The use of repetitive element PCR fingerprinting (rep-PCR) for genetic subtyping of German field isolates of Paenibacillus larvae subsp. larvae

Abstract

-Studies using the repetitive element PCR fingerprinting technique (rep-PCR) revealed that BOX A1R-, MBO REP1-and ERIC-primers reproducibly generate distinctive DNA fingerprints from Paenibacillus larvae subsp. larvae. Four different genetic subtypes of P. larvae were identified in Germany using the three primers. Correlating the results from genetic and morphological subtyping, the flat and transparent morphotype could be assigned to one genetic group. Geographic evaluation of our results demonstrated that the different genetic subtypes appeared in clusters correlating with different outbreaks of American foulbrood. Taken together, our results indicated that rep-PCR performed with a combination of BOX A1R-and MBO REP1-primers will be an effective tool for establishing a molecular epidemiology of P. l. larvae.American foulbrood / Paenibacillus larvae larvae / DNA-fingerprinting / genetic subtyping / molecular epidemiology

Biochemical characterization of different genotypes of Paenibacillus larvae subsp. larvae, a honey bee bacterial pathogen

“…By analysing German P. l. larvae isolates using rep-PCR fingerprinting performed with primers BOX A1R and MBO REP1, three BOX A1R and two MBO REP1 patterns (A, a, a and B, b, respectively) were characterized recently (Genersch & Otten, 2003). Differential combinations of these patterns were found, resulting in the identification of the four genetic subgroups AB, Ab, ab and aB (Genersch & Otten, 2003).…”

“…Recent genotyping of German field isolates of P. l. larvae revealed at least four different genotypes, named AB, Ab, ab and aB (Genersch & Otten, 2003). Here we present data on the further characterization of the genotypes AB, Ab and ab, and of the reference strain DSM 7030/ATCC 9545.…”

“…Honey samples had been stored at 4 uC until P. l. larvae was cultivated for scientific purposes from these samples on bacterial plates. Two representatives of the recently identified P. l. larvae genotype AB (Genersch & Otten, 2003) have been deposited at the DSMZ (accession numbers DSM 16115, DSM 16116).…”

“…The culture of P. l. larvae from honey samples was performed essentially as previously described (Genersch & Otten, 2003). Briefly, for growth of spore-forming bacteria, honey samples were solubilized overnight at 37 uC.…”

“…Preparation of bacterial DNA for fingerprinting and subsequent rep-PCR analysis of bacterial isolates was performed essentially as described previously (Genersch & Otten, 2003). In brief, P. l. larvae DNA suitable for rep-PCR DNA fingerprinting was isolated from colonies grown on culture plates using 6 % InstaGene matrix (Bio-Rad) following the instructions of the manufacturer.…”