Effect of storage conditions from the lone star tick Amblyomma americanum samples in RNA extraction quality

The lone star tick (Amblyomma americanum) is widely distributed in the eastern and central USA and Mexico. A. americanum is a known vector of a number of pathogens, including Ehrlichia chaffeensis, E. ewingii, Francisella tularensis, Rickettsia parkeri, Heartland virus, Bourbon virus, and Southern tick-associated rash illness (STARI), that have significant impacts on human and animal health. Sampling of tick populations is vital to understand the prevalence of these pathogens, but the conditions in which samples are stored from collection through processing and analysis can dramatically affect the accuracy of detection of microorganisms. Thus, in this work, we compared the concentration and quality of RNA extracted from non-fed adult A. americanum ticks stored either dry or in DNA/RNA-shield at different temperatures (Room-T°/4°C/-20°C/80°C) to determine optimal conditions for collecting high-quality nucleic acid samples for downstream applications. Nine female and 17 male ticks were obtained from the Oklahoma State tick rearing facility and kept in different storage conditions before RNA extraction (between one week and ten days after storage). Our results indicate that the RNA quality and purity from ticks stored in DNA/RNA-shield were significantly higher, independent of temperature, compared to ticks that were dry-preserved at different temperatures. Also, cold temperatures (-20°C/-80°C) tended to increase the purity and quantity of isolated RNAs even without DNA/RNA-shield. Our results show that using an RNA preservation solution increases RNA quality; however, in the absence of this product, maintaining the specimens in colder temperatures improves the quality of nucleic acid isolations.