Table 1 Mutations with moderate and high effect acquired during the infections and passages.

Strain	Mutation*	bp	Type	Gene	Encoded product	Group	Lineage
NCTC8325	SNP	27,460	Missense	walK	Sensor protein kinase WalK	GR \| TM \| ST
NCTC8325	SNP	98,945	Missense	SAOUHSC_00092	Conserved hypothetical protein	HP \| TM \| SE	c222
NCTC8325	SNP	120,749	Missense	cap5B	Capsular polysaccharide synthesis enzyme Cap5B	SE \| HP
NCTC8325	SNP	24,5287	Missense	tarI	Conserved hypothetical protein	SE \| HP
NCTC8325	SNP	280,449	Missense	essB	ESAT-6 secretion machinery protein EssB	TM \| SE \| P
NCTC8325	SNP	393,634	Missense	SAOUHSC_00390	Conserved hypothetical protein	HP \| P
NCTC8325	SNP	420,058	Missense	SAOUHSC_00417	Conserved hypothetical protein	TM
NCTC8325	SNP	464,931	Missense	rnmV	Ribonuclease M5	TM
NCTC8325	SNP	541,242	Missense	SAOUHSC_00535	Epimerase/dehydratase	TM
NCTC8325	SNP	546,984	Missense	azo1	FMN-dependent NADPH-azoreductase	TM
NCTC8325	SNP	568,990	Missense	SAOUHSC_00560	Conserved hypothetical protein		c222
NCTC8325	SNP	588,150	Missense	SAOUHSC_00586	Conserved hypothetical protein	SE
NCTC8325	SNP	706,223	Missense	SAOUHSC_00723	Conserved hypothetical protein	TM	c222
NCTC8325	SNP	721,884	Missense	SAOUHSC_00736	Putative lipid kinase	TM	c222
NCTC8325	SNP	778,092	Missense	SAOUHSC_00795	Glyceraldehyde-3-phosphate dehydrogenase	TM \| HP	c222
NCTC8325	SNP	813,228	Missense	metN2	Methionine import ATP-binding protein MetN2	TM
NCTC8325	SNP	1,030,094	Missense	SAOUHSC_01064	Pyruvate carboxylase	TM
NCTC8325	SNP	1,067,459	Missense	sdhB	Iron-sulfur subunit of succinate dehydrogenase%2C putative	TM
NCTC8325	SNP	1,121,026	Missense	carB	Carbamoyl phosphate synthase large subunit	TM
NCTC8325	SNP	1,138,999	Missense	SAOUHSC_01187	Conserved hypothetical protein	SE
NCTC8325	SNP	1,550,169	Stop-gain	SAOUHSC_01628	Conserved hypothetical protein	SE
NCTC8325	SNP	1,727,352	Missense	SAOUHSC_01821	Conserved hypothetical protein	TM	c222
NCTC8325	SNP	1,757,024	Missense	ccpA	Catabolite control protein A	GR \| P	c222
NCTC8325	SNP	1,773,253	Missense	SAOUHSC_01866	Conserved hypothetical protein	TM
NCTC8325	SNP	2,033,426	Missense	scni	Staphylococcal complement inhibitor	HP \| P \| SE
NCTC8325	SNP	2,045,511	Missense	yqbO	Tail length tape measure protein		c222
NCTC8325	SNP	2,095,322	Missense	agrC	Accessory gene regulator protein C	GR \| ST \| P \| SE
NCTC8325	SNP	2,249,603	Missense	SAOUHSC_02420	Conserved hypothetical protein	SE	c222
NCTC8325	SNP	2,286,918	Missense	hysA	Hyaluronate lyase	TM \| CWA \| P	c222
NCTC8325	SNP	2,311,214	Missense	rpsH	Ribosomal protein S8	TM	c222
NCTC8325	SNP	2,624,646	Missense	SAOUHSC_02849	Putative pyruvate oxidase	TM
NCTC8325	SNP	2,756,289	Missense	asp2	Accessory Sec system protein Asp2	TM \| SE
NCTC8325	Ins	487,126	Frameshift	SAOUHSC_00488	Cysteine synthase	TM
NCTC8325	Del	513,207	Frameshift	cysE	Serine acetyltransferase	TM
NCTC8325	Del	624,930	Disruptive	SAOUHSC_00634	ABC transporter	TM \| SE
NCTC8325	Ins	658,822	Disruptive	SAOUHSC_00670	Conserved hypothetical protein	TM \| SE	c222
