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Biomedical subjects

R López

Publications and source records attributed to R López.

At least 19 recordsLinked to original sources

Characterization of the galU gene of Streptococcus pneumoniae encoding a uridine diphosphoglucose pyrophosphorylase: a gene essential for capsular polysaccharide biosynthesis.

The galU gene of Streptococcus pneumoniae has been cloned and sequenced. Escherichia coli cells harboring the recombinant plasmid pMMG2 (galU) overproduced a protein that has been shown to correspond to a uridine 5'-triphosphate:glucose-1-phosphate uridylyltransferase (uridine diphosphoglucose [UDP-Glc] pyrophosphorylase) responsible for the synthesis of UDP-Glc, a key compound in the biosynthesis of polysaccharides. A gene very similar to the S. pneumoniae galU has been found in a partial nucleotide sequence of the Streptococcus pyogenes genome. Knockout galU mutants of type 1 pneumococci are unable to synthesize a detectable capsule. An identical result was found in type 3 S. pneumoniae cells in spite of the fact that these bacteria contain a type-specific gene (cap3C) that also encodes a UDP-Glc pyrophosphorylase. Since eukaryotic UDP-Glc pyrophosphorylases appear to be completely unrelated to their prokaryotic counterparts, we postulate that GalU may be an appropriate target for the search of new drugs to control the pathogenicity of bacteria like pneumococcus and S. pyogenes.

Amino Acid Sequence

Molecular structure of the gene cluster responsible for the synthesis of the polysaccharide capsule of Streptococcus pneumoniae type 33F.

The organization and nucleotide sequence of the capsular gene cluster involved in the biosynthesis of the type 33F capsular polysaccharide of Streptococcus pneumoniae have been determined. The complete type 33F operon (cap33f) is composed of 14 potential open reading frames where the last ten genes are group-specific. Putative functions have been assigned to several gene products by sequence comparison with the proteins included in the databases. A functional promoter located immediately upstream from the first gene of the cap33f gene cluster has been demonstrated. A 20 kb DNA fragment containing the cap33f genes and the operon promoter was sufficient to transform a S. pneumoniae type 3 unencapsulated mutant to the type 33F capsule.

Bacterial Capsules

Molecular characterization of PcpA: a novel choline-binding protein of Streptococcus pneumoniae.

The gene pcpA that encodes a novel pneumococcal choline-binding protein has been cloned and characterized. Northern blot analysis revealed that pcpA is expressed during the exponential phase of growth of pneumococci as a monocistronic transcript of about 2.3 kb. The transcription start site has been located 132 bp upstream of the start codon and the proposed -35 and -10 boxes that are highly similar to those of the typical sigma 70 promoters from Escherichia coli. This gene encodes a putative 79 kDa protein that contains a typical C-terminal choline-binding domain (ChBD). The ChBD of PcpA is built up by 11 identical motifs of 20 amino acids plus a tail of 19 amino acids, which represents the longest ChBD that has been characterized so far. Interestingly, two tandem arrays of five characteristic amphipathic leucine reach repeats (LRRs) of 22-26 amino acids in length have been found in the N-terminal region of PcpA. Since LRRs have been proposed to be involved in protein-protein and protein-lipid interactions our findings suggests a role for PcpA in pneumococcal adhesion.

Amino Acid Sequence

[Epidemiologic situation of tuberculosis under the influence of HIV in the health area of Ibiza and Formentera (1987-1993)].

OBJECTIVES: To analyse the epidemiology of Tuberculosis and determine the characteristics of patients infected by HIV vs those not infected. DESIGN: A descriptive-retrospective study. SETTING: Ibiza and Formentera Health Area. PATIENTS: 268 patients over 16 diagnosed with TB between 7/1/1987 and 12/31/1993. MEASUREMENTS AND MAIN RESULTS: The highest accumulated annual incidence was detected in 1989 (82.4/100,000 inhabitants), but had decreased considerably by 1993 (42.8/100,000). The proportion of HIV+ cases increased to 37.0% of the total in 1993. 75% of the total were male and 57.5% between 20 and 39 (x = 38.1; SD = 15.4). 63.1% had pulmonary TB, 60.8% producing bacilli, 19.8% were HIV+, of which 47.2% were intravenous drug users and 20.8% homosexuals. There was a 75.4% cure rate, 4.9% who left treatment and 16.1% lost during follow-up. CONCLUSIONS: During the study period a progressive increase in the proportion of cases with HIV infection was observed. Control and follow-up of Tuberculosis was more difficult in these patients.

AIDS-Related Opportunistic Infections

Evidence for horizontal transfer from Streptococcus to Escherichia coli of the kfiD gene encoding the K5-specific UDP-glucose dehydrogenase.

