Transmission of Antimicrobial-Resistant Staphylococcus Aureus Clonal Complex 9 Between Pigs and Humans, United States

Pranay R. Randad; Jesper Larsen; Hülya Kaya; Nora Pisanic; Carly Ordak; Lance B. Price; Maliha Aziz; Maya L. Nadimpalli; Sarah Rhodes; Jill R. Stewart; Dave C. Love; David Mohr; Meghan F. Davis; Lloyd S. Miller; Devon Hall; Karen C. Carroll; Trish M. Perl; Christopher D. Heaney

Disclosures

Emerging Infectious Diseases. 2021;27(3):740-748. 

In This Article

Results

All 49 isolates from the North Carolina collection used in WGS analysis were classified as sequence type 9 by MLST. Among 81 S. aureus CC9 isolates analyzed, 95% (77/81) were located in 3 major clades, C1, C2, and C3 (Figure 1; Appendix Tables 3, 4). Despite the small number of pig isolates, each clade contained both pig and human isolates (Figure 1; Appendix Table 3).

Figure 1.

Maximum-likelihood tree demonstrating population structure of Staphylococcus aureus clonal complex (CC) 9 isolates from humans and livestock in North Carolina, USA, and reference sequences. A total of 81 S. aureus CC9 isolates from human and livestock specimens were included in this midpoint-rooted maximum-likelihood phylogeny based on 3,847 core genome single-nucleotide polymorphisms. S. aureus isolates belonged to 3 phylogeographically distinct clades (C1–C3). All the North Carolina collection isolates were included in C3. IEC genes are shown in columns 1, scn; 2, sak; and 3, chp. MRSA is shown in column 4. AMR genes are shown in columns 5, mecA; 6, tet(K); 7, tet(L); 8, tet(T); 9, erm(A); 10, erm(B); 11, erm(C); 12, vga(A)LC; 13, lnu(A); 14, lnu(B); 15, str; 16, spc; 17, aadD; 18, aac(6); 19, ant(6)-1a; 20, dfrG; and 21, dfrK. Scale bar indicates nucleotide substitutions per site. AMR, antimicrobial resistance; Chick, chicken; COO, country of origin; IEC, immune evasion cluster; MRSA, methicillin-resistant S. aureus; MSSA, methicillin-susceptible S. aureus; NA, not applicable.

Among C1–C3 isolates, 61% (47/77) contained tetracycline resistance genes. By contrast, only 1 (1.3%) of the isolates in C1–C3 contained IEC genes (Figure 1; Appendix Table 4). The presence of pig isolates coupled with the absence of IEC genes and presence of tetracycline resistance genes in C1–C3 suggest that C1–C3 isolates may be members of a larger LA-SA CC9 clade. LA-MRSA CC9, which harbored the mecA gene, was present in C1 and C2 but absent from C3.

C1 was composed of isolates primarily originating from Asia (12/13 isolates; 92%), of which 46% (6/13) were from China and 46% (6/13) were from Taiwan (Figure 1; Appendix Table 3). All of C2 (14/14 isolates) was composed of isolates originating from Europe, of which 71% (10/14) were from Germany, 21% (3/14) were from the Netherlands, and 7% (1/14) were from Denmark (Figure 1; Appendix Table 3). C3 included 100% (49/49) of the North Carolina isolates, which made up 98% (49/50) of all C3 isolates (Figure 1; Appendix Table 3). Only 2 isolates grouped into a clade that did not correspond to the continent of predominance within the clade. A single isolate from Colombia (South America) grouped into C3 with isolates from North Carolina, and a single isolate from the Netherlands grouped into C1 with isolates from Asia (Figure 1; Appendix Table 3).

High-resolution phylogenetic analysis of C3 revealed multiple distinct subclades, one of which contained all 6 IHO-1 pig isolates from North Carolina (Figure 2). The pairwise SNP distance among IHO pig isolates from IHO-1 ranged from 0–43 SNPs. Thus, we used 43 SNPs as the threshold to identify putative transmission clusters and found that 19 isolates fell into 2 distinct putative transmission clusters (Figure 2). The minimum pairwise SNP distance between IHO pig and human isolates within putative transmission clusters ranged from 12–34 SNPs.

