Simultaneous PCR detection of Paenibacillus larvae targeting insertion sequence IS256 and Melissococcus plutonius targeting pMP1 plasmid from hive specimens.
American foulbrood
Apis mellifera
European foulbrood
Melissococcus plutonius
Multiplex PCR
Paenibacillus larvae
Journal
Folia microbiologica
ISSN: 1874-9356
Titre abrégé: Folia Microbiol (Praha)
Pays: United States
ID NLM: 0376757
Informations de publication
Date de publication:
05 Jan 2024
05 Jan 2024
Historique:
received:
08
02
2023
accepted:
11
12
2023
medline:
5
1
2024
pubmed:
5
1
2024
entrez:
5
1
2024
Statut:
aheadofprint
Résumé
Paenibacillus larvae and Melissococcus plutonius represent the most threatening bacterial diseases of honeybee (Apis mellifera)-American and European foulbrood, respectively. For efficient control of those diseases, rapid and accurate detection of the pathogens is crucial. Therefore, we developed a novel multiplex PCR method simultaneously detecting both pathogens. To design and optimize multiplex PCR reaction, four strains of P. larvae representing four ERIC genotypes I-IV (strain DSM 7030-ERIC I, DSM 25430-ERIC II, LMG 16252-ERIC III, DSM 3615-ERIC IV) were selected. Those strains were fully sequenced using long-read sequencing (Sequel I, Pacific Biosciences). For P. larvae, the multicopy insertion sequence IS256 identified in all genotypes of P. larvae was selected to provide high sensitivity. M. plutonius was detected by plasmid pMP1 sequence and the virulence verified by following detection of ETX/MTX2 toxin responsible for pore formation in the cell membrane. As an internal control, a gene encoding for major royal jelly protein 1 specific for honeybees was selected. The method was validated on 36 clinical specimens collected from the colonies suffering from American and European foulbrood in the Czech Republic. Based on the results, sensitivity of PCR was calculated to 93.75% and specificity to 100% for P. larvae diagnosed from hive debris and 100% sensitivity and specificity for honeybee workers and larval scales as well as for diseased brood infected by M. plutonius.
Identifiants
pubmed: 38180723
doi: 10.1007/s12223-023-01125-0
pii: 10.1007/s12223-023-01125-0
doi:
Types de publication
Journal Article
Langues
eng
Sous-ensembles de citation
IM
Subventions
Organisme : Ministerstvo Zemědělství
ID : QK1710228
Organisme : Ministerstvo Školství, Mládeže a Tělovýchovy
ID : LX22NPO5103
Informations de copyright
© 2024. The Author(s).
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