Genotypic diversity of Salmonella ser. Abortusequi isolates from Argentina

Equine Veterinary Journal ISSN 0425-1644
DOI: 10.1111/evj.13123
Genotypic diversity of Salmonella ser. Abortusequi isolates from
Argentina
C. P. BUSTOS†‡§* , M. MORONI#, M. I. CAFFER#, A. IVANISSEVICH–, M. HERRERAU, A. R. MOREIRA**,
N. GUIDA§ and P. CHACANA‡
†
cnicas (CONICET), Ciudad Auto
noma de Buenos Aires, Buenos Aires, Argentina
Consejo Nacional de Investigaciones Cientıficas y Te
Instituto Nacional de Tecnologıa Agropecuaria (INTA), CICVyA, Instituto de Patobiologıa, Hurlingham, Buenos Aires, Argentina
§
noma de Buenos Aires,
Universidad de Buenos Aires (UBA), Facultad de Ciencias Veterinarias, C
atedra de Enfermedades Infecciosas, Ciudad Auto
Buenos Aires, Argentina
#
n Nacional de Laboratorios e Institutos de Salud (ANLIS) “Dr. Carlos G. Malbran”, Instituto Nacional de Enfermedades Infecciosas (INEI),
Administracio
noma de Buenos Aires, Buenos Aires, Argentina
Departamento de Bacteriologıa, Servicio de Enterobacterias, Ciudad Auto
–
CRESAL VETERINARIA S.A., Pilar, Buenos Aires, Argentina
U
Servicio Nacional de Sanidad y Calidad Agroalimentaria (SENASA), DiLab, Departamento de Salmonelosis, Martınez, Buenos Aires, Argentina
n Experimental Agropecuaria Balcarce, Buenos Aires, Argentina.
**Instituto Nacional de Tecnologıa Agropecuaria (INTA), Estacio
‡
*Correspondence email: carlabustos@fvet.uba.ar; Received: 16.07.18; Accepted: 09.04.19
Summary
Background: Salmonella enterica subsp. enterica serovar Abortusequi (S. Abortusequi) is a serotype restricted to equines, which produces abortion
outbreaks. Nowadays the disease is being reported in different countries including Argentina thus generating an important impact in the equine
industry. Molecular characterization of the 95 kb virulence plasmid and the spvC gene of S. Abortusequi demonstrated their importance in the
pathogenicity of the serotype. In the last decades, high clonality of S. Abortusequi was identified in Japan, Mongolia and Croatia.
Objectives: The aim of this work was to characterize S. Abortusequi isolates obtained in Argentina between 2011 and 2016 by virulence-gene profiling
and pulsed-field gel electrophoresis.
Study design: Case report.
Methods: S. Abortusequi isolates were studied by virulence-gene profiling and pulsed-field gel electrophoresis.
Results: Four virulence profiles and nine pulsed-field gel electrophoresis pulsotypes were identified among the 27 isolates included in the study.
Different strains were found in the same outbreak and/or farm suggesting the presence of different sources of infection or mutation of isolates.
Main limitations: The number of related and nonrelated strains. More isolates may be necessary for a more intensive study.
Conclusions: Most strains presented the same virulence profile, being positive for all the studied genes except gipA and sopE1, which are involved in
intestinal virulence. Only few isolates showed different results in the same outbreak or farm. Unlike other studies, our results demonstrate a
considerable diversity of S. Abortusequi pulsed-field gel electrophoresis pulsotypes, which suggests that different sources of infection may be involved
within the same outbreak.
