Today, we are happy to share a new publication! Dr. Nirmala and colleagues from the University of Minnesota report on a viral RNA extraction method that can be used to detect both the porcine reproductive and respiratory syndrome virus (PRRSV) and influenza A virus (IAV) from used farm air filters. This work is published in the Journal of Aerosol Science. Take a look at it!
- Extracting RNA using TRIzol reagent from filter material ground with liquid nitrogen can be used to detect PRRSV and IAV from air filters.
- IAV was detected in more filters than PRRSV, though PRRSV could be detected in used filters from farms with PRRSV negative status at the time of filter removal.
- The obtained results shows likely evidence of between farm aerosol spread and the methods derived from this study open up avenues to further investigate regional airborne transmission and risk of virus introduction into farms.
It is of great interest to identify and characterize the viruses that are captured by air filters in swine farms. The biggest challenge in identifying the viruses from the filters is to efficiently sample and elute the viral particles for RNA extraction. Therefore, the main goal of this study was to evaluate the feasibility of identifying PRRSV and IAV on used air filters retrieved from swine farms. The objectives of this study were to 1) identify the best combination of sample elution and RNA extraction methods to detect PRRSV from Minimum Efficiency Reporting Values (MERV) filters aerosolized with PRRSV under experimental conditions and 2) exploit the best method identified to detect PRRSV and IAV from used filters from farms located in pig dense areas of the Midwestern USA.
Materials and Methods
A first pilot study was run to evaluate if PRRSV could be identified from MERV14 filters and understand the detectable limits. With this purpose, one square inch area of brand new MERV14 filters were spiked with 1000 μL of varying dilutions (10^-1 to 10^-8) of VR-2332 PRRSV reference strain. Filters were subjected to four different methods to elute the viral particles and RNA was extracted using TRIzol reagent. A second pilot study was run to identify aerosolized PRRSV in the filters. In this case, the filters were loaded by aerosolizing three different concentrations of a VR-2332 derived PRRSV modified live vaccine virus into MERV14 filters. In this case, the samples were subjected to three different elution methods and two different RNA extraction strategies. Lastly, a set of 44 filters, originating from 13 different farms and 4 different agricultural companies were tested for both PRRSV and IAV. RNA was extracted using TRIzol reagent.
Please, read the full manuscript for details of the materials and procedures used.
Results and Discussion
From the first pilot study, PRRSV was successfully detected by all the four elution methods namely elbow shake, vortex, filter thawing and grinding with liquid nitrogen. This is not surprising since the filters were infiltrated with a high concentration of the virus inoculum. However, grinding filters with liquid nitrogen was reported as the best strategy to detect PRRSV from air filters, as it yielded lower Ct values. Afterwards, three elution methods (freeze thaw, vortex, and liquid nitrogen grinding) were evaluated by aerosolizing PRRSV into MERV14 commercial filters to mimic how air filters become loaded under normal operating conditions. PRRSV was detected by all the elution and extraction methods. Nevertheless, the liquid nitrogen/TRIzol combination had the lowest Ct for all three vaccine dilutions used. Then, exploiting the liquid nitrogen grinding + TRIzol combination, a set of 44 used filters from swine farms were evaluated for PRRSV and IAV. Both PRRSv and IAV were identified from MERV14, 15 and 16 rated filters. PRRSV was identified in 27% (12/44) of the used filters from 31% (4/13) of the farms, while IAV was identified in 66% (29/44) of the filters from 77% (10/13) of the farms. Interestingly, PRRSV was also detected on used filters from farms with PRRSv negative status at the time of filter removal.
To read the results in more detail, refer to the full manuscript here.
Filtering the air entering swine farms has significantly reduced the incidence of porcine reproductive and respiratory syndrome virus (PRRSV) infections in Midwestern breeding herds. Despite the significant investment made in installation of air filters, the nature and type of viruses trapped by these filters are not yet clear because of the unavailability of reliable methods to elute and detect these viruses. Here we report that eluting viral particles from the air filters by grinding filter specimens with liquid nitrogen, coupled with TRIzol reagent to extract RNA can detect both PRRSV and influenza A virus (IAV) from used minimum efficiency reporting value (MERV) 14, 15 and 16 rated air filters. PRRSv was detected in 27% (12/44) and IAV was detected in 66% (29/44) of filters that had been in installation between 08/12/13 and 07/12/17. Interestingly, PRRSV was also detected on used filters from farms with PRRSv negative status at the time of filter removal. Presence of PRRSv and IAV in the used filters show likely evidence of between herd aerosol spread for these viruses. The methods derived from this study open up avenues to further investigate airborne transmission and risk of virus introduction into farms contributing to the control of diseases in swine.