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| Food Microbiology | Research Article
Unveiling the genomic landscape of Salmonella enterica
serotypes Typhimurium, Newport, and Infantis in Latin American
surface waters: a comparative analysis
Zhao Chen,1 Magaly Toro,1,2 Andrea I. Moreno-Switt,3 Aiko D. Adell,4 Enrique J. Delgado-Suárez,5 Raquel R. Bonelli,6 Celso J. B.
Oliveira,7 Angélica Reyes-Jara,2 Xinyang Huang,1,8 Brett Albee,9 Christopher J. Grim,9 Marc Allard,9 Sandra M. Tallent,9 Eric W.
Brown,9 Rebecca L. Bell,9 Jianghong Meng1,8
AUTHOR AFFILIATIONS See aliation list on p. 16.
ABSTRACT Surface waters are considered ecological habitats where Salmonella enterica
can persist and disseminate to fresh produce production systems. This study aimed
to explore the genomic proles of S. enterica serotypes Typhimurium, Newport, and
Infantis from surface waters in Chile, Mexico, and Brazil collected between 2019 and
2022. We analyzed the whole genomes of 106 S. Typhimurium, 161 S. Newport, and
113 S. Infantis isolates. Our phylogenetic analysis exhibited distinct groupings of isolates
by their respective countries except for a notable case involving a Chilean S. Newport
isolate closely related to two Mexican isolates, showing 4 and 13 single nucleotide
polymorphisms of dierence, respectively. The patterns of the most frequently detected
antimicrobial resistance genes varied across countries and serotypes. A strong correla
tion existed between integron carriage and genotypic multidrug resistance (MDR) across
serotypes in Chile and Mexico (R > 0.90, P < 0.01), while integron(s) were not detected
in any of the Brazilian isolates. By contrast, we did not identify any strong correlation
between plasmid carriage and genotypic MDR across diverse countries and serotypes.
IMPORTANCE Unveiling the genomic landscape of S. enterica in Latin American surface
waters is pivotal for ensuring public health. This investigation sheds light on the
intricate genomic diversity of S. enterica in surface waters across Chile, Mexico, and
Brazil. Our research also addresses critical knowledge gaps, pioneering a comprehensive
understanding of surface waters as a reservoir for multidrug-resistant S. enterica. By
integrating our understanding of integron carriage as biomarkers into broader MDR
control strategies, we can also work toward targeted interventions that mitigate the
emergence and dissemination of MDR in S. enterica in surface waters. Given its potential
implications for food safety, this study emphasizes the critical need for informed policies
and collaborative initiatives to address the risks associated with S. enterica in surface
waters.
KEYWORDS surface water, Salmonella enterica, whole-genome sequencing, multidrug
resistance, integron, biomarker, genetic relatedness
Salmonella, an enteric pathogen with over 2,600 serotypes, has long been associated
with foodborne illnesses and public health challenges (1). Among these, Salmonella
enterica subspecies enterica encompasses more than 1,500 serotypes, which is responsi
ble for more than 99% of human salmonellosis (2). Historically, the primary concern
surrounding S. enterica infections has revolved around contaminated food products (3).
However, our understanding of the ecological adaptability of S. enterica has broadened
in recent years to encompass surface waters, which include a broad spectrum of water
May 2024 Volume 12 Issue 5 10.1128/spectrum.00047-24 1
Editor Xianqin Yang, Agriculture and Agri-Food
Canada, Lacombe, Canada
Address correspondence to Zhao Chen,
zhchen29@umd.edu.
The authors declare no conict of interest.
See the funding table on p. 17.
Received 5 January 2024
Accepted 6 March 2024
Published 28 March 2024
Copyright © 2024 Chen et al. This is an open-access
article distributed under the terms of the Creative
Commons Attribution 4.0 International license.
bodies such as rivers, lakes, ponds, and irrigation canals (4–6). These aquatic ecosystems
have emerged as signicant ecological niches where S. enterica can survive and
potentially contribute to its ongoing presence in fresh produce production systems (7).
Remarkably, S. enterica can persist in diverse aquatic ecosystems, adapting to varying
temperatures, nutrient levels, and the presence of competing microorganisms (8). This
adaptability allows S. enterica to maintain a reservoir in surface waters. The recognition
of the presence of S. enterica in surface waters has given rise to emerging public health
issues. Surface waters are not only habitats for S. enterica but also are interconnected
with agricultural production systems (5, 9). The presence of S. enterica in surface waters
is a matter of concern due to its potential implications for food safety, particularly in the
context of fresh produce. As surface waters are commonly used for irrigation and may
come into contact with fresh produce, the risk of contamination with S. enterica increases
(10).
S. enterica strains found in surface waters may possess distinctive genomic fea
tures, possibly altering their antimicrobial resistance (AMR) and pathogenicity (11).
In Latin America, a region characterized by rich biodiversity and varying environmen
tal conditions, the presence of S. enterica in surface waters has become a subject
of heightened interest. Prior research has unveiled the high prevalence and genetic
variability of S. enterica in surface waters in Latin America, including Chile, Mexico,
and Brazil (12–17). These three countries are not only major food producers but also
leading exporters of fresh produce in the region (18). The reliance on surface waters for
agricultural practices creates a critical interface between S. enterica, the environment,
and the food supply chain. S. enterica originating from contaminated surface waters
in these countries can potentially contaminate fresh produce, aecting not only local
populations but also posing risks to international trade and worldwide public health.
It is, therefore, imperative to extend our knowledge of S. enterica circulating in these
waters, as they have been less studied compared to traditional food sources. The study
of S. enterica in surface waters becomes pivotal, considering that these waters serve as
a reservoir for the pathogen and may contribute to its dissemination through the food
web.
In an era marked by advances in genomics and molecular epidemiology, whole-
genome sequencing (WGS) has emerged as a powerful tool for unraveling the genomic
intricacies of human pathogens (19). Our study, therefore, employed WGS to shed light
on the genomic diversity of S. Typhimurium, Newport, and Infantis from surface waters
in Chile, Mexico, and Brazil. These serotypes were among the top 10 most prevalent
serotypes for each country (our unpublished data). When considering isolates from
three countries collectively, they emerged as the top three most prevalent serotypes.
Moreover, by comparing the genomic attributes of S. enterica in surface waters from
distinct Latin American regions, this research also delved into the genetic relationships
among isolates of each serotype. In addition, it can contribute to future strategies for
mitigating the risks associated with S. enterica in surface waters, ultimately aiding in the
development of more eective surveillance and intervention measures.