Keywords: horse; Salmonella ser. Abortusequi; equine abortion; genotypic diversity; Pulsed-field gel electrophoresis (PFGE); virulence genes
Introduction
Salmonella enterica subsp. enterica serovar Abortusequi (S. Abortusequi)
is a serovar adapted and restricted to equines and associated to abortion
in mares, orchitis in stallions and septicaemia and polyarthritis in foals
[1,2]. Between the 1950s and 1970s, mares with reproductive pathologies
caused by S. Abortusequi were described in Argentina, the United States
and several countries from Europe (Albania, Italy and Croatia), Asia (India
and Japan) and Africa (Cameroon) [1–5]. The disease has been involved in
abortion storms causing large economical loses in Argentina and Japan
[3,6–8]. Since 2011, equine paratyphoid abortion has been affecting
Argentine equine industry as a re-emerging disease causing abortion
storms involving large numbers of infected mares in short periods of time
[6–8] and many foals were born infected and manifesting septic arthritis
(Ivanissevich, 2016). While the microorganism is considered as an
eradicated pathogen in the United States, its isolation in Europe seems to
be sporadic. Recently, Stritof et al. [4] reported outbreaks of equine
paratyphoid abortion in Croatia and Grandolfo et al. [9] have associated
high mortality in foals with S. Abortusequi in Italy.
Several virulence factors of Salmonella have been studied, including
those which are clustered within Salmonella pathogenicity islands (SPIs)
and encoded in plasmids [10–12]. S. Abortusequi is transmitted by the
98
fecal-oral route [2] and consequently genes associated with intestinal
epithelial cell interactions (gipA and sopE1) may be involved in its
virulence. Virulence-gene profiling is an useful tool that may provide
information regarding bacterial virulence and its relationship with
pathogenicity. Recently, the sequence of the genome of a
representative strain isolated in Japan in 1978 has been determined to
be genetically close to host-adapted and host-restricted serotypes [13].
Furthermore, molecular characterization of plasmids containing spv as
well as trials conducted in mice showed that these plasmids were
homologous and may confer virulence to S. Abortusequi [14].
Epidemiological studies by pulsed-field gel electrophoresis (PFGE)
macro-restriction suggest high clonality among strains isolated in Japan,
Mongolia and Croatia [3,15]. However, these studies included strains
isolated between 1987 and 1995 and only one from 2007 and thus
these findings may not be representative for recent outbreaks. Due to
the increased prevalence of this serotype involved in reproductive
problems in equines during the last years, the aim of this work was
to characterize S. Abortusequi isolates from Argentina obtained from
2011 to 2016 to elucidate epidemiological relationships among the
strains by PFGE and to compare their virulence-gene profiles. This is
the first work that has studied the clonality of S. Abortusequi isolates
in South America.
Equine Veterinary Journal 52 (2020) 98–103 © 2019 EVJ Ltd
Salmonella ser. Abortusequi characterization
C. P. Bustos et al.
Materials and methods
Isolates
In this study, 27 isolates of S. Abortusequi obtained between 2011 and
2016 from clinical cases in Argentina were included (Supplementary Item
1). The microorganism was isolated from different samples (organs from
aborted equine fetuses; placenta, vagina and uterine of mares and joint
fluid of a foal). Twenty-five isolates were obtained from 13 outbreaks that
occurred mainly in Buenos Aires province, one isolate from a foal with
arthritis and the other one from a single case of abortion. All isolates were
identified as Salmonella sp. by standard biochemical test and PCR
(detection of invA) [16,17]. All the isolates were serotyped as Salmonella
ser. Abortusequi (4,12:–:e,n,x) according to the White-Kauffmann-Le Minor
scheme [18]. Farm and animal information and epidemiological data were
recorded (Supplementary Item 1).
Virulence-gene profiling
Isolates were incubated in brain heart infusion at 37°C overnight and then
1 mL was centrifuged at 14,000 g for 10 min, and the cell pellet was
resuspended in 300 lL of 19 TE buffer (Tris-HCl 1 mol/L, EDTA 0.5 mol/L,
ultra pure water, pH 8). Bacterial suspensions were incubated at 100°C for
10 min and immediately cooled for 5 min. Thereafter, suspensions were
centrifuged at 14,000 g for 5 min and supernatant was stored at 20°C
until use as DNA template. Overall, 10 virulence genes were amplified by
PCR: six targets (avrA, ssaQ, mgtC, siiD and sopB) located on the SPIs 1–5,
three targets (gipA, sodC1 and sopE1) on prophages, one (spvC) on the
Salmonella virulence plasmid and one (bcfC) on a fimbrial cluster. Forward
and reverse primers were used according to Huehn et al. [12].
