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Article

An Evaluation of the Pathogenic Potential, and the Antimicrobial Resistance, of Salmonella Strains Isolated from Mussels

by
Antonio Lozano-León
1,2,
Carlos García-Omil
1,
Rafael R. Rodríguez-Souto
1,
Alexandre Lamas
3 and
Alejandro Garrido-Maestu
4,*
1
Laboratorio ASMECRUZ, Playa de Beluso s/n, 36939 Bueu, Spain
2
Group CI8, Biomedical Research Center (CINBIO), Campus Universitario de Vigo, University of Vigo, 36310 Vigo, Spain
3
Department of Analytical Chemistry, Nutrition and Bromatology, University of Santiago de Compostela, 27002 Lugo, Spain
4
Food Quality and Safety Research Group, International Iberian Nanotechnology Laboratory, Av. Mestre José Veiga s/n, 4715-330 Braga, Portugal
*
Author to whom correspondence should be addressed.
Microorganisms 2022, 10(1), 126; https://doi.org/10.3390/microorganisms10010126
Submission received: 7 December 2021 / Revised: 30 December 2021 / Accepted: 5 January 2022 / Published: 7 January 2022
(This article belongs to the Section Food Microbiology)
Browse Figures
Versions Notes

Abstract

:
Salmonella spp. and antimicrobial resistant microorganisms are two of the most important health issues worldwide. In the present study, strains naturally isolated from mussels harvested in Galicia (one of the main production areas in the world), were genetically characterized attending to the presence of virulence and antimicrobial resistance genes. Additionally, the antimicrobial profile was also determined phenotypically. Strains presenting several virulence genes were isolated but lacked all the antimicrobial resistance genes analyzed. The fact that some of these strains presented multidrug resistance, highlighted the possibility of bearing different genes than those analyzed, or resistance based on completely different mechanisms. The current study highlights the importance of constant surveillance in order to improve the safety of foods.

1. Introduction

Salmonella spp. is one of the most important foodborne pathogens worldwide, as highlighted by figures of salmonellosis in Europe, for which 52,702 confirmed cases were reported in 2020. In Spain, 3526 human salmonellosis were reported that year. At the distribution level, the highest number of Salmonella-positive samples was reported to be from meat products intended to be cooked (broiler, turkey and even pig and bovine meat) [1]. Regardless of these figures, Salmonella spp. is specifically regulated by the European Regulation 2073/2005 in a plethora of foodstuffs, including live mollusks [2]. In line with this Regulation, is the fact that the European Food Safety Authority (EFSA) reported that 2.01% of the outbreaks linked to the consumption of fish and fishery products were caused by Salmonella spp. [3]. In addition to this, another major health problem is the increased resistance of the microorganisms to antibiotics [4]. It was estimated that, every year, antimicrobial resistant microorganisms (ARMs) cause more than 23,000 deaths in the United States [5]. It has been previously reported that Salmonella spp. can be multidrug resistant (MDR) to antibiotics, thus becoming an even more important threat [6]. The EFSA data on antimicrobial resistance in 2018/2019, reported a 25.4% MDR in isolates of Salmonella spp. of human origin. Regarding the isolates from the food production chain, approximately 38% of the strains obtained from chicken, turkey and swine farms were MDR, but the highest number of MDR was found in pig carcasses, from which 43.3% of the Salmonella spp. recovered were MDR [7].
Spain is the third largest mussel producer worldwide, and Galicia accounts for 98% of the Spanish production [8]. In this region, the production has been developed in five different Rias: Vigo, Pontevedra, Arousa, Ares-Betanzos and Muros-Noia, but mainly in Arousa [9,10,11]. Previous studies have already reported the presence of pathogenic bacteria in this economically important mussel production area. These studies included pathogenic Vibrio and Campylobacter species, as well as Shiga toxin-producing Escherichia coli, thus highlighting the importance of screening for additional microorganisms, which can pose a risk for consumers [12,13,14]. In addition to these studies, Salmonella spp. was also detected and characterized in this area more than 10 years ago. All these facts highlight the importance of analyzing this food product [15,16]. For these reasons, the aim of the present study is to determine the presence of virulence genes in Salmonella spp. Strains, naturally isolated from mussels harvested in the Galician Rias (NW Spain), determine their antimicrobial resistance profiles, and compare these, if possible, with the results observed in previous studies conducted in the same geographical area and in other parts of the world.

2. Materials and Methods

2.1. Sampling

Mussel samples, harvested in the Rias of Arousa and Vigo (Figure 1), were analyzed from 2012 to 2016. The samples were collected, as previously reported [12]. Briefly, the mussels were collected from the rafts, placed in sterile bags and transported under refrigeration to the laboratory. Upon arrival, they were washed with tap water, opened and collected under aseptic conditions (dead or broken mussels were discarded). Twenty-five grams of mussel (tissue and liquor), obtained from a minimum of 15 individuals, were weighted and diluted 1/10 in 225 mL of buffered peptone water (BPW, BioMérieux, S.A., Marcy l’Etoile, France), homogenized for 90 s and incubated at 37 °C for 18 ± 2 h. Once the incubation was completed, the DNA was extracted and analyzed by qPCR, as described below.

2.2. DNA Extraction Protocol

DNA extraction was performed with the PrepSEQ™ Rapid Spin Sample Preparation Kit (Applied Biosystems, Foster City, CA, USA) from enrichment broths, following the manufacturer’s instructions. Briefly, 750 µL was loaded in a sterile tube and centrifuged for 3 min at 14,000 rpm. The supernatant was discarded and the pellet was resuspended in sterile distilled water and loaded in a spin column previously placed in a sterile tube and centrifuged again in the same conditions. The column was discarded as well as the supernatant. The pellet was resuspended in 50 µL of Lysis Buffer plus 5 µL of proteinase K solution (Sigma-Aldrich, St. Louis, MO, USA), and homogenized by vortexing. Then, the samples were incubated at 95 °C for 15 min. After incubation, the samples were left at room temperature for 3 min and were centrifuged for 1 min at 14,000 rpm. Finally, 250 µL of milli-Q water was added and the samples were centrifuged for 1 min at 14,000 rpm. All extracts were stored at −80 °C until qPCR analysis.

2.3. Screening of Salmonella spp. by qPCR in Mussel Samples

The detection of Salmonella spp. was performed with the MicroSEQ® (Applied Biosystems, Foster City, CA, USA) kit following the supplier’s instructions. The DNA extraction from the pre-enriched cultures was performed, as described in M & M 2.3, and 5 µL of DNA was used as a template. The reactions were performed in a final reaction volume of 20 µL, and run in a 7500 Fast Real Time PCR System Thermal Cycler (Applied Biosystems, Foster City, CA, USA) with the following thermal profile: 95 °C for 5 min (Hot-Start) followed by 40 cycles of denaturation at 95 °C for 5 s and annealing–extension at 60 °C for 30 s. Positive qPCR results were confirmed streaking the pre-enrichment culture on CHROMID® Salmonella ELITE agar, and incubated at 37 °C for 18 ± 2 h. Typical colonies were subcultured on triptic soy agar (TSA), incubated at 37 °C for 18 ± 2 h and then biochemically identified with API20E (BioMérieux, S.A., Marcy l’Etoile, France).