MATERIALS AND METHODS
S. Typhimurium, Newport, and Infantis isolates
S. Typhimurium (n = 349), Newport (n = 339), and Infantis (n = 301) isolates were
collected from various surface water sources such as rivers, dams, lakes, ponds, and
irrigation canals in Chile, Mexico, and Brazil from 2019 to 2022. The sampling strat
egies and the characteristics of the sampling sites were described by Toro et al. (17)
and Ballesteros-Nova et al. (15). The isolates were collected as part of a collaborative
surveillance initiative involving the University of Maryland, partner universities in Chile,
Mexico, and Brazil, and the United States Food and Drug Administration (FDA). To
avoid biases in subsequent genomic analyses, clonal S. Typhimurium, Newport, and
Infantis isolates were identied and excluded from this study. Clonal isolates were
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dened as those derived from the same sample and grouped in the same single
nucleotide polymorphism (SNP) cluster according to the National Center for Biotechnol
ogy Information (NCBI) Pathogen Detection database (15). In addition, in alignment with
the criteria set by the FDA, these isolates were mandated to have an SNP distance of ≤20
(20). This cut-o was established based on a thorough examination of the published
literature, particularly focusing on the maximum pairwise SNPs observed in investiga
tions related to foodborne outbreaks and contamination events. After excluding clonal
isolates, we included 106 S. Typhimurium, 161 S. Newport, and 112 S. Infantis isolates
from surface waters in Chile, Mexico, and Brazil in the present study (Table 1; Table S1).
DNA extraction
S. Typhimurium, Newport, and Infantis isolates were streaked onto trypticase soy agar
(TSA; Fisher Scientic Inc., Hampton, NH). After a 24-h incubation at 35°C, one single
colony was transferred into tryptic soy broth (TSB; Fisher Scientic Inc.). The TSB culture
was then allowed to grow overnight at 35°C. Genomic DNA was extracted using the
Maxwell RSC cultured cells DNA kit (Promega Corporation, Madison, WI) on the Maxwell
RSC 48 instrument (Promega Corporation). Once extracted, the genomic DNA samples
were stored at 4°C until they were ready for use. The concentration of DNA in each
sample was measured using the Qubit 1× dsDNA broad range assay kit (Fisher Scientic
Inc.) on the Qubit 3.0 uorometer (Fisher Scientic Inc.).
Library preparation and WGS
Libraries were prepared on the Sciclone G3 NGSx iQ workstation (PerkinElmer, Inc.,
Waltham, MA) using the Illumina DNA prep kit in conjunction with IDT for Illumina
DNA/RNA UD indexes (Illumina Inc., SanDiego, CA). WGS was performed on the NextSeq
2000 platform (Illumina Inc.) with the NextSeq 1000/2000 P2 reagents (300 Cycles)
(Illumina Inc.) with 2 × 150 bp paired-end chemistry.
Data pre-processing and genome assembly
Raw reads were subjected to trimming using Trimmomatic 0.39 (21). Specically,
we employed the SLIDINGWINDOW operation with a window size of four bases for
averaging and a minimum average quality score of 20. Subsequently, the trimmed reads
were utilized for genome assembly, which was performed using SPAdes 3.15.5 (22),
following the default settings. This assembly process involved the use of k-mers at sizes
21, 33, and 55, along with careful correction to improve accuracy. To examine the quality
of each assembly, a thorough quality check was carried out using QUAST 5.2.0 (23).
Contigs with short lengths (<1,000 bp) and/or low coverages (<30×) were excluded from
each assembly to minimize the inclusion of potential contaminants.
Identications of AMR determinants, plasmids, integrons, and virulence
genes
AMRFinderPlus 3.11.14 was used to detect AMR determinants [AMR genes (ARGs)
and point mutations] (24), with a minimum identity threshold of −1 and a minimum
coverage of 50%. To streamline the terminology, we dened “genotypically antimicro
bial-resistant” isolates with at least one AMR determinant as “resistant.” Similarly, for
TABLE 1 Numbers of Salmonella enterica serotypes Typhimurium, Newport, and Infantis isolates from
Latin American surface waters
Serotype Number of isolates
Chile Mexico Brazil Combined
Typhimurium 62 28 16 106
Newport 51 73 37 161
Infantis 72 22 18 112
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“genotypically multidrug-resistant” isolates, we dened them as “multidrug-resistant”
when they exhibited AMR determinants associated with at least three distinct antimi
crobial classes. Plasmids were identied using Staramr 0.9.1 (25), which compared the
sequences to known plasmid sequences integrated with the PlasmidFinder database
(26). This process utilized a minimum identity of 98% and a minimum coverage of
60%. Integron identication was carried out using IntegronFinder 2.0.2 (27), with a
clustering threshold of 4 kb. The analysis included a lter for the clusters of attC sites
lacking integron integrases (CALINs) with a specied threshold, and the attC size was
constrained to a maximum of 200 bp and a minimum of 40 bp for accurate detection.
We conducted the Pearson correlation analysis using SigmaPlot 15 (Systat Software Inc.,
San Jose, CA) to assess (i) the correlation between the presence/absence of plasmid(s)
or integron(s) and genotypic AMR or multidrug resistance (MDR) for each isolate and (ii)
the correlation between the proportion of plasmid- or integron-carrying isolates and the
proportion of resistant or multidrug-resistant isolates among all isolates. The correlation
matrix was plotted using the “ggplot2” 3.4.4 (28), “corrplot” 0.92 (29), “ggplotify” 0.1.2
(30), and “ggcorrplot” 0.1.4.1 (31) R packages (R 4.3.2). Linear regression was conducted
and visualized with the “ggplot2” and “ggrepel” 0.9.4 R packages (32) when a strong
correlation existed [correlation coecient (R) ≥ 0.80 or ≤−0.80, P < 0.05] (33). For
the detection of virulence genes, we employed ABRicate 1.0.0, utilizing known gene
sequences from the Virulence Factors Database (VFDB) (34). The criteria for this detection
included a minimum identity of 80% and a minimum coverage of 60%.
Multilocus sequence typing
We conducted multilocus sequence typing (MLST) using mlst 2.23.0 (35; https://
github.com/tseemann/mlst). This tool integrates components from the PubMLST
database and performs scans on whole genomes against traditional PubMLST typing
schemes that rely on seven housekeeping genes. Specic criteria were applied for the
analysis, including a minimum identity threshold for the full allele of 95%, a minimum
coverage requirement for the partial allele of 10%, and a minimum score to match a
scheme of 50. As a result, mlst also reported the sequence types (STs) obtained from the
analysis.
Whole-genome phylogeny
SNPs were called and ltered for each serotype using the CFSAN SNP pipeline (36).
S. Typhimurium LT2 (RefSeq assembly accession: GCF_000006945.2), Newport CDC
2010K-2159 (RefSeq assembly accession: GCF_000973685.2), and Infantis FSIS1502916
(RefSeq assembly accession: GCF_001931575.1) served as the reference genomes for the
whole-genome phylogenetic analysis of S. Typhimurium, Newport, and Infantis isolates,
respectively. Following SNP calling, we employed FastTree 2.1.11 (37) to construct
whole-genome maximum-likelihood phylogenetic trees based on the generalized
time-reversible model. Each inferred whole-genome phylogeny was then visualized as
a rooted rectangular phylogram using iTOL 6.7.4 (38).