Capsular polysaccharides are important virulence factors both in Gram-positive and Gram-negative bacteria. A similar cluster organization of the genes involved in the synthesis of bacterial exopolysaccharides has been postulated in both cases, suggesting that these clusters evolved by module assembly. Horizontal gene transfer has been postulated to explain the polymorphism found in these cellular polymers. The cap1 K and cap3A genes coding for the pneumococcal type 1 and type 3 UDP-glucose dehydrogenases, respectively, have been compared with other UDP-sugar dehydrogenases. We have observed that the evolutionary distance between Cap1K and Cap3A is approximately equal to that found between Cap1K (or Cap3A) and other UDP-GlcDH of families evolutionarily distant like KfiD, the dehydrogenase from Escherichia coli K5. On the basis of comparisons of G + C content, patterns of synonymous and nonsynonymous substitutions, dinucleotide frequencies, and codon usage bias, we conclude that the kfiD gene has been introduced into E. coli from an exogenous source, probably from a streptococcal species.

Bacterial Capsules

Endogenous cholecystokinin enhances postprandial gastroesophageal reflux in humans through extrasphincteric receptors.

BACKGROUND & AIMS: Exogenous cholecystokinin (CCK) decreases lower esophageal sphincter (LES) pressure and increases transient LES relaxations (TLESRs) in humans. The aims of this study were to determine whether endogenous CCK increases gastroesophageal reflux in humans and whether this is a direct effect on the LES. METHODS: Esophageal pH, LES pressure, and gallbladder volume were measured in 8 healthy volunteers after ingestion of a 181-kcal meal alone or adding 12 g cholestyramine to increase endogenous CCK release. In 7 additional volunteers, the effect of cholestyramine was studied during intravenous perfusion of saline or the CCK-A receptor antagonist loxiglumide. In circular LES strips from 9 transplant donors, we measured the effect of CCK-8 (10(-11) to 3 x 10(-8) mol/L) on basal tension and on electrical field-induced relaxation. RESULTS: Cholestyramine increased gallbladder emptying, reflux episodes, TLESRs, and time of esophageal pH of <4. Loxiglumide inhibited postprandial gallbladder emptying, reflux episodes, TLESRs, and time of pH of <4 and prevented the decrease in LES pressure induced by cholestyramine. In vitro, CCK-8 contracted LES strips through a tetrodotoxin-insensitive pathway but did not modify electrical field-induced LES relaxations. CONCLUSIONS: Endogenous CCK enhances postprandial gastroesophageal reflux in humans by increasing the rate of TLESRs and reduces postprandial LES pressure. These actions seem mediated by extrasphincteric CCK-A receptors that override a direct contractile effect of CCK on the LES muscle.

Adult

Levels of IgM and IgA circulating immune complexes in dogs with leishmaniasis.

The concentrations of IgM and IgA circulating immune complexes (CIC) were determined in 82 dogs with naturally acquired leishmania infection and in a control group of 25 healthy dogs. The mean serum IgM and IgA CIC concentration in infected dogs were significantly (IgM; P < 0.004; IgA; P < 0.000) higher than in the control group. An increase in IgM and IgA CIC concentration was found in 47.6 and 95.1%, respectively, of the leishmania-infected dogs. The serum IgM and IgA CIC concentrations in infected dogs showing hypercreatininaemia were not statistically higher than those of sick dogs with normal creatininaemia. When hypercreatininaemia (> or = 1.30 mg/dl) was used as an indicator for CIC disease, the positive predictive value obtained with 0.368 for IgM CIC and 0.894 for IgA CIC, indicating that renal function impairment was associated with the high serum IgM and IgA CIC concentrations in 36.8 and 89.4%, respectively, of the infected dogs.

Animals

Use of allopurinol for maintenance of remission in dogs with leishmaniasis.

Current treatments for infected dogs with leishmaniasis do not always provide long-term control of the disease and clinical relapses are common. In this study, the usefulness of long-term allopurinol administration in the maintenance of clinical remission in canine leishmaniasis was evaluated. Fifteen dogs with natural leishmania infection were subjected to an initial treatment based on the simultaneous administration of meglumine antimoniate (100 mg/kg/day) and allopurinol (30 mg/kg/day). Once clinical remission was achieved, a maintenance treatment with allopurinol (20 mg/kg/day) administered for one week a month was instituted. Results were compared with those of a retrospective control group comprising 15 infected dogs which only followed the induction treatment. Relapses occurred in 86 per cent of control dogs within 14 months of discontinuing treatment. In contrast, those dogs on intermittent oral allopurinol administration were successfully maintained in clinical remission for a follow-up period of 10 to 44 months. In this latter group, specific antibody titres decreased or were unchanged, no side effects directly attributable to allopurinol were seen and treatment was well accepted by the owners. It is concluded that long-term intermittent administration of allopurinol is an effective way of maintaining clinical remission in dogs with leishmaniasis.