2.4. Genetic Characterization of the Isolates

2.4.1. Screening for Virulence Genes

A total of five virulence genes were screened (Table 1). These were: invA (first gene in an operon which is thought to trigger the internalization of Salmonella spp.); hilA (important in the regulation of the Type III secretion system); sopB (encodes for Salmonella outer protein B, part of the Type III secretion system); pefA (plasmid encoded fimbriae) and spvC, encoded in a virulence plasmid, whose expression helps the replication of Salmonella in the host cell reticuloendothelial system, especially in cells of the liver and spleen [17]. The detection of these genes was performed, as previously described [18,19,20,21,22].

2.4.2. Screening for Antibiotic Resistance Genes

In addition to these virulence factors, genes implicated in the resistance to aminoglusides and β-lactams were also screened. The selected genes were armA, rmtA, rmtB, rmtC, rmtD and npmA for the aminoglucosides, while blaCTX-M-15 was targeted to assess resistance to β-lactams. The detection of all these genes was performed, as previously described [23,24]. A detailed list of the primers used is provided in Table 1. Additionally, the specific thermal profiles for each genetic target is provided in Table 2. All primers were purchased from Sigma-Aldrich (St. Louis, MO, USA).

2.5. PCR Tests for Isolate Characterization

The detection of the different virulence and antimicrobial resistance genes was performed by PCR in a final reaction volume of 25 µL, with a final primer concentration of 1000 nM and 2 µL of template. To this end, a PTC-200 (MJ Research, Waltham, MA, USA) thermocycler was used, programed with the specific thermal profile detailed in Table 2.
After amplification, the fragments were visualized after electrophoresis in a 1.8 % agarose gel (Promega, Madison, WI, USA) prepared with 1 X Tris-Borate buffer (TBE 1 X, Tris-base 89 mM, boric acid 89 mM, 2 mM EDTA, pH 8.3), to which 5 µL of a 1 % ethidium bromide solution (PanReac-AppliChem, Barcelona, Spain) was added. A total of 5 µL of the PCR product was mixed with another 5 µL of acridine orange loading buffer (PanReac-AppliChem, Barcelona, Spain), and fully loaded in the gel. Additionally, 2 µL of 100 bp DNA ladder (Nippon Genetics Europe, Düren, Germany) was mixed with 8 µL of loading buffer. The electrophoresis was performed in a kuroGel Midi Plus 15 (VWR, Radnor, PA, USA) cuvette with a Model 300 V programable power source (VWR, Radnor, PA, USA), and the fragments were migrated for 1 h and 30 min at 60 V after what was visualized under UV light with a Benchtop UV transilluminator (UVP, Upland, CA, USA).

2.6. Antimicrobial Resistance Test

The antimicrobial resistance tests were performed on a VITEK®2 (BioMérieux, S.A., Marcy l’Etoile, France) using the AST-N244 cards (cefalotin; cefditoren; nitrofurantoin; tobramycin; fosfomycin; ampicillin; amoxicillin/clavulanic acid; piperacillin/tazobactam; cefuroxime; cefuroxime/axetil; cefoxitin; cefotaxim; ceftazidime; cefepime; ertapenem; imipenem; amikacin; gentamicin; nalidixic acid; ciprofloxacin; tigecycline; and trimethoprim/sulfamethoxazole). The classification as sensitive (S), intermediate (I) or resistant (R) was based on the criteria determined by the “Clinical and Laboratory Standards Institute” [25].

3. Results and Discussion

3.1. Prevalence and Serotypes of Salmonella enterica

Over the period of study, a total of 27 Salmonella spp. isolates were recovered from the mussel samples. A detailed list of the strains isolated, along with the sampling site, is provided in Table 3. Martinez et al. previously evaluated the presence of foodborne pathogens by PCR, in bivalve mollusks from the Ria de Arousa, over a period of 18 months [26]. In the mentioned study, 22 mussel sample batches were negative for invA, thus negative for Salmonella spp. The low prevalence of Salmonella in this type of product, along with the small number of batches tested, can justify these results. In line with this observation, Lozano-Leon et al. reported 19 Salmonella positive samples from 5907 mussel batches, representing a prevalence of 0.3% [27]. This prevalence contrasts with that observed in other countries where, for instance, Zahli et al. found a prevalence of Salmonella of 19.15% in mussels collected from Moroccan markets; likewise, Mannas et al. and Setti et al. found a Salmonella prevalence of 15.4% and 10% respectively, in mussel samples collected from the Atlantic coastline of Morocco [28,29,30]. Although an attempt was made to establish a correlation between the environmental factors, such as rainfall and fecal contamination with the prevalence of Salmonella, the results were contradictory [29,30].
S. typhimurium was the most prevalent serotype (4/27), followed by S. rissen (3/27) and S. senftenberg (3/27). The information available about the serotypes present in mussel samples is limited, but the presence of S. senftenberg in mussels has to be pointed out as this serotype was associated with the persistent contamination in high saline environments in mussel facilities, between 1998 and 2002 [31]. Curiously, Setti et al. detected the presence of this serotype in the seawater and sediment of Morocco’s Atlantic coast, but not in mussels [29]. Previous studies carried out in the Rias of Vigo, Pontevedra, Arousa and Muros-Noia, between 1998 and 2002, in mussels, seawater and mollusk depuration plants, found that S. senftenbergt was the most prevalent Salmonella serotype [15,32]. In addition to this, the isolates of this serotype were higher in the Ria de Arousa in comparison to the other Rias. In the present study, all the isolates of S. senftenberg were from Ria de Arousa. The data from both previous studies mentioned, and the fact that this serotype is still isolated in mussel samples from Ria de Arousa, seem to indicate that this serotype can be endemic to this area. The S. senftenberg isolates characterized by Martinez-Urtaza et al., presented a rugose morphotype, which is associated with a high capacity to produce biofilm [31,33]. This can explain the persistence of these strains in mussel production plants, and that they are a permanent source of contamination in Ria de Arousa. Likewise, the fact that this serotype is isolated in marine samples from other parts of the world, seems to indicate a high adaptation of this serotype to high salt concentrations.
S. typhimurim was also one of the most prevalent serotypes isolated from the Galician marine environments, in previous studies conducted by Martinez-Urtaza et al. [15,16]. The presence of this serotype in mussels, represents an important concern for public health since, together with S. enteritidis, it is one of the most pathogenic serotypes of Salmonella. Factors, such as densely populated areas and inefficient wastewater treatments, can be related to the prevalence of Salmonella in the marine environment. In line with this hypothesis, a study carried out on the coast of Colombia established a relationship between inefficient wastewater treatment systems and the presence of Salmonella in beach waters [34]. S. typhimurium is the second serotype most commonly associate with human salmonellosis in the European Union, and therefore highly populated coastal regions can suffer a potential contamination with human pathogenic Salmonella serotypes [35]. On the other hand, Galicia is an important farming region with a large number of cattle, swine and poultry farms. These farms can contaminate the nearby aquifers and rivers with bacterial pathogens that can end up in the sea. It is worth noting that the serotype Senftenberg was also isolated from Galician poultry farms, between 2011–2015 [36]. Finally, it is also important to mention that S. agona was isolated in this study, and also showed a high prevalence of molluscan shellfish by Martinez-Urtaza et al. [15]. Isolates belonging to this serotype have been characterized by their high capacity to form biofilms, which can partly explain why this serotype can be a permanent source of contamination [37].

