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Biomedical subjects

A Hurtado

Publications and source records attributed to A Hurtado.

At least 37 records · Page 2Linked to original sources
PubMed2001

Mutations in genes involved in the flagellar export apparatus of the solvent-tolerant Pseudomonas putida DOT-T1E strain impair motility and lead to hypersensitivity to toluene shocks.

Pseudomonas putida DOT-T1E is a solvent-tolerant strain able to grow in the presence of 1% (vol/vol) toluene in the culture medium. Random mutagenesis with mini-Tn5-'phoA-Km allowed us to isolate a mutant strain (DOT-T1E-42) that formed blue colonies on Luria-Bertani medium supplemented with 5-bromo-4-chloro-3-indolylphosphate and that, in contrast to the wild-type strain, was unable to tolerate toluene shocks (0.3%, vol/vol). The mutant strain exhibited patterns of tolerance or sensitivity to a number of antibiotics, detergents, and chelating agents similar to those of the wild-type strain. The mutation in this strain therefore seemed to specifically affect toluene tolerance. Cloning and sequencing of the mutation revealed that the mini-Tn5-'phoA-Km was inserted within the fliP gene, which is part of the fliLMNOPQRflhBA cluster, a set of genes that encode flagellar structure components. FliP is involved in the export of flagellar proteins, and in fact, the P. putida fliP mutant was nonmotile. The finding that, after replacing the mutant allele with the wild-type one, the strain recovered the wild-type pattern of toluene tolerance and motility unequivocally assigned FliP a function in solvent resistance. An flhB knockout mutant, another gene component of the flagellar export apparatus, was also nonmotile and hypersensitive to toluene. In contrast, a nonpolar mutation at the fliL gene, which encodes a cytoplasmic membrane protein associated with the flagellar basal body, yielded a nonmotile yet toluene-resistant strain. The results are discussed regarding a possible role of the flagellar export apparatus in the transport of one or more proteins necessary for toluene tolerance in P. putida DOT-T1E to the periplasm.

Alkaline Phosphatase↗
PubMed2000

Maternal stress responses and anxiety during pregnancy: effects on fetal heart rate.

This study examined the effect of an acute maternal stress response and anxiety on fetal heart rate. Seventeen healthy, 3rd-trimester pregnant women (mean age = 26 +/- 6 years) were instrumented for continuous electrocardiography, blood pressure (BP), respiration, and fetal heart rate (HR). Subjects completed the state anxiety subscale of the State Trait Personality Inventory (STPI), then rested quietly in a semirecumbent position for a 5-min baseline period, followed by either a 5-min arithmetic or Stroop color-word task. Over the entire 5-min stress period and when averaged across all subjects, the stressors led to significant increases in maternal systolic BP and respiratory rate but changes in maternal HR, diastolic BP, and fetal HR were not significant. However, when subjects were dichotomized into groups that had above or below average anxiety scores [ANX(+) and ANX(-)], both groups had similar respiration rate increases to the stressors, but the BP and fetal heart rate (FHR) responses were significantly different. Women in the ANX(-) group had significantly greater BP responses compared to women in the ANX(+) group whereas the fetuses of ANX(+) women showed significant HR increases and the fetuses of ANX(-) women exhibited nonsignificant decreases. These findings suggest that women's acute emotional reactivity during pregnancy can influence fetal HR patterns and that a stress-induced increase in maternal BP is not the primary signal by which a women's stress response is transduced to her fetus. The results are consistent with the hypothesis that maternal psychological variables may shape the neurobehavioral development of the fetus.

Adult↗
PubMed2000

Distinct patterns of glomerular disease in Lima, Peru.

