Longitudinal characterization of antimicrobial resistance genes in feces shed from cattle fed different subtherapeutic antibiotics.
BACKGROUND
Environmental transmission of antimicrobial-resistant bacteria and resistance gene determinants originating from livestock is affected by their persistence in agricultural-related matrices. This study investigated the effects of administering subtherapeutic concentrations of antimicrobials to beef cattle on the abundance and persistence of resistance genes within the microbial community of fecal deposits. Cattle (three pens per treatment, 10 steers per pen) were administered chlortetracycline, chlortetracycline plus sulfamethazine, tylosin, or no antimicrobials (control). Model fecal deposits (n = 3) were prepared by mixing fresh feces from each pen into a single composite sample. Real-time PCR was used to measure concentrations of tet, sul and erm resistance genes in DNA extracted from composites over 175 days of environmental exposure in the field. The microbial communities were analyzed by quantification and denaturing gradient gel electrophoresis (DGGE) of PCR-amplified 16S-rRNA.


RESULTS
The concentrations of 16S-rRNA in feces were similar across treatments and increased by day 56, declining thereafter. DGGE profiles of 16S-rRNA differed amongst treatments and with time, illustrating temporal shifts in microbial communities. All measured resistance gene determinants were quantifiable in feces after 175 days. Antimicrobial treatment differentially affected the abundance of certain resistance genes but generally not their persistence. In the first 56 days, concentrations of tet(B), tet(C), sul1, sul2, erm(A) tended to increase, and decline thereafter, whereas tet(M) and tet(W) gradually declined over 175 days. At day 7, the concentration of erm(X) was greatest in feces from cattle fed tylosin, compared to all other treatments.


CONCLUSIONS
The abundance of genes coding for antimicrobial resistance in bovine feces can be affected by inclusion of antibiotics in the feed. Resistance genes can persist in feces from cattle beyond 175 days with concentrations of some genes increasing with time. Management practices that accelerate DNA degradation such as frequent land application or composting of manure may reduce the extent to which bovine feces serves as a reservoir of antimicrobial resistance.
Agriculture and Agri-Food Canada Research Centre, Lethbridge, Alberta T1J 4B1, Canada.
Trevor W
TW
Alexander
Jay L
JL
Yanke
Tim
T
Reuter
Ed
E
Topp
Ronald R
RR
Read
Brent L
BL
Selinger
Tim A
TA
McAllister
D000821:Animal Feed; D000818:Animals; D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D001426:Bacterial Proteins / Q000235:genetics; D002417:Cattle; D002751:Chlortetracycline / Q000494:pharmacology; D025141:Drug Resistance, Fungal / Q000235:genetics*; D005243:Feces / Q000382:microbiology*; D016133:Polymerase Chain Reaction; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D013418:Sulfamethazine / Q000494:pharmacology; D015645:Tylosin / Q000494:pharmacology
D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D012336:RNA, Ribosomal, 16S; D013418:Sulfamethazine; D002751:Chlortetracycline; D015645:Tylosin
Appl Environ Microbiol. 2009 Nov;75(22):7125-34
19801481
FEMS Microbiol Lett. 2005 Apr 15;245(2):195-203
15837373
J Environ Qual. 2006 Aug 09;35(5):1844-54
16899756
J Sci Food Agric. 2010 Mar 15;90(4):650-7
20355094
Environ Microbiol. 2005 Jul;7(7):933-46
15946290
Nutr Res Rev. 2000 Dec;13(2):279-99
19087443
Appl Environ Microbiol. 2006 Jul;72(7):4583-8
16820447
J Anim Sci. 2008 Sep;86(9):2385-91
18469042
Antimicrob Agents Chemother. 1999 Dec;43(12):2823-30
10582867
Microb Ecol. 2006 Apr;51(3):267-76
16598633
Antimicrob Agents Chemother. 1991 Jul;35(7):1273-6
1929281
Appl Environ Microbiol. 1996 Jul;62(7):2273-8
8779565
Appl Environ Microbiol. 2007 Jan;73(1):278-88
17071787
BMC Microbiol. 2009 Oct 02;9:210
19799792
Appl Environ Microbiol. 2008 Jul;74(14):4405-16
18502931
Microb Drug Resist. 2009 Sep;15(3):159-66
19728772
Environ Microbiol. 2007 Jan;9(1):143-51
17227419
Appl Environ Microbiol. 1999 Nov;65(11):4799-807
10543789
J Environ Qual. 2009 Apr 27;38(3):1086-108
19398507
Appl Environ Microbiol. 2007 Jul;73(14):4407-16
17496134
J Microbiol Methods. 2007 Sep;70(3):565-9
17683816
Environ Microbiol. 2007 Mar;9(3):657-66
17298366
ISME J. 2007 Jul;1(3):191-203
18043630
Chemosphere. 2005 Apr;59(4):457-65
15788168
Br J Pharmacol. 2008 Mar;153 Suppl 1:S347-57
18193080
Microbiology (Reading). 2002 Jan;148(Pt 1):257-266
11782518
Can J Public Health. 2005 Jul-Aug;96(4):254-8
16625790
FEMS Microbiol Ecol. 2002 Jan 1;39(1):33-9
19709182
J Environ Qual. 2007 Oct 16;36(6):1695-703
17940270
Mol Cell Probes. 2001 Aug;15(4):209-15
11513555
Appl Environ Microbiol. 2008 Oct;74(20):6178-86
18723654
Int J Antimicrob Agents. 2000 May;14(4):327-35
10794955
Appl Environ Microbiol. 2007 Dec;73(24):7917-25
17951435
Appl Environ Microbiol. 2007 Jan;73(1):193-202
17085701
Vet Microbiol. 2003 Jan 2;91(1):73-84
12441233
Environ Microbiol. 2007 Mar;9(3):703-15
17298370
Appl Environ Microbiol. 2007 Sep;73(17):5486-93
17616622
Appl Environ Microbiol. 2008 Aug;74(16):5063-7
18552189