Allopurinol

Functional analysis of the two-gene lysis system of the pneumococcal phage Cp-1 in homologous and heterologous host cells.

The two lysis genes cph1 and cpl1 of the Streptococcus pneumoniae bacteriophage Cp-1 coding for holin and lysozyme, respectively, have been cloned and expressed in Escherichia coli. Synthesis of the Cph1 holin resulted in bacterial cell death but not lysis. The cph1 gene was able to complement a lambda Sam mutation in the nonsuppressing E. coli HB101 strain to produce phage progeny, suggesting that the holins encoded by both phage genes have analogous functions and that the pneumococcal holin induces a nonspecific lesion in the cytoplasmic membrane. Concomitant expression of both holin and lysin of Cp-1 in E. coli resulted in cell lysis, apparently due to the ability of the Cpl1 lysozyme to hydrolyze the peptidoglycan layer of this bacterium. The functional analysis of the cph1 and cpl1 genes cloned in a pneumococcal mutant with a complete deletion of the lytA gene, which codes for the S. pneumoniae main autolysin, provided the first direct evidence that, in this gram-positive-bacterium system, the Cpl1 endolysin is released to its murein substrate through the activity of the Cph1 holin. Demonstration of holin function was achieved by proving the release of pneumolysin to the periplasmic fraction, which strongly suggested that the holin produces a lesion in the pneumococcal membrane.

Amino Acid Sequence

Characterization of IS1515, a functional insertion sequence in Streptococcus pneumoniae.

We describe the characterization of a new insertion sequence, IS1515, identified in the genome of Streptococcus pneumoniae I41R, an unencapsulated mutant isolated many years ago (R. Austrian, H. P. Bernheimer, E. E. B. Smith, and G. T. Mills, J. Exp. Med. 110:585-602, 1959). A copy of this element located in the cap1EI41R gene was sequenced. The 871-bp-long IS1515 element possesses 12-bp perfect inverted repeats and generates a 3-bp target duplication upon insertion. The IS encodes a protein of 271 amino acid residues similar to the putative transposases of other insertion sequences, namely IS1381 from S. pneumoniae, ISL2 from Lactobacillus helveticus, IS702 from the cyanobacterium Calothrix sp. strain PCC 7601, and IS112 from Streptomyces albus G. IS1515 appears to be present in the genome of most type 1 pneumococci in a maximum of 13 copies, although it has also been found in the chromosome of pneumococcal isolates belonging to other serotypes. We have found that the unencapsulated phenotype of strain 141R is the result of both the presence of an IS1515 copy and a frameshift mutation in the cap1EI41R gene. Precise excision of the IS was observed in the type 1 encapsulated transformants isolated in experiments designed to repair the frameshift. These results reveal that IS1515 behaves quite differently from other previously described pneumococcal insertion sequences. Several copies of IS1515 were also able to excise and move to another locations in the chromosome of S. pneumoniae. To our knowledge, this is the first report of a functional IS in pneumococcus.

Amino Acid Sequence

Pneumococcal bacteriophage Cp-1 encodes its own protease essential for phage maturation.

The major capsid protein of the pneumococcal phage Cp-1 that accounts for 90% of the total protein found in the purified virions is synthesized by posttranslational processing of the product of the open reading frame (ORF) orf9. Cloning of different ORFs of the Cp-1 genome in Escherichia coli and Streptococcus pneumoniae combined with Western blot analysis of the expressed products led to the conclusion that the product of orf13 is an endoprotease that cleaves off the first 48 amino acid residues of the major head protein. This protease appears to be a key enzyme in the morphopoietic pathway of the Cp-1 phage head. To our knowledge, this is the first case of a bacteriophage infecting gram-positive bacteria that encodes a protease involved in phage maturation.

DNA-Binding Proteins

Effect of discontinuing prophylaxis with norfloxacin in patients with hematologic malignancies and severe neutropenia. A matched case-control study of the effect on infectious morbidity.

