FbpA--a bacterial transferrin with more to offer.

BACKGROUND: Gram negative bacteria require iron for growth and virulence. It has been shown that certain pathogenic bacteria such as Neisseria gonorrhoeae possess a periplasmic protein called ferric binding protein (FbpA), which is a node in the transport of iron from the cell exterior to the cytosol. SCOPE OF REVIEW: The relevant literature is reviewed which establishes the molecular mechanism of FbpA mediated iron transport across the periplasm to the inner membrane. MAJOR CONCLUSIONS: Here we establish that FbpA may be considered a bacterial transferrin on structural and functional grounds. Data are presented which suggest a continuum whereby FbpA may be considered as a naked iron carrier, as well as a Fe-chelate carrier, and finally a member of the larger family of periplasmic binding proteins. GENERAL SIGNIFICANCE: An investigation of the molecular mechanisms of action of FbpA as a member of the transferrin super family enhances our understanding of bacterial mechanisms for acquisition of the essential nutrient iron, as well as the modes of action of human transferrin, and may provide approaches to the control of pathogenic diseases. This article is part of a Special Issue entitled Transferrins: Molecular mechanisms of iron transport and disorders.

Distribution of genes encoding virulence factors ompB2, ompCD, ompE, ß-lactamase and serotype in pathogenic and colonizing strains of Moraxella catarrhalis.

BACKGROUND AND AIMS: A total of 115 Moraxella catarrhalis isolates from patients with lower respiratory tract and otorhinolaryngeal infections, as well as healthy carriers, were collected to study the prevalence of outer membrane virulence and resistance encoding genes and lipooligosaccharide (LOS) serotypes. METHODS: PCR technique was used to determine the presence of genes ompB2 (encoding uptake of vital iron), ompCD and ompE (for adhesion, complement resistance and transporter proteins), bro1 and bro2 (for ß-lactamases). Serotyping was carried out by monoclonal antibodies (MAb) against LOS serotypes A, B and C. RESULTS: The frequency of genes determining virulence factors were ompE 82.61%, ompCD 70.43%, ompB2 43.48%, and bro 98.26%. Dissemination of virulence genes varied according to the type of infections and carrier state. CONCLUSIONS: The present study revealed the presence of a greater number of virulence factors in isolates from patients compared to that in strains from healthy individuals. In the strains of patients with bronchopulmonary infections, a combination of ompB2-ompCD-ompE genes was predominant. In patients suffering from otorhinolaryngeal infections, isolates with combination ompCD-ompE prevailed. Isolates from the control group revealed single ompCD or ompE genes or none of the virulence genes tested. The greatest proportion (91.30%) of ß-lactamases was encoded from bro1 genes. Serotype A was the most frequent (70.43%) serotype. No correlation was found between the strain serotype and presence of the virulence and ß-lactamase encoding genes tested.

Receptores solubles de transferrina como mejor indicador bioquímico para definir deficiencia de hierro / Soluble transferrin receptors as best biochemical test for iron deficiency / Receptores solúveis de transferrina como o melhor indicador bioquímico para definir deficiência de ferro

Con el propósito de valorar el mejor análisis bioquímico como indicador del perfil de hierro en niños preescolares sin anemia se analizaron 149 muestras de niños y niñas con una edad promedio de 4 años de una comunidad urbana marginal y otra rural de Costa Rica a los que se les realizó análisis de hemoglobina, ferritina, receptores solubles de transferrina, protoporfirina eritrocitaria y proteína C reactiva. El 42% de las muestras presentaron un perfil de hierro dentro de los intervalos de referencia. Sin embargo, se detectó deficiencia de hierro en el 30,8% utilizando receptores solubles de transferrina, en un 14% utilizando la protoporfirina zinc eritrocitaria y en un 10% mediante la ferritina sérica. Además, el 16,8% de las muestras mostraron una elevación inespecífica de la ferritina debido a un proceso infeccioso o inflamatorio agudo y el 5% elevación de la protoporfirina zinc eritrocitaria. Se puede concluir que si se cuantifica únicamente ferritina sérica para evaluar perfil de hierro se estaría diagnosticando mal a una proporción importante de la población (16,8%). Si se considera únicamente la protoporfirina eritrocitaria aumentarían en un 19% las muestras deficientes en hierro pero con un 5% de falsas disminuciones. En cambio, si se evalúan los receptores solubles de transferrina se estaría detectando un número mayor de muestras, un 30,8% con perfil bajo de hierro. Por lo tanto, en la experiencia de estos autores resultó de mayor utilidad usar los receptores solubles de tranferrina como mejor indicador bioquímico para valorar perfil de hierro, ya que la ferritina y la protoporfirina eritrocítica son sensibles a los cambios circadianos y a la presencia de procesos agudos inespecíficos. Asimismo, de acuerdo a los datos obtenidos, no se consideró necesario utilizar intervalos de referencia según sexo y lugar de residencia en niños y niñas de 4 años ya que no hubo diferencia estadísticamente significativa entre sexo y zona urbana marginal y rural para ningún análisis estudiado.