Core-genome MLST
The core-genome MLST (cgMLST) analysis was conducted using cgMLSTFinder 1.2 (39;
https://cge.cbs.dtu.dk/services/cgMLSTFinder/). This analysis utilized the core-genome
database for Salmonella retrieved from EnteroBase (40), which encompasses 3,002 loci.
Subsequently, a minimum spanning tree, based on the allelic proles for each serotype,
was constructed using GrapeTree 1.5.0 (41).
Pan-genome
Whole genomes were annotated using Prokka 1.14.5 (42), with a locus tag counter
increment of one, a minimum contig size of 200, and a similarity e-value cut-o of
0.000001. Afterward, we conducted pan-genome analysis utilizing Roary 3.13.0 (43),
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with a minimum percentage identity for blastp of 95% and a maximum limit of 50,000
clusters. We utilized the annotated genomes as input for Roary, enabling us to determine
the quantities of core and accessory (soft-core, shell, and cloud) genes: core genes
are present in 99% ≤ n ≤ 100%; soft-core genes are present in 95% ≤ n < 99%; shell
genes are present in 15% ≤ n < 95%; cloud genes are present in 0% ≤ n < 15%.
Heatmaps displaying the counts of core and accessory genes were generated with the
“pheatmap” 1.0.12 R package (44). Area-proportional Venn diagrams illustrating the core
and accessory genes were created using the “VennDiagram” 1.7.3 R package (45). Parsnp
1.7.4 was used to execute core-genome SNP alignment (46). This procedure involved
the automated recruitment of the reference sequence and required the inclusion of
all genomes for the analysis. The pan-genome results were visualized employing the
Roary plots module to construct a matrix showcasing the presence/absence of core and
accessory genes in the context of the core-genome phylogenetic tree. In addition, a
pan-genome pie chart was generated to provide insights into the composition of core,
soft-core, shell, and cloud genes for each serotype. The t-test was conducted using
SigmaPlot 15 to determine if signicant dierences (P < 0.05) existed among countries.
RESULTS AND DISCUSSION
AMR determinants
The presence of ARGs and point mutation among S. Typhimurium isolates from Chile,
Mexico, and Brazil was observed in 17.7 (11/62), 64.3 (18/28), and 12.5% (2/16) of cases,
respectively (Fig. 1). For S. Newport isolates from these countries, the presence of ARGs
and point mutation was 2.0 (1/51), 35.6 (26/73), and 2.7% (1/37), respectively. In the case
of S. Infantis isolates, 72.6 (52/72), 50.0 (11/22), and 5.6% (1/18) from Chile, Mexico, and
Brazil contained these features, respectively. The predominant ARGs exhibited variations
among countries and serotypes (Table 2).
A point mutation in gyrA_S83Y was the sole observed mutation in the quinolone
resistance-determining region (QRDR) for S. Typhimurium and Infantis isolates from
Chile and Mexico. None of the S. Newport isolates, regardless of the country of origin,
FIG 1 Genotypic antimicrobial resistance (AMR) of Salmonella enterica serotypes Typhimurium, Newport, and Infantis isolates from Latin American surface
waters. The number noted on each category indicates the number of isolates in the category.
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exhibited the mutation (Chile: 0/1; Mexico: 0/26; Brazil: 0/1), although gyrA mutations
in S. Newport have previously been reported (47–49). The mutation was present in 3.2
(2/62) and 10.7% (3/28) of S. Typhimurium isolates from Chile and Mexico, respectively.
For S. Infantis isolates from Chile and Mexico, the mutation was detected in 72.2 (52/72)
and 27.3% (6/22) of cases, respectively. By contrast, none of the S. Typhimurium (0/2) and
Infantis (0/1) isolates from Brazil carried the mutation. Meanwhile, 9.1 (1/11) and 16.7%
(3/18) of resistant S. Typhimurium isolates from Chile and Mexico exhibited the mutation.
By contrast, among resistant S. Infantis isolates, the mutation was present in all Chilean
isolates and 54.5% (6/11) of Mexican isolates.
Among resistant isolates, S. Typhimurium isolates exhibited MDR proportions of
45.5% (5/11) in Chile, 88.9% (16/18) in Mexico, and 50.0% (1/2) in Brazil (Fig. 1). For S.
Newport isolates, these rates were 100.0% (1/1) in Chile, 80.8% (21/26) in Mexico, and
0.0% (0/1) in Brazil. In the case of S. Infantis isolates, the prevalence of MDR was 100.0%
(52/52) in Chile, 81.8% (9/11) in Mexico, and 100.0% (1/1) in Brazil, signifying a high
level of MDR among these isolates. A point mutation in gyrA_S83Y was present in 40.0
(2/5) and 18.8% (3/16) of multidrug-resistant S. Typhimurium isolates from Chile and
Mexico, respectively. By contrast, multidrug-resistant S. Infantis isolates were observed
to have a higher mutation prevalence, with 100.0 (52/52) and 66.7% (6/9) for Chile and
Mexico, respectively. The mutation was not detected in multidrug-resistant S. Newport
isolates (Chile: 0/1; Mexico: 0/21). None of the multidrug-resistant Brazilian isolates with
genotypic AMR had the mutation (S. Typhimurium: 0/1; S. Infantis: 0/1). Our results reveal
that MDR, an important concern for public health, is notably high among resistant S.
Infantis isolates from all countries. While S. Typhimurium isolates from Mexico had a high
proportion of MDR, S. Newport isolates from Brazil displayed the lowest MDR rates.
The substantial occurrence of tet(A) in S. enterica isolates from Chile, Mexico, and
Brazil aligns with earlier reports that underscore the widespread prevalence of tetracy
cline resistance in bacterial populations, including S. enterica, within surface waters in
these regions (50–53). However, our results also demonstrate that the prevalence of
AMR determinants varied signicantly among both serotypes and within isolates of the
same serotype across Chile, Mexico, and Brazil, highlighting the complex interaction
of serotype-, strain-, and country-specic AMR proles among these isolates. Chile has
been identied as one of the top ve countries contributing signicantly to global
antimicrobial consumption in animal production in 2020 (54). In addition, projections
indicate that by 2030, Mexico and Brazil are expected to join the top ve countries with
the largest shares of global antimicrobial consumption in animal production (55).
