Differential diagnosis of the honey bee trypanosomatids Crithidia mellificae and Lotmaria passim

Trypanosomatids infecting honey bees have been poorly studied with molecular methods until recently.
After the description of Crithidia mellificae (Langridge and McGhee, 1967) it took about forty years until
molecular data for honey bee trypanosomatids became available and were used to identify and describe a
new trypanosomatid species from honey bees, Lotmaria passim (Evans and Schwarz, 2014). However, an
easy method to distinguish them without sequencing is not yet available. Research on the related bumble
bee parasites Crithidia bombi and Crithidia expoeki revealed a fragment length polymorphism in the inter-
nal transcribed spacer 1 (ITS1), which enabled species discrimination. In search of fragment length poly-
morphisms for differential diagnostics in honey bee trypanosomatids, we studied honey bee
trypanosomatid cell cultures of C. mellificae and L. passim. This research resulted in the identification of
fragment length polymorphisms in ITS1 and ITS1-2 markers, which enabled us to develop a diagnostic
method to differentiate both honey bee trypanosomatid species without the need for sequencing.
However, the amplification success of the ITS1 marker depends probably on the trypanosomatid infection
level. Further investigation confirmed that L. passim is the dominant species in Belgium, Japan and
Switzerland. We found C. mellificae only rarely in Belgian honey bee samples, but not in honey bee sam-
ples from other countries. C. mellificae was also detected in mason bees (Osmia bicornis and Osmia cor-
nuta) besides in honey bees. Further, the characterization and comparison of additional markers from
L. passim strain SF (published as C. mellificae strain SF) and a Belgian honey bee sample revealed very
low divergence in the 18S rRNA, ITS1-2, 28S rRNA and cytochrome b sequences. Nevertheless, a variable
stretch was observed in the gp63 virulence factor.


Jorgen Ravoet, Ryan S. Schwarz, Tine Descamps, Orlando Yañez, Cansu Ozge Tozkar, Raquel Martin-Hernandez,  Carolina Bartolomé, Lina De Smet, Mariano Higes , Tom Wenseleers, Regula Schmid-Hempel,  Peter Neumann, Tatsuhiko Kadowaki, Jay D. Evans, Dirk C. de Graaf (2015). Differential diagnosis of the honey bee trypanosomatids Crithidia mellificae and Lotmaria passim. Journal of Invertebrate Pathology 130 (2015) 21–27.

Supported by

Ricola Foundation


University of Bern

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