With the aim to evaluate the best biochemical analysis of iron status in preschool children without anemia, a hundred and fortynine samples of boys and girls with an average age of 4 years from an urban marginal community and a rural área from Costa Rica hemoglobin, ferrítin, soluble receptors oí transferrin, erythrocyte protoporphyrln and C reactive proteln were analysed. Forty-two per cent oí the samples presented iron status between the reference interval. Nevertheless, iron deficiency was detected in 30.8% oí the cases using soluble transferrin receptor, 14% with erythrocyte protoporphyrln and 10% with serie ferrítin. In addition, 16.8% of the samples had an unspecified increase in ferrítin due to acute infectious or inflammatory process, and 5% of erythrocyte protoporphyrín. It can be concluded that if serum ferrítin is quantified solely to evalúate iron status, an ímportant portíon of the population (16.8%) would be wrongly díagnosed. If erythrocyte protoporphyrln was considered alone, it would increase the iron deficient samples by 19%, but with 5% of false reductions. However, evaluating the soluble receptors of transferrin we would be detecting a greater number of samples-30.8% with low iron status. In the experience of the authors, it was of major utility use to use the soluble receptors of tranferrin as better biochemical indicators to assess iron status, since ferrítin and erythrocyte protoporphyrln are sensible to circadian changes and to the presence of unspecific acute processes. Because of the data obtained, it was not considered necessary to use different reference valúes for sex or place of residence in four year-old boys and girís because there are no significant statistical differences between sex or resideney for any analysis studied.

Staphylococcus aureus siderophore-mediated iron-acquisition system plays a dominant and essential role in the utilization of transferrin-bound iron.

Staphylococcus aureus is known to be capable of utilizing transferrin-bound iron, via both siderophore- and transferrin-binding protein (named IsdA)-mediated iron-acquisition systems. This study was designed in order to determine which iron-acquisition system plays the essential or dominant role with respect to the acquisition of iron from human transferrin, in the growth of S. aureus. Holotransferrin (HT) and partially iron-saturated transferrin (PT), but not apotransferrin (AT), were found to stimulate the growth of S. aureus. S. aureus consumed most of the transferrin-bound iron during the exponential growth phase. Extracellular proteases were not, however, involved in the liberation of iron from transferrin. Transferrin-binding to the washed whole cells via IsdA was not observed during the culture. The expression of IsdA was observed only in the deferrated media with AT, but not in the media supplemented with PT or HT. In contrast, siderophores were definitely produced in the deferrated media with PT and HT, as well as in the media supplemented with AT. The siderophores proved to have the ability to remove iron directly from transferrin, but the washed whole cells expressing IsdA did not. In the bioassay, the growth of S. aureus on transferrin-bound iron was stimulated by the siderophores alone. These results demonstrate that the siderophore-mediated iron-acquisition system plays a dominant and essential role in the uptake of iron from transferrin, whereas the IsdA-mediated iron-acquisition system may play only an ancillary role in the uptake of iron from transferrin.

Construction and characterization of Moraxella catarrhalis mutants defective in expression of transferrin receptors.

We have previously reported the construction of an isogenic mutant defective in expression of OmpB1, the TbpB homologue, in Moraxella catarrhalis 7169. In this report, we have extended these studies by constructing and characterizing two new isogenic mutants in this clinical isolate. One mutant is defective in expression of TbpA, and the other mutant is defective in expression of both TbpA and TbpB. These isogenic mutants were confirmed by using PCR analysis, sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and sequencing. In vitro growth studies, comparing all three mutants, demonstrated that the tbpA mutant and the tbpAB mutant were severely limited in their ability to grow with human holotransferrin as the sole source of iron. In contrast, the ompB1 (tbpB) mutant was capable of utilizing iron from human transferrin, although not to the extent of the parental strain. While affinity chromatography with human holotransferrin showed that each Tbp was capable of binding independently to transferrin, solid-phase transferrin binding studies using whole cells demonstrated that the tbpA mutant exhibited binding characteristics similar to those seen with the wild-type bacteria. However, the ompB1 (tbpB) mutant exhibited a diminished capacity for binding transferrin, and no binding was detected with the double mutant. These data suggest that the M. catarrhalis TbpA is necessary for the acquisition of iron from transferrin. In contrast, TbpB is not essential but may serve as a facilitory protein that functions to optimize this process. Together these mutants are essential to provide a more thorough understanding of iron acquisition mechanisms utilized by M. catarrhalis.

Use of an isogenic mutant constructed in Moraxella catarrhalis To identify a protective epitope of outer membrane protein B1 defined by monoclonal antibody 11C6.

Moraxella catarrhalis-induced otitis media continues to be a significant cause of infection in young children, prompting increased efforts at identifying effective vaccine antigens. We have previously demonstrated that M. catarrhalis expresses specific outer membrane proteins (OMPs) in response to iron limitation and that this organism can utilize transferrin and lactoferrin for in vitro growth. One of these proteins, which binds human transferrin, is OMP B1. As the human host presents a naturally iron-limited environment, proteins, like OMP B1, which are expressed in response to this nutritional stress are potential vaccine antigens. In this study, we have developed monoclonal antibody (MAb) 11C6, which reacts to a surface-exposed epitope of OMP B1 expressed by M. catarrhalis 7169. This antibody was used to clone ompB1, and sequence analysis suggested that OMP B1 is the M. catarrhalis homologue to the transferrin binding protein B described for pathogenic Neisseriaceae, Haemophilus influenzae, Actinobacillus pleuropneumoniae, and M. catarrhalis. Expression of recombinant OMP B1 on the surface of Escherichia coli confers transferrin binding activity, confirming that this protein is likely involved in iron acquisition. In addition, ompB1 was used to construct an isogenic mutant in M. catarrhalis 7169. This mutant, termed 7169b12, was used as the control in bactericidal assays designed to determine if OMP B1 elicits protective antibodies. In the presence of MAb 11C6 and human complement, wild-type 7169 demonstrated a 99% decline in viability, whereas the ompB1 isogenic mutant was resistant to this bactericidal activity. Further analysis with MAb 11C6 revealed the presence of this OMP B1 epitope on 31% of the clinical isolates tested. These data suggest that OMP B1 is a potential vaccine antigen against M. catarrhalis infections.