American foulbrood (AFB) is a dangerous disease of honeybees (Apis mellifera) caused by the spore-forming bacterium Paenibacillus larvae. According to the ERIC (enterobacterial repetitive intergenic consensus) classification, five genotypes are distinguished, i.e., I, II, III, IV, and V, which differ in their virulence and prevalence in colonies. In the Czech Republic, AFB prevalence is monitored by the State Veterinary Administration; however, the occurrence of specific P. larvae genotypes within the country remains unknown. In this study, our aim was to genotype field P. larvae strains collected in the Czech Republic according to the ERIC classification. In total, 102 field isolates from colonies with AFB clinical symptoms were collected from various locations in the Czech Republic, and the PCR genotypization was performed using ERIC primers. We confirmed the presence of both ERIC I and II genotypes, while ERIC III, IV, and V were not detected. The majority of samples (n = 82, 80.4%) were identified as ERIC II, while the ERIC I genotype was confirmed only in 20 samples (19.6%). In contrast to other European countries, the ERIC II genotype is predominant in Czech honeybee colonies. The ERIC I genotype was mostly detected in border regions close to Poland, Slovakia, and Austria.

Department of Biochemistry Faculty of Science Palacký University Olomouc Olomouc Czechia

Department of Geoinformatics Faculty of Science Palacký University Olomouc Olomouc Czechia

Experimental Zooprophylactic Institute in Lombardy and Emilia Romagna Brescia Italy

State Veterinary Institute Olomouc Olomouc Czechia

Front Vet Sci. 2021 Dec 06;8:807222. doi: 10.3389/fvets.2021.807222. PubMed

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Genersch E. American Foulbrood in honeybees and its causative agent, Paenibacillus larvae. J Invertebr Pathol. (2010) 103:S10–19. 10.1016/j.jip.2009.06.015 PubMed DOI

Oie W.O.F.a.H . American Foulbrood of Honey Bees (Infection of Honey Bees With Paenibacillus larvae). In OIE Terrestrial Manual, ed. Anonymous. World Organisation for Animal Health (2018).

White GF, United S, Bureau Of E, United S, Department Of A . The Bacteria of the Apiary: With Special Reference to Bee Diseases. Washington: G.P.O; (1906). 10.5962/bhl.title.87503 DOI

Forsgren E, Locke B, Sircoulomb F, Schäfer MO. Bacterial diseases in honeybees. Curr Clin Microbiol Reports. (2018) 5:18–25. 10.1007/s40588-018-0083-0 DOI

Stephan JG, De Miranda JR, Forsgren E. American foulbrood in a honeybee colony: spore-symptom relationship and feedbacks. BMC Ecol. (2020) 20. 10.1186/s12898-020-00285-8 PubMed DOI PMC

Brødsgaard CJ, Ritter W, Hansen H. Response of in vitro reared honey bee larvae to various doses of Paenibacillus larvae larvae spores. Apidologie. (1998) 29:569–78. 10.1051/apido:19980609 DOI

Beims H, Bunk B, Erler S, Mohr KI, Spröer C, Pradella S, et al. . Discovery of Paenibacillus larvae ERIC V: Phenotypic and genomic comparison to genotypes ERIC I-IV reveal different inventories of virulence factors which correlate with epidemiological prevalences of American Foulbrood. Int J Med Microbiol. (2020) 310:151394. 10.1016/j.ijmm.2020.151394 PubMed DOI

Genersch E, Forsgren E, Pentikainen J, Ashiralieva A, Rauch S, Kilwinski J, et al. . Reclassification of Paenibacillus larvae subsp. pulvifaciens and Paenibacillus larvae subsp larvae as Paenibacillus larvae without subspecies differentiation. Int J Syst Evol Microbiol. (2006) 56:501–11. 10.1099/ijs.0.63928-0 PubMed DOI

Forsgren E, Stevanovic J, Fries I. Variability in germination and in temperature and storage resistance among Paenibacillus larvae genotypes. Vet Microbiol. (2008) 129:342–9. 10.1016/j.vetmic.2007.12.001 PubMed DOI