AIM: We performed both a retrospective and prospective study to elucidate the types of glomerular diseases present in adults in Lima, Peru. MATERIAL AND METHODS: In the retrospective study, we analyzed 1,263 renal biopsies over a 10-year period (1985 -1995) that were processed at a central reference renal pathology laboratory in Lima. 101 cases were examined in the prospective study. RESULTS: The most common glomerular diseases observed were those due to systemic lupus erythematosus (30.2%), membranoproliferative glomerulonephritis (MPGN, 14.8%), and focal and segmental glomerulosclerosis (FSGS, 13.9%). Although mesangial-proliferative nephritis was observed in 9.5% of cases, IgA nephropathy was rare (0.9%). Examination of the year to year frequency showed that MPGN has tended to decrease in frequency with time whereas FSGS has been increasing. Although there is known to be a high frequency of infections in Peru, only 4.2% of the cases in the retrospective study were associated with infection. Furthermore, in the prospective study, only one case of hepatitis C and no cases of hepatitis B viral infection were detected, including in the 11 cases of MPGN observed. CONCLUSION: We conclude that the epidemiology of glomerular disease in Lima, Peru, is distinct from most areas of the world, but has similarities to certain regions in Africa, in that MPGN is common whereas IgA nephropathy appears to be rare. Further studies are necessary to elucidate the reasons why the patterns of glomerular disease are different from that observed in other parts of the world.

Adult↗
PubMed1999

Minor displacements in the insertion site provoke major differences in the induction of antibody responses by chimeric parvovirus-like particles.

An antigen-delivery system based on hybrid virus-like particles (VLPs) formed by the self-assembly of the capsid VP2 protein of canine parvovirus (CPV) and expressing foreign peptides was investigated. In this report, we have studied the effects of inserting the poliovirus C3:B epitope in the four loops and the C terminus of the CPV VP2 on the particle structure and immunogenicity. Epitope insertions in the four loops allowed the recovery of capsids in all of the mutants. However, only insertions of the C3:B epitope in VP2 residue 225 of the loop 2 were able to elicit a significant anti-peptide antibody response, but not poliovirus-neutralizing antibodies, probably because residue 225 is located in an small depression of the surface. To fine modulate the insertion site in loop 2, a cassette-mutagenesis was carried out to insert the epitope in adjacent positions 226, 227, and 228. The epitope C3:B inserted into these positions was well recognized by the specific monoclonal antibody C3 by immunoelectron microscopy. BALB/c mice immunized with these chimeric C3:B CPV:VLPs were able to elicit an strong neutralizing antibody response (>3 log(10) units) against poliovirus type 1 (Mahoney strain). Therefore, minor displacements in the insertion place cause dramatic changes in the accessibility of the epitope and the induction of antibody responses.

Amino Acid Sequence↗
PubMed1999

Parvovirus-like particles as vaccine vectors.

A wide array of systems have been developed to improve "classic" vaccines. The use of small polypeptides able to elicit potent antibody and cytotoxic responses seems to have enormous potential in the design of safer vaccines. While peptide coupling to large soluble proteins such as keyhole limpet hemocyanin is the current method of choice for eliciting antibody responses and insertion in live viruses for cytotoxic T-lymphocyte responses, alternative cheaper and/or safer methods will clearly be required in the future. Virus-like particles constitute very immunogenic molecules that allow for covalent coupling of the epitopes of interest in a simple way. In this article, we detail the methodology employed for the preparation of efficient virus vectors as delivery systems. We used parvovirus as the model for the design of new vaccine vectors. Recently parvovirus-like particles have been engineered to express foreign polypeptides in certain positions, resulting in the production of large quantities of highly immunogenic peptides, and to induce strong antibody, helper-T-cell, and cytotoxic T-lymphocyte responses. We discuss the different alternatives and the necessary steps to carry out this process, placing special emphasis on the flow of decisions that need to be made during the project.

Animals↗
PubMed1999

Engineering parvovirus-like particles for the induction of B-cell, CD4(+) and CTL responses.

An antigen delivery system based on hybrid recombinant parvovirus-like particles (VLPs) formed by the self-assembly of the capsid VP2 protein of porcine (PPV) or canine parvovirus (CPV) expressed in insect cells with the baculovirus system has been developed. PPV:VLPs containing a CD8(+) epitope from the LCMV nucleoprotein evoked a potent CTL response and were able to protect mice against a lethal infection with the virus. Also, PPV:VLPs containing the C3:T epitope from poliovirus elicited a CD4(+)3 log(10) units) against poliovirus. The possibility of combining different types of epitopes in different positions of a single particle to stimulate different branches of the immune system paves the way to the production of more potent vaccines in a simple and cheap way.

