Novel TaqMan PCR assay for the quantification of Paenibacillus larvae spores in bee-related samples

Kušar, Darja (avtor), Papić, Bojan (avtor), Zajc, Urška (avtor), Zdovc, Irena (avtor), Golob, Majda (avtor), Žvokelj, Lucija (avtor), Knific, Tanja (avtor), Avberšek, Jana (avtor), Ocepek, Matjaž (avtor), Pislak, Metka (avtor)

Paenibacillus larvae is the causative agent of American foulbrood (AFB), a devastating disease of honeybees. P. larvae spore counts in bee-related samples correlate with the presence of AFB symptoms and may, therefore, be used to identify at-risk colonies. Here, we constructed a TaqMan-based real-time PCR (qPCR) assay targeting a single-copy chromosomal metalloproteinase gene for reliable quantification of P. larvae. The assay was calibrated using digital PCR (dPCR) to allow absolute quantification of P. larvae spores in honey and hive debris samples. The limits of detection and quantification were 8 and 58 spores/g for honey and 188 and 707 spores/mL for hive debris, respectively. To assess the association between AFB clinical symptoms and spore counts, we quantified spores in honey and hive debris samples originating from honeybee colonies with known severity of clinical symptoms. Spore counts in AFB-positive colonies were significantly higher than those in asymptomatic colonies but did not differ significantly with regard to the severity of clinical symptoms. For honey, the average spore germination rate was 0.52% (range = 0.04–6.05%), indicating poor and inconsistent in vitro germination. The newly developed qPCR assay allows reliable detection and quantification of P. larvae in honey and hive debris samples but can also be extended to other sample types.

• Jezik: Angleški jezik
• Ključne besede: American foulbrood (AFB), Paenibacillus larvae, real-time or quantitative PCR (qPCR), digital PCR (dPCR), plate counting, spore germination rate, honey, hive debris, Polymerase Chain Reaction
• Vrsta gradiva: Članek v reviji
• Tipologija: 1.01 - Izvirni znanstveni članek
• Organizacija: VF - Veterinarska fakulteta
• Status publikacije: Objavljeno
• Različica publikacije: Objavljena publikacija
• Leto izida: 2021
• Št. strani: 18 str.
• Številčenje: Vol. 12, iss. 11, art. 1034
• PID: 20.500.12556/RUL-133255
• UDK: 577
• ISSN pri članku: 2075-4450
• DOI: 10.3390/insects12111034
• COBISS.SI-ID: 85522179
• Datum objave v RUL: 18.11.2021

Gradivo je del revije

• Naslov: Insects
• Skrajšan naslov: Insects
• Založnik: MDPI
• ISSN: 2075-4450
• COBISS.SI-ID: 519122457

Licence

• Licenca: CC BY 4.0, Creative Commons Priznanje avtorstva 4.0 Mednarodna
• Povezava: http://creativecommons.org/licenses/by/4.0/deed.sl
• Opis: To je standardna licenca Creative Commons, ki daje uporabnikom največ možnosti za nadaljnjo uporabo dela, pri čemer morajo navesti avtorja.
• Začetek licenciranja: 17.11.2021