Next Article in Journal
Towards Cell-Permeable Hepatitis B Virus Core Protein Variants as Potential Antiviral Agents
Previous Article in Journal
Response of Escherichia coli to Acid Stress: Mechanisms and Applications—A Narrative Review
Previous Article in Special Issue
Exploring Propolis as a Sustainable Bio-Preservative Agent to Control Foodborne Pathogens in Vacuum-Packed Cooked Ham
 
 
Search for Articles:
Title / Keyword
Author / Affiliation / Email
Journal
Article Type
 
 
Advanced Search
 
Section
Special Issue
Volume
Issue
Number
Page
 
Journals
Microorganisms
Volume 12
Issue 9
10.3390/microorganisms12091775
microorganisms-logo
Submit to this Journal Review for this Journal Propose a Special Issue
► Article Menu

Article Menu

share Share announcement Help format_quote Cite question_answer Discuss in SciProfiles thumb_up
...
Endorse textsms
...
Comment
first_page
settings
Order Article Reprints
Font Type:
Arial Georgia Verdana
Font Size:
Aa Aa Aa
Line Spacing:
Column Width:
Background:
Article

Microbiological Quality and Antibiotic Resistance of Relevant Bacteria from Horsemeat

by
Elena Gonzalez-Fandos
* and
Jessica da Silva Guedes
Food Technology Department, CIVA Research Center, University of La Rioja, Madre de Dios 53, 26006 Logroño, La Rioja, Spain
*
Author to whom correspondence should be addressed.
Microorganisms 2024, 12(9), 1775; https://doi.org/10.3390/microorganisms12091775
Submission received: 9 August 2024 / Revised: 20 August 2024 / Accepted: 24 August 2024 / Published: 28 August 2024
(This article belongs to the Special Issue Evaluation of Risks of Microbiological Origin Associated with Food Consumption, Third Edition)
Browse Figures
Versions Notes

Abstract

:
The aim of this work was to assess the microbiological safety and quality of horsemeat. A total of 19 fresh horsemeat samples were analysed. Mesophile counts were 4.89 ± 1.08 log CFU/g, and Enterobacteriaceae, Staphylococcus spp., and enterococci were only isolated from 36.84%, 21.05%, and 15.79% of the samples, respectively. Neither Staphylococcus aureus nor Escherichia coli were found in any sample. Listeria spp. and Listeria monocytogenes were detected in 31.58% and 21.05% of the samples, respectively. Campylobacter jejuni was not detected in any sample. The dominant bacteria were lactic acid bacteria. Seven different Staphylococcus spp. were identified, the most common being S. delphini, S. saprophyticus, and S. warneri. S. delphini showed resistance against mupirocin and cefoxitin. All the L. monocytogenes strains showed resistance against ampicillin, cefotaxime, and oxacillin. Multi-resistant Yersinia enterocolitica, Stenotrophomonas maltophilia, and Vagococcus. fluvialis strains were found, with resistance to 11, 7, and 8 antibiotics, respectively, causing significant concern. Therefore, specific actions should be taken to decrease the contamination of horsemeat.

