mental conditions (Zander1909;Chene

mental conditions (Zander
1909
;Chenetal.
2009
).
identify QTLs associated with low N. ceranae
In contrast, N. ceranae, which was originally found
spore load. In a subsequent step, we quantified the
in the Asian honeybee A. cerana (Fries et al.
1996
),
expression level of a candidate gene identified in
is a newly established parasite of A. mellifera (Fries
the QTL analyses in the same mapping population.
et al.
2006
; Higes et al.
2006
). Although there were
reports suggesting that N. ceranae had a high
2. MATERIALS AND METHODS
virulence in A. mellifera at both the colony and the
individual level (Higes et al.
2008
,
2009
), more
2.1. Instrumental insemination of queen
recent studies suggest a moderate virulence of N.
bees
ceranae similar to that of N. apis (Forsgren and
Fries
2010
;Gisderetal.
2010
).
Nine virgin queens of the selected Danish strain were
In Denmark, honeybee colonies have been
provided by the Department of Integrated Pest Manage-
selected for the absence of Nosema infections
ment Research Centre Flakkebjerg, Denmark. The
(Nosemosis) for decades (Traynor
2008
). Within
queens were all artificially inseminated with the same
the Danish honeybee breeding scheme, worker
mixed sperm (Moritz
1984
) of 30 drones of an unselected
samples from colonies were used to determine the
strain kept at the Laboratoire de Biologie et Protection de
Nosema infection level. If the samples were found
l’abeille, INRA Avignon, France. The inseminated
infected by Nosema, the queen of the colony
queens were introduced into small colonies composed
would be replaced with one reared from a colony
of 4,000 freshly emerged workers at the apiary of the
without Nosema infection. Although this mode of
Martin-Luther-University Halle-Wittenberg, Germany.
selection was at the colony level based on the
We reared F1 hybrid queens from the inseminated
presence of Nosema in the worker samples, the
queens and treated the F1 queens with CO
to initiate
2
beekeepers physically exchanged queens. This
ovary activation without mating, so that the F1 queens
breeding scheme resulted in a honeybee strain in
exclusively produced unfertilized eggs developing into
which Nosema infections are rarely found and
drones. A single F1 queen, who first started to lay eggs,
individual bees showed a high tolerance towards
was chosen to produce the mapping population. The
experimental N. ceranae infections (Huang et al.
drones were reared in drone frames in full-sized colonies.
2012
). Since the biological mechanisms underly-
ing the colony level tolerance are unknown so far,
2.2. N. ceranae infection
the identification of associated genes (quantitative
trait locus, QTL) that had been selected by the
Workers freshly infected with N. ceranae were
beekeepers might help to better understand the
provided by the Laboratoire de Biologie et Protection
actual tolerance mechanisms.
de l’abeille, INRA Avignon France, as the source of
In the honeybee, QTL mapping has been widely
N. ceranae spores for subsequent infection. The
used to address quantitative traits and complex
abdomens of infected workers were homogenized in
social behavior (Hunt et al.
1998
; Oxley et al.
distilled water, filtered through filtering paper and
2010
; Behrens et al.
2011
). In the case of Nosema
centrifuged at 3,220×g for 10 min. The pellet was
tolerance, we can take advantage of haploid drones
re-dissolved in distilled water and centrifuged at
for QTL mapping because the trait is also
8,700×g for5mintopurifytheN. ceranae
expressed in the male sex. The male haploid
spores. Spores were counted using a Fuchs–
genetic system is particularly suited for QTL
Rosenthal hemocytometer and the Nosema species
mapping studies as interpretational problems
was verified by a standard PCR protocol (Hamiduzzaman
resulting from dominance interactions between
et al.
2010
).
the alleles on different homologous chromosomes
Frames of sealed drone brood of the single F1
cannot occur. We here used the drone offspring of a
hybrid queen were kept in an incubator (34±1 °C,
single hybrid queen which resulted from a cross of
60 % rel. humidity). Freshly emerged drones were
the queen of the Nosema-resistant selected Danish
collected daily from the brood frames to provide age
strain with drones of an unselected French strain to
standardized individuals (0–24 h). Freshly emerged

النتائج (العربية) 1:

الشروط العقلية (زاندر1909; شنتال.2009).تعريف قتلس المرتبطة بانخفاض ceranae أ.وفي المقابل، أ. ceranae، الذي عثر عليه أصلاًتحميل بوغ. وفي خطوة لاحقة، نحن كمياًفي سرنا نحل العسل الآسيوي ألف (فرايز et al.عام 1996),مستوى التعبير للجينات المرشحة المحددة فيطفيلي المنشأ حديثا من A. mellifera (فرايزتحليل QTL في نفس وضع خرائط السكان.وآخرون.2006; Higes et al.2006). على الرغم من أن هناكالتقارير التي تفيد بأن ceranae أ. كانت عالية2-المواد والأساليبالفوعة في A. mellifera في كل مستعمرةالمستوى الفردي (Higes et al.2008,2009)، أكثر2-1 دور فعال التلقيح للملكةتشير الدراسات الأخيرة إلى ضراوة معتدلة من أ.النحلceranae مماثلة لتلك التي أ. واجهات برمجة التطبيقات (فورسجرين والبطاطا المقلية2010; جيسديريتال.2010).كانت الملكات العذراء تسعة من سلالة الدانمركية مختارةوفي الدانمرك، كانت مستعمرات نحل العسلقدمتها إدارة إدارة "الآفات المتكاملة"-المحدد لعدم وجود التهابات المسقمهمنه مركز البحوث فلاكيبجيرج، الدانمرك. أن(نوسيموسيس) للعقود (Traynor2008). ضمنكانت تلقح الملكات كلها مصطنعة مع نفسهنظام تربية النحل الدانمركية، وعاملالحيوانات المنوية مختلطة (موريتزعام 1984) من 30 طائرات بدون طيار لغير محددواستخدمت عينات من المستعمرات تحديدسلالة أبقى في بيولوجي دي مختبر et de الحمايةمستوى العدوى المسقمه. إذا تم العثور على العيناتلعبيلي، "معهد أفينيون، فرنسا". تلقحinfected by Nosema, the queen of the colonyqueens were introduced into small colonies composedwould be replaced with one reared from a colonyof 4,000 freshly emerged workers at the apiary of thewithout Nosema infection. Although this mode ofMartin-Luther-University Halle-Wittenberg, Germany.selection was at the colony level based on theWe reared F1 hybrid queens from the inseminatedpresence of Nosema in the worker samples, thequeens and treated the F1 queens with COto initiate2beekeepers physically exchanged queens. Thisovary activation without mating, so that the F1 queensbreeding scheme resulted in a honeybee strain inexclusively produced unfertilized eggs developing intowhich Nosema infections are rarely found anddrones. A single F1 queen, who first started to lay eggs,individual bees showed a high tolerance towardswas chosen to produce the mapping population. Theexperimental N. ceranae infections (Huang et al.drones were reared in drone frames in full-sized colonies.2012). Since the biological mechanisms underly-ing the colony level tolerance are unknown so far,2.2. N. ceranae infectionthe identification of associated genes (quantitativetrait locus, QTL) that had been selected by theWorkers freshly infected with N. ceranae werebeekeepers might help to better understand theprovided by the Laboratoire de Biologie et Protectionactual tolerance mechanisms.de l’abeille, INRA Avignon France, as the source ofIn the honeybee, QTL mapping has been widelyN. ceranae spores for subsequent infection. Theused to address quantitative traits and complexabdomens of infected workers were homogenized insocial behavior (Hunt et al.1998; Oxley et al.distilled water, filtered through filtering paper and2010; Behrens et al.2011). In the case of Nosemacentrifuged at 3,220×g for 10 min. The pellet wastolerance, we can take advantage of haploid dronesre-dissolved in distilled water and centrifuged atfor QTL mapping because the trait is also8,700×g for5mintopurifytheN. ceranaeexpressed in the male sex. The male haploidspores. Spores were counted using a Fuchs–genetic system is particularly suited for QTLRosenthal hemocytometer and the Nosema speciesmapping studies as interpretational problemswas verified by a standard PCR protocol (Hamiduzzamanresulting from dominance interactions betweenet al.2010).the alleles on different homologous chromosomesFrames of sealed drone brood of the single F1cannot occur. We here used the drone offspring of ahybrid queen were kept in an incubator (34±1 °C,single hybrid queen which resulted from a cross of60 % rel. humidity). Freshly emerged drones werethe queen of the Nosema-resistant selected Danishcollected daily from the brood frames to provide agestrain with drones of an unselected French strain tostandardized individuals (0–24 h). Freshly emerged