Risk assessment and monitoring of Legionella by culture and q-PCR in a newly built block of flats associated with a small outbreak of legionnaires´ disease

During the investigation of a small outbreak of legionnaires’ disease in a newly built block of flats, several risk factors were uncovered. The outbreak encompassed two men (age 44 and 65) of which the oldest one died. Two interventions (heat treatments of 70ºC for 12 and 24 hours respectively) and permanent changes of the flow and temperature were conducted to overcome the high concentrations of Legionella in the water. Water samples (104 in total, both cold and warm water) from taps and shower hoses were collected and cultivated. The samples consisted of: A samples: the first one litre, B samples: one litre collected after flushing until constant temperature and water from shower hoses at a temperature of 38ºC. The survey pointed at important risk factors 1) low temperature of the hot water in some of the most distant taps 2) low flow of the water circulating system because of too small pipe dimensions and too low pump capacity 3) high numbers of Legionella in the shower hoses 4) stagnancy of water before residents move in and in unoccupied apartments and 5) lack of or inappropriate control measures. Following cultivation, DNA from all water samples was extracted by a simple method with Chelex-100. The samples will be investigated by an in-house q-PCR (Quantitative Real Time PCR). The q-PCR is a Taq-Man based assay with 5S primers detecting Legionella spp and mip primers detecting Legionella pneumophila. The two Legionella q-PCR assays have been validated according to the Afnor standard (T90 471) with standards calibrated according to the international standard from Legionelles centre National de Référence (SRM_LEGDNA_01 and CQE_LEGDNA_01). The results from culture (CFU/L) and PCR (GU/L) will be compared, and on this background it will be evaluated if q-PCR could have been used for fast risk assessment in the initial phase of the investigation and to follow the effect of the interventions in the later phase of the investigation.