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\begin{document}

\title{Characterization and microsatellite marker development for a common bark
and ambrosia beetle associate, Geosmithia obscura}



\author[1]{Grace  Pietsch}%
\author[1]{William Klingeman}%
\author[1]{Matthew Huff}%
\author[2]{Miroslav Kolarik}%
\author[1]{Denita Hadziabdic}%
\author[3]{Romina Gazis}%
\author[4]{Margaret (Meg) Staton}%
\affil[1]{The University of Tennessee}%
\affil[2]{Czech Academy of Sciences}%
\affil[3]{University of Florida}%
\affil[4]{University of Tennessee}%


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  \date{May 23, 2024}


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\begin{abstract}
Background. Symbioses between Geosmithia fungi and wood-boring and bark
beetles seldom result in disease induction within the plant host. Yet
exceptions exist such as Geosmithia morbida, the causal agent of
Thousand Cankers Disease (TCD) of walnuts and wingnuts, and Geosmithia
sp. 41, the causal agent of Foamy Bark Canker disease of oaks. Isolates
of G. obscura were recovered from black walnut trees in eastern
Tennessee and at least one isolate induced cankers following artificial
inoculation. Due to the putative pathogenicity and lack of recovery of
G. obscura from natural lesions, a molecular diagnostic screening tool
was developed using microsatellite markers mined from the G. obscura
genome. Results. A total of 3,256 candidate microsatellite markers were
identified (2236, 789, 137 di-, tri-, and tetra- motifs were identified,
respectively), with 2011, 703, 101 di-, tri-, and tetra- motifs
containing markers with primers. From these, 75 microsatellite markers
were randomly selected, screened, and optimized, resulting in 28
polymorphic markers that yielded single, consistently recovered bands
which were used in downstream analyses. Five of these microsatellite
markers were found to be specific to G. obscura and did not
cross-amplify into other, closely related species. Although the
remaining tested markers could be useful, they cross-amplified within
different Geosmithia species, making them not reliable for G. obscura
detection. Conclusion. Five novel microsatellite markers (GOBS9, GOBS10,
GOBS41, GOBS43, GOBS50) were developed based on the G. obscura genome.
These species-specific microsatellite markers are available as a tool
for use in molecular diagnostics and can assist future surveillance
studies.%
\end{abstract}%



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