The PCR was performed in a final volume of 25 lL containing 2.5 lL of
109 Reaction Buffer TASa, 1.5 lL of 25 mmol/L MgCl2a, 2 lL of 10 mmol/L
deoxynucleotide triphosphatea, 1 lL of 10 pmol/lL each primerb, 0.25 lL
of 5000 U/lL Taq DNA polymerasea, 13.8 lL of DNAse/RNAse free watera
and 4 lL of DNA template. PCR was carried out using a first step of 95°C
for one minute followed by 30 cycles of 95°C for 30 s, 58°C for 30 s and
72°C for 30 s and finally 72°C for 4 min. Those conditions were used for all
targets except bcfC PCR, which annealing temperature was 53°C [12]. After
electrophoresis at 90 V for 40 min, amplification products were stained
with ethidium bromide and observed in 1.5% agarose gela. PCR products of
control strains were purified by alcohol-EDTA method and sequenced by
capillary electrophoresis to compare the results with sequences uploaded
in GenBank (BLAST-NCBI). Once confirmed the identity of sequences, the
strain S. Abortusequi UBA1174/15 was used as positive control for avrA,
ssaQ, mgtC, siiD, sopB, sodC1, spvC and bcfC, the strain S. Enteritidis
INTA86/360 for sopE1 and the strain S. Typhimurium SENASA64 for gipA.
PFGE analysis
The isolates were subtyped by PFGE following the standardised laboratory
protocol of the International Pulse Net (Center for Disease Control and
Prevention, CDC) [19]. The results were analysed using BioNumerics
softwarec, and the dendrograms were constructed applying the DICE
coefficient and UPGMA (Unweighted Pair Group Method with Arithmetic
Mean) with a band position tolerance window of 1.5% as established by
PulseNet. The genetic relationship among isolates was evaluated according
to the variability in the serotype under study, and the epidemiological
context and the frequency of the patterns identified based on information
from the national database. Also, Tenover et al. [20] criteria were used to
assign relatedness categories based on the number of bands among the
isolates.
Results
Virulence-gene profiling
Ten virulence genes previously identified in Salmonella were selected to
characterize Argentinian S. Abortusequi isolates. Low variability regarding
the presence of the studied genes was found among the strains. The avrA,
ssaQ, mgtC, siiD, sopB, sodC1 and bcfC were detected in all the isolates
Equine Veterinary Journal 52 (2020) 98–103 © 2019 EVJ Ltd
included in this study. Four virulence profiles (named A, B, C and D) were
identified among the isolates being A (avrA+/ssaQ+/mgtC+/siiD+/sopB+/
gipA/sodC1+/sopE1/spvC+/bcfC+) the most frequent (Table 1). Most of
the isolates were positive for svpC (25/27; 92,59%) and negative for gipA
(22/27; 81,481%) or sopE1 (25/27; 92,59%).
In our study, spvC was negative in only two strains that were
isolated from an aborted fetus and from the vagina of a mare. The
gipA, located on a Gifsy-1 bacteriophage and considered as Peyer0 s
patch-specific virulence factor, was absent in most of the isolates.
Curiously, CRESAL913B/13 strain (gipA negative) was isolated together
with CRESAL913A/13 strain (gipA and sopE1 negative). Furthermore,
CRESAL21324/14 strain (gipA negative) was isolated from the same
abortion storm as two other isolates. One of them gipA positive and
sopE1 negative and the other gipA and sopE1 negative.
PFGE analysis
Nine PFGE pulsotypes (1–9) were identified among the isolates with up to
eight bands of difference. Similarity among the pulsotypes is shown in the
dendrogram (Fig 1). Two different clusters were found (Fig 1) and except
for two stables (farm 5 and 11) isolates obtained in each farm belonged to
the same cluster.
