Characterization of Monoclonal Antibodies to Avian Escherichia coli Iss

Abstract

Colibacillosis accounts for annual multimillion dollar losses in the poultry industry, and control of this disease is hampered by limited understanding of the virulence mechanisms used by avian pathogenic Escherichia coli (APEC). Previous work in our laboratory has found that the presence of the increased serum survival gene (iss) is strongly associated with APEC but not commensal E. coli, making iss and the protein it encodes (Iss) candidate targets of colibacillosis-control procedures. Previously, we produced monoclonal antibodies (MAbs) against Iss to be used as a reagent in studies of APEC virulence and colibacillosis pathogenesis. Unfortunately, the utility of these MAbs was limited because these MAbs exhibited nonspecific binding. It was thought that the lack of specificity might be related to the fact that these MAbs were of the immunoglobulin M (IgM) isotype. In the present study, new MAbs were produced using a different immunization strategy in an effort to generate MAbs of a different isotype. Also, because Iss bears strong similarity to Bor, a lambda-derived protein that occurs commonly among E. coli, MAbs were assessed for their ability to distinguish Iss and Bor. For these studies, the bor gene from an APEC isolate was cloned into an expression vector. The fusion protein expressed from this construct was used to assess the potential of the anti-Iss MAbs produced in the past and present studies to distinguish Bor and Iss. The MAbs produced in this study were of the IgG 1 isotype, which appeared to bind more specifically to Iss than previously generated antibodies in certain immunologic procedures. These results suggested that the MAbs generated in this study might prove superior to the previous MAbs as a reagent for study of APEC. However, both MAbs recognized recombinant Iss and Bor, suggesting that any results obtained using anti-Iss MAbs would need to be interpreted with this cross-reactivity in mind. BioOne sees sustainable scholarly publishing as an inherently collaborative enterprise connecting authors, nonprofit publishers, academic institutions, research libraries, and research funders in the common goal of maximizing access to critical research. Characterization of Monoclonal Antibodies to Avian Escherichia coli IssAuthor (s) SUMMARY. Colibacillosis accounts for annual multimillion dollar losses in the poultry industry, and control of this disease is hampered by limited understanding of the virulence mechanisms used by avian pathogenic Escherichia coli (APEC). Previous work in our laboratory has found that the presence of the increased serum survival gene (iss) is strongly associated with APEC but not commensal E. coli, making iss and the protein it encodes (Iss) candidate targets of colibacillosis-control procedures. Previously, we produced monoclonal antibodies (MAbs) against Iss to be used as a reagent in studies of APEC virulence and colibacillosis pathogenesis. Unfortunately, the utility of these MAbs was limited because these MAbs exhibited nonspecific binding. It was thought t...

CARACTERIZACIÓN Y DISTRIBUCIÓN DE CEPAS DE Escherichia coli POTENCIALMENTE PATÓGENAS AISLADAS DE POLLOS BROILER DE EXPLOTACIONES AVÍCOLAS EN EL PERÚ.

“…De igual manera, el gen iss está fuertemente relacionado al plásmido ColV ya que se ubica dentro de esta sección del genoma (Lynne et al, 2006). Dada la alta presentación de este plásmido en los aislamientos, se podría sospechar una incidencia similar del gen iss en conjunto con el gen estructural de la Colicina V, tal como se dio en este estudio.…”

“…Las vacunas experimentales están dirigidas contra serogrupos específicos, muchos de los cuales sólo presentan protección contra desafíos homólogos (Kariyawasan et al, 2003;Roland et al, 2004), lo cual es una seria limitación debido a la variedad tan extensa que poseen los APEC. Lynne et al (2006) sugieren la creación de vacunas contra las estructuras comunes a la mayoría de cepas APEC, como los genes codificadores de virulencia. Sus estudios revelan que la creación de una vacuna experimental a partir de la proteína Iss, codificada por el gen iss, fue capaz de conferir protección ante el desafío heterólogo.…”

“…Las cepas patógenas o potencialmente patógenas, además de asociarse a causas de mortalidad embrionaria (Jeffrey et al, 2002;Skyberg et al, 2003;Gibbs et al, 2004), pueden ser identificadas por su resistencia al suero aviar debido a la lipoproteína externa de membrana ISS, codificada por el gen iss (Lynne et al, 2005(Lynne et al, , 2006Johnson et al, 2002Johnson et al, , 2008. El crecimiento bacteriano en suero aviar de cepas mutantes pAPEC-O2-colV issse vio notablemente reducido, pero se restituyó al insertar nuevamente el gen iss en el genoma de la cepa mutante, dando a entender la importancia de este factor para la supervivencia de la bacteria en el suero (Lynne et al, 2007).…”

Detection of Iss and Bor on the surface of Escherichia coli

“…This similarity is of concern, as it makes generation of Iss‐specific monoclonal antibodies (Mabs) for use as reagents in the study of APEC and control of avian colibacillosis problematic (Foley et al. 2003; Lynne et al. 2006a).…”

“…2003). These were considered undesirable because of their nonspecific binding of negative controls (Lynne et al. 2006a).…”