Noticeably, antimicrobial usage (AMU) in agricultural practices, especially animal
husbandry, can directly impact the prevalence of AMR (56–59). Resistant strains may
emerge in response to the selective pressure imposed by routine AMU in veterinary
medicine. Hence, distinct AMR patterns among isolates of the same serotype from Chile,
Mexico, and Brazil potentially reect the variation in local practices in AMU, which
emphasizes the importance of considering geographical origin when evaluating the
risk of resistant S. enterica. For instance, Mexican isolates exhibited a high prevalence
of ARGs linked to phenicol, trimethoprim, and sulfonamide resistance. The approval of
trimethoprim-sulfamethoxazole as a broad-spectrum antimicrobial for treating bacterial
infections in livestock in Mexico aligns with our frequent detection of corresponding
TABLE 2 Predominant antimicrobial resistance genes among genotypically antimicrobial-resistant Salmonella enterica serotypes Typhimurium, Newport, and
Infantis isolates from Latin American surface waters
Serotype Predominant antimicrobial resistance genes
Chile Mexico Brazil Combined
Typhimurium sul2 (5/11) oR (15/18) aph(3'')-Ib, aph(6)-Id, sul2,
tet(A), qnrB19 (1/2)
aph(3'')-Ib, aph(6)-Id, sul2, tet(A)
(19/31)
Newport aadA2, blaCARB-2, dfrA1, oR, mph(A),
qacE, qnrA1, sul1, tet(A) (1/1)
dfrA1, oR, mph(A), qacEdelta1,
sul1, tet(A) (21/26)
qnrB19 (1/1) dfrA1, oR, mph(A), qacEdelta1,
sul1, tet(A) (22/28)
Infantis tet(A) (53/53) aadA1, qacEdelta1, tet(A) (9/11) blaTEM-1, oR, tet(A) (1/1) tet(A) (63/65)
aData in parentheses indicate the number of isolate(s) harboring the antimicrobial resistance gene(s)/the number of genotypically antimicrobial-resistant isolate(s).
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ARGs, including dfrA1 and sul1 (60). Moreover, despite the discontinuation of chloram
phenicol, other phenicol-based antimicrobials, such as orfenicol, remain registered and
utilized in Mexico.
Nevertheless, it is crucial to exercise caution when attempting to correlate AMU
with observed AMR patterns, primarily due to the signicant challenge posed by the
limited availability of comprehensive and specic data on local AMU within the sampled
regions. Obtaining accurate information about the types, quantities, and frequencies of
antimicrobials used in animal husbandry in specic regions within Chile, Mexico, and
Brazil is often challenging. The multifaceted nature of AMR involves intricate interactions
between environmental, genetic, and anthropogenic factors, making it even harder to
pinpoint the direct impact of AMU on AMR patterns without detailed and consistent
usage data. There is, thus, an urgent need for enhanced monitoring systems to track
AMU in animal husbandry in these countries, which is crucial for assessing its impact on
public health and developing targeted interventions.
Notably, the AMR features also diered among serotypes and within isolates of
the same serotype in each country. Each S. enterica serotype may have distinct
genetic characteristics, including the presence of specic AMR determinants, leading
to variations in AMR proles (61). The genetic diversity observed in isolates of the same
serotype may be attributed, in part, to the wide geographical distribution of the sampled
areas within each country. By collecting samples from various regions, including both
urban and rural environments, we aimed to capture the diverse ecological niches where
S. enterica may persist. The inclusion of samples from dierent locales increases the
likelihood of encountering distinct bacterial populations, contributing to the observed
genetic diversity. The variations in AMR features within isolates of the same serotype in
each country could, thus, be due to local selection pressures. Factors such as dieren-
ces in AMU practices, agricultural practices, and environmental conditions may create
unique selective environments favoring the emergence of specic AMR mechanisms.
Plasmid(s)
Notably, a signicant proportion of S. Typhimurium isolates in all three countries were
found to carry plasmids (Chile: 96.8%, 60/62; Mexico: 89.3%, 25/28; Brazil: 100.0%, 16/16),
with the highest prevalence observed in Brazil (Table S2). Conversely, plasmid occur
rence in S. Newport isolates was notably lower (Chile: 7.8%, 4/51; Mexico: 41.1%, 30/73;
Brazil: 2.7%, 1/37), with the least frequency in Brazilian isolates (Table S2). However,
the prevalence of plasmids in S. Infantis isolates exhibited variation (Chile: 76.4%,
55/72; Mexico: 36.4%, 8/22; Brazil: 5.6%, 1/18) (Table S2). These ndings suggest that
the carriage of plasmids varied not only among serotypes but was also inuenced
by geographical factors. The high prevalence of plasmids in S. Typhimurium isolates,
especially in Chile and Brazil, is a noteworthy observation.
It is important to highlight that all resistant S. Typhimurium (Chile: 11/11; Mexico:
18/18; Brazil: 2/2) and Newport (Chile: 1/1; Mexico: 26/26; Brazil: 1/1) isolates from the
three countries carried plasmid(s). All resistant S. Infantis isolates from Chile carried
plasmid(s) (52/52), while 72.7% (8/11) of the resistant S. Infantis isolates from Mexico
exhibited plasmid presence. However, plasmids were not present in the sole resistant
S. Infantis isolate from Brazil. Our Pearson correlation analysis did not reveal any strong
correlation between plasmid carriage and genotypic AMR across various countries and
serotypes (Fig. S1A and B). Isolates carrying plasmid(s) that bear ARGs are more likely
to exhibit resistance to antimicrobials targeted by those genes (62). In instances where
isolates carry plasmid(s) devoid of ARGs specic to certain antimicrobials, they may
remain susceptible to those antimicrobials. A comprehensive study examining 150,767
S. enterica genomes across 1,204 distinct serotypes revealed that most plasmids in S.
enterica are not involved in the dissemination of ARGs (63).
All multidrug-resistant S. Typhimurium (Chile: 5/5; Mexico: 16/16; Brazil: 1/1) and
Newport (Chile: 1/1; Mexico: 21/21) isolates from the three countries were found to
contain plasmid(s). All S. Infantis isolates from Chile (52/52) and 66.7% (6/9) of S. Infantis
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isolates from Mexico with MDR were found to harbor plasmid(s). However, plasmids
were absent in the only multidrug-resistant S. Infantis isolates from Brazil. Our Pearson
correlation analysis did not identify any strong correlation between plasmid carriage
and genotypic MDR across diverse countries and serotypes. Our study relied on Illumina
short reads, and the draft-genome nature of the data hindered the ability to accurately
pinpoint the specic genomic locations of ARGs (64). Further investigations should
necessitate long-read sequencing techniques to obtain complete genomes, which would
enable in-depth exploration of the structural attributes and locations (chromosome- or
plasmid-borne) of ARGs.
Integron(s)
Integron(s) were detected in 3.2% (2/62) and 57.1% (16/28) of S. Typhimurium isolates
from Chile and Mexico, respectively (Table S3). For S. Newport isolates, integron(s) were
found in 2.0% (1/51) and 17.8% (13/73) from Chile and Mexico, respectively (Table S3).
In the case of S. Infantis isolates, integron(s) were identied in 72.2% (52/72) and 40.9%
(9/22) from Chile and Mexico, respectively (Table S3). None of the Brazilian isolates
harbored integron(s), regardless of serotype.