Animals↗
PubMed1999

Cardiorespiratory responses to bidirectional tilts in infants.

Prior research in newborns has shown that head-up and head-down tilting elicits sustained increases and decreases in heart rate, respectively. Other studies in older infants have suggested that the pattern of heart rate responses to head-up tilting varies with risk for sudden infant death syndrome (SIDS). In this study, heart and respiratory rate changes following bidirectional tilting were recorded in sleeping infants on Day 1 or 2 of life, and during the period of maximum risk for SIDS, at 2 and 4 months of age. Newborns show increases in heart rate following 30 degrees head-up tilts and decreases in heart rate to 300 head-down tilting. Respiratory rates decreased to head-up tilting but did not change significantly to head-down tilting. While respiratory rate changes at 2 and 4 months of age are comparable to those of newborns, and decreases in heart rate to head-down tilting are similar across ages, sustained elevations in heart rate following head-up tilting are no longer apparent at the older ages. These results are consistent with the hypothesis that, during the period of maximum risk for SIDS, infants may have reduced ability to compensate for challenges that lead to decreases in blood pressure.

Age Factors↗
PubMed1999

Campylobacter coli strains with enlarged flagellin genes isolated from river water.

A group of campylobacters isolated from river water were found to possess unusually large flagellin genes. Both phenotype and serology were consistent with identification as Campylobacter coli. Phylogenetic analysis of small (16S, rrs) and large subunit (23S, rrl) rRNA genes of a representative strain, NCTC 13006, demonstrated high levels of relatedness with C. jejuni and C. coli (99.1 and 98.3% similarity for 16S; 99.3 and 99.4% similarity for 23S). Large flagellin proteins were demonstrated by SDS-PAGE analysis. The flaA and flaB genes were sequenced and aligned with known campylobacter flagellin amino acid sequences. The encoded FlaA protein of the new group exhibited a high degree of divergence from other Campylobacter species. Within the central variable region of FlaA, a further hypervariable domain was identified containing characteristic repeated motifs. Separate pairwise alignments performed for the variable regions of the polypeptide indicated these large fla genes were more closely related to those of C. upsaliensis than to those of C. coli or C. jejuni.

Amino Acid Sequence↗
PubMed1999

Accessory DNA in the genomes of representatives of the Escherichia coli reference collection.

Different strains of the Escherichia coli reference collection (ECOR) differ widely in chromosomal size. To analyze the nature of the differential gene pool carried by different strains, we have followed an approach in which random amplified polymorphic DNA (RAPD) was used to generate several PCR fragments. Those present in some but not all the strains were screened by hybridization to assess their distribution throughout the ECOR collection. Thirteen fragments with various degrees of occurrence were sequenced. Three of them corresponded to RAPD markers of widespread distribution. Of these, two were housekeeping genes shown by hybridization to be present in all the E. coli strains and in Salmonella enterica LT2; the third fragment contained a paralogous copy of dnaK with widespread, but not global, distribution. The other 10 RAPD markers were found in only a few strains. However, hybridization results demonstrated that four of them were actually present in a large selection of the ECOR collection (between 42 and 97% of the strains); three of these fragments contained open reading frames associated with phages or plasmids known in E. coli K-12. The remaining six fragments were present in only between one and four strains; of these, four fragments showed no similarity to any sequence in the databases, and the other two had low but significant similarity to a protein involved in the Klebsiella capsule synthesis and to RNA helicases of archaeal genomes, respectively. Their percent GC, dinucleotide content, and codon adaptation index suggested an exogenous origin by horizontal transfer. These results can be interpreted as reflecting the presence of a large pool of strain-specific genes, whose origin could be outside the species boundaries.

Bacteriology↗
PubMed1999

Clinicopathologic correlations in lupus nephritis in Lima, Peru.