Figure 2.

High-resolution population structure of clade 3 livestock-associated Staphylococcus aureus clonal complex (CC) 9 isolates from humans and livestock in North Carolina, USA, and reference isolates. A subset of 50 livestock-associated S. aureus CC9 isolates that were collected from IHO pigs, IHO workers, IHO minors, and CR adults were included in this midpoint-rooted maximum-likelihood phylogeny based on 1,198 core genome single-nucleotide polymorphisms. A single subclade, denoted as the IHO pig cluster, included only pig isolates from IHO-1 and was used to set a threshold of 43 SNPs for identifying transmission clusters; clusters of IHO pig and human isolates separated by ≤43 SNPs are considered transmission clusters. Two subclades included intermingled human and IHO pig isolates with a high degree of phylogenetic relatedness and were considered transmission clusters. IEC isolates are shown in columns 1, scn, 2, sak, and 3, chp. AMR genes are shown in columns 4, tet(K); 5, tet(L); 6, tet(T); 7, erm(A); 8, erm(C); 9, vga(A)LC; 10, lnu(A); 11, spc; 12, aadD; and 13, aac(6). MDRSA is shown in column 14. Antimicrobial drug resistance is shown in columns 15, fluoroquinolone resistance, considered phenotypic resistance to moxifloxacin; 16, lincosamide resistance, considered phenotypic resistance to clindamycin; 17, macrolide resistance, considered phenotypic resistance to erythromycin; 18, aminoglycoside resistance, considered phenotypic resistance to gentamicin; 19, tetracycline resistance, considered phenotypic resistance to tetracycline; and 20, penicillin resistance. Scale bar indicates nucleotide substitutions per site. AMR, antimicrobial resistance; CR, community resident, a person with no known exposure to livestock; IHO, industrial hog operation; MDRSA, multidrug resistant S. aureus; SSTI, skin and soft tissue infection.

Almost all (94.7%) transmission cluster isolates were classified as MDRSA (Figure 2). Among 19 putative transmission cluster isolates, 14 were recovered from IHO workers, all of which were classified as MDRSA. Among IHO worker isolates, 2 differed from an IHO pig isolate by only 12 SNPs. An IHO worker isolate that was associated with a recent SSTI differed from an IHO pig isolate by only 20 SNPs (Figure 2). One transmission cluster isolate was from an adult community resident with no known exposure to livestock; this isolate also was classified as MDRSA (Figure 2). The minimum SNP distance between this isolate and the closest IHO pig isolate was 25 SNPs and it was 22 SNPs from the closest IHO worker isolate. Among 3 isolates from minors, 2 were identical (0 SNP differences) to an isolate from an IHO worker in the same household (Figure 2); 1 of the isolates from a minor was collected at the same sampling time as the IHO worker isolate. Among C3 isolates, we noted genetic determinants conferring resistance to tetracyclines, including tet(K), tet(L), tet(T); macrolides, including erm(A), erm(C); lincosamides, including lnu(A); aminoglycosides, aac6'-aph2", spc, and aadD; and streptogramins, including vga(A)LC (Figure 2).

We noted abundant genetic determinants conferring resistance to several antimicrobial classes among C3 isolates, including tetracyclines in 50% (25/50), macrolides in 56% (28/50), and aminoglycosides in 62% (31/50) of C3 isolates (Figure 2; Appendix Table 4). Among LA-SA CC9 clades, 50% (25/50) of C3 isolates were uniquely enriched for erm(A) genes, 16% (8/50) for vga(A)LC, 42% (21/50) for lnu(A), and 54% (27/50) for spc (Figure 1; Appendix Table 4). The mecA gene was absent from C3 but common among C1 and C2 isolates.

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