3.2. Genetic Characterization of the Isolates

3.2.1. Screening for Virulence Genes

All the strains isolated were positive for invA (100%); 7 were positive for hilA and sopB (25.9%), 6 for spvC (22.2%) and only 1 was positive for pefA (3.7%). The strains analyzed can be classified into 7 different groups, attending to the virulence genes presented, which would be: P1 (invA); P2 (invA/hilA/sopB); P3 (invA/spvC); P4 (invA/hilA/spvC); P5 (invA/hilA/sopB/spvC/pefA); P6 (invA/sopB) and P7 (invA/hilA/sopB/spvC), as depicted in Figure 2 and summarized in Table 4, and only one isolated (S. typhimurium 268) presented all the virulence genes analyzed. It is not surprising that all the strains were positive for invA, as this gene has been extensively used for the specific detection of Salmonella spp. by a wide variety of DNA amplification methods, being also selected for interlaboratory validation studies [38,39,40]. However, in the present study, only 25.9% of the isolates were positive for the main virulence transcriptional regulator of Samonella Pathogenicity Island 1 (SPI-1) hilA, and 59.3 % of isolates did not present any gene other than invA [41]. A study previously conducted by Campioni et al., reported higher percentages of sopB and spvC. It is noteworthy that the study focused on spvB, but these two genes belong to the same operon, spvRABCD [42]. These differences can be explained either by a difference in the pathogenic potential of the strains, or directly by the fact that these genes can be conserved in certain serovars (Campioni’s study was restricted to Salmonella enteritidis). In this sense, Lamas et al. observed that spvC was present in only 44.8% of the Salmonella strains isolated from Galician poultry farms. However, that gene was mainly present in S. typhimurium and S. enterica subsp arizonae, in which this gene is chromosomally encoded, but almost absent in the other serotypes of S. enterica subsp. enterica [43]. Ammar et al., like Campioni, analyzed the strains related with human disease, and obtained high percentages of hilA (88%), sopB (41.2%) and pefA (41.2%); the only discrepancy among them was for spvC, as only 5.9% of Ammar’s strains possessed the gene [44]. The results obtained in the present work are aligned with those reported by Gharieb et al., who indicated the percentages of 10%, 30 % and 16.7–30% for pefA, hilA and sopB respectively [45].
Attending to the geographical distribution, it is worth commenting that 25.9% of the isolates that presented more than one virulence gene belonged to the Ria of Arousa, while 14.8% belonged to the Ria of Vigo. Of particular importance is the fact that P7 (all 5 virulence genes) was obtained from a sample harvested in Vigo, and P5 (4 virulence genes) belonged to a sample harvested in Arousa. Caution must be taken when extracting conclusions related to the abundance as, due to the fact that certain areas are more productive, more samples were analyzed. In line with this, in previous studies, a bias existed towards the Ria of Arousa, due to a higher number of samples harvested and analyzed from this area, as a consequence of having a higher production [12,13]. The current study adds to the others previously published, informing the presence of different pathogens in this economically important area, and highlighting the importance of depuration for the safety of the consumers.

3.2.2. Screening for Antimicrobial Resistance Genes

Regarding the presence of antimicrobial resistance genes, none of the seven genetic targets selected were detected in any of the isolates obtained. The fact that none of the genes screened were positive, but the isolates presented phenotypic resistance (see below) to many aminoglycosides, suggests the presence of additional mechanisms of resistance to these compounds, or the presence of other genes rather than those screened, such as blaSHV, blaTEM or other bla, instead of M-15 in ESBL strains or aacC for the resistance of aminoglycosides [46,47,48].