Farm 5 showed unexpected results. Although the same pulsotype was
identified in all the isolates from 2013, isolates from 2014 differed: two of
them were identical but the third showed seven bands of difference
(78.89% of similarity) (Fig 2a). Similar results were found in farm 6, where
different pulsotypes were identified in two isolates although both strains
were involved in the same outbreak in 2015. The CRESAL22230/15 strain
was isolated in June 2015, whereas the UBA1163/15 strain was isolated
1 month later showing two additional bands.
The isolate obtained from a large outbreak in the farm 7 (CRESAL145/15
strain) had three bands less than the pulsotype observed in the INTA-IP80/
16 strain (pulsotype 7), which was isolated in 2016 from the unique
abortion at the farm (Fig 2b).
The unique isolate from the farm 9 located in Balcarce presented a
pulsotype similar to other isolates from the same location, whereas the
unique isolate (CRESAL22472/15) from an outbreak in 2015 situated in
Lujan was clustered with other isolates obtained in 2014 and 2015 from
different distant locations. Farm 11 showed the most striking results. An
abortion storm occurred in 2015 on that farm and thereafter many of the
pregnant mares that did not abort gave birth septicaemic newborns. The
CRESAL517/15 strain isolated from an aborted fetus during the outbreak
and the CRESAL821/16 strain isolated from joint fluid of a foal with arthritis
presented two different PFGE pulsotypes (pulsotypes 2 and 7 respectively).
The comparison of both showed 91.43% of similarly (Fig 2c) but belonging
to two different clusters (Fig 1).
Discussion
Despite the importance of equine paratyphoid abortion, there is little
published information regarding molecular epidemiology and characterization of S. Abortusequi strains isolated from different outbreaks.
Most Salmonella virulence genes are located in the SPIs and on virulenceassociated plasmids [10–12,21,22] and virulence profiling is a tool not
