Nanobodies Selectively Binding to the Idiotype of a Dengue Virus Neutralizing Antibody Do Not Necessarily Mimic the Viral Epitope.

Vaccination against dengue virus is challenged by the fact that a generic immune response can induce antibody-dependent-enhancement (ADE) in secondary infections. Only some antibodies targeting a quaternary epitope formed by the dimerization of the virus protein E possess sufficient neutralizing capacity. Therefore, the immunization with anti-idiotypic antibodies of neutralizing antibodies might represent a safe vaccination strategy. Starting from a large pre-immune library, we succeeded in isolating a wide set of anti-idiotypic nanobodies characterized by selective and strong binding to the paratope of the neutralizing antibody 1C10. However, the mice immunized with such constructs did not produce effective antibodies, despite at least some of them eliciting an immune response selective for the nanobody variable regions. The results suggest that complex conformational epitopes might be difficult to be recreated by anti-idiotypic structures. The selection process of the anti-idiotypic candidates might be optimized by applying epitope mapping and modeling approaches aimed at identifying the key residues that is necessary to bind to trigger selective immune response.

Native llama Nanobody Library Panning Performed by Phage and Yeast Display Provides Binders Suitable for C-Reactive Protein Detection.

C-reactive protein (CRP) is an inflammation biomarker that should be quantified accurately during infections and healing processes. Nanobodies are good candidates to replace conventional antibodies in immunodiagnostics due to their inexpensive production, simple engineering, and the possibility to obtain higher binder density on capture surfaces. Starting from the same pre-immune library, we compared the selection output resulting from two independent panning strategies, one exclusively exploiting the phage display and another in which a first round of phage display was followed by a second round of yeast display. There was a partial output convergence between the two methods, since two clones were identified using both panning protocols but the first provided several further different sequences, whereas the second favored the recovery of many copies of few clones. The isolated anti-CRP nanobodies had affinity in the low nanomolar range and were suitable for ELISA and immunoprecipitation. One of them was fused to SpyTag and exploited in combination with SpyCatcher as the immunocapture element to quantify CRP using electrochemical impedance spectroscopy. The sensitivity of the biosensor was calculated as low as 0.21 µg/mL.

Estudio de marcadores fenotípicos y de susceptibilidad antimicrobiana en cepas de Escherichia coli entéricas / Study of phenotypical and antimicrobial susceptibility markers in enteric Escherichia coli strains

Se analizaron 41 cepas de Escherichia coli, aisladas en niños menores de 5 años con diarreas agudas, procedentes de las diferentes provincias del país. Fueron probados 4 importantes determinantes fenotípicos: sorbosa, sorbitol, enterohemolisina y la serología de O157: H7, para seleccionar las cepas de la categoría enterohemorrágica o productora de toxina shiga. De igual modo fueron caracterizadas por el método de biotipaje y de susceptibilidad antimicrobiana. El uso de las pruebas fenotípicas mostró 6 cepas con características presuntivas, de estas, 4 mostraron mayor probabilidad de pertenecer a la categoría productora de toxina shiga. En el ensayo de susceptibilidad antimicrobiana las cepas mostraron elevada resistencia, sobre todo para la ampicilina y el trimetropin-sulfametoxasol. Otro hallazgo de interés fueron los valores de resistencia y de susceptibilidad intermedia obtenidos para el augmentin, aztreonan y ceftriaxona. Se alcanzaron 12 patrones de resistencia antimicrobiana y de ellos 10 resaltaron por ser multirresistentes.

Forty strains of Escherichia coli isolated from children under 5 years of age with acute diarreas, coming from different provinces of the country , were analyzed. Four important phenotypical determinants were tested: sorbosa, sorbitol, enterohemolysin and 0157: H7 serology, in order to select those strains from enterohemorrhagic or Shiga toxin-producing category. Likewise, they were characterized by biotyping and antimicrobial susceptibility methods. The use of phenotypical tests showed six strains with presumptive characteristics, four of which were most likely to be Shiga toxin-producing strains. In antimicrobial susceptibility test, the strains showed high resistance mainly to ampicillin and trimethrophin-sulfamethoxasole. Another interesting finding were intermediate resistance and susceptibility values to augmentin, aztreonan and ceftriaxone. There were 12 antimicrobial resistance patterns of which 10 were multi-resistant.

Estudio de marcadores fenotípicos y de susceptibilidad antimicrobiana en cepas de Escherichia coli entéricas / Study of phenotypical and antimicrobial susceptibility markers in enteric Escherichia coli strains

Se analizaron 41 cepas de Escherichia coli, aisladas en niños menores de 5 años con diarreas agudas, procedentes de las diferentes provincias del país. Fueron probados 4 importantes determinantes fenotípicos: sorbosa, sorbitol, enterohemolisina y la serología de O157: H7, para seleccionar las cepas de la categoría enterohemorrágica o productora de toxina shiga. De igual modo fueron caracterizadas por el método de biotipaje y de susceptibilidad antimicrobiana. El uso de las pruebas fenotípicas mostró 6 cepas con características presuntivas, de estas, 4 mostraron mayor probabilidad de pertenecer a la categoría productora de toxina shiga. En el ensayo de susceptibilidad antimicrobiana las cepas mostraron elevada resistencia, sobre todo para la ampicilina y el trimetropin-sulfametoxasol. Otro hallazgo de interés fueron los valores de resistencia y de susceptibilidad intermedia obtenidos para el augmentin, aztreonan y ceftriaxona. Se alcanzaron 12 patrones de resistencia antimicrobiana y de ellos 10 resaltaron por ser multirresistentes(AU)

Forty strains of Escherichia coli isolated from children under 5 years of age with acute diarreas, coming from different provinces of the country , were analyzed. Four important phenotypical determinants were tested: sorbosa, sorbitol, enterohemolysin and 0157: H7 serology, in order to select those strains from enterohemorrhagic or Shiga toxin-producing category. Likewise, they were characterized by biotyping and antimicrobial susceptibility methods. The use of phenotypical tests showed six strains with presumptive characteristics, four of which were most likely to be Shiga toxin-producing strains. In antimicrobial susceptibility test, the strains showed high resistance mainly to ampicillin and trimethrophin-sulfamethoxasole. Another interesting finding were intermediate resistance and susceptibility values to augmentin, aztreonan and ceftriaxone. There were 12 antimicrobial resistance patterns of which 10 were multi-resistant(AU)

[Study of phenotypical and antimicrobial susceptibility markers in enteric Escherichia coli strains]. / Estudio de marcadores fenotípicos y de susceptibilidad antimicrobiana en cepas de Escherichia coli entéricas.

Forty strains of Escherichia coli isolated from children under 5 years of age with acute diarreas, coming from different provinces of the country , were analyzed. Four important phenotypical determinants were tested: sorbosa, sorbitol, enterohemolysin and 0157:H7 serology, in order to select those strains from enterohemorrhagic or Shiga toxin-producing category. Likewise, they were characterized by biotyping and antimicrobial susceptibility methods. The use of phenotypical tests showed six strains with presumptive characteristics, four of which were most likely to be Shiga toxin-producing strains. In antimicrobial susceptibility test, the strains showed high resistance mainly to ampicillin and trimethrophin-sulfamethoxasole. Another interesting finding were intermediate resistance and susceptibility values to augmentin, aztreonan and ceftriaxone. There were 12 antimicrobial resistance patterns of which 10 were multi-resistant.