3.3. Antimicrobial Resistance

All 27 isolated strains presented MDR, as all them were resistant to at least 4 antimicrobials. Furthermore, 11 were resistant to 5 antimicrobials, and 3 to 9 of the drugs tested. None of the isolates were sensitive to all of the antimicrobials tested. All the isolates were resistant to cefuroxime and cefuroxime/axetil, a second generation cephalosporin. Similar to previous presentations, all 27 strains can be classified into 8 different profiles attending to their antimicrobial resistance characteristics. This would be: P1 (cefuroxime, cefuroxime/axetil, cefalotin, gentamicin and amikacin); P2 (cefalotin, cefuroxime, cefuroxime/axetil, cefoxitin, gentamicin and tobramycin); P3 (ampicillin, cefuroxime, cefuroxime/axetil, cefoxitin, gentamicin and amikacin); P4 (ampicillin, cefuroxime, cefuroxime/axetil, cefoxitin, gentamicin, trimethoprim/sulfamethoxazole and amikacin); P5 (ampicillin, cefalotin, cefuroxime, cefuroxime/axetil, cefoxitin, gentamicin, tobramycin, nalidixic acid and ciprofloxacin); P6 (ampicillin, cefuroxime, cefuroxime/axetil and trimethoprim/sulfamethoxazole); P7 (ampicillin, cefalotin, cefuroxime, cefuroxime/axetil, cefoxitin, gentamicin, tobramycin and trimethoprim/sulfamethoxazole) and P8 (ampicillin, amoxicillin/clavulanic acid, cefuroxime, cefuroxime/axetil, cefoxitin, cefotaxime, gentamicin, trimethoprim/sulfamethoxazole and amikacin). Most of the isolates tested belonged to P1, as depicted in Figure 3 and summarized in Table 4.
Previous studies, conducted by Martinez-Urtaza et al., on Salmonella isolated in the same area of study, indicated a higher incidence of strains resistant to certain antibiotics, such as ceftazidime, or in agreement with the findings reported in the present work, such as the incidence in amoxicillin/clavulanic acid resistance [49]. However, it is worth commenting that further serotyping information would be needed for a better comparison, as the cited study only focused on S. senftenberg, and variability in the resistance profiles can exist linked to specific serovars. Martinez-Urtaza et al. also evaluated the resistance profile of S. typhimurium isolates from bivalve mollusks, from 1998 to 2002 [16]. They found that 69.6% of isolates were sensitive to all the antimicrobials tested while, in the present study, the 4 S. Typhimurium presented multidrug resistance. These results follow the trend of the last few years, in which an increase in resistance has been observed in non-typhoidal Salmonella [50]. For example, Giacometti et al. observed an increased trend in the antimicrobial resistance profile of S. typhimurium strains isolated from bivalve mollusks collected from Ferrara, Italy, between 2001 and 2017 [51]. In Spain, the same type of trend was observed in strains of S. enterica isolated from chicken samples, in which the average number of resistances increased from 3.98% in 1993 to 5.00% in 2006, and an increased incidence in the resistance to cephalothin, enrofloxacin and tetracycline was also observed [52].
It is worth noting that resistance among these isolates was not limited to one type of antibiotic. As it can be observed in pattern 8 of Figure 3, three isolates were resistant to β-lactams (ampicillin, cefalotin, cefuroxime, cefuroxime/axetil and cefoxitin), aminoglycosides (gentamicin and tobramycin) and quinolones (nalidixic acid and ciprofloxacin), covering three different classes of antibiotics. It is also important to note that isolates within this resistance profile, namely AMC 270 and AMC 281, belonged to the virulence profiles 6 (invA/sopB) and 7 (invA/spvC/hilA/sopB) respectively, thus presenting additional virulence factors, and possibly posing an added risk.
Giacometti et al. reported low resistance to third generation cephalosporins and quinolones on isolates of S. typhimurium. These finding were in line to those reported in the present study. Contrary to their results, 96.3% of the isolates, included in the current study, were resistant to gentamicin, while Giacometti et al. only observed 1.9% [51]. These results also differ from those observed by Zahli et al., who did not report any isolate resistant to gentamicin from mollusks from the North Coast of Morocco [28]. Additionally, Martinez-Urtaza et al. found only one isolate of S. typhimurium that was resistant to gentamicin, among the ones studied from marine environments between 1998 and 2002 [16]. Interestingly, the high resistance to gentamicin has not been observed in Salmonella isolated from other points of the food chain in Galicia. Lamas et al. found that all the Salmonella isolated from poultry farms from 2011–2015 were sensitive to gentamicin [36]. In the EU report on the antimicrobial resistance in zoonotic bacteria in 2015, the data from Spain showed resistance to gentamicin in 3.8% and 6.3% of isolates of Samonella from humans and calves, respectively [53].
High resistance to second-generation cephalosporins was observed in this study. In humans, cefuroxime is the fourth most consumed antibiotic per inhabitant in Galicia [54]. This usage could be partly responsible for the high resistance observed in this pathogen; all the isolates were resistant to cefuroxime. This group of antimicrobials are widely used in animal production in Spain, which can cause an environmental pressure that selects bacteria resistant to these antibiotics [55]. Finally, 37.0% of the isolates were resistant to ampicillin. This is one of the most common resistances observed in Salmonella [53]. Penicillins are the most widely used group of antibiotics in animal production, which makes these resistances common in foodborne pathogens [55].
Finally, it is also important to highlight the data on intermediate resistances. These were observed for amoxicillin/clavulanic acid, ceftazidime, cefepime, nitrofurantoin and piperacillin-tazobactam. The WHO has classified these substances as critically important antimicrobials (CIAs), with the exception of nitrofurantoin [56]. Of particular interest is the antimicrobial profile of Salmonella AMC 92, which, in addition to the resistance to nine different antimicrobials, also presented three intermediate resistances to ceftazidime, cefotaxime and piperacillin-tazobactam. All isolates were sensitive to: cefditoren, ceftazidime, cefepime, ertapenem, imipenem, fosfomycin, nitrofurantoin, tigecycline and peperacicline/tazobactam; thus, these antimicrobials still represent a reliable way to treat possible infections associated with these bacteria.

4. Conclusions

S. typhimurim and S. senftenberg are serotypes commonly isolated from mussels and the latter seems to be endemic to the Ria de Arousa. Several strains isolated from mussel samples were positive for more than 1 virulence gene. All isolates were negative for the antimicrobial resistance genes tested, but presented phenotypic resistance to more than 4 antibiotics. Even though the multiresistance seems widespread, the strains under study were still susceptible to another 9 different antibiotics, which can represent the future first line treatment. The authors would like to highlight that depuration procedures are efficient in eliminating Salmonella spp. from mussels, and, if detected in this product, it would most likely be the result of a post-depuration contamination rather than the inefficacy of the treatment.

Author Contributions

A.L.-L.: conceptualization, methodology, validation and writing—review and editing; C.G.-O.: investigation; R.R.R.-S.: investigation and writing—review and editing; A.L.: data curation and writing—review and editing; A.G.-M.: data curation and writing—original draft preparation. All authors have read and agreed to the published version of the manuscript.

Funding

A.L. has a postdoctoral fellowship from Xunta de Galicia (Axudas de apoio á etapa de formación posdoutoral IN606B (Modalidade A)).

Institutional Review Board Statement

Not applicable.

Informed Consent Statement

Not applicable.

Conflicts of Interest

The authors declare no conflict of interest.