previously used for S. Abortusequi that may help to understand virulence
mechanisms of the bacterium and its pathogenicity. All the isolates carried
the targets located on the SPIs 1–5 that are related to intestinal epithelial
cell invasion through the type III secretion system (invA and ssaQ) and
macropinocytosis (sopB), toxin production by type I secretion system (siiD),
intracellular survival at limited Mg++ concentrations (mgtC) and control of
Salmonella-induced inflammation (avrA). These results, together with the
fact that the fimbrial bcfC located in the chromosome was detected in
100% of the isolates, support previous studies that indicate that virulence
determinants located in the SPIs as well as bcfC are highly conserved
among serotypes [10–12,22]. The sodC1, located on bacteriophage and
encoding superoxide dismutase, was detected in all the isolates included in
this study. In previous reports, this gene was found in all S. Typhimurium
and S. Dublin isolates [10,12,22] but was not detected in S. Infantis,
Virchow, Hadar, Chagoua, Hindmarch or Inganda [12,22]. On the other
hand, the presence of the sodC1 was variable in other serotypes, being
99
Salmonella ser. Abortusequi characterization
C. P. Bustos et al.
TABLE 1: Results of the virulence-gene profiling of the S. Abortusequi isolates
mgtC
siiD
sopB
gipA
sodC1
sopE1
spvC
bcfC
Virulence profile
+
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A
A
D
B
B
A
B
A
B
A
C
B
C
A
A
A
A
A
A
A
D
A
A
A
A
A
A
100
ssaQ
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95
avrA
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90
invA
UBA-SABQ1563/11
UBA-SABQ1564/11
CRESAL 622/11
INTA-SABQ1138/12
INTA-SABQ1191/12
INTA-SABQ1192/12
CRESAL 20740
CRESAL 2312/12
SABQ77/15
CRESAL 913 A
CRESAL 913 B
CRESAL 1134/14
CRESAL 21324
CRESAL 21367
UBA 1163/15
UBA 1166/15
UBA 1168/15
UBA1169/15
UBA 1171/15
UBA 1174/15
CRESAL 22472/15
CRESAL 517/15
INTA-SABQ68/15
CRESAL 145/15
CRESAL 22230/15
CRESAL 821/16
INTA-IP80/16
85
Isolate
Isolate
CRESAL1134/14
CRESAL145/15
CRESAL22472/15
CRESAL22230/15
CRESAL821/16
INTA-IP80/16
UBA1163/15
UBA1166/15
UBA1169/15
UBA1171/15
UBA1174/15
CRESAL622/11
CRESAL394/11
CRESAL2312/12
CRESAL913A/13
CRESAL913B/13
INTA-SABQ1138/12
INTA-SABQ1191/12
INTA-SABQ1192/12
INTA-SABQ77/12
INTA-SABQ68/15
CRESAL21324/14
CRESAL21367/14
CRESAL20740/12
CRESAL517/15
UBA-SABQ1563/11
UBA-SABQ1564/11
Pulsotype
8
8
8
5
7
7
7
7
7
7
7
3
3
1
1
1
1
1
1
6
4
9
9
2
2
2
2
Farm
5
7
10
6
11
7
6
8
8
8
8
2
2
1
5
5
3
3
3
4
9
5
5
1
11
1
1
Fig 1: Dendrogram constructed applying the DICE coefficient and UPGMA with a band position tolerance window of 1.5%.
positive in most S. Enteritidis isolates and negative in most isolates of
S. Eppendorf [12,22,23].
The majority of the S. Abortusequi isolates were positive for svpC and
negative for gipA and sopE1, which is similar to the findings in other
100
serotypes [10–12,22,23]. Considering that the svpC is relevant for the rapid
growth and survival of the microorganism within the host and essential for
the virulence of S. Abortusequi in murine models, it is not surprising that
most of the isolates carried this gene. Anzai et al. [24] also demonstrated
Equine Veterinary Journal 52 (2020) 98–103 © 2019 EVJ Ltd
Salmonella ser. Abortusequi characterization
C. P. Bustos et al.
a)
819.49
672.28
680.22
519.47
375.60
349.76
307.71
287.61
351.08
313.38
289.32
234.34
224.88
241.29
131.24
120.88
98.39
84.68
78.35
69.85
59.24
41.74
38.31
33.39
20.58
133.01
122.57
99.76
79.36
71.25
43.74
35.20
20.36
CRESAL21367/14 CRESAL134/14
Strain
Strain
b)
681.15
660.54
503.57
509.75
343.81
344.72
303.24
282.97
239.41
299.82
281.27
227.61
220.23
121.98
115.24
95.29
83.29
76.38
67.98
58.15
41.40
34.84
23.00
127.52
120.26
98.47
78.84
69.50
43.93
34.96
21.29
INTA-IP80/16 CRESAL145/15
Strain
Strain
c)
857.62
679.88
693.96
509.47
374.53
346.78
342.03
299.44
279.31
302.39
280.81
228.33
216.82
229.62
219.37
122.93
116.08
95.49
82.93
74.28
66.25
55.30
39.55
33.77
18.44
124.73
115.60
95.54
83.55
76.60
66.51
56.71
40.75
34.65
22.32
CRESAL821/16 CRESAL517/15
Strain
Strain
Fig 2: Pulsotypes comparison. a) CRESAL21367/14 and CRESAL134/14 strains.
b) CRESAL145/15 and INTAIP80/16 strains. c) CRESAL517/15 and CRESAL821/16
strains. Red arrows indicate variant bands and blue arrows indicate constant
bands. The size of each band is indicated in kb.
