Table 1.

Summary of studies included in this review.

Ref.	Country(ies)	Sample size	Method(s)		ARG (N)a	Pipeline(s)	Reference Database(s)	Key findings	Limitations
4	Canada	24	SeqMeta		13,218	RAY Meta 2.2.1	MERGEM	Culture-enrichment in prior to sequencing allows identification of core and accessory ARGs in rare gut microbial species.	Enrichment step has eliminated otherwise prevalent Akkermansia and Prevotella.
5	Demark, Spain	663	SeqMeta		6,095	Prodigal, CD-HIT v4.5.7	Resfinder, ARG-ANNOT, Lahey Clinic, RED-DB	The predicted ARGs had 29.8% amino acid identity to known ARGs, supporting ARG transfer between microbes is rare.	ARGs in the subdominant microbial species and that non-homologous to the known ARG classes may be missed.
6	Tanzania, Italy	38	SeqMeta		2,570	TBLASTN	ARDB; ResFams	The gut resistome of the Hadza hunter-gatherer resembles the soil, with less syntenic mobile elements than that of the Italian.	Survey of ARGs was limited to known genes. Functional validation was not performed.
7	Syria, Iraq, Iran, Eritrea, Afghanistan, Germany	606	Mf-PCR		42	NA	NA	Refugees had more than five ARGs, with high prevalence of blaCTX-M, blaSHV, blaoxa-1, vanC1, and qnrB. German had three or less ARGs.	Mobility of the ARGs were not investigated and thus their transferability was unclear.
8	China, Denmark, Spain	162	SeqMeta; FunMeta		1,093	BLASTx	ARDB	Chinese had the highest abundance of ARGs, which are phylogenetically distant from others.	The limited samples may not reflect population-wide profile.
13	USA, China, Spain, Denmark	1,267	SeqMeta		112	BLASTx, SOAP	ARDB, ARG-ANNOT	ARG composition was similar among all population studied. Chinese had the highest ARG abundance than others.	Insufficient epidemiologic data that supported gut ARG being affected by antibiotic policy.
14	The Netherlands	122	PCR		10	NA	NCBI	The prevalence of blaCTX-M, qnrB and qnrS increased from 6.6% to 9% before travel to 33.6% to 55.7% after international travel.	Small study population size precluded risk analysis.
15	China	77	Microarray		2,915	NA	ARDB	ARG diversity is related to age.	False-positive or -negative hybridization signals may exist.
16	The Netherlands	10	Mf-PCR		81	NA	ResFinder, NCBI	The catA and tetW genes were more abundant in healthy subjects than the critically ill.	The findings may not apply to countries where resistant bacteria is prevalent.
17	Canada	70	Microarray		254	NA	CARD, ResFinder, ARG-ANNOT	Gut resistome in healthy adults showed prompt recovery to baseline in one week after antibiotic perturbation.	The microarray did not include all known ARGs.
18	The Netherlands	1,179	SeqMeta	11		ShortBRED	CARD	Females had higher diversity of ARGs and prevalence of lnu genes.	Small number of male subjects may be underpowered.
19	Peru	263	SeqMeta	1,100		ShortBRED	CARD, Lahey	The resistome from human gut and their associated animals and environment are shared	No functional validation of the ARGs identified.
20	China, Europe, USA	2,768	SeqMeta	19,569		SOAP-denovo	ResFinder	Mobile resistomes are mainly found, in order of frequency, in Proteobacteria (399 genes), Firmicutes (86), Bacteroidetes (46), and Actinobacteria (40).	Nucleotide identity cutoff was set at 95% against ResFinder.
21	Caucasian male	12	SeqMeta	NA		CARD-RGI	CARD, ResFams,	Healthy subjects with glycopeptide or aminoglycoside resistance genes were prone to de novo colonization upon antibiotic exposure.	The annotation algorithm was conservative.
22	UK, France, Italy, Finland, Norway, Scotland	22	Microarray; FunMeta	NA		Chromas, DNAMAN	NCBI	The tet(W) gene and transposons Tn4451 and Tn1549 were highly prevalent in fecal metagenomes.	Only tetracycline and erythromycin resistance genes were investigated.
23	Finland, France, Italy, Norway, UK	100	PCR; Microarray; FunMeta	97		NA	NA	ARGs identified by microarray were not recovered by functional screen. Combining these methods increase the identifiable ARG repertoire.	The cloned ARG in functional screen needs to be expressive in the heterologous host.
25	11 countries	180	SeqMeta	1,209		ARGs-OAP	SARG	The ermF was the representative macrolide resistant gene in the Chinese population.	Bacterial hosts of ARGs were predicted by network analysis.
28	Denmark, Spain, USA	252	SeqMeta	3,496		MOCAT	ARDB	The most prevalent human gut ARGs were those that encoded resistance to antibiotics used in animals in a country-specific manner.	Only three developed countries were involved.
29	USA	2	FunMeta	NA		NA	NCBI	Half of the resistance genes recovered from aerobic gut isolates were identical to that in pathogens.	The majority of gut commensals was anaerobic.
30	Germany	2	SeqMeta	NA		DIAMOND	CARD	The gut resistome in two persons changed differently in response to ciprofloxacin treatment.	Given sequence reads did not span the whole ARG length, its frequency could not be identified.
31	USA	29	SeqMeta	21		Prodigal, BLASTP	DeepARG	Sixty four ARGs were shared amongst the gut resistomes from chickens, pigs and humans.	The genetic contexts of the ARGs were unclear.
32	Malawi, USA, Venezuela	110	SeqMeta	70		BLAST+	ARDB, ResFams	Antibiotic resistance and virulence genes are correlated and may be co-selected.	Results were based on sequence analysis without downstream laboratory validation.

^aNumber of antibiotic resistance genes (ARG) surveyed or annotated; FunMeta, functional metagenomics; Marilyn, Marilyn Robert’s website for macrolides and tetracycline resistance genes; MERGEM, Mobile Elements and Resistance Genes Enhanced for Metagenomics; Mf-PCR, microfluidic PCR; PATRIC, Pathosystems Resource Integration Center; Ref, reference; SeqMeta, sequence-based metagenomics; SOAP, short oligonucleotide analysis package.