Rapid quantification of Legionella in agricultural air purification systems from fattening pig houses with culture-independent methods

Abstract

Fattening pig houses often utilize biological agricultural exhaust air purification systems (APS) that employ an active microbiome to degrade nitrogen. Consequently, disinfection cannot be applied, and a neutral pH value must be maintained. However, the biofilm in biotrickling filters and a higher temperature can potentially facilitate the growth of Legionella spp. To investigate the occurrence of Legionella spp. or even the pathogen Legionella pneumophila in these systems, traditional cultivation methods proved impractical due to overgrowth and long turnaround times from sampling in the field to results in the laboratory. Therefore, innovative concepts for rapid and cultivation-independent analysis of Legionella spp. are highly demanded. In this study, two rapid analysis methods were applied using a standard addition qPCR method for the detection of L. pneumophila Sg1 and Legionella spp. as well as flow cytometry coupled with immunomagnetic separation (IMS-FCM) for the detection of viable L. pneumophila. Three APS were monitored over a period of more than a year during summer, winter, and intermediate seasons. While cultivation failed to quantify any Legionella spp., the standard addition qPCR quantified 230 to 9500 Legionella spp. cells per m³ in air passing through the APS (clean gas). In process water that is used for circulating washing of the APS a high occurrence of 10⁴ to 2.9 × 10⁵ Legionella spp. cells/mL was measured. By IMS-FCM it was confirmed that viable L. pneumophila in concentrations higher than 100 cells/mL for process water and higher than 100 cells/m³ in clean gas were found in all seasons. In contrast, Legionella spp. or L. pneumophila were rarely found in air directly from the barn (raw gas). We see no risk coming from the closed barn itself but there is a health risk, because the contamination of viable L. pneumophila in process water is not efficiently reduced in clean gas.