the presence of 95-kb virulence plasmid contained spvC in a high
percentage of S. Abortusequi isolates (93.4%). They also observed that
isolates without this plasmid were avirulent for mice and horses and
therefore the authors highlight the relevance of the plasmid for the equine
Equine Veterinary Journal 52 (2020) 98–103 © 2019 EVJ Ltd
paratyphoid abortion. Anyhow, Astolfi-Ferreira et al. [25] proposed that
other virulence factors may be important since they found that spvCnegative strains were still capable of causing invasive infections in
chickens. Unfortunately, in our study it is not possible to assess if any
another related isolate was positive for the spvC as reported by other
researchers [24]. Also, several authors did not detect this gene in
serotypes such as Enteritidis, Typhimurium, Dublin, Chagoua, Hindmarch,
Inganda, Infantis or Eppendorf [10,22,23]. However, Huehn et al. [12]
identified medium and high percentage of gipA positive isolates of
S. Typhimurium and S. Virchow respectively. In our study, a high
percentage of the isolates were negative for sopE1 which is associated
with intestinal epithelial cell membrane ruffling and disruption of tight
junctions. The extremely low prevalence of the gipA and the sopE1 genes
found in our study may be related to their function. Even if these genes are
mostly involved in the interaction of the microorganism with the gut
environment, in S. Abortusequi they may be not essential for the invasion
and the reproductive manifestation of abortion, although their hypothetical
importance for the fecal-oral route of dissemination. Similarly, Suez et al.
[26] reported that the presence of gipA and sopE1 genes in nontyphoidal
Salmonella may be irrelevant for invasive manifestations in human
salmonellosis.
PFGE is a useful subtyping molecular technique to be included in
epidemiological studies of serotypes of Salmonella [11,19]. Despite the
crucial information that PFGE results and epidemiological data may provide
to understand the behaviour of the equine paratyphoid abortion, only a
few Japanese, Mongolian and Croatian isolates of S. Abortusequi were
analysed by PFGE. Unlike previous studies on S. Abortusequi, XbaI
digestion showed genomic heterogeneity among the isolates analysed
[3,15]. The diversity among isolates from Argentine equine industry
suggests that equine paratyphoid has been endemic in Argentina before
2011. Also elucidating clonal relations among the strains may help to
reveal failures in the management of the disease. For instance, all isolates
from the same outbreak in farm 3, that produces Criollo horses in Sierra de
la Ventana, belong to the same pulsotype (1) and thus they may be
considered the same strain. Likewise, the isolate from farm 4 in Balcarce
showed a single pulsotype, which was similar to another isolate previously
found in the same location. In these farms, it is likely that infections are
caused by strains that persist in the environment locally and thus more
efforts are needed to control the disease by disinfection, vaccination or
other biosecurity strategies.
On the other hand, in many of the farms that we studied strains
belonging to more than a single pulsotype were isolated from the same
outbreak. For example from farm 1, which is in Las Flores and produces
Criollo horses, we obtained 2 isolates in 2011 and 2 isolates in 2012.
Isolates from 2011 had the same pulsotype, whereas isolates from 2012
belonged to two different pulsotypes, one identical to the previous year
(CRESAL20740/12 strain, pulsotype 2) and other with two additional bands
(CRESAL2312/12 strain, pulsotype 1) (Fig 1). High similarity between the
isolates (94.12%) was observed and according to Tenover et al. criteria
(1995), those isolates should be considered closely related since they
differ only by a single genetic event, for example a spontaneous mutation
that removed a chromosomal restriction site. One of the 2012 isolates
from farm 1 was the same pulsotype as the isolates from farm 3 and farm
5 that also produce Criollo horses. However, Farm 3 is in Sierra de la
Ventana, more than 350 km from farm 1, and farm 5 is in Lujan, 200 km
from farm 1, and therefore the epidemiological relation between the
isolates cannot be explained by geographical proximity.