BACKGROUND: We assessed whether immunohistologic markers for glomerular or tubulointerstitial injury might provide better correlations with ongoing renal function and disease activity as compared with the WHO classification or the NIH activity and chronicity indices in lupus nephritis. METHODS: Thirty-three patients with clinically defined systemic lupus underwent renal biopsy over a 1-year period at Hospital Loayza in Lima, Peru. Biopsy specimens were evaluated for macrophages, proliferating cells, alpha-actin expression, and type IV collagen deposition in both glomeruli and the tubulointerstitium and the results compared with the current WHO and NIH classifications in relation to the clinical presentation. RESULTS: Patients with WHO class IV lupus nephritis were more likely to have lower serum complements, greater proteinuria and hematuria, and worse renal function. An elevated NIH activity index correlated with microhematuria, proteinuria, and impaired renal function, whereas an elevated chronicity index correlated with renal function, hypertension, and microhematuria, but not with proteinuria. The presence of glomerular macrophages correlated with both glomerular alpha-actin expression and type IV collagen deposition, but did not correlate with renal function or proteinuria. In contrast, interstitial macrophages correlated not only with interstitial collagen deposition and myofibroblast accumulation, but also correlated with both renal function and the presence of nephrotic syndrome. CONCLUSIONS: Both the WHO classification and the NIH activity/chronicity indices correlate with clinical manifestations of lupus nephritis. While glomerular macrophage accumulation correlates with mesangial cell activation (alpha-actin expression) and collagen deposition, and interstitial macrophage accumulation correlates with interstitial fibroblast activation and collagen deposition, only interstitial macrophages correlate with renal function. Of particular interest will be future studies to determine whether these markers correlate with the prognosis.

Actins↗
PubMed1997

Sequence similarities between large subunit ribosomal RNA gene intervening sequences from different Helicobacter species.

When the 23S rRNA genes from several Helicobacter species were amplified by PCR and compared with similar amplicons derived from H. pylori, they were seen to be enlarged in size. Sequencing of these enlarged genes from H. mustelae, H. canis (two strains) and H. muridarum identified insertions of novel sequence (intervening sequences, IVSs) sized between 93 and 377 bp located at nt 545, in place of an 8-nt sequence in the conventionally sized H. pylori gene. These IVSs were not present elsewhere in the genome. All strains with such IVSs lacked intact 23S rRNA which was replaced by two fragment whose sizes were consistent with cleavage at either side of the particular IVS. The predicted secondary structures of the four IVSs were characterised by base pairing at the 5' and 3' ends to form a stem. The four IVSs exhibited significant sequence inter-relationships. Further relationships were also observed between them and similar elements in both small and large subunit rRNA genes of other Helicobacter and Campylobacter species. Alignment of each IVS with the other such elements identified blocks of related sequence consistent with insertion/deletion events, indicating possible evolutionary relationships.

Base Sequence↗
PubMed1997

A molecular scheme based on 23S rRNA gene polymorphisms for rapid identification of Campylobacter and Arcobacter species.

23S rRNA gene PCR amplicons from 118 strains representing 15 species of Campylobacter and four species of Arcobacter were consecutively digested with HpaII, CfoI, and HinfI. A reproducible and discriminatory identification scheme based on combined restriction patterns was developed by using reference strains and was applied to the identification of a variety of isolates.

Campylobacter↗
PubMed1997

[Prevalence of neurocysticercosis in individuals affected by epilepsy].

