Inactivation, storage, and PCR detection of Mycoplasma on FTA filter paper.
We evaluated the feasibility of using Flinders Technology Associates (FTA) filter paper for the inactivation and storage of mycoplasma DNA templates and their detection by the polymerase chain reaction (PCR). FTA paper is a cotton-based cellulose membrane containing lyophilized chemicals that lyses most types of bacteria and viruses. Mycoplasma gallisepticum (MG) and Mycoplasma synoviae (MS) cultures were spotted at various volumes on the filter paper and stored at different temperatures for various periods of time before performing PCR. MG and MS were readily detected at all time frames (1-60 days) independent of the volume applied (1-100 microl) or storage temperature (4 C-41 C). Sensitivity and specificity of the FTA-PCR were comparable to the standard diagnostic PCR, allowing the detection of MG/MS in field samples without interference by nontargeted mycoplasma. Analysis of 193 field samples by both methods showed nearly 100% agreement with serology and culture results. The long-term DNA stability at a wide range of temperatures makes the FTA cards a good alternative for collecting and simultaneously inactivating mycoplasma. It also offers the convenience of storage and transport of DNA in a cost-effective manner for further molecular analysis, such as restriction enzyme length polymorphism and nucleotide sequencing.
Moscoso
Hugo
H
Poultry Diagnostic and Research Center, Department of Avian Medicine, University of Georgia, Athens, GA 30602-4875, USA.
Thayer
Stephan G
SG
Hofacre
Charles L
CL
Kleven
Stanley H
SH
D001431:Bacteriological Techniques / Q000295:instrumentation*; D004269:DNA, Bacterial / Q000737:chemistry*; D005374:Filtration / Q000295:instrumentation*; D009174:Mycoplasma / Q000302:isolation & purification*; D009175:Mycoplasma Infections / Q000175:diagnosis / Q000662:veterinary; D010209:Paper; D016133:Polymerase Chain Reaction / Q000379:methods; D012150:Polymorphism, Restriction Fragment Length; D012680:Sensitivity and Specificity; D013048:Specimen Handling / Q000295:instrumentation
D004269:DNA, Bacterial