References

  1. EFSA; ECDC. The European Union One Health 2020 Zoonoses Report. EFSA J. 2021, 19, e6971. [Google Scholar]
  2. European Commission Regulation (EC) No 2073/2005. Microbiological Criteria for Foodstuffs; European Union: Brussels, Belgium, 2005.
  3. EFSA. Foodborne Outbreaks Reported in 2019. Available online: https://app.powerbi.com/view?r=eyJrIjoiY2FmNmUzYWEtZjg4Zi00ODJiLTkwMDMtOGUzYTY0YzYxMWY5IiwidCI6ImM0ODdkZDVhLTM3NjktNDQyYy1hYjc3LTI5MTkwODFkODVmYyIsImMiOjl9 (accessed on 7 December 2021).
  4. Mir, R.A.; Weppelmann, T.A.; Johnson, J.A.; Archer, D.; Morris, J.G.; Jeong, K.C.C. Identification and characterization of cefotaxime resistant bacteria in beef cattle. PLoS ONE 2016, 11, e0163279. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  5. Mir, R.A.; Weppelmann, T.A.; Teng, L.; Kirpich, A.; Elzo, M.A.; Driver, J.D.; Jeong, K.C. Colonization Dynamics of Cefotaxime Resistant Bacteria in Beef Cattle Raised Without Cephalosporin Antibiotics. Front. Microbiol. 2018, 9, 500. [Google Scholar] [CrossRef]
  6. Chen, S.; Zhao, S.; White, D.G.; Schroeder, C.M.; Lu, R.; Yang, H.; McDermott, P.F.; Ayers, S.; Meng, J. Characterization of Multiple-Antimicrobial-Resistant Salmonella Serovars Isolated from Retail Meats. Appl. Environ. Microbiol. 2004, 70, 1–7. [Google Scholar] [CrossRef] [Green Version]
  7. EFSA; ECDC (European Food Safety Authority and European Centre for Disease Prevention and Control). The European Union Summary Report on Antimicrobial Resistance in zoonotic and indicator bacteria. EFSA J. 2021, 18. [Google Scholar]
  8. Ferreira, M.; Lago, J.; Vieites, J.M.; Cabado, A.G. Chapter 8: World Production of Bivalve Mollusks and Socioeconomic Facts Related to Teh Impact of Marine Biotoxins, in Seafood and Freshwater Toxins: Pharmacology, Physiology, and Detection, 3rd ed.; Botana, L.M., Ed.; Taylor & Francis Group: Abingdon, UK, 2014; ISBN 978-1-4665-0514-8. [Google Scholar]
  9. Caballero Miguez, G.; Garza Gil, M.D.; Varela Lafuente, M.M. The institutional foundations of economic performance of mussel production: The Spanish case of the Galician floating raft culture. Mar. Policy 2009, 33, 288–296. [Google Scholar] [CrossRef]
  10. Caballero-Miguez, G.; Garza-Gil, M.D.; Varela-Lafuente, M.M. Legal change, property rights system and institutional stability: The case of the floating raft culture in the Galician mussel sector. Ocean Coast. Manag. 2012, 55, 84–93. [Google Scholar] [CrossRef]
  11. Costas-Rodríguez, M.; Lavilla, I.; Bendicho, C. Classification of cultivated mussels from Galicia (Northwest Spain) with European protected designation of origin using trace element fingerprint and chemometric analysis. Anal. Chim. Acta 2010, 664, 121–128. [Google Scholar] [CrossRef] [PubMed]
  12. Rodriguez-Souto, R.R.; Garrido-Maestu, A.; Pastoriza-Fontan, A.; Lozano-Leon, A. Investigation and characterization of Shiga toxin-producing Escherichia coli present in mussels from harvesting areas in Galician southern Rias (NW Spain). J. Food Saf. 2017, 37, e12367. [Google Scholar] [CrossRef]
  13. Garrido-Maestu, A.; Lozano-León, A.; Rodríguez-Souto, R.R.; Vieites-Maneiro, R.; Chapela, M.-J.; Cabado, A.G. Presence of pathogenic Vibrio species in fresh mussels harvested in the southern Rias of Galicia (NW Spain). Food Control 2016, 59, 759–765. [Google Scholar] [CrossRef]
  14. Lozano-León, A.; Rodríguez-Souto, R.R.; González-Escalona, N.; Llovo-Taboada, J.; Iglesias-Canle, J.; Álvarez-Castro, A.; Garrido-Maestu, A. Detection, molecular characterization, and antimicrobial susceptibility, of Campylobacter spp. isolated from shellfish. Microb. Risk Anal. 2021, 18, 100176. [Google Scholar] [CrossRef]
  15. Martinez-Urtaza, J.; Saco, M.; Hernandez-Cordova, G.; Lozano, A.; Garcia-Martin, O.; Espinosa, J. Identification of Salmonella Serovars Isolated from Live Molluscan Shellfish and Their Significance in the Marine Environment. J. Food Prot. 2003, 66, 226–232. [Google Scholar] [CrossRef]
  16. Martinez-Urtaza, J.; Liebana, E.; Garcia-Migura, L.; Perez-Pin, P.; Perez-Piñeiro, P.; Saco, M. Characterization of Salmonella enterica serovar Typhimurium from marine environments in coastal waters of Galicia (Spain). Appl. Environ. Microbiol. 2004, 70, 4030–4034. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  17. Van Asten, A.J.A.M.; Van Dijk, J.E. Distribution of “classic” virulence factors among Salmonella spp. FEMS Immunol. Med. Microbiol. 2005, 44, 251–259. [Google Scholar] [CrossRef] [PubMed]
  18. Swamy, S.C.; Barnhart, H.M.; Lee, M.D.; Dreesen, D.W. Virulence determinants invA and spvC in Salmonellae isolated from poultry products, wastewater, and human sources. Appl. Environ. Microbiol. 1996, 62, 3768–3771. [Google Scholar] [CrossRef] [Green Version]
  19. Rahn, K.; De Grandis, S.A.; Clarke, R.C.; McEwen, S.A.; Galan, J.E.; Ginocchio, C.; Curtiss, R.; Gyles, C.L. Amplification of an invA gene sequence of Salmonella Typhimurium by polymerase chain reaction as a specific method of detection of Salmonella. Mol. Cell. Probes 1992, 6, 271–279. [Google Scholar] [CrossRef]
  20. Murugkar, H.V.; Rahman, H.; Dutta, P.K. Distribution of virulence genes in Salmonella serovars isolated from man & animals. Indian J. Med. Res. 2003, 117, 66–70. [Google Scholar]
  21. Cardona-Castro, N.; Restrepo-Pineda, E.; Correa-Ochoa, M. Detection of hilA gene sequences in serovars of Salmonella enterica subspecies enterica. Mem. Inst. Oswaldo Cruz 2002, 97, 1153–1156. [Google Scholar] [CrossRef] [Green Version]
  22. Rahman, H. Prevalence & phenotypic expression of sopB gene among clinical isolates of Salmonella enterica. Indian J. Med. Res. 2006, 123, 83. [Google Scholar]