1. Introduction

Horsemeat has a high protein content, high levels of n-3 fatty acids, and it is a good source of minerals and vitamins [1,2]. Moreover, horsemeat is low fat and has a high unsaturated fatty acid content, as well as a high iron content compared with that of other meats, and as such, it is considered as a healthy alternative to other meats [1]. Due to its high iron content, it is of interest for use in the treatment of human iron-deficiency anaemia diseases [3]. Furthermore, horsemeat has attractive sensory characteristics [2]. However, the consumption of horsemeat is limited, as it is affected by cultural and ethical biases [1]. The highest consumption of horsemeat is found in countries in Central Asia, some countries in Europe and Mexico [4]. In 2020, 0.8 million tonnes of horsemeat were produced worldwide [4]. In the European Union, Spain is one of the largest producers of horsemeat, followed by Poland and Italy [4]. In 2022, the production of horsemeat in Spain amounted to 8246 tonnes [5].
Since horsemeat is not a common food in many countries, studies on the microbiological quality and safety of this type of meat are scarce [6,7,8]. Most of the studies are focused on lactic acid bacteria, pseudomonas, and Enterobacteriaceae [6,7,8,9]. Pathogenic bacteria such as Escherichia coli and Listeria monocytogenes can be present in horsemeat [9,10,11]. Since Yersinia enterocolitica and Staphylococcus aureus have often been found in horses, contamination of meat can occur during processing, depending on the hygienic conditions of the processing plant [10,12,13].
Since food-borne bacteria that are resistant to antibiotics could have an impact on human health, bacterial monitoring is crucial [14]. There is a great concern regarding methicillin-resistant S. aureus (MRSA), vancomycin-resistant enterococci (VRE), colistin-resistant Enterobacteriaceae, as well as extended-spectrum-lactamase (ESBL) and carbapenemase-producing Enterobacteriaceae [15,16,17]. There is no information available on the prevalence of these bacteria in horsemeat. However, ESBL-producing E. coli and methicillin-resistant S. aureus (MRSA) have been found in horses, and they could be transferred to horsemeat during processing [10,18,19].
The aim of this work was to assess the microbiological safety and quality of horsemeat, including the antimicrobial resistance of relevant bacteria.

2. Materials and Methods

2.1. Horsemeat Samples and Microbiological Determinations

A total of 19 fresh horsemeat samples were collected in Logroño (Spain) from two hypermarkets (12 samples from A and 7 samples from B) between June and September 2020 (Table S1, Supplementary Materials). They were transferred to the university facilities under refrigeration and analysed as quickly as possible.
For the microbiological analysis, 10 grams were obtained and blended, as described previously [20]. The microbial groups evaluated, as well as the media used and conditions employed, are shown in Table 1. The methodology used has been described previously [20,21,22,23]. Also, Enterobacteriaceae colistin resistant was determined in ChromID Colistin agar.

2.2. Isolation and Identification

A total of 3 to 5 typical colonies were chosen from each culture medium inoculated with horsemeat samples. Strains were purified and maintained as described previously [20]. Bacterial identification was performed using the MALDI-TOF Biotyper (Bruker, Daltonik, Bremen, Germany).

2.3. Phenotypic Antimicrobial Resistance of Y. enterocolitica Isolates

The resistance of Y. enterocolitica strains was evaluated, using the disk diffusion method, against the following antibiotics (Oxoid, Basingstoke, Hampshire, UK): cefpodoxime (CPD, 10 µg), cefepime (FEP, 30 µg), ceftriaxone (CRO, 30 µg), cefoxitin (FOX, 30 µg), ceftazidime (CAZ, 30 µg), cefotaxime (CTX, 30 µg), ampicillin (AMP, 10 µg), aztreonam (ATM, 30 µg), ampicillin/surbactam (SAM, 10/10 µg), amoxicillin-clavulanate (AUG, 20/10 µg), imipenem (IPM, 10 µg), ertapenem (ETP, 10 µg), doripenem (DOR, 10 µg), meropenem (MEM, 10 µg), trimethoprim/sulfamethoxazole (SXT, 1.25/23.75 µg), piperacillin (PRL, 100 µg), trimethoprim (W, 5 µg), chloramphenicol (C, 30 µg), sulfadiazine (SUZ, 300 µg), minocycline (MH, 30 µg), tetracycline (TE, 30 µg), doxycycline (DO, 30 µg), tigecycline (TE, 30 µg), levofloxacin (LEV, 5 µg), enrofloxacin (ENR, 5 µg), norfloxacin (NOR, 10 µg), ciprofloxacin (CIP, 5 µg), nalidixic acid (NA, 30 µg), gatifloxacin (GAT, 5 µg), amikacin (AK, 30 µg), kanamycin (K, 30 µg), gentamicin (CN, 10 µg), streptomycin (S, 10 µg), nitrofurantoin (F, 300 µg), and tobramycin (TOB, 10 µg). The strains were classified as resistant, susceptible, or intermediate, according to the CLSI guidelines [24].

