الفهرس 
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Abstract
American Foulbrood (AFB) is still among the most deleterious bee diseases. This bacterial disease not only kills infected larvae but also potentially lethal to infected colonies.
American foulbrood has traditionally been controlled by artificial swarming and burning infected colonies. In order to lessen this financial loss to beekeepers, Antibiotics were used for foulbrood prevention and control. Not surprisingly, antibiotic resistant strains of the causative bacteria, Paenibacillus larvae have evolved. Another problem with this practice is residues of antibiotics in honey and other hive products. They also threaten the beneficial microbial communities within the colony, such as, Lactic acid bacteria (LAB) which proved in our and previous studies to assemble in honey bee heath.
Treatment with antibiotics is not allowed and completely banned in the European Union, but is common in, for example, USA and Canada where preventive treatments with antibiotics is considered a routine procedure to avoid outbreaks of AFB.
In this study, a new practical aspect was investigated to control one of the most serious honey bee diseases, AFB. Our in vitro and field results strongly suggest that the probiotic LAB linked to the honey bee stomach have important implications for honey bee pathology in general and for AFB tolerance in particular.
1. Isolation and identification of the probiotic lactic acid bacteria (LAB):
Some of the honey bee gut microbial diversity of worker bees using 560 amplicon assays of the 16S rRNA gene was investigated. Our findings revealed that Lactobacillus spp. is an abundant isolate from the honey bee workers’ gut. PCR results and direct sequencing recorded the presence of nine anaerobic lactic acid bacteria. Four of the LABs sequences results, are carefully related to four different strains of Lactobacillus plantarum species. Two of the LABs sequences results are closely identical to two different strains of Lactobacillus kunkeei species. One of the LABs is closely related to a strain of Lactobacillus pentosus species. Two of LABs sequences results are matching two different strains of Lactobacillus sp..
2. Lactic Acid Bacteria (LAB); as a control agent of American Foulbrood disease:
The isolated LAB from the gut of the honey bees were estimated as a new control agent for AFB bacterial pathogen, Paenibacillus larvae larvae (P. l. larvae) spores.
Lactic acid bacteria inhibition assays on agar plates were investigated, to evaluate the effects of the extracted honey bee LAB on P. l. larvae growth in vitro. Our results demonstrated a strong inhibitory effect of the honey bee stomach LAB flora. Nine LAB phylotypes, individually, partially inhibited the represented P. l. larvae genotype. They gave inhibition zones ranges from 0.4 to 1.8 cm.
The honey bee LAB was tested in an in vivo system, particularly on honey bee larvae, against P. l. larvae. Artificial infection was accompanied by the administration of a mixture of five of the most effective endogenous LAB, previously tested for their inhibitory effect on agar plates in vitro. The honey bee endogenous LAB inhibited P. l. larvae. Adding the LAB mixture to the larval food in honey bee colonies significantly reduced number of infected larvae when pooled data from all experiments were analyzed (P ≃ 0.000, P < 0.001).
Checking colonies for clinical symptoms were carried out by visual inspection of the brood combs. The clinical symptoms of AFB are typical and clear only in artificially infected untreated colonies (+ve control) and completely absent from treated colonies with LAB. Checking of colonies results were illustrated by a followed culturing of larvae on J-agar media which proved our field observations and absence of P. l. larvae from treated colonies by LAB, in spite of its heavy appearance in cultures from infected colonies. Further field proceedings for verification of field efficacy of our treatment; showed that after 2 months post treatment with LAB, colonies retained their healthy situation and colony affairs were all absolutely alright. On the other hand, infected untreated colonies were completely destroyed and dead.
This study represented an applicable, reliable, promising and highly effective treatment for AFB disease in honey bee colonies. New avenues were pointed for the prophylactic or therapeutic treatment of honey bee diseases.
Finally, some information was provided about novel nonpathogenic LAB in the honey bee gut of Apis mellifera found in Egypt. It recorded the first data about the effectiveness of application of such probiotic LAB through field experiment.