63ª Reunião Anual da SBPC

C. Ciências Biológicas - 10. Microbiologia - 3. Microbiologia

Evaluation of Modified Moore Swab and Tangential Flow Filtration for Concentration of Salmonella spp. and Escherichia coli O157:H7 from Larges Volumes of Water

Camilla Alves Pereira Rodrigues 1 Michelle D. Danyluk 2

1. Especialista- Lab. de Controle Higiênico Sanitário de Alimentos- FANUT/UFG 2. Profa. Dra./Orientadora-Citrus Research & Education Center- UF/IFAS

INTRODUÇÃO:

This study evaluated and compared the efficacy of Modified Moore Swabs (MMS) and Tangential Flow Filtration (TFF) for concentration and subsequent detection of Escherichia coli O157:H7 and Salmonella spp. from large volumes of water (10L). Saline was spiked with a 3 strain cocktail of either Escherichia coli O157:H7 or Salmonella spp., all of which were previously transformed to express resistance to ampicillin and a fluorescence protein. The objective this study was evaluate the efficacy of two filters methods for concentrations of E. coli O157:H7 and Salmonella spp. in a saline model system for agricultural and produce wash water.

METODOLOGIA:

A three strain cocktail of Escherichia coli O157:H7 (MDD36pGFP, MDD37DSRed and MDD38pAmCyan) and a three strain cocktail of Salmonella spp. (MDD22pGFP,MDD237pDsRed, and MDD314pAMCyan) were prepared. The cocktail was prepared after take the strains from freezer - 80°C put in plates TSA+AMP (Trypticase Soy Agar and Ampicillin concentration 20mg/l) for 24h/37°C. After this pick the bacteria for TSB+AMP (Trypticase Soy Broth and Ampicillin concentration 20mg/l) for 24h/37°C and pick again in TSB+AMP for 24h/37°C. After this, was take 1 ml each E. coli spp or Salmonella spp put in Vortex ® for 10 seconds. Inoculum levels used were:101,102,103 and 104 CFU/100ml. One ml of inoculum in lambda buffer was added to 10 l of non-sterile normal saline. Two filter types were used: tangential flow filter (TFF), and modified moore swab(MMS). Samples were run through (300 ml min-1); subsequently the gauze was removed, placed in a stomacher bag and squeezed from the outside to release a maximum volume of 25-35 ml. Retentate water was passed through a Millipore filter. The filters were placed on TSA+AMP plates, and incubated 24h/37C. For two lowest spiking levels, 100 µL will be transferred to 5 mL TSB+AMP incubated and tested via Reveal® Salmonella or E. coli O157:H7 lateral flow device.

RESULTADOS:

For E. coli inoculum levels 10 1 and 10 3 had the same behavior where MMS were positive and only one positive. For the inoculum102 two out of the three of the samples positive for both MMS and for TF. All samples were positive for MMS and TF for the inoculum 104. Salmonella spp. For the inoculum 104 all samples were positive for MMS and TF. In inoculums 10 1,102 and 103 the two out of the three samples tested were positive for the filter MMS. No sample was positive for the inoculum 10 1 for the filter TF. Only one sample was positive in 102 for the filter TF. All samples were positive for inoculum 10 3 for TF.

CONCLUSÃO:

In this study we evaluated MMS and TFF for concentration of Salmonella spp. and E. coli O157: H7 from large volumes of water, and established they were equally efficient for concentration of these two pathogens.

Palavras-chave: Salmonella spp., E. coli, water.