2.4. Phenotypic Antimicrobial Resistance of Staphylococcus spp.

The phenotypic antimicrobial resistance of Staphylococcus spp. was evaluated, using the disk diffusion method, against the following antibiotics: cefoxitin (FOX, 30 µg), ceftaroline (CPT, 30 µg), clindamycin (CMN, 2 µg), penicillin (P, 10 UI), fusidic acid (FAD, 10 µg), trimethoprim -sulfamethoxazole (SXT 1.25:23.75 µg), trimethoprim (W, 5 µg), tetracycline (TE, 30 µg), minocycline (MH, 30 µg), enrofloxacin (ENR, 5 µg), ciprofloxacin (CIP, 5 µg), gatifloxacin (GAT, 5 µg), doxycycline (DO, 30 µg), norfloxacin (NOR, 5 µg), levofloxacin (LEV, 5 µg), gentamicin (CN, 10 µg), amikacin (AK, 30 µg), kanamycin (K, 30 µg), erythromycin (ERY, 15 µg), streptomycin (S, 10 UI), tobramycin (TOB,10 µg), sulfadiazine (SUZ, 300 µg), mupirocin (PUM, 200 µg), tylosin (TY, 30 µg), lincomycin (MY, 15 µg), nitrofurantoin (F, 300 µg), chloramphenicol (C, 30 µg), tedizolid (TZD, 2 µg), linezolid (LZD, 30 µg), vancomycin (VA, 30 µg), and rifampicin (RD, 5 µg). The strains were classified as resistant, susceptible, or intermediate, according to the CLSI guidelines [24].

2.5. Phenotypic Antimicrobial Resistance of Enterococci

The resistance of enterococci was evaluated, using the disk diffusion method, against the following antibiotics: norfloxacin (NOR, 5 µg), levofloxacin (LEV, 5 µg), enrofloxacin (ENR, 5 µg), ciprofloxacin (CIP, 5 µg), vancomycin (VA, 30 µg), chloramphenicol (C, 30 µg), doxycycline (DO, 30 µg), tetracycline (TE, 30 µg), teicoplanin (TEC, 30 µg), tigecycline (TGC, 15 µg), linezolid (LZD, 30 µg), gentamicin (CN, 120 µg), ampicillin (AMP, 10 µg), nitrofurantoin (F, 300 µg), imipenem (IPM, 5 µg), and minocycline (MH, 30 µg). The strains were classified as resistant, susceptible, or intermediate, according to the CLSI guidelines [24].

2.6. Phenotypic Antimicrobial Resistance of L. monocytogenes

The phenotypic antimicrobial resistance of of L. monocytogenes was evaluated, using the disk diffusion method, against the following antibiotics: amikacin (AK, 30 µg), streptomycin (S, 10 µg), amoxycillin/clavulanic acid (AUG, 30 µg), gentamicin (CN, 10 µg), ampicillin (AMP, 2 µg), tobramycin (TOB, 10 µg), penicillin G (PNG, 10 µg), cefotaxime (CTX, 30 µg), oxacillin (OX, 1 µg), ceftaroline (CPT, 30 µg), meropenem (MEM, 10 µg), imipenem (IPM, 10 µg), ciprofloxacin (CIP, 5 µg), levofloxacin (LEV, 5 µg), enrofloxacin (ENR, 5 µg), norfloxacin (NOR, 10 µg), minocycline (MH, 30 µg), doxycycline (DO, 30 µµg), tetracycline (TE, 30 µg), teicoplanin (TEC, 30 µg), tigecycline (TGC, 15 µg), chloramphenicol (C, 30 µg), linezolid (LZD, 30 µg), vancomycin (VA, 30 µg), erythromycin (ERY, 15 µg), quinupristin/dalfopristin (QD, 15 µg), nitrofurantoin (F, 300 µg), trimethoprim/sulphamethoxazole 1:19 (SXT, 25 µg), and rifampicin (RD, 5 µg). The strains were classified as resistant, susceptible, or intermediate, according to the CLSI guidelines [24].

2.7. Phenotypic Antimicrobial Resistance of Stenotrophomonas maltophilia

The phenotypic antimicrobial resistance of Stenotrophomonas maltophilia was evaluated, using the disk diffusion method, against the following antibiotics: ampicillin (AMP, 10 µg), aztreonam (ATