Cost-effective bead-based method for high-throughput homogenization of individual small arthropods.

of vector-borne diseases in both the hosts and vectors of these agents. A pre-requisite for such analyses is effective methods for the extraction of high quality, inhibition-free nucleic acids. Arthropods are the vectors of multiple infectious disease agents and the extraction of nucleic acids from arthropods requires adequate disruption of the exoskeleton and maceration of tissues to release nucleic acids from the cells. Automation of small arthropod maceration is particularly problematic as these specimens have buoyant bodies and hydrophobic exoskeletons which make them difficult targets for standard beads used in many mechanical tissue disruption methods. Incomplete homogenization results in reduced nucleic acid yields and limits the ability to detect infecting organisms. The use of efficient high-throughput processing techniques for disease vectors is critical for the screening of field collected or laboratory test specimens. The homogeneous disruption of individual small arthropod vectors of zoonotic diseases such as biting midges (Diptera), fleas (Siphonaptera), or ticks (Ixodida) is typically done manually with pestles in microcentrifuge tubes, or with Dounce tissue grinders1–3. Both the methods are labor intensive and can result in: (a) inconsistent sample homogenization; (b) significant volume loss due to adhesion of sample and buffer to the pestle that is then removed from the tube; and (c) introduce the possibility of sample cross contamination as homogenization takes place in open tubes. Other methods capable of homogenizing single small arthropod in high-throughput formats include mechanical agitation with either silica (400 to 800 μm) or zirconium (200 μm) beads4, 5, solid stainless steel ball bearings6, 7, tungsten carbide beads (Qiagen)8, and goldplated tungsten hollow core beads9 (Spirit River Inc.). The silica and zirconium beads require multiple beads per well and vigorous agitation. Use of single solid tungsten carJ Vector Borne Dis 50, March 2013, pp. 62–64