Isolation, characterization, and genetic diversity of Paenibacillus larvae from AFB suspected specimens in the Central and Eastern Black Sea Regions

American foulbrood is an important bacterial disease affecting the larvae of honey bees (Apis mellifera L.) caused by Paenibacillus larvae. Due to easy transmission of disease and the ability of bacteria to create spores, it is a bacterium resistant to both physical and chemical conditions. The study aims to isolate, perform microbiological analyses, and determine biochemical properties and genotypes P. larvae strains from AFB samples collected from Turkey's Central and Eastern Black Sea regions. An isolation study was conducted on adult bees, larvae, honey, and primary honeycomb samples from suspected colonies in the regions under study. After the purification of bacterial isolates from samples, P. larvae strains were identified using biochemical and molecular methods. The genetic diversity and ERIC types of P. larvae isolates were determined by rep-PCR DNA genotyping using BOX A1R and MBO REP1 primers and multiplex-PCR. A phylogenetic tree of P. larvae strains was constructed in the study. All P. larvae isolates were determined as ERIC I type. According to the rep-PCR results of P. larvae strains, 15 of the 28 isolates were Ab genotype (54%), 7 (25%) Aβ genotype, 4 (14%) AB genotype, 1 (3.5%) αB genotype, and 1 (3.5%) ab genotype. From an epidemiological viewpoint, it was determined that Ab and Aβ genotypes were widely distributed, while other genotypes (AB, αB, and ab) showed less spread. The results of the study will guide researchers in taking relevant measures to prevent and control American foulbrood. © 2023, The Author(s), under exclusive licence to Plant Science and Biodiversity Centre, Slovak Academy of Sciences (SAS), Institute of Zoology, Slovak Academy of Sciences (SAS), Institute of Molecular Biology, Slovak Academy of Sciences (SAS)

Antibacterial activity of bryophyte species against Paenibacillus larvae isolates

This study was performed to determine the antibacterial activity of methanol extracts of 23 bryophyte species against Paenibacillus larvae isolates that cause American foulbrood diseases in honeybee larvae. The honey and larva samples were collected from nine different locations of Rize in Turkey. A total of 22 gram-positive spore-forming bacteria were isolated from the larva and honey samples. According to the results of morphological, biochemical, and molecular (16S rRNA gene sequencing) tests, 10 isolates of the 22 gram-positive spore-forming bacteria were identified as P. larvae. A total of 10 bryophyte species (Polytrichum formasum,Polytrichum commune, Calliergonella cuspitada, Calliergonella lindbergi, Metzgeria conjugata, Isothecium alopecuroides, Syntrichia calcicola, Syntrichia intermedia, Tortella densa,and Grimmia alpestris) among 23 bryophytes showed good antimicrobial activity against P. larvae isolates according the results of agar-well diffusion method and minimal inhibition concentration experiments

Characterization and antibiotic sensitiveness of gram positive bacteri̇a isolated from American foulbrood suspect bee (apis mellifera) and bee products, with genetic di̇versity of paenibacillus larvae isolates