NCTC8325	Del	764,412	Disruptive	lgt	Prolipoprotein diacylglyceryl transferase	TM \| SE
NCTC8325	Del	1,323,611	Frameshift	oppC2	Oligopeptide transporter putative membrane permease domain	TM \| SE
NCTC8325	Del	2,096,058	Frameshift	agrA	Accessory gene regulator protein A	GR \| ST \| P
NCTC8325	Ins	2,188,845	Frameshift	SAOUHSC_02366	Conserved hypothetical protein	TM
N315	SNP	105,983	Missense	SA_RS00630	Lipoprotein	SE
N315	SNP	274,258	Stop-gain	SA_RS01330	Acetyl-CoA/acetoacetyl-CoA transferase	TM	n222
N315	SNP	377,083	Missense	SA_RS01835	PTS lactose transporter subunit IIB	TM
N315	SNP	403,331	Missense	metE	5-Methyltetrahydropteroy-ltriglutamate— homocysteine methyltransferase	TM
N315	SNP	572,540	Missense	cysS	Cysteine—tRNA ligase	TM
N315	SNP	580,166	Missense	rpoB	DNA-directed RNA polymerase subunit beta	TM
N315	SNP	758,669	Missense	SA_RS03785	Hypothetical protein	SE
N315	SNP	863,049	Missense	osmC	Organic hydroperoxide resistance protein	HP
N315	SNP	949,546	Missense	LysR TR	LysR family transcriptional regulator	GR
N315	SNP	1,343,567	Missense	sbcC	Nuclease SbcCD subunit C	TM
N315	SNP	1,471,383	Missense	norB	Quinolone resistance protein NorB	TM \| P \| SE
N315	SNP	1,486,091	Missense	recU	Holliday junction resolvase RecU	TM
N315	SNP	1,735,479	Missense	pykA	Pyruvate kinase	TM	n222
N315	SNP	1,817,943	Missense	leuS	Leucine—tRNA ligase	TM
N315	SNP	1,948,268	Missense	vraS	Two-component sensor histidine kinase	GR \| ST \| SE \| P
N315	SNP	2,035,898	Stop-gain	SA_RS10285	Hypothetical protein		n222
N315	SNP	2,193,397	Missense	SA_RS11160	EVE domain–containing protein	TM
N315	SNP	2,320,771	Missense	acr	AcrB/AcrD/AcrF family protein	TM \| SE
N315	SNP	2,393,851	Missense	SA_RS12225	MOSC domain–containing protein	TM
N315	SNP	2,575,229	Missense	SA_RS13145	Hypothetical protein	CWA \| HP
N315	SNP	2,626,125	Missense	SA_RS13405	Membrane protein	SE
N315	SNP	2,637,600	Missense	SA_RS13450	N-succinyldiaminopimelate aminotransferase	TM	n222
N315	SNP	2,743,605	Stop-gain	SA_RS13980	Hypothetical protein	CWA \| SE
N315	SNP	2,751,067	Missense	asp2	Accessory Sec system protein Asp2	TM \| SE	n222
N315	Del	90,901	Frameshift	SA_RS00555	Anion membrane transporter	TM \| SE
N315	Del	1,085,977	Frameshift	SA_RS05425	Hypothetical protein	SE
N315	Del	1,473,471	Frameshift	tdcB	l-Threonine dehydratase catabolic TdcB	TM
N315	Ins	1,475,240	Frameshift	ald1	Alanine dehydrogenase	TM \| CWA \| SE
N315	Del	1,947,711	Frameshift	vraS	Two-component sensor histidine kinase	GR \| ST \| P \| SE
N315	Del	2,568,566	Disruptive	fnbA	Fibronectin-binding protein A	CWA \| HP \| SE \| P

*Only mutations with moderate effect (missense SNPs and disruptive indels) and high effect (stop-gained and frameshifts) are listed because they are more likely to play a role in host adaptation. Mutations include SNPs, deletions (Del), and insertions (Ins). Group: HP (host-pathogen interaction), GR (gene regulation), ST (signal transduction), TM (transport and metabolism), P (pathogenesis), CWA (cell wall associated), and SE (surface associated/exported). Lineage indicates the fittest clone in which mutations were identified.