TABLE 3.
Initial resistances in soil, manure slurry, and hydrated lime alkaline stabilized (HLAS) manure slurry assessed by multiplex PCR of tetracycline resistance genes and quantitative PCR of select resistance elements.
| Mulitplex PCR, % positive† | Soil | Raw manure | HLAS-treated manure | Resistance mechanism/action |
| tet(A) | 0 | 100 | 100 | Efflux pump |
| tet(B) | 0 | 100 | 100 | Efflux pump |
| tet(C) | 0 | 100 | 100 | Efflux pump |
| tet(D) | 100 | 0 | 0 | Efflux pump |
| tet(E) | 0 | 0 | 0 | Efflux pump |
| tet(G) | 100 | 100 | 100 | Efflux pump |
| tet(K) | 0 | 0 | 0 | Efflux pump |
| tet(L) | 100 | 0 | 0 | Efflux pump |
| tet(A/P) | 0 | 0 | 0 | Efflux pump |
| tet(M) | 0 | 100 | 100 | Ribosome protection |
| tet(O) | 0 | 100 | 100 | Ribosome protection |
| tet(Q) | 0 | 100 | 100 | Ribosome protection |
| tet(S) | 0 | 0 | 0 | Ribosome protection |
| tet(X) | 0 | 100 | 100 | Antibiotic metabolism |
| Quantitative PCR‡ | ||||
| Per gram dry matte2r | ||||
| 16S rRNA gene | 8.42 (0.08) A | 8.43 (0.02) A | 8.68 (0.04) B | Protein synthesis |
| intI1 | 3.48 (0.10) A | 6.11 (0.02) B | 6.15 (0.01) B | Integrase gene |
| blaCTX–M–32 | 3.80 (0.27) A | 5.03 (0.03) B | 4.76 (0.01) C | Antibiotic metabolism/ring cleavage |
| sul(I) | 4.39 (0.07) A | 6.70 (0.03) B | 6.84 (0.03) C | Alternative dihydropteroate synthase |
| erm(B) | <1.7 A | 6.65 (0.04) B | 7.15 (0.02) C | Ribosome protection/dimethylation |
| Log abundance normalized per 16S rRNA gene | ||||
| intI1 | −4.94 (0.05) A | −2.31 (0.04) B | −2.53 (0.05) B | |
| blaCTX–M–32 | −4.78 (0.29) A | −3.40 (0.04) B | −3.92 (0.10) C | |
| sul(I) | −4.02 (0.05) A | −1.73 (0.05) B | −1.84 (0.07) B | |
| erm(B) | <−6.70 A | −1.78 (0.06) B | −1.53 (0.05) B | |
| Abundance normalized per106 16S rRNA gene | ||||
| intI1 | 12 A | 4900 B | 2980 B | |
| blaCTX–M–32 | 17 A | 400 B | 120 C | |
| sul(I) | 96 A | 18900 B | 14700 B | |
| erm(B) | <0.2 A | 16800 B | 29800 B | |
†Results of multiplex PCR expressed as percentage of PCR-positive soil or manure samples (n = 3).
‡For quantitative PCR, each sample was amplified three times and averaged. The average log abundance (±SE) for three samples is reported. “ABC” denotes a difference (P < 0.05) between sample types (i.e., soil, manure, and HLAS-treated manure).