Integrons are genetic elements that can capture and express gene cassettes
containing ARGs, which play a signicant role in the dissemination of AMR among
bacteria (65, 66). Most importantly, integron-mediated ARGs have previously been
reported to contribute to the MDR of S. enterica (67, 68). It should be noted that
all Chilean (S. Typhimurium: 2/2; S. Newport: 1/1; S. Infantis: 52/52) and Mexican (S.
Typhimurium: 16/16; S. Newport: 13/13; S. Infantis: 9/9) isolates containing integron(s)
were multidrug-resistant across serotypes (Fig. 1). Noticeably, our Pearson correlation
analysis underscores a strong positive correlation between integron carriage and
genotypic AMR, especially MDR, spanning diverse countries and serotypes (R > 0.80,
P < 0.05) (Fig. 2A and B). We found a linear correlation between the proportion of S.
Typhimurium, Newport, and Infantis isolates carrying integron(s) and the proportion
of those with genotypic AMR (R² =0.95) (Fig. 2C). Interestingly, a more robust positive
correlation existed between the proportion of S. Typhimurium, Newport, and Infantis
isolates waters carrying integron(s) and the proportion of those with genotypic MDR
across diverse countries and serotype (R²=0.98) (Fig. 2).
In this study, the observed strong correlation between integron carriage and
genotypic MDR suggests a potential role of integrons in mediating MDR in S. enterica.
The presence of integrons was consistently associated with genotypic MDR in S.
Typhimurium, Newport, and Infantis isolates from diverse geographical regions. This
correlation implies that the acquisition and maintenance of integrons can contribute
signicantly to the accumulation of ARGs in S. enterica, leading to a higher likelihood of
MDR.
Recent studies have underscored the signicance of integrons as predictive biomark
ers for AMR in various settings. Barraud et al. (69) highlighted the potential of integrons
as predictive markers for detecting AMR in acute sepsis, emphasizing their role in Gram-
negative bacteria-positive blood cultures (70). Azizi et al. (71) evaluated integrons in
Acinetobacter baumannii and identied class 1 integrons as biomarkers for MDR pheno
types in clinical situations. Similarly, the study by Hsiao et al. (72) associated class 1
integrons with ARG cassettes in Pseudomonas aeruginosa. Drawing parallels with these
ndings, our study emphasizes the crucial role of monitoring integron carriage as
biomarkers to comprehend the potential for MDR development in S. enterica popula
tions. This knowledge is pivotal for developing eective strategies to mitigate the spread
of multidrug-resistant strains. Given the implications of integron carriage, our ndings
advocate for their incorporation into future surveillance initiatives. Specically targeting
integrons as biomarkers in surveillance strategies will enable the monitoring of integron-
mediated MDR prevalence and evolution over time. In addition, optimizing water
examination protocols becomes imperative to curtail the dissemination of integron-
bearing S. enterica. Exploring specic gene cassettes within integrons and their
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correlations with MDR proles can also provide valuable insights into the mechanisms
driving MDR in S. enterica. The primary limitation in our study regarding the precise
localization of ARGs and integrons stems from the use of Illumina short reads. Future
research utilizing long-read sequencing techniques to obtain complete genomes has the
potential to shed light on the structures and physical proximity of specic ARGs and
integrons.
Virulence genes
A total of 116 virulence genes were identied in S. Typhimurium isolates from
all countries (Fig. S1A). Interestingly, we observed that some virulence genes were
exclusively present in Chilean and Brazilian isolates. Specically, astA encoding the
heat-stable enterotoxin 1 was solely detected in one Brazilian isolate. Also, cesT encoding
FIG 2 Correlations between integron carriage and genotypic antimicrobial resistance (AMR) or multidrug resistance (MDR) of Salmonella enterica serotypes
Typhimurium, Newport, and Infantis isolates from Latin American surface waters: (A) Pearson correlation between the presence/absence of plasmid(s) or
integron(s) and genotypic AMR or MDR for each isolate; (B) Pearson correlation between the proportion of isolates carrying plasmid(s) or integron(s) and the
proportion of those with genotypic AMR or MDR among all isolates. The correlation coecient (R) is presented in an ellipse by a coloring scheme from red
(negative correlation) to blue (positive correlation). The size of each ellipse is negatively correlated with R. The combination labeled with a green asterisk shows a
signicant dierence (P < 0.05)); (C) linear regression between the proportion of isolates carrying integron(s) and the proportion of those with genotypic AMR or
MDR.
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multi-eector chaperone was exclusively found in one Chilean isolate, and gspI encoding
the general secretion pathway protein I was identied in just one Chilean isolate.
Furthermore, it is noteworthy that sspH1 encoding the type III secretion system eector
SspH1 E3 ubiquitin ligase was carried by only two Chilean isolates and three Mexican
isolates. In addition, shdA encoding the AIDA autotransporter-like protein was only
identied in seven Chilean isolates, four Mexican isolates, and one Brazilian isolate.
Noticeably, two isolates from Brazil and Mexico did not possess some virulence genes
that were universally present in other isolates. Specically, one Brazilian isolate lacked
two genes consistently found in other isolates, including sodCI encoding the superoxide
dismutase precursor (Cu-Zn) and sseI/srfH encoding the type III secretion system eector
SseI/SrfH cysteine protease. In addition, one Mexican isolate did not contain ssaI, the
gene encoding the type III secretion system inner rod protein SsaI.
In S. Newport isolates from all countries, up to 106 virulence genes were detected
(Fig. S1B). It is worth noting the specic distribution of virulence genes among these
isolates. For instance, astA was exclusively identied in 13 Chilean isolates. Also, cdtB
encoding the cytolethal distending toxin B was found in only one isolate from Chile,
and cheY encoding the chemotaxis protein CheY was observed in a sole isolate from
Mexico. A subset of isolates from the three countries was devoid of genes consistently
identied in other isolates. Specically, pipB encoding the type III secretion system
eector PipB was not detected in two Mexican isolates, and ratB encoding the putative
outer membrane protein was notably absent in six Mexican and one Brazilian isolate.
Furthermore, sicP encoding the chaperone for SptP and sinH encoding the intimin-like
protein were not found in one Chilean isolate and three Mexican isolates, respectively.
Lastly, sspH2 encoding the type III secretion system eector SspH2 E3 ubiquitin ligase
was lacking in ve Mexican isolates.
Figure 1C illustrates that a combined total of 111 virulence genes were detected in S.
Infantis isolates from all the countries. Notably, some isolates from Chile and Brazil lacked
genes universally found in other isolates. Specically, two Chilean isolates did not carry
ssek1 encoding the type III secretion system eector SseK1, while ratB was not detected
in one isolate from Brazil.
The presence of genotypic MDR (aminoglycoside, beta-lactam, bleomycin, fosfomy
cin, lincosamide, phenicol, quaternary ammonium, quinolone, sulfonamide, tetracycline,
and trimethoprim) and the absence of specic virulence genes in certain S. Typhimurium
isolates, including six Chilean isolates and 17 Mexican isolates, indicate the potential
tness costs imposed by either ARGs or virulence genes (Fig. S1A). While the develop
ment of AMR may confer survival advantages in the presence of antimicrobials, it can
lead to selective disadvantages in terms of bacterial virulence (73). The underlying
mechanisms of such tness costs are complex and multilayered (74). Although increased
resistance to aminoglycoside, beta-lactam, and quinolone has been documented to be
associated, either directly or indirectly, with attenuated virulence attributes of S. enterica
(75–77), the current body of literature still lacks in-depth coverage of these biological
compromises in S. enterica.
The missing virulence genes in these S. Typhimurium isolates encompass gogB (solely
absent in Mexican isolates) responsible for encoding the type III secretion system eector
GogB, grvA related to the Gifsy-2 prophage, the pef gene cluster encoding the plasmid-
encoded mbriae, rck involved in resistance to complement killing, and the spv gene
cluster responsible for Salmonella plasmid virulence. Notably, several missing virulence
genes are linked to mobile genetic elements (MGEs) such as prophages and plasmids.
This process may be inuenced by the movement of MGEs through horizontal gene
transfer, where their loss or acquisition can result in changes in bacterial traits (78).
The connection between missing virulence genes and their associated MGEs suggests
a dynamic interplay in the genetic makeup of S. enterica, inuenced by factors such as
AMR, bacterial evolution, and the transfer of genetic materials. This dynamic nature of
virulence genes and MGEs is a critical area for further research on bacterial adaptation
and survival. To gain a comprehensive understanding of the fundamental mechanisms,
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additional virulence assessment would necessitate an in-depth exploration of whether
the loss of these virulence genes could lead to actual virulence attenuation.
It is signicant to mention that the specic tness costs can vary depending on
the strains and the types of antimicrobials encountered. The compromises were not
always consistent since some resistant strains still maintained their virulence genes,
while others experienced the loss of virulence genes. Furthermore, it was observed that
multidrug-resistant S. Newport and Infantis isolates retained a substantial portion of the
virulence genes. Some genotypically sensitive S. Infantis isolates were also consistently
devoid of certain virulence genes, including fyuA, the irp gene cluster, and the ybt gene
cluster. Understanding these complicated adaptations in S. enterica is crucial for public
health eorts, as it highlights the need for science-based AMU and surveillance to track
the emergence of AMR.
Whole-genome phylogeny
The whole-genome maximum-likelihood phylogenetic trees with sequence types, and
plasmid, integron, AMR, and virulence patterns of S. Typhimurium, Newport, and Infantis
isolates from Latin American surface waters are shown in Fig. S1. Figure S1A illustrates
the formation of two major well-dened clades (Clades I and II) on the phylogenetic
tree comprising 106 S. Typhimurium isolates, exhibiting a broad range of SNPs from 0
to 1,552. Clade I consisted of 42 isolates, encompassing 32 Chilean isolates, ve Mexican
isolates, and ve Brazilian isolates, all sharing the same ST (19) (Table S4). The SNPs
among isolates in Clade I ranged from 1 to 1,015. The largest SNP dierence (1,015) was
detected between two Chilean isolates. Clade II comprised 64 isolates, with 30 Chilean
isolates, 23 Mexican isolates, and 11 Brazilian isolates, representing a diverse range of STs,
including 19, 34, 99, 213, and 2072 (Table S4). The range of SNPs among isolates in Clade
II extended from 0 to 1,221. In a manner similar to Clade I, the highest SNP variation
in Clade II (1,221) was also identied between two Chilean isolates. Interestingly, 23 S.
Typhimurium isolates from Chile and Mexico exhibiting potential tness costs imposed
by either ARGs or virulence genes formed a single cluster on the tree. This cluster
included three distinct STs, including 19, 34, and 213 (Table S4). The SNP variation among
isolates in this cluster spanned from 0 to 443. The evolutionary sacrice events appeared
to have shaped the genetic similarity within the cluster. This clustering further highlights
a shared phenomenon where acquiring ARGs has led to the selective loss or reduced
presence of specic virulence genes.
The phylogenetic tree with 161 S. Newport isolates reveals the presence of two major
clades, denoted as Clades I and II (Fig. S1B). These clades exhibit a signicant diversity
in SNPs, ranging from 0 to 1,167. Clade I included 79 isolates, comprising 15 Chilean
isolates, 34 Mexican isolates, and 31 Brazilian isolates, showcasing a wide variety of STs,
including 118, 164, and 2370 (Table S4). The SNPs among isolates in Clade I ranged
from 0 to 102. The largest SNP dierence (102) was detected between one Mexican
isolate and one Brazilian isolate. Clade II included 82 isolates, consisting of 36 Chilean
isolates, 39 Mexican isolates, and 7 Brazilian isolates, representing a diverse spectrum
of STs, such as 31, 45, 132, and 7815 (Table S4). The SNPs among isolates in Clade
II ranged from 0 to 956. The largest SNP dierence (956) was detected between one
Chilean isolate and one Mexican isolate. Multidrug-resistant S. Newport isolates formed
a single cluster of 30 isolates, including one Chilean isolate and 29 Mexican isolates. All
isolates within this cluster shared the same ST (132) (Table S4). Notably, we detected
a genetic relatedness between one Chilean isolate (CFSAN125066) and two Mexican
isolates (CFSAN115844 and CFSAN121391) in this cluster, with a surprisingly minimal
genetic distance of four and 13 SNPs, respectively. Intriguingly, the closely related
isolates also exhibited congruent MDR, plasmid (IncR), and virulence patterns. It should
be emphasized that the sampling site for CFSAN115844 is not directly connected to the
one for CFSAN121391 (11 SNPs). According to the NCBI Pathogen Detection database,
these isolates were identied within the SNP cluster PDS000007781.917 and displayed a
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close relationship with two Mexican isolates originating from beef-based dog food (bully
stick) samples (CFSAN125066 and CFSAN115844: two SNPs; CFSAN121391: 11 SNPs).
CFSAN125066 was collected from a sampling site located in the small urban area
of Talagante, which is surrounded by agricultural zones and situated within a slum on
the riverbed of the Mapocho River. Livestock such as cows and horses are frequently
observed drinking water from the Mapocho River in this area. CFSAN115844 was
collected from a canal located in the Xochimilco municipality of Mexico City. The canal
is not only a tourist attraction but also vital to producing owers and vegetables for
human consumption. Treated wastewater from the city is used to replenish the canal.
However, it is possible that the treatment may not be eective enough to prevent
water contamination. In addition, the presence of animals, particularly pets and birds,
with direct access to these waters could also contribute to potential contamination.
CFSAN121391 was obtained from a river in Tlaxcala State, located more than 100 km
to the south of Mexico City. This area is in close proximity to extensive agricultural
regions. While Tlaxcala primarily emphasizes vegetable production, there could be some
small-scale livestock farms in this area. This site is easily accessible to animals for drinking
water.