Similarly, isolates from farm 6 in 2015 belong to pulsotypes 5 and 7
with two different bands between them, whereas isolates from farm 8
in the same year belonged also to pulsotype 7. Although both farms
produce Polo horses, they are separated by 280 km and a likely
explanation for the spread of a single PFGE pulsotype in both farms
may involve contamination by fomites or movement of mares carrying
the pathogen. Isolates from farm 5 in 2014 belonged to two unrelated
pulsotypes (Fig 2a) according to Tenover et al. criteria [20]. Future
studies should consider origin of the animals, transportation, persistent
contamination of the environment and other reservoirs. The isolate
INTA-IP80/16 was involved in an abortion storm occurred in 2016 in
farm 7 in Cordoba province but belonged to a different pulsotype than
the strain CRESAL145/15, isolated from a previous outbreak in the farm
101
Salmonella ser. Abortusequi characterization
that also affected mares of 4 months of gestation. Pulsotype of strain
INTA-IP80/16 only had three additional bands than the strain CRESAL145/
15 (Fig 2b) but both isolates had the same virulence profile (A). If a
single source of infection is considered, a likely explanation may be that
the strain mutated while persisting in the mares or in the farm
environment. Interesting results were also found in farm 11. The strain
CRESAL517/15, isolated from an aborted fetus during an outbreak,
presented the pulsotype 2 and virulence profile A. The CRESAL821/16
strain, isolated from a joint fluid from a foal with arthritis, showed the
same virulence profile but a different pulsotype with three different
bands respect to the outbreak strain. According to Tenover et al.
criteria [20] these strains are closely related so these findings suggest
that the strain mutated in the foal or the mother. The fact that INTAIP80/16 strain showed although farms 7 and 8 are dedicated to
production of different breeds, suggests that the movement of recipient
mares used for embryo transfer may contribute to the dissemination of
the pathogen among nonrelated farms, a risk factor which should be
considered in the equine industry.
In conclusion, most of the studied strains had virulence-gene profiles
positive for all genes except gipA and sopE1, involved in the pathogenesis
of Salmonella in the gut. However, few isolates showed dissimilar results
regarding gipA, sopE1 and spvC, even when they were associated with the
same outbreak or farm. Unlike previous studies, our results showed a
significant diversity of PFGE pulsotypes for S. Abortusequi strains, which
suggests that different sources of infection are involved in the same
outbreak. These findings also suggest that equine paratyphoid has been
endemic in Argentina for a considerable time and additional studies
including genome sequencing of relevant strains are essential for a better
understanding of the equine paratyphoid abortion epidemiology in
Argentina. Serological surveillance would be informative and also, the
dynamics and reservoirs of S. Abortusequi should be investigated in order
to develop rational control measures to reduce the impact of the disease
on the equine industry.
Authors’ declaration of interests
No competing interests have been declared.
Ethical animal research
Research ethics committee oversight not currently required by this journal:
the study was performed on material collected previously during clinical
procedures.
Owner informed consent
Explicit owner informed consent for participation in this study was not
stated.
Sources of funding
This research was supported by INTA National Animal Health Program
Research Project. Carla Bustos was recipient of a Postdoctoral Grant from
the Consejo Nacional de Investigaciones Cientıficas y T
ecnicas (CONICET),
Argentina.
Acknowledgements
We thank veterinarian practitioners for providing samples and information.
Also, we are grateful to our laboratory co-workers, especially M. Mesplet,
~oz, J. Gallardo, G. Retamar, N. Lanza, A. Alcain and A. Velilla.
A. Mun
Authorship
The study was designed by C.P. Bustos, M. Moroni, P. Chacana, N. Guida
and M.I. Caffer. Laboratory work was performed by C.P. Bustos and
102
C. P. Bustos et al.
M. Moroni. All authors contributed to data analysis and interpretation.
C.P. Bustos prepared the initial manuscript draft and all authors
contributed to the manuscript revision and approved the final version.
Manufacturers’ addresses
a
Inbio Highway, Buenos Aires, Argentina.
Biodynamics, Buenos Aires, Argentina.
c
Applied Maths, Texas, USA.
b
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Supporting Information
Additional Supporting Information may be found in the online version
of this article at the publisher’s website:
Supplementary Item 1: S. Abortusequi isolates: isolate name, origin of
isolates (samples), date and place of isolation, information on farms and
epidemiological data.
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