  23. Fritsche, T.R.; Castanheira, M.; Miller, G.H.; Jones, R.N.; Armstrong, E.S. Detection of methyltransferases conferring high-level resistance to aminoglycosides in Enterobacteriaceae from Europe, North America, and Latin America. Antimicrob. Agents Chemother. 2008, 52, 1843–1845. [Google Scholar] [CrossRef] [Green Version]
  24. Leflon-Guibout, V.; Jurand, C.; Bonacorsi, S.; Espinasse, F.; Guelfi, M.C.; Duportail, F.; Heym, B.; Bingen, E.; Nicolas-Chanoine, M.H. Emergence and spread, of three clonally related virulent isolates of CTX-M-15-producing Escherichia coli with variable resistance to aminoglycosides and tetracycline in a French geriatric hospital. Antimicrob. Agents Chemother. 2004, 48, 3736–3742. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  25. CLSI. Methods for Dilution Antimicrobial Susceptibility Tests for Bacteria That Grow Aerobically, 9th ed.; CLSI: Pittsburgh, PA, USA, 2012; Volume 32, ISBN 1562387839.
  26. Martínez, O.; Rodríguez-Calleja, J.M.; Santos, J.A.; Otero, A.; García-López, M.L. Foodborne and indicator bacteria in farmed molluscan shellfish before and after depuration. J. Food Prot. 2009, 72, 1443–1449. [Google Scholar] [CrossRef] [PubMed]
  27. Lozano-Leon, A.; Garcia-Omil, C.; Dalama, J.; Rodriguez-Souto, R.; Martinez-Urtaza, J.; Gonzalez-Escalona, N. Detection of colistin resistance mcr-1 gene in Salmonella enterica serovar Rissen isolated from mussels, Spain, 2012 to 2016. Eurosurveillance 2019, 24, 1–5. [Google Scholar] [CrossRef] [PubMed]
  28. Zahli, R.; Soliveri, J.; Abrini, J.; Copa-Patiño, J.L.; Nadia, A.; Scheu, A.-K.; Nadia, S.S. Prevalence, typing and antimicrobial resistance of Salmonella isolates from commercial shellfish in the North coast of Morocco. World J. Microbiol. Biotechnol. 2021, 37, 170. [Google Scholar] [CrossRef]
  29. Setti, I.; Rodriguez-Castro, A.; Pata, M.P.; Cadarso-Suarez, C.; Yacoubi, B.; Bensmael, L.; Moukrim, A.; Martinez-Urtaza, J. Characteristics and dynamics of Salmonella contamination along the coast of agadir, Morocco. Appl. Environ. Microbiol. 2009, 75, 7700–7709. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  30. Mannas, H.; Mimouni, R.; Chaouqy, N.; Hamadi, F.; Martinez-Urtaza, J. Occurrence of Vibrio and Salmonella species in mussels (Mytilus galloprovincialis) collected along the Moroccan Atlantic coast. Springerplus 2014, 3, 1–11. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  31. Martinez-Urtaza, J.; Peiteado, J.; Lozano-León, A.; Garcia-Martin, O. Detection of Salmonella Senftenberg Associated with High Saline Environments in Mussel Processing Facilities. J. Food Prot. 2004, 67, 256–263. [Google Scholar] [CrossRef]
  32. Martinez-Urtaza, J.; Saco, M.; De Novoa, J.; Perez-Piñeiro, P.; Peiteado, J.; Lozano-Leon, A.; Garcia-Martin, O. Influence of Environmental Factors and Human Activity on the Presence of Salmonella Serovars in a Marine Environment. Appl. Environ. Microbiol. 2004, 70, 2089–2097. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  33. Obe, T.; Nannapaneni, R.; Sharma, C.S.; Kiess, A. Homologous stress adaptation, antibiotic resistance, and biofilm forming ability of Salmonella enterica serovar Heidelberg ATCC8326 on different food-contact surfaces following exposure to sublethal chlorine concentrations 1. Poult. Sci. 2018, 97, 951–961. [Google Scholar] [CrossRef]
  34. Soto-Varela, Z.E.; Rosado-Porto, D.; Bolívar-Anillo, H.J.; González, C.P.; Pantoja, B.G.; Alvarado, D.E.; Anfuso, G. Preliminary microbiological coastal water quality determination along the department of atlántico (Colombia): Relationships with beach characteristics. J. Mar. Sci. Eng. 2021, 9, 122. [Google Scholar] [CrossRef]
  35. EFSA; ECDC. The European Union summary report on trends and sources of zoonoses, zoonotic agents and food-borne outbreaks in 2016. EFSA J. 2017, 15, e05077. [Google Scholar] [CrossRef] [Green Version]
  36. Lamas, A.; Fernandez-No, I.C.; Miranda, J.M.; Vázquez, B.; Cepeda, A.; Franco, C.M. Prevalence, molecular characterization and antimicrobial resistance of Salmonella serovars isolated from northwestern Spanish broiler flocks (2011–2015). Poult. Sci. 2016, 95, 2097–2105. [Google Scholar] [CrossRef] [PubMed]
  37. Díez-García, M.; Capita, R.; Alonso-Calleja, C. Influence of serotype on the growth kinetics and the ability to form biofilms of Salmonella isolates from poultry. Food Microbiol. 2012, 31, 173–180. [Google Scholar] [CrossRef]
  38. Cheng, C.M.; Lin, W.; Van, K.T.; Phan, L.; Tran, N.N.; Farmer, D. Rapid Detection of Salmonella in Foods Using Real-Time PCR. J. Food Prot. 2008, 71, 2436–2441. [Google Scholar] [CrossRef] [PubMed]
  39. Hara-Kudo, Y.; Yoshino, M.; Kojima, T.; Ikedo, M. Loop-mediated isothermal amplification for the rapid detection of Salmonella. FEMS Microbiol. Lett. 2005, 253, 155–161. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  40. Malorny, B.; Cook, N.; D’Agostino, M.; De Medici, D.; Croci, L.; Abdulmawjood, A.; Fach, P.; Karpiskova, R.; Aymerich, T.; Kwaitek, K.; et al. Multicenter validation of PCR-based method for detection of Salmonella in chicken and pig samples. J. AOAC Int. 2004, 87, 861–866. [Google Scholar] [CrossRef] [Green Version]
  41. Boddicker, J.D.; Knosp, B.M.; Jones, B.D. Transcription of the Salmonella invasion gene activator, hilA, requires HilD activation in the absence of negative regulators. J. Bacteriol. 2003, 185, 525–533. [Google Scholar] [CrossRef] [Green Version]
  42. Campioni, F.; Moratto Bergamini, A.M.; Falcão, J.P. Genetic diversity, virulence genes and antimicrobial resistance of Salmonella Enteritidis isolated from food and humans over a 24-year period in Brazil. Food Microbiol. 2012, 32, 254–264. [Google Scholar] [CrossRef]