The striking close relatedness of these three isolates from Chile and Mexico, as
exemplied by their nearly identical genomes, raises intriguing questions about the
potential mechanisms underpinning their genetic similarity. Most importantly, these
isolates shared the noteworthy feature of MDR. The presence of isolates from Chile
and Mexico within the same SNP cluster, as cataloged in the NCBI Pathogen Detec
tion database, hints at the potential global dissemination of these multidrug-resistant
isolates. This observation challenges the traditional understanding of geographical
divergence in bacterial populations. Our ndings, therefore, emphasize the importance
of a comprehensive approach to S. enterica surveillance, as factors beyond geographical
boundaries, such as international trade, human travels, animal movements, overlapping
ecological niches, and potentially shared sources of contamination, may collectively play
roles in shaping the genomic landscape of S. enterica (79). Ultimately, the genomic
relatedness of S. enterica isolates from dierent countries underscores the need for
further investigation into these factors inuencing the global distribution and genomic
relatedness of this pathogen.
In Fig. S1C, we identied two major clades (I and II) on the phylogenetic tree of 112
S. Infantis isolates, showcasing a range of SNPs from 0 to 411. Clade I consisted of 24
isolates, encompassing 6 Chilean isolates and 18 Brazilian isolates, all with STs of 32 and
1032 (Table S4). The SNP variation among isolates in Clade I spanned from 1 to 286,
with the greatest SNP divergence (286) observed between two Brazilian isolates. Clade
II comprised 88 isolates, with 66 from Chile and 22 from Mexico, sharing STs of 32 and
9835. Within Clade II, the range of SNP variation among isolates extended from 0 to
270, with the most notable SNP dierence (270) observed between two Mexican isolates.
A total of 58 S. Infantis isolates with genotypic MDR clustered together, including 52
Chilean and 6 Mexican isolates. Within this cluster, the range of SNPs among isolates
varied from 0 to 103. Interestingly, these isolates in the MDR cluster possessed unique
virulence genes not found in the other 55 isolates. These genes included fyuA, which
encodes the pesticin/yersiniabactin receptor protein, the irp gene cluster responsible for
encoding the yersiniabactin biosynthetic protein, and the ybt gene cluster associated
with the yersiniabactin siderophore biosynthetic protein.
cgMLST
Our cgMLST analysis not only reinforced the clustering patterns observed in the
whole-genome phylogenetic analysis but also allowed for a more focused examination
of serotype-specic genetic diversity (Fig. 3). Specically, within each of the cgMLST-
based minimum-spanning trees in Fig. 3, we observed two major clades for each
serotype. This observation suggests that the genetic variations and relatedness of
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isolates within each serotype were maintained across both analyses, highlighting the
consistency and accuracy of our ndings.
Pan-genome
Our pan-genome analysis unveiled distinctive genomic proles among the isolates
derived from Chile, Mexico, and Brazil, shedding light on the potential genetic adapta
tions to local environments. As revealed in Fig. 4A, higher counts of core genes were
found in S. Typhimurium, Newport, and Infantis isolates from Brazil, with 4,266, 3,932,
and 4,095 core genes, respectively. This observation suggests that Brazilian isolates may
share a more conserved genomic core. It is important to note that core genes typically
encode fundamental cellular functions (80), and their higher presence among these
isolates may indicate that these functions are essential for adapting to and surviving in
the local conditions prevalent in those regions. The genetic diversity observed among
the isolates from dierent countries led to the identication of distinct sets of core genes
for each country. While core genes are generally conserved across isolates, the variations
in genomic content among isolates from dierent geographical regions can result in
the identication of unique sets of core genes for each country. This diversity can be
inuenced by factors such as regional dierences in bacterial populations, environmental
FIG 3 cgMLST-based minimum spanning tree of Salmonella enterica serotypes Typhimurium (A), Newport (B), and Infantis (C) isolates from Latin American
surface waters. The node area in each tree is proportional to the number of isolates in the area.
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conditions, and evolutionary processes (81). In summary, dierent core genes observed
in Chile, Mexico, and Brazil reect the genomic diversity within the isolates collected
from these specic geographical locations. The identication of country-specic core
genes allows us to explore the unique genomic features of S. enterica populations in
each region. While our study provides valuable insights into the pan-genome diversity of
S. Typhimurium, Newport, and Infantis isolates from Latin American surface waters, it is
crucial to acknowledge the potential inuence of sample size on our ndings. Notably,
the smaller number of isolates from Brazil (n = 71) in comparison to Chile (n = 185) and
FIG 4 Numbers of core and accessory genes (A), and area-proportional Venn diagrams of core (B, C,
and D, respectively) and accessory (E, F, and G, respectively) genes of Salmonella enterica serotypes
Typhimurium, Newport, and Infantis isolates from Latin American surface waters. The intersection area in
each diagram is proportional to the percentage of isolates in the area.
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Mexico (n = 123) may have contributed to an overestimation of the count of core genes.
This limitation underscores the need for a cautious interpretation of these results.
By contrast, S. Typhimurium and Newport isolates from Chile and Infantis isolates
from Brazil exhibited higher counts of accessory genes (2,193, 3,819, and 1,518,
respectively) (Fig. 4A). The elevated number of accessory genes in these isolates suggests
that the isolates may have promoted the acquisition and retention of genes that oer
the isolates unique traits that facilitate niche-specic adaptations to surface waters
and thrive in the local environment. The proportions of overlapped and distinct core
and accessory genes between isolates from each pair of the three countries did not
show signicant dierences (P > 0.05). Nonetheless, higher proportions of overlapped
accessory genes were observed between S. Typhimurium isolates from Chile and Mexico
(318, 11.9%), S. Newport isolates from Chile and Mexico (1,221, 27.5%), and S. Infantis
isolates from Chile and Brazil (297, 17.1%) (Fig. 4B through G).
As shown in Fig. 5, the results from our pan-genome analysis involving isolates from
the three countries unveil interesting patterns. S. Typhimurium and Infantis isolates
had a greater number of core genes (4,042 and 4,145, respectively) than S. Newport
FIG 5 Pan-genome compositions (I), and core-genome phylogenetic trees aligned with the matrices of the presence and absence of core and accessory genes
(II) of Salmonella enterica serotypes Typhimurium (A), Newport (B), and Infantis (C) isolates from Latin American surface waters.
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isolates (3,857). Meanwhile, S. Typhimurium and Infantis isolates had higher proportions
of overlapped core (2,253, 37.4% and 2,313, 39.9%, respectively) and accessory (1,262,
47.4% and 947, 54.6%, respectively) genes among the three countries (Fig. 4B through
G). This implies that S. Typhimurium and Infantis isolates shared a more stable genomic
core across countries, highlighting their conserved genetic elements that likely play
pivotal roles in their survival and adaptability. Conversely, S. Newport isolates contained
more accessory genes (6,971) than S. Typhimurium and Infantis isolates (4,192 and 3,400,
respectively). In our investigation, the observed larger number of accessory genes in S.
