Data Sheet 2_Phylogenomic insights into LA-MRSA from Argentine pig farm environments: novel OptrA variant and regional emergence of an ST9 lineage co-circulating with international CC398 lineages.pdf

IntroductionStaphylococcus aureus (SA) colonizes both humans and animals. The spread of livestock-associated methicillin-resistant S. aureus (LA-MRSA) in farms and the environment poses a public health concern. While CC398 is globally predominant, regional variation exists, and data from Argentina remain scarce. This study investigated the presence, antimicrobial resistance (AMR), and genomic features of pig-associated MRSA in central Argentina, including transmission dynamics and the global phylogenetic context of LA-MRSA lineages CC398 and CC1/ST9.
MethodsBetween February and March 2022, 41 MRSA isolates were recovered from 50 fecal and effluent samples collected from 8 of 10 (80%) farrow-to-finish pig farms in Córdoba province, Argentina. Phenotypic susceptibility was assessed by disk diffusion and Vitek2 (CLSI 2023). Genotypic characterization included PCR for antimicrobial resistance genes (ARGs) and the immune evasion cluster (IEC), as well as PFGE, spa typing, SCCmec typing, and MLST. Twenty-four isolates underwent whole-genome sequencing (Illumina NovaSeq 6000) for resistome, virulome, and phylogenomic analysis.
ResultsAntimicrobial use was extensive, with most farms reporting use of more than seven drug classes and up to 18 compounds in the previous six months. Two predominant IEC¯ LA-MRSA lineages were identified, mostly associated to SCCmecV: CC398 (53.7%, n:22), comprising ST398 (n:17, spa-types t034/t571) and ST8814 (n:5, t571); and CC1/ST9 (41.5%, n:17, spa-t16964). One IEC+ CA-MRSA lineage (CC8/ST72, n:2, SCCmecIVa) was also detected. All isolates were multidrug-resistant (MDR), with LA-MRSA showing broader resistance across 4–7 antimicrobial classes. Multiple ARGs matched phenotypic resistance, except optrA, found in ST398 (n:5) and ST9 (n:1) despite borderline linezolid MICs (2–4 µg/mL), representing the first report of optrA in S. aureus in Argentina. WGS revealed a novel optrA variant (OptrA/FDKFP), likely linked to a mobile genetic element. Phylogenetic analysis revealed the local emergence of a distinct ST9 clade and multiple introductions of CC398 within internationally recognized subgroups (C6/AAP and C6/EP4).
DiscussionThese findings underscore pig farms as emerging reservoirs of multidrug-resistant MRSA both locally and globally, reinforcing the need for targeted surveillance and control strategies at the animal–human interface.

引言：金黄色葡萄球菌（Staphylococcus aureus, SA）可定殖于人类与动物体内。畜牧相关耐甲氧西林金黄色葡萄球菌（livestock-associated methicillin-resistant S. aureus, LA-MRSA）在养殖场及环境中的传播已构成公共卫生隐患。尽管CC398在全球范围内占据主导地位，但存在区域差异，且阿根廷相关研究数据仍较为匮乏。本研究针对阿根廷中部猪源MRSA的携带情况、抗菌药物耐药性（antimicrobial resistance, AMR）及基因组特征展开调查，涵盖传播动力学以及LA-MRSA谱系CC398与CC1/ST9的全球系统发育背景。
材料与方法：2022年2月至3月期间，研究人员从阿根廷科尔多瓦省10家自繁自育猪场中的8家（占比80%）采集的50份粪便与污水样本中，分离得到41株MRSA分离株。采用纸片扩散法与Vitek2系统（CLSI 2023）进行表型药敏试验。基因分型鉴定涵盖抗菌药物耐药基因（antimicrobial resistance genes, ARGs）与免疫逃逸簇（immune evasion cluster, IEC）的PCR检测，以及脉冲场凝胶电泳（pulsed-field gel electrophoresis, PFGE）、spa分型、SCCmec分型与多位点序列分型（multilocus sequence typing, MLST）。选取24株分离株进行全基因组测序（Illumina NovaSeq 6000），以开展耐药组、毒力组及系统基因组学分析。
结果：抗菌药物使用情况普遍，多数养殖场在过去6个月内使用了7种以上药物类别，最多达18种化合物。研究鉴定出2种优势的IEC阴性LA-MRSA谱系，主要与SCCmecV型相关：分别为CC398（占比53.7%，n=22），包含ST398（n=17，spa型别为t034/t571）与ST8814（n=5，spa型别为t571）；以及CC1/ST9（占比41.5%，n=17，spa型别为t16964）。同时还检测到1种IEC阳性社区获得性MRSA（community-acquired MRSA, CA-MRSA）谱系CC8/ST72（n=2，SCCmecIVa型）。所有分离株均为多重耐药（multidrug-resistant, MDR），其中LA-MRSA的耐药谱更广，可耐受4~7类抗菌药物。多数抗菌药物耐药基因（ARGs）与表型耐药结果相符，但optrA基因例外：该基因在ST398（n=5）与ST9（n=1）中被检出，尽管对应的利奈唑胺最低抑菌浓度（minimum inhibitory concentration, MIC）处于临界范围（2~4 μg/mL），这也是阿根廷首次在金黄色葡萄球菌中报道optrA基因。全基因组测序结果显示存在一种新型optrA变异株（OptrA/FDKFP），其可能与移动遗传元件相关。系统发育分析表明，本地出现了独特的ST9进化分支，且CC398多次从国际公认的亚群（C6/AAP与C6/EP4）引入。
讨论：本研究结果证实，无论在本地还是全球范围内，养殖场均为多重耐药MRSA的新兴储存宿主，这进一步强调了在动物-人类接触界面开展针对性监测与防控策略的必要性。