was used to follow the enrichment of the polysaccharide during the last steps of purification. A model mixture of the component polysaccharide sugars provided an adequate and economic standard to construct the calibration curve for this assay, with absorbance reading either in the reaction tubes or in a microplate. A similar procedure may be applied to the determination of other bacterial polysaccharides as well.

Bacterial Capsules↗

PubMed2003

Larval behavioral diapause regulates life cycle of Amblyomma cajennense (Acari: Ixodidae) in Southeast Brazil.

Previous studies have determined that Amblyomma cajennense (F.) produces one generation per year in Southeastern Brazil. Larvae predominate from April to July, nymphs from July to October, and adults from October to March. In the current study, we observed engorged females, their eggs, and respective larvae in the field. Engorged females were released in grass plots under natural conditions, at monthly intervals, from October 2000 to March 2001. Preoviposition periods lasted 1-2 wk, and mean egg incubation periods lasted 5-7 wk. Consequently, larvae hatched at different periods from early December 2000 to early April 2001. However, all hatched larvae stayed confined to the plot ground, under the vegetation, until late April or early May 2001, when some larvae were seen questing on top of vegetation for the first time. Thereafter, larvae were always seen on this questing position until August 2001, when all larvae had died on the plots. In addition, premolt period data of engorged larvae and nymphs held in laboratory at different temperatures were used to construct a premolt mathematical model, which was applied to reported seasonal dynamic data of A. cajennense in the field. The premolt model indicated that the field intervals between larvae and nymphs and between nymphs and adults are controlled solely by nondiapause events, such as ground temperature. We concluded that the 1-yr generation pattern of A. cajennense in Southeastern Brazil is primarily controlled by larval behavioral diapause, shown by larvae born during spring/summer months, before their activity period at next autumn.

Animals↗

PubMed2002

Seasonal dynamics of ticks (Acari: Ixodidae) on horses in the state of São Paulo, Brazil.

Natural tick infestations were assessed every 14 days on horses over a 2-year period. Amblyomma cajennense adult ticks were counted individually, without detachment from the horses. Larvae and nymphs of A. cajennense were collected using a rubber scraper that scratched engorged immature ticks from the host. Adult females of Anocentor nitens larger than 4mm length were counted on the horses. Blood samples were also obtained from the horses every 14 days and macroclimatic data were obtained for the study period. Infestations of A. cajennense demonstrated distinct peaks of activity for each of the three parasitic stages over each 12-month period, showing a 1-year generation pattern. Larvae predominated from April to July and nymphs from June to October. Adults predominated from October to March with a greater number of adult males than females. Although other studies on seasonal dynamics in the states of Rio de Janeiro and Minas Gerais were performed with the free-living stages of A. cajennense on pastures, the present study in the state of São Paulo, performed with the parasitic stages of A. cajennense on horses, showed similar results to those observed in other states. Infestations by A. nitens demonstrated distinct peaks of activity of adult females (>4 mm), suggesting different tick generations during the year. Infestation with A. nitens was much higher in the first year than the second year which may have been related to horse nutritional status and stocking rate. Although several climatic variables showed statistical significant correlation (r) with tick counts, the determination coefficients (R(2)) were always lower than 0.40, suggesting that any single significant variable (i.e. mean temperature) would not explain the tick distribution pattern over the year. The highest peaks of A. nitens females (>4 mm) were significantly associated with decrease in horse packed cell volumes (R(2)=0.603). The ears and the perineum, tail and groin region accounted for around 70% of all A. nitens females counted on the horses.

Animals↗

PubMed2002

myo-Inositol hexakisphosphate is a major component of an extracellular structure in the parasitic cestode Echinococcus granulosus.

myo-Inositol hexakisphosphate (IP(6)) is an abundant intracellular component of animal cells. In this study we describe the presence of extracellular IP(6) in the hydatid cyst wall (HCW) of the larval stage of the cestode parasite Echinococcus granulosus. The HCW comprises an inner cellular layer and an outer, acellular (laminated) layer up to 2 mm in thickness that protects the parasite from host immune cells. A compound, subsequently identified as IP(6), was detected in and purified from an HCW extract on the basis of its capacity to inhibit complement activation. The identification of the isolated compound was carried out by a combination of NMR, MS and TLC. The majority of IP(6) in the HCW was found in the acellular layer, with only a small fraction of the compound being extracted from cells. In the laminated layer, IP(6) was present in association with calcium, and accounted for up to 15% of the total dry mass of the HCW. IP(6) was not detected in any other structures or stages of the parasite. Our results imply that IP(6) is secreted by the larval stage of the parasite in a polarized fashion towards the interface with the host. This is the first report of the secretion of IP(6), and the possible implications beyond the biology of E. granulosus are discussed.

Animals↗