Newport compared to S. Typhimurium and Infantis can be attributed to the presence
of three distinct lineages within S. Newport (82). The extended evolutionary divergence
among these lineages can contribute to a more expansive accessory genome, reect-
ing the genetic diversity accumulated over their respective evolutionary histories. This
distinction aligns with the complexities introduced by the diverse lineages within S.
Newport, providing a nuanced perspective on the observed genomic dierences among
S. enterica serotypes. Moreover, we acknowledge that the observed abundance of
accessory genes in S. Newport compared to the other two serotypes, S. Typhimurium
and Infantis, can also be attributed, in part, to the larger number of S. Newport isolates in
our study. The inuence of sample size on pan-genome analysis is a crucial consideration
and the unequal representation of serotypes may introduce variability in accessory gene
prevalence.
The outcomes of our pan-genome analysis provide insights into the genomic
diversity among S. Typhimurium, Newport, and Infantis isolates from Chile, Mexico,
and Brazil, oering a glimpse into potential genetic adaptations to their respective
environments. The variations in core and accessory gene count among isolates from
dierent countries and serotypes suggest that S. enterica has undergone distinct genetic
adaptations to their local environments, reecting the complexity and diversity of
surface water ecosystems across Chile, Mexico, and Brazil. These ndings warrant further
investigation to uncover the specic genetic traits and ecological factors contributing to
these observed patterns.
Conclusions
Our comprehensive genomic analysis of S. Typhimurium, Newport, and Infantis from
surface waters across Chile, Mexico, and Brazil has unveiled a complex landscape of
genomic diversity. Our ndings highlight the critical role that environmental reservoirs of
S. enterica play in public health, reinforcing the importance of continued surveillance and
good agricultural practices aimed at minimizing the transmission of this pathogen from
surface waters to humans through various pathways. By revealing the genetic makeup of
these isolates, we also gain insights into potential risks for MDR dissemination. For future
studies, it would be valuable to explore the genetic relatedness and potential transmis
sion routes between surface water and clinical isolates of S. Typhimurium, Newport, and
Infantis, which can provide insights into the public health implications of environmental
reservoirs in the epidemiology of salmonellosis.
ACKNOWLEDGMENTS
This research was supported by the FDA of the U.S. Department of Health and Human
Services (HHS) as part of federal award U01FDU001418.
AUTHOR AFFILIATIONS
1Joint Institute for Food Safety and Applied Nutrition and Center for Food Safety and
Security Systems, University of Maryland, College Park, Maryland, USA
2Instituto de Nutrición y Tecnología de los Alimentos, Universidad de Chile, Santiago,
Chile
3Escuela de Medicina Veterinaria, Facultad de Ciencias Biológicas, Ponticia Universidad
Católica de Chile, Santiago, Chile
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4Escuela de Medicina Veterinaria, Facultad de Ciencias de la Vida, Facultad de Agronomía
y Sistemas Naturales, Facultad de Ciencias Biológicas y Facultad de Medicina, Universidad
Andrés Bello, Santiago, Chile
5Facultad de Medicina Veterinaria y Zootecnia, Universidad de Nacional Autónoma de
México, Mexico City, Mexico
6Instituto de Microbiologia Paulo de Góes, Universidade Federal do Rio de Janeiro, Rio de
Janeiro, Brazil
7Departamento de Zootecnia, Universidade Federal da Paraíba, Areia, Brazil
8Department of Nutrition and Food Science, University of Maryland, College Park,
Maryland, USA
9Center for Food Safety and Applied Nutrition, United States Food and Drug Administra
tion, College Park, Maryland, USA
AUTHOR ORCIDs
Zhao Chen http://orcid.org/0000-0002-1784-167X
Enrique J. Delgado-Suárez http://orcid.org/0000-0001-5380-8095
Raquel R. Bonelli http://orcid.org/0000-0003-4537-5458
Sandra M. Tallent http://orcid.org/0000-0002-2971-8345
FUNDING
Funder Grant(s) Author(s)
HHS | U.S. Food and Drug Administration (FDA) U01FDU001418 Jianghong Meng
AUTHOR CONTRIBUTIONS
Zhao Chen, Conceptualization, Data curation, Formal analysis, Investigation, Method
ology, Project administration, Software, Supervision, Validation, Visualization, Writing
– original draft, Writing – review and editing | Magaly Toro, Project administration,
Resources, Writing – review and editing | Andrea I. Moreno-Switt, Resources, Writing –
review and editing | Aiko D. Adell, Resources, Writing – review and editing | Enrique
J. Delgado-Suárez, Resources, Writing – review and editing | Raquel R. Bonelli, Resour
ces | Celso J. B. Oliveira, Resources | Angélica Reyes-Jara, Resources | Xinyang Huang,
Investigation | Brett Albee, Investigation | Christopher J. Grim, Investigation, Resources,
Writing – review and editing | Marc Allard, Writing – review and editing | Sandra M.
Tallent, Resources | Eric W. Brown, Resources | Rebecca L. Bell, Project administration,
Resources, Writing – review and editing | Jianghong Meng, Funding acquisition, Project
administration, Resources, Writing – review and editing
DATA AVAILABILITY
Raw reads were deposited into the Sequence Read Archive (SRA) database hosted by the
NCBI under BioProject accession numbers PRJNA186035 and PRJNA560080.
ADDITIONAL FILES
The following material is available online.
Supplemental Material
Figure S1A (Spectrum00047-24-s0001.docx). Whole-genome maximum-likelihood
phylogenetic tree with sequence types, and plasmid, integron, genotypic antimicrobial
resistance, and virulence patterns of Salmonella enterica serotype Typhimurium isolates.
Figure S1B (Spectrum00047-24-s0002.docx). Whole-genome maximum-likelihood
phylogenetic tree with sequence types, and plasmid, integron, genotypic antimicrobial
resistance, and virulence patterns of Salmonella enterica serotype Newport isolates.
Research Article Microbiology Spectrum
May 2024 Volume 12 Issue 5 10.1128/spectrum.00047-2417
Figure S1C (Spectrum00047-24-s0003.docx). Whole-genome maximum-likelihood
phylogenetic tree with sequence types, and plasmid, integron, genotypic antimicrobial
resistance, and virulence patterns of Salmonella enterica serotype Infantis isolates.
Table S1 (Spectrum00047-24-s0004.xlsx). Metadata of Salmonella enterica serotypes
Typhimurium, Newport, and Infantis isolates.
Table S2 (Spectrum00047-24-s0005.xlsx). Plasmid presence and absence of Salmonella
enterica serotypes Typhimurium, Newport, and Infantis isolates.
Table S3 (Spectrum00047-24-s0006.xlsx). Integron presence and absence of Salmonella
enterica serotypes Typhimurium, Newport, and Infantis isolates.
Table S4 (Spectrum00047-24-s0007.xlsx). Sequence types of Salmonella enterica
serotypes Typhimurium, Newport, and Infantis isolates.
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