The use of fluorinated quinolones for prophylaxis of infections in neutropenic cancer patients has led to a reduction of infections with gram-negative enteric bacilli, but there is concern about the emergence of antibiotic-resistant entero-bacterial infections and a rise of gram-positive bacteremias. Due to these concerns, in mid-1995 the use of prophylactic norfloxacin was discontinued in our unit. In order to evaluate the impact of this measure on the infectious morbidity in our unit, 91 severe neutropenic episodes in 58 patients with hematologic malignancies who did not receive norfloxacin prophylaxis (NO group) were closely matched to 91 episodes in 60 patients who received norfloxacin prophylaxis (NORFLO group). There were no differences in the incidence of febrile neutropenia, fever of unknown origin or bacteremia during the first febrile episode. There was a trend for a higher rate of coagulase-negative staphylococcal bacteremia in the NORFLO group (5 vs. 11 cases in the NO and NORFLO groups, respectively, p = NS). Enterobacterial bloodstream infections were more frequent in the NO group (13 vs. 2 cases, respectively, p = 0.01), especially Escherichia coli (9 vs. 1 case, respectively, p = 0.01). Twelve of 13 enterobacterial isolates in the NO group were sensitive to the fluoroquinolones vs. 0/2 in the NORFLO group (p = 0.07). We conclude that the abrupt discontinuation of norfloxacin prophylaxis in our ward led to a rapid increase in the rate of fluoroquinolone-susceptible enterobacterial infections, with a scarce impact on infectious morbidity. This suggests that the selection of resistant flora in an inpatient ward by prophylactic antimicrobials may be reversible following the discontinuation of the prophylactic agent(s).

Adolescent

[Iron deficiency and pica].

PURPOSE: To study the relationship between pica and iron-lack anaemia in a series of iron-deficiency patients in order to establish the pathogenesis of such relationship. PATIENTS AND METHODS: Four-hundred and thirty-three patients were analysed. Pica was studied by introducing certain diet queries into the clinical history. All patients received oral iron and were periodically controlled with the usual clinico-haematological procedures. RESULTS: Pica was present in 23 patients (5.3%). Eight nourishing (namely, coffee grains, almonds, chocolate, ice, lettuce, carrots, sunflower seeds and bread) and 2 non-nourishing (clay and paper) substances were involved. A second episode of pica appeared in 9 cases upon relapsing of iron deficiency. Both anaemia and pica were cured by etiologic and substitutive therapy in all instances. No clear correlation was found with either socio-economic status or pathogenetic causes of iron deficiency and pica, and no haematological differences were seen between patients with pica and those without this alteration. CONCLUSIONS: (1) The pathogenesis of pica is unclear, although it appears unrelated to the degree of iron deficiency. (2) According to the findings in this series, pica seems a consequence of iron deficiency rather than its cause. (3) Adequate therapy can cure both conditions, although pica may reappear upon relapse of iron deficiency.

Adolescent

[The use of anorectal manometry and dynamic proctography in patients for diagnosis of solitary rectal ulcer syndrome].

BACKGROUND: Solitary rectal ulcer syndrome is caused by rectal prolapse and/or anismus. The diagnosis by physical examination, or using endoscopy or radiology alone is difficult. AIM: To evaluate the diagnostic utility of videoproctography and anorectal manometry in solitary rectal ulcer syndrome. PATIENTS AND METHODS: We studied six patients (5 women/1 man; 21-66 years) with rectal lesions suggestive of solitary rectal ulcer. By videoproctography, we measured the changes in the anorectal angle and the shape of the rectal wall during voiding. By manometry, we measured the resting pressure of the anal canal, and the pressure during maximal voluntary contraction and during straining. RESULTS: Proctography showed a rectal prolapse in 5 patients (non-exteriorized in 4). Both manometry and proctography demonstrated the normal relaxation of puborectalis and external sphincter during straining in 4 patients, manometry found sphincteric weakness in two of these patients. Both manometry and proctography evidenced anismus in a patient with rectal prolapse. CONCLUSION: The high prevalence of rectal prolapse in our patients allows us to recommend proctography in patients with morphological lesions suggestive of solitary rectal ulcer. The manometric examination identifies patients at risk of developing incontinence. Both techniques contribute to ascertain the presence of anismus and are indispensable in the selection of the adequate treatment for patients with solitary rectal ulcer.

Adult

[Cardiorespiratory failure and diabetic autonomic neuropathy: a case report].

Advanced autonomic neuropathy in diabetic patients is associated with homeostatic cardiovascular response. Sympathetic compensation diminishes and the organism becomes more susceptible to the depressant effect of certain drugs. The incidence of undesirable cardiovascular effects (bradycardia, hypotension and sudden heart failure) during anesthesia is higher in such patients. We report the case of a woman with diabetes who suffered two episodes of sudden heart failure after brachial plexus block through the axilla, followed one year later by sudden heart failure during general anesthesia for kidney transplantation. The possible contributing mechanisms are described.

Adult