Bu çalışmada, Amerikan Yavru Çürüklüğü hastalığı şüpheli arı ve arı ürünlerinden izolen edilen Gram-pozitif bakterilerin karakterizasyonu, antibiyotik duyarlılıkları ve Paenibacillus larvae izolatlarının genetik çeşitliliğinin belirlenmesi amaçlanmıştır. 64 izolatın fiziksel ve biyokimyasal özelliklerinin yanında moleküler tanımlama için 16S rRNA gen bölgesi sekans analizi metodu kullanıldı ve mikroorganizmaların tür tayini yapıldı. İzolatların18'i Paenibacillus larvae, 15'i P. dentriformis, 1'i P. alvei, 8'i Bacillus subtilis, 3'ü B. pumilus, 2'si B. licheniformis, 1'i B. amyloliquefaciens, 3'ü Staphylococcus warneri olarak tür düzeyinde tanımlandı. İzolatların 10'u Bacillus sp., 2'si Kocuria sp. olarak sadece cins seviyesinde tanımlandı. Çalışmada izole edilen 63 izolatın 54'ünün (%86) kullanılan antibiyotiklerden en az birine dirençli olduğu tespit edildi. İzolatların 38'i (%60) kullanılan iki ve daha fazla antibiyotiğe karşı dirençli olduğu ve çoklu direnç taşıdıkları tespit edildi. Diğer yandan tanımlanan Paenibacillus larvae izolatlarının genetik çeşitliliğini belirlemek için BOX A1R ve MBO REP1 primerleri vasıtasıyla rep-PZR DNA parmak izi yöntemi kullanıldı. Yapılan rep-PZR sonuçlarına göre 18 izolatın 8'i (%53) Ab genotipinde iken, 7'si (%39) Aß, 2'si (%11) AB ve 1'i (%6) ab genotipinde olduğu tespit edildi. Çalışmada izole edilen 18 P. larvae izolatının her birinin 1 adet plazmit DNA içerdiği tespit edildi. Aynı zamanda tetrasiklin ve oksitetrasiklin dirençli olan 8 P. larvae izolatında tet(K), tet(L), tet(M) ve tet(W) genleri, plazmit DNA'lar kalıp olarak kullanılarak tarandı. Yapılan çalışmada 7 izolatın tet(W) geni yönünden pozitif olduğu tespit edilirken, 8 izolatın tet(M) geni yönünden pozitif olduğu tespit edildi. 2017, 101 sayfa Anahtar Kelimeler: Amerikan Yavru Çürüklüğü (AYÇ), Paenebacillus larvae, rep-PZR DNA parmak izi, tetrasiklin direnç genleri. In this study, it was aimed to determine the characterization of Gram-positive bacteria which were isolated from bee and bee products suspected the American Foulbrood disease, their susceptibility to antibiotics and the genetic diversity of Paenibacillus larvae isolates. In addition to the physical and biochemical properties of 64 isolates, the 16S rRNA gene sequences were used for molecular identification and species identification of the isolates were performed. The isolates were identified at species level as Paenibacillus larvae (18 isolates), P. dentriformis (15 isolates), P. alvei (1 isolate), Bacillus subtilis (8 isolates), B. pumilus (3 isolates), B. licheniformis (2 isolates), B. amyloliquefaciens (1 isolate) and Staphylococcus warneri (3 isolates). Ten isolates were identified Bacillus sp. and two isolates were identified as Kocuria sp. at genus level. It was determined that 54 of the 63 isolates (86%) used in this study were found to be resistant to at least one of the antibiotics used. It was found that 38 isolates (60%) were resistant to two or more antibiotics used and had multiple resistance. On the other hand, the rep-PCR DNA fingerprinting method was used to determine the genetic diversity of the identified Paenibacillus larvae isolates via BOX A1R and MBO REP1 primers. According to the rep-PCR results, it was determined that 8 of 18 isolates (53%) were Ab genotype, 7 (39%) were Ab, 2 (11%) were AB and 1 (6%) was ab genotype. It was determined that each of 18 P. larvae isolates used in the study contained 1 plasmid DNA. At the same time, tet(K), tet(L), tet(M) and tet(W) genes were screened using plasmid DNAs as templete DNA in 8 P. larvae isolates which were resistant to tetracycline and oxytetracycline and the tet(W) gene was detected in 7 isolates while tet(M) gene was detected in 8 isolates. 2017, 101 pages Keywords: American Foulbrood, Paenibacillus larvae, rep-PCR DNA fingerprinting, tetracycline resistance genes