INTRODUCTION: In this investigations, was carried out a neurocysticercosis (NC) prevalence study during seven months in the Instituto Neurológico de Antioquia with the purpose of known neurocysticercosis frequency as cause of epilepsy in patients older than ten years that we attended in our institute. MATERIAL AND METHODS: Computerized tomographies (CT) were made to 503 patients, with epilepsy, 24.7% of them were CT positive for NC. Cysticercosis enzyme linked immunoelectrotransfer blot (EITB) and enzyme linked immunoabsorbent assay (ELISA) test were made to 178 patients, 19.6% were EITB positive for NC and 5% ELISA positive for NC. Results. From this result it is possible to infer that about 8% of the 503 patients with epilepsy had cysticercosis, according to EITB that is the golden assay for NC. The CT and ELISA test had 94.3% and 27.7% sensitivity, respectively, according to EITB. The specificity of the CT for NC was 49.2% and specificity for ELISA test was 100% as compared to EITB. The multivariate analysis with logistic regression allowed to establish association of positive EITB with factors such as male sex, eating pork, headaches and multiple lesions in CT. CONCLUSIONS: Prevention and education actions are necessary for the interruption of the neurocysticercosis transmission chain in order to diminish the high prevalence of epilepsy in the country and its complication and consequences.

Adult↗
PubMed1996

Identification of domains in canine parvovirus VP2 essential for the assembly of virus-like particles.

Canine parvovirus capsids are composed of 60 copies of VP2 and 6 to 10 copies of VPl. To locate essential sites of interaction between VP2 monomers, we have analyzed the effects of a number of VP2 deletion mutants representing the amino terminus and the four major loops of the surface, using as an assay the formation of virus-like particles (VLPs) expressed by recombinant baculoviruses. For the amino terminus we constructed three mutants with progressively larger deletions, i.e., 9, 14, and 24 amino acids. Deletions of 9 and 14 amino acids did not affect the morphology and assembly capabilities of the mutants. However, the mutant with the 24-amino-acid deletion did not show hemagglutination properties or correct VLP morphology, stressing again the relevance of the RNER domain in canine parvovirus functionality. Three of the four mutants with deletions in the loops failed to make correct VLPs, indicating that these regions are essential for correct capsid assembly and morphology. Only the mutant with the deletion in loop 2 was able to assemble in regular VLPs, suggesting that this loop has little or no effect in capsid morphogenesis. Further research has demonstrated that this region can tolerate the insertion of foreign epitopes that are correctly exposed in the surface of the capsid. This result opens the door to the use of these VLPs for antigen delivery.

Amino Acid Sequence↗
PubMed1994

Genetic diversity of the Helicobacter pylori haemagglutinin/protease (hap) gene.

Helicobacter pylori has been associated with the etiology of gastritis, gastric and duodenal ulcers and gastric cancer in man. In this study we confirm the presence of the haemagglutinin/protease (hap) gene in ten geographically diverse strains of H. pylori, but the Southern analysis of genomic DNA suggested that the location of the hap gene was different in each strain. We suggest that the hap gene is subject to a high rate of mutation and subsequent polymorphisms by virtue of a mobile, genetic element located upstream of the hap gene in the H. pylori genome. Simple transformation experiments suggested that this polymorphism may be mediated by a transposon-like element.

Blotting, Southern↗
PubMed1994

Flagellin gene profiling of Helicobacter pylori infecting symptomatic and asymptomatic individuals.

Diversity within and around the flagellin (fla) A gene of Helicobacter pylori was studied by polymerase chain reaction/restriction fragment length polymorphism (PCR/RFLP) analysis and genomic Southern blot hybridization profiling. Four distinct pattern types were identified by DdeI restriction analysis of the 1.5-kb flaA amplicon of 55 strains. Most strains (73%) had the same flaA RFLP type, but subtypic variation was evident in some strains. No consistent associations were observed for selected strain subsets between the DdeI flaA profiles and phenotype (motility and cytotoxicity), urease gene profile or patient symptomatology. A subset of seven (F-1 profile) and four (F-2 profile) strains with identical HindIII digest patterns provided further evidence that the flaA gene was relatively highly conserved within H. pylori. By contrast, the flaA gene blot hybridization profiles were more diverse and consistent with greater variation at restriction sites in adjacent regions of the genome. We conclude that analyses of polymorphisms within the flaA gene provide limited discrimination between strains of H. pylori. The flaA genomic blot profiles offer greater potential for molecular typing purposes, although no associations with other pathogenicity factors or disease symptoms could be deduced.

Blotting, Southern↗