High mobility group (HMG-box) genes in the honeybee fungal pathogen Ascosphaera apis
ABSTRACT The genome of the honeybee fungal pathogen Ascosphaera apis (Maassen) encodes three putative high mobility group (HMG-box) transcription factors. The predicted proteins (MAT1-2, STE11 and HTF), each of which contain a single strongly conserved HMG-box, exhibit high similarity to mating type proteins and STE11-like transcription factors previously identified in other ascomycete fungi, some of them important plant and human pathogens. In this study we characterized the A. apis HMG-box containing genes and analyzed the structure of the mating type locus (MAT1-2) and its flanking regions. The MAT1-2 locus contains a single gene encoding a protein with an HMG-box. We also have determined the transcriptional patterns of all three HMG-box containing genes in both mating type idiomorphs and discuss a potential role of these transcription factors in A. apis development and reproduction. A multiplex PCR method with primers amplifying mat1-2-1 and Ste11 gene fragments is described. This new method allows for identification of a single mating type idiomorph and might become an essential tool for applied and basic research of chalkbrood disease in honeybees.
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ABSTRACT: The objective of this study was to isolate and characterize bacterial strains associated with the gut of the hybrid Carniolan honey bee, Apis mellifera carnica, and to determine their in vitro and in vivo potential against Ascosphaera apis, the causal organism of chalkbrood disease, with the purpose of exploring feasible biological control. Six bacterial strains were isolated from healthy worker honey bees by culture-dependent methods. Six fungal strains (A3, A4, A7, A8, A9, and A15) of A. apis were isolated from larvae suffering from chalkbrood disease on Yeast–Glucose–Starch agar (YGPSA) medium. All bacteria were identified by a combination of morphology, Gram stain, and 16S rRNA sequence analysis, and fungal strains were identified by morphology and 5.8S rRNA. In vitro and in vivo inhibition assays were carried out to determine the ability of bacterial isolates to inhibit A. apis, the causal agent of chalkbrood disease. The analysis of 16S rRNA sequences revealed that four bacterial strains (B2, B4, B10, and B100) belong to Bacillus subtilis species, and two strains (P1 and P5) belong to Pseudomonas fluorescence. Significant differences in antagonistic activity of all bacterial strains were observed. B. subtilis isolate B2 showed the highest antagonistic activity, as measured by the inhibition zone against A. apis, followed by the P1 strain of P. fluorescence. SEM analysis also supports the antagonistic activity of these bacteria against A. apis. This study provides a theoretical basis for biological control of honey bee chalkbrood diseaseJournal of Apicultural Science 05/2014; 58(1):17-27. · 0.82 Impact Factor
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ABSTRACT: Fungi have evolved a remarkable diversity of reproductive strategies. Some of these, most notably those of the model fungi, have been well studied but others are poorly understood. The latter is also true for uni-directional mating type switching, which has been reported in only five fungal genera, including Ceratocystis. Mating type switching allows a self-fertile fungal isolate to produce both self-fertile and self-sterile offspring. This study considered the molecular nature of uni-directional mating type switching in the type species of Ceratocystis, C. fimbriata. To do this, the genome of C. fimbriata was first examined for the presence of mating type genes. Three mating genes (MAT1-1-1, MAT1-2-1 and MAT1-1-2) were found in an atypical organisation of the mating type locus. To study the effect that uni-directional switching has on this locus, several self-sterile offspring were analysed. Using a combination of next generation and conventional Sanger sequencing, it was shown that a 3581 base pair (bp) region had been completely deleted from the MAT locus. This deletion, which includes the entire MAT1-2-1 gene, results in the permanent loss of self-fertility, rendering these isolates exclusively self-sterile. Our data also suggest that the deletion mechanism is tightly controlled and that it always occurs at the same genomic position. Two 260 bp direct repeats flanking the deleted region are strongly implicated in the process, although the exact mechanism behind the switching remains unclear.PLoS ONE 03/2014; 9(3):e92180. DOI:10.1371/journal.pone.0092180 · 3.53 Impact Factor
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ABSTRACT: Chalkbrood and stonebrood are two fungal diseases associated with honey bee brood. Chalkbrood, caused by Ascosphaera apis, is a common and widespread disease that can result in severe reduction of emerging worker bees and thus overall colony productivity. Stonebrood is caused by Aspergillus spp. that are rarely observed, so the impact on colony health is not very well understood. A major concern with the presence of Aspergillus in honey bees is the production of airborne conidia, which can lead to allergic bronchopulmonary aspergillosis, pulmonary aspergilloma, or even invasive aspergillosis in lung tissues upon inhalation by humans. In the current chapter we describe the honey bee disease symptoms of these fungal pathogens. In addition, we provide research methodologies and protocols for isolating and culturing, in vivo and in vitro assays that are commonly used to study these host pathogen interactions. We give guidelines on the preferred methods used in current research and the application of molecular techniques. We have added photographs, drawings and illustrations to assist bee-extension personnel and bee scientists in the control of these two diseases.Journal of Apicultural Research 01/2013; 52(1). DOI:10.3896/IBRA.126.96.36.199 · 1.36 Impact Factor