Abstract
Bacteriophage endolysins present enormous biotechnological potentials and have been successfully used to control and detect bacterial pathogens. Endolysins targeting Gram-positive bacteria are modular, displaying a cell binding (CBD) and an enzymatically active domain. The CBD of phage endolysins are recognized by their high specificity and host affinity, characteristics that make them promising diagnostic tools. No CBD able to bind Paenibacillus larvae has been identified so far. P. larvae is a Gram-positive spore forming bacteria that causes the American Foulbrood. This highly contagious infection leads to honeybee larvae sepsis and death, resulting in an adverse impact on pollination and on the beekeeping industry. In this work, the first CBD targeting P. larvae was identified and its core binding sequence was investigated. Moreover, it was shown that the domain is highly specific, targeting exclusively P. larvae cells from all ERIC genotypes. The identification of such a domain represents a step forward for the development of effective methods to detect and control this pathogen.
Highlights
Their intrinsic characteristics together with the successful results obtained enabled cell binding (CBD) to emerge as promising diagnostic tools[4,5]
An up-to-date bioinformatic analysis of the annotated lysins from the currently sequenced 25 Paenibacillus phage genomes deposited at the NCBI was still unable to identify a functional domain at their C-terminus
The observation of green-decorated cells by fluorescence microscopy after incubation with the fusion protein green fluorescent protein (GFP)-cell binding-containing fragment (CBCF), and the inability of GFP alone to decorate the same cells, indicates that the CBCF peptide stretch contains a CBD (Fig. 2). This corresponds to the first identification of a lysin CBD able to bind Paenibacillus larvae cells
Summary
Their intrinsic characteristics together with the successful results obtained enabled CBDs to emerge as promising diagnostic tools[4,5]. The economic impact in the beekeeping industry and the critical role of honeybees in crop pollination[10,15] encourages the development of new strategies for the detection and control of AFB. Molecular methods, as those based on PCR, have overcome many of the cultivation-based limitations to detect bacteria due to their specificity, high sensitivity and enrichment culturing avoidance. The false-positive results obtained from DNA of dead bacteria can compromise the success of such methods[16]. The identification of new proteins as bio-recognition elements able to bind to this bacterium will play a critical role in the design of new detection and control methods for P. larvae
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