Poppinga L, Janesch B, Fünfhaus A, Sekot G, Garcia-Gonzalez E, Hertlein G, et al. . Identification and functional analysis of the S-Layer protein SplA of paenibacillus larvae, the causative agent of american foulbrood of honey bees. PLoS Pathog. (2012) 8:e1002716. 10.1371/journal.ppat.1002716 PubMed DOI PMC

Funfhaus A, Poppinga L, Genersch E. Identification and characterization of two novel toxins expressed by the lethal honey bee pathogen Paenibacillus larvae, the causative agent of American foulbrood. Environ Microbiol. (2013) 15:2951–65. 10.1111/1462-2920.12229 PubMed DOI

Yue D, Nordhoff M, Wieler LH, Genersch E. Fluorescence in situ hybridization (FISH) analysis of the interactions between honeybee larvae and Paenibacillus larvae, the causative agent of American foulbrood of honeybees (Apis mellifera). Environ Microbiol. (2008) 10:1612–20. 10.1111/j.1462-2920.2008.01579.x PubMed DOI

Bassi S, Formato G, Milito M, Trevisiol K, Salogni C, Carra E. Phenotypic characterization and ERIC-PCR based genotyping of Paenibacillus larvae isolates recovered from American foulbrood outbreaks in honey bees from Italy. Vet Q. (2015) 35:27–32. 10.1080/01652176.2014.993095 PubMed DOI

Loncaric I, Derakhshifar I, Oberlerchner JT, Koglberger H, Moosbeckhofer R. Genetic diversity among isolates of Paenibacillus larvae from Austria. J Invertebr Pathol. (2009) 100:44–6. 10.1016/j.jip.2008.09.003 PubMed DOI

Antúnez K, Piccini C, Castro-Sowinski S, Rosado AS, Seldin L, Zunino P. Phenotypic and genotypic characterization of Paenibacillus larvae isolates. Vet Microbiol. (2007) 124:178–83. 10.1016/j.vetmic.2007.04.012 PubMed DOI

Alippi AM, Lopez AC, Aguilar OM. A PCR-based method that permits specific detection of Paenibacillus larvae subsp. larvae, the cause of American Foulbrood of honey bees, at the subspecies level. Lett Appl Microbiol. (2004) 39:25–33. 10.1111/j.1472-765X.2004.01535.x PubMed DOI

Krongdang S, Evans JD, Chen Y, Mookhploy W, Chantawannakul P. Comparative susceptibility and immune responses of Asian and European honey bees to the American foulbrood pathogen, Paenibacillus larvae. Insect Sci. (2019) 26:831–42. 10.1111/1744-7917.12593 PubMed DOI

Versalovic J, Schneider GM, Bruijn F, Lupski JR. Genomic fingerprint of bacteria using repetitive sequence-based polymerase chain reaction. Methods Mol Cell Biol. (1994) 5:25–40.

Hulton CS, Higgins CF, Sharp PM. ERIC sequences: a novel family of repetitive elements in the genomes of Escherichia coli, Salmonella typhimurium and other enterobacteria. Mol Microbiol. (1991) 5:825–34. 10.1111/j.1365-2958.1991.tb00755.x PubMed DOI

SVA . Mor včelího plodu. State Veterinary Administration. (2021). Available online at: https://www.svscr.cz/zdravi-zvirat/mor-vceliho-plodu/ (accessed February 25, 2021).

Bzdil J. Detection of Paenibacillus larvae spores in the debris and wax of honey bee by the tween 80 method. Acta Veterinaria Brno. (2007) 76:643–8. 10.2754/avb200776040643 DOI

Morrissey BJ, Helgason T, Poppinga L, Funfhaus A, Genersch E, Budge GE. Biogeography of Paenibacillus larvae, the causative agent of American foulbrood, using a new multilocus sequence typing scheme. Environ Microbiol. (2015) 17:1414–24. 10.1111/1462-2920.12625 PubMed DOI PMC

Spivak M, Reuter GS. Resistance to American foulbrood disease by honey bee colonies Apis mellifera bred for hygienic behavior. Apidologie. (2001) 32:555–65. 10.1051/apido:2001103 DOI

Screening of Honey Bee Pathogens in the Czech Republic and Their Prevalence in Various Habitats