  43. Libby, S.J.; Lesnick, M.; Hasegawa, P.; Kurth, M.; Belcher, C.; Fierer, J.; Guiney, D.G. Characterization of the spv locus in Salmonella enterica serovar Arizona. Infect. Immun. 2002, 70, 3290–3294. [Google Scholar] [CrossRef] [Green Version]
  44. Ammar, A.M.; Mohamed, A.A.; El-Hamid, M.I.A.; El-Azzouny, M.M. Virulence genotypes of clinical Salmonella serovars from broilers in Egypt. J. Infect. Dev. Ctries. 2016, 10, 337–346. [Google Scholar] [CrossRef] [Green Version]
  45. Gharieb, R.M.; Tartor, Y.H.; Khedr, M.H.E. Non-Typhoidal Salmonella in poultry meat and diarrhoeic patients: Prevalence, antibiogram, virulotyping, molecular detection and sequencing of class I integrons in multidrug resistant strains. Gut Pathog. 2015, 7, 34. [Google Scholar] [CrossRef] [Green Version]
  46. Ho, P.L.; Wong, R.C.; Lo, S.W.; Chow, K.H.; Wong, S.S.; Que, T.L. Genetic identity of aminoglycoside-resistance genes in Escherichia coli isolates from human and animal sources. J. Med. Microbiol. 2010, 59, 702–707. [Google Scholar] [CrossRef] [PubMed]
  47. Cloeckaert, A.; Praud, K.; Doublet, B.; Bertini, A.; Carattoli, A.; Butaye, P.; Imberechts, H.; Bertrand, S.; Collard, J.M.; Arlet, G.; et al. Dissemination of an extended-spectrum-β-lactamase blaTEM-52 gene-carrying IncI1 plasmid in various Salmonella enterica serovars isolated from poultry and humans in Belgium and France between 2001 and 2005. Antimicrob. Agents Chemother. 2007, 51, 1872–1875. [Google Scholar] [CrossRef] [Green Version]
  48. Wachino, J.I.; Shibayama, K.; Kurokawa, H.; Kimura, K.; Yamane, K.; Suzuki, S.; Shibata, N.; Ike, Y.; Arakawa, Y. Novel plasmid-mediated 16S rRNA m1A1408 methyltransferase, NpmA, found in a clinically isolated Escherichia coli strain resistant to structurally diverse aminoglycosides. Antimicrob. Agents Chemother. 2007, 51, 4401–4409. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  49. Martinez-Urtaza, J.; Liebana, E. Use of pulsed-field gel electrophoresis to characterize the genetic diversity and clonal persistence of Salmonella Senftenberg in mussel processing facilities. Int. J. Food Microbiol. 2005, 105, 153–163. [Google Scholar] [CrossRef] [PubMed]
  50. Michael, G.B.; Schwarz, S. Antimicrobial resistance in zoonotic nontyphoidal Salmonella: An alarming trend? Clin. Microbiol. Infect. 2016, 22, 968–974. [Google Scholar] [CrossRef] [PubMed]
  51. Giacometti, F.; Pezzi, A.; Galletti, G.; Tamba, M.; Merialdi, G.; Piva, S.; Serraino, A.; Rubini, S. Antimicrobial resistance patterns in Salmonella enterica subsp. enterica and Escherichia coli isolated from bivalve molluscs and marine environment. Food Control 2021, 121, 107590. [Google Scholar] [CrossRef]
  52. Álvarez-Fernández, E.; Alonso-Calleja, C.; García-Fernández, C.; Capita, R. Prevalence and antimicrobial resistance of Salmonella serotypes isolated from poultry in Spain: Comparison between 1993 and 2006. Int. J. Food Microbiol. 2012, 153, 281–287. [Google Scholar] [CrossRef]
  53. EFSA. The European Union summary report on antimicrobial resistance in zoonotic and indicator bacteria from humans, animals and food in 2015. EFSA J. 2017, 15, e04694. [Google Scholar] [CrossRef] [Green Version]
  54. Prioritarias, T. Boletín Epidemiolóxico de Galicia; Galician Healthcare Service: Galicia, Spain, 2018; Volume 27, pp. 1–23. [Google Scholar]
  55. European Medicines Agency. Sales of Veterinary Antimicrobial Agents in 31 European Countries in 2018: Trends from 2010–2018; European Medicines Agency: Amsterdam, The Netherlands, 2019; ISBN 9789291550685.
  56. Collignon, P.C.; Conly, J.M.; Andremont, A.; McEwen, S.A.; Aidara-Kane, A.; Griffin, P.M.; Agerso, Y.; Dang Ninh, T.; Donado-Godoy, P.; Fedorka-Cray, P.; et al. World Health Organization Ranking of Antimicrobials According to Their Importance in Human Medicine: A Critical Step for Developing Risk Management Strategies to Control Antimicrobial Resistance from Food Animal Production. Clin. Infect. Dis. 2016, 63, 1087–1093. [Google Scholar] [CrossRef] [Green Version]
Microorganisms 10 00126 g001 550
Figure 1. Geographical area for mussel sample collection.
Figure 1. Geographical area for mussel sample collection.
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Microorganisms 10 00126 g002 550
Figure 2. Virulence profiles obtained for the 27 isolates analyzed, based on the presence of the genes invA, hilA sopB, pefA and spvC.
Figure 2. Virulence profiles obtained for the 27 isolates analyzed, based on the presence of the genes invA, hilA sopB, pefA and spvC.
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Microorganisms 10 00126 g003 550
Figure 3. Antibiotic resistance profiles obtained from the 27 isolates.
Figure 3. Antibiotic resistance profiles obtained from the 27 isolates.
Microorganisms 10 00126 g003
Table
Table 1. List of primers for virulence and antimicrobial resistance genes.
Table 1. List of primers for virulence and antimicrobial resistance genes.
PrimersTarget GeneSequence (5′-3′)Reference
invA-finvAGTG AAA TTA TCG CCA CGT TCG GGC AA[19]
invA-rTCA TCG CAC CGT CAA AGG AAC C
spvC-1spvCCGG AAA TAC CAT CTA CAA ATA[18]
spvC-2CCC AAA CCC ATA CTT ACT CTG
pefA1 pefATGT TTC CGG GCT TGT GCT[20]
pefA2CAG GGC ATT TGC TGA TTC TTC C
hilA DShilACGG AAG CTT ATT TGC GCC ATG CTG AGG TAG[21]
hilA USGCA TGG ATC CCC GCC GGC GAG ATT GTG
sopB PRSB1sopBCCA CCG TTC TGG GTA AAC AAG AC[22]
sopB PRSB2AGG ATT GAG CTC CTC TGG CGA T
armA-farmATAT GGG GGT CTT ACT ATT CTG CCTAT[23]
armA-rTCT TCC ATT CCC TTC TCC TTT
rmtA-f rmtACTA GCG TCC ATC CTT TCC TC
rmtA-rTTT GCT TCC ATG CCC TTG CC
rmtB-frmtBTCA ACG ATG CCC TCA CCT C