Antibacterial effects of anatolian propolis on paenibacillus larvae

Paenibacillus larvae (P. larvae) is a pathogenic bacterium causing American Foulbrood Disease (AFB) in honeybee larvae. It is necessary to develop alternative strategies for the control of this disease due to the serious honeybee colonies loses and the use of antibiotics. Recent studies are aimed at the investigating natural fighting agents against P. larvae. In our study, it was aimed to demonstrate potential antibacterial efficacy of ethanol extract of Anatolian Propolis (EAP) against P. larvae strains PB35 and SV35 which were isolated in Turkey. The total phenolic content (TPC) and flavonoid content (TFC) of EEAP were determined as 181.73±5.20 mg Gallic Acid Equivalents (GAE)/g, and 42.33±1.40 mg Quercetin Equivalents (QE)/g, respectively. It was found that EAP contains different amounts of ferulic, caffeic, coumaric acids, rutin, and caffeic acid phenethyl ester (CAPE). The antibacterial activity of the EAP was determined by using agar-well diffusion, microdilution, and Bioscreen C techniques. The Minimal Inhibition Concentration (MIC) values of the EAP were determined as 74.87 ?g/ml against strain PB35 and SV35 using both microdilution and Bioscreen C teqnique. In both techniques, Minimal Bactericidal Concentration (MBC) values of the EAP were evaluated as 149 and 598.4 ?g/ml against strain PB35 and SV35, respectively. The fact that EAP shows low concentrations of bacteriostatic (MIC) and bactericide (MBC) activity values against P. larvae strains, spore-forming bacilli, which are agents of AFB disease, suggests that it may be a potential source in AFB disease control. © The Author(s), 2021.Recep Tayyip Erdogan Üniversitesi, RTEU: RTEU-BAP 2015.53001.102.03.04Funding: This work was supported by the Recep Tayyip Erdoğan University research fund (RTEU-BAP 2015.53001.102.03.04. Thanks for their financial supports.Funding: This work was supported by the Recep Tayyip Erdo?an University research fund (RTEU-BAP 2015.53001.102.03.04. Thanks for their financial supports.2-s2.0-8511957887

Antibacterial activity of bryophyte species against Paenibacillus larvae isolates

This study was performed to determine the antibacterial activity of methanol extracts of 23 bryophyte species against Paenibacillus larvae isolates that cause American foulbrood diseases in honeybee larvae. The honey and larva samples were collected from nine different locations of Rize in Turkey. A total of 22 gram-positive spore-forming bacteria were isolated from the larva and honey samples. According to the results of morphological, biochemical, and molecular (16S rRNA gene sequencing) tests, 10 isolates of the 22 gram-positive spore-forming bacteria were identified as P. larvae. A total of 10 bryophyte species (Polytrichum formasum,Polytrichum commune, Calliergonella cuspitada, Calliergonella lindbergi, Metzgeria conjugata, Isothecium alopecuroides, Syntrichia calcicola, Syntrichia intermedia, Tortella densa,and Grimmia alpestris) among 23 bryophytes showed good antimicrobial activity against P. larvae isolates according the results of agar-well diffusion method and minimal inhibition concentration experiments.This study was performed to determine the antibacterial activity of methanol extracts of 23 bryophyte species against Paenibacillus larvae isolates that cause American foulbrood diseases in honeybee larvae. The honey and larva samples were collected from nine different locations of Rize in Turkey. A total of 22 gram-positive spore-forming bacteria were isolated from the larva and honey samples. According to the results of morphological, biochemical, and molecular (16S rRNA gene sequencing) tests, 10 isolates of the 22 gram-positive spore-forming bacteria were identified as P. larvae. A total of 10 bryophyte species (Polytrichum formasum,Polytrichum commune, Calliergonella cuspitada, Calliergonella lindbergi, Metzgeria conjugata, Isothecium alopecuroides, Syntrichia calcicola, Syntrichia intermedia, Tortella densa,and Grimmia alpestris) among 23 bryophytes showed good antimicrobial activity against P. larvae isolates according the results of agar-well diffusion method and minimal inhibition concentration experiments