TY - JOUR
T1 - Determining the optimal number of individual samples to pool for quantification of average herd levels of antimicrobial resistance genes in Danish pig herds using high-throughput qPCR
AU - Clasen, Julie
AU - Mellerup, Anders
AU - Olsen, John Elmerdahl
AU - Angen, Øystein
AU - Folkesson, Anders
AU - Halasa, Tariq
AU - Toft, Nils
AU - Birkegard, Anna Camilla
PY - 2016/6/30
Y1 - 2016/6/30
N2 - The primary objective of this study was to determine the minimum number of individual fecal samples to pool together in order to obtain a representative sample for herd level quantification of antimicrobial resistance (AMR) genes in a Danish pig herd, using a novel high-throughput qPCR assay. The secondary objective was to assess the agreement between different methods of sample pooling. Quantification of AMR was achieved using a high-throughput qPCR method to quantify the levels of seven AMR genes (ermB,ermF, sulI, sulII, tet(M), tet(O) and tet(W)). A large variation in the levels of AMR genes was found between individual samples. As the number of samples in a pool increased, a decrease in sample variation was observed. It was concluded that the optimal pooling size is five samples, as an almost steady state in the variation was observed when pooling this number of samples. Good agreement between different pooling methods was found and the least time-consuming method of pooling, by transferring feces from each individual sample to a tube using a 10 mu l inoculation loop and adding 3.5 ml of PBS, approximating a 10% solution, can therefore be used in future studies.
AB - The primary objective of this study was to determine the minimum number of individual fecal samples to pool together in order to obtain a representative sample for herd level quantification of antimicrobial resistance (AMR) genes in a Danish pig herd, using a novel high-throughput qPCR assay. The secondary objective was to assess the agreement between different methods of sample pooling. Quantification of AMR was achieved using a high-throughput qPCR method to quantify the levels of seven AMR genes (ermB,ermF, sulI, sulII, tet(M), tet(O) and tet(W)). A large variation in the levels of AMR genes was found between individual samples. As the number of samples in a pool increased, a decrease in sample variation was observed. It was concluded that the optimal pooling size is five samples, as an almost steady state in the variation was observed when pooling this number of samples. Good agreement between different pooling methods was found and the least time-consuming method of pooling, by transferring feces from each individual sample to a tube using a 10 mu l inoculation loop and adding 3.5 ml of PBS, approximating a 10% solution, can therefore be used in future studies.
KW - Pooling
KW - Antimicrobial resistance gene
KW - Pig
KW - High-throughput qPCR
KW - Sample size
KW - Herd level
U2 - 10.1016/j.vetmic.2016.04.017
DO - 10.1016/j.vetmic.2016.04.017
M3 - Journal article
C2 - 27259826
SN - 0378-1135
VL - 189
SP - 46
EP - 51
JO - Veterinary Microbiology
JF - Veterinary Microbiology
ER -