rmtB-rGCA GGG CAA AGG TAA AAT CC
rmtC-frmtCGCC AAA GTA CTC ACA AGT GG
rmtC-rCTC AGA TCT GAC CCA AC AAG
rmtD-frmtDCTG TTT GAA GCC AGC GGA ACG C
rmtD-rGCG CCT CCA TCC ATT CGG AAT AG
npmA-fnpmACTC AAA GGA ACA AAG ACG G
npmA-rGAA ACA TGG CCA GAA ACT C
CTX-M-15-F1blaCTX-M-15ATA AAA CCG GCA GCG GTG[24]
CTX-M-15-F2GAA TTT TGA CGA TCG GGG
Table
Table 2. Thermal profiles set for each genetic target.
Table 2. Thermal profiles set for each genetic target.
GeneHot-StartCyclesDenaturalizationHybridizationExtensionFinal ExtensionFragment Size (bp)
invA94 °C/5 min3093 °C/1 min42 °C/1 min72 °C/2 min72 °C/4 min284
spvC669
pefA94 °C/5 min2594 °C/55 s55 °C/55 s72 °C/55 s72 °C/10 min700
hilA94 °C/3 min3094 °C/1 min65 °C/1 min72 °C/1 min72 °C/10 min854
sopB94 °C/5 min3094 °C/1 min55 °C/1 min72 °C/2 min72 °C/10 min1348
armA94 °C/5 min3094 °C/1 min55 °C/1 min72 °C/2 min72 °C/10 min514
rmtA635
rmtB459
rmtC752
rmtD375
npmA641
blaCTX-M-15483
Table
Table 3. Salmonella strains isolated in the present study.
Table 3. Salmonella strains isolated in the present study.
CodeStrainYear of IsolationOrigin
AMC 28S. montevideo *2012Ria de Arousa
AMC 90S. rissen *2014Ria de Arousa
AMC 92Salmonella spp.2014Ria de Arousa
AMC 93Salmonella spp.2014Ria de Arousa
AMC 200S. wentworth *2014Ria de Arousa
AMC 238S. typhimurium *2015Ria de Arousa
AMC 239S. rissen *2015Ria de Arousa
AMC 240S. rissen *2015Ria de Arousa
AMC 256Salmonella spp.2015Ria de Arousa
AMC 257S. offa *2015Ria de Arousa
AMC 265S. montevideo *2015Ria de Arousa
AMC 266S. senftenberg *2015Ria de Arousa
AMC 267S. senftenberg *2015Ria de Arousa
AMC 268S. typhimurium *2015Ria de Vigo
AMC 270S. agona *2015Ria de Vigo
AMC 281Salmonella spp.2015Ria de Arousa
AMC 287Salmonella spp.2015Ria de Arousa
AMC 288Salmonella spp.2015Ria de Arousa
AMC 289S. senftenberg *2015Ria de Arousa
AMC 290 Salmonella spp.2015Ria de Arousa
AMC 291 S. typhimurium *2015Ria de Arousa
AMC 294 S. typhimurium *2015Ria de Arousa
AMC 299 S. typhimurium *2015Ria de Arousa
AMC 300 Salmonella spp.2015Ria de Vigo
AMC 301 S. bredeney *2015Ria de Arousa
AMC 303 Salmonella spp.2016Ria de Vigo
AMC 327 S. liverpool *2016Ria de Vigo
* Serotype information obtained from Lozano-Leon et al. [27] in this reference additional molecular information may be obtained for the designated strains.
Table
Table 4. Summary of the antibiotic resistance and virulence genes for each of the strains isolated.
Table 4. Summary of the antibiotic resistance and virulence genes for each of the strains isolated.
AntibioticsVirulence Genes
StrainAMC CodeAreaAMPAMCCEFCXMCXM/axetilFOXCDNCTXCAZFEPETPIPMGENTOBNALCIPFOFNITSXTAMKTGCTZPinvAspvCpefAhilAsopBarmArmtArmtBrmtCrmtDnpmAblaCTX-M-15
S. montevideo *28ASSRRRRSSSSSSRRSSSSSNNN+
S. rissen *90ARSRRRRSSSSSSRRSSSSRNNN+
Salmonella spp.92ARRNRRRNRIISSRNSSNNRRSI+
Salmonella spp.93ARINRRRNSSSSSRNSSNNRRSS+
S. wentworth *200ASSRRRRSSSSSSRRSSSSSNNN+
S. typhimurium *238ARSNRRRNSSSSSRNSSNNSRSS+
S. rissen *239ARSNRRRNSSSSSRNSSNNSRSS+
S. rissen *240ARSNRRRNSSSSSRNSSNNSRSS+
Salmonella spp.256ARSNRRRNSSSSSRNSSNNRRSS+ ++
S. offa *257ASSNRRRNSSSSSRNSSNNSRSS+
S. montevideo *265ASSNRRRNSSSSSRNSSNNSRSS+
S. senftenberg *266ASSRRRRSSSSSSRRSSSSSNNN+ ++
S. senftenberg *267ASSRRRRSSSSSSRRSSSSSNNN++ +
S. typhimurium *268VSSRRRRSSSSSSRRSSSSSNNN+++++
S. agona *270VRSRRRRSSSSSSRRRRSISNNN+ +
Salmonella spp.281ARSRRRRSSSSSSRRRRSISNNN++ ++
Salmonella spp.287ASSNRRRNSSSSSRNSSNNSRSS++
Salmonella spp.288ASSNRRRNSSSSSRNSSNNSRSS+
S. senftenberg *289ASSNRRRNSSSSSRNSSNNSRSS++
Salmonella spp.290ASSNRRRNSSSSSRNSSNNSRSS+ ++
S. typhimurium *291VSSRRRRSSSSSSRRSSSSSNNN++
S. typhimurium *294ARINRRSNSSSSSSNSSNNRSSI+
S. typhimurium *299ASSNRRRNSSSSSRNSSNNSRSS+
Salmonella spp.300VSSNRRRNSSSSSRNSSNNSRSS+
S. bredeney *301ASSNRRRNSSSSSRNSSNNSRSS+
Salmonella spp.303VSSNRRRNSSSSSRNSSNNSRSS+ ++
S. liverpool *327VSSNRRRNSSSSSRNSSNNSRSS+
“S”: Sensitive, “R”: Resistant, “I”: Intermediate and “N”: Not tested. “+”: positive result for the corresponding gene. * Serotype information obtained from Lozano-Leon et al. [27]. In this reference, additional molecular information can be obtained for the designated strains AMP: ampicillin; AMC: Amoxicillin/clavulanic acid; CEF: cephalothin; CXM: cefuroxime; FOX: cefoxitin; CDN: cefditoren; CTX: cefotaxime; CAZ: ceftazidime; FEP: cefepime; ETP: ertapenem; IPM: imipenem; GEN: gentamicin; TOB: tobramycin; NAL: nalidixic acid; CIP: ciprofloxacin; FOF: fosfomycin; NIT: nitrofurantoin; SXT: trimethoprim-sulfamethoxazole; AMK: amikacin; TGC: tigecycline and TZP: piperacillin-tazobactam.
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Lozano-León, A.; García-Omil, C.; Rodríguez-Souto, R.R.; Lamas, A.; Garrido-Maestu, A. An Evaluation of the Pathogenic Potential, and the Antimicrobial Resistance, of Salmonella Strains Isolated from Mussels. Microorganisms 2022, 10, 126. https://doi.org/10.3390/microorganisms10010126

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Lozano-León A, García-Omil C, Rodríguez-Souto RR, Lamas A, Garrido-Maestu A. An Evaluation of the Pathogenic Potential, and the Antimicrobial Resistance, of Salmonella Strains Isolated from Mussels. Microorganisms. 2022; 10(1):126. https://doi.org/10.3390/microorganisms10010126

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Lozano-León, Antonio, Carlos García-Omil, Rafael R. Rodríguez-Souto, Alexandre Lamas, and Alejandro Garrido-Maestu. 2022. "An Evaluation of the Pathogenic Potential, and the Antimicrobial Resistance, of Salmonella Strains Isolated from Mussels" Microorganisms 10, no. 1: 126. https://doi.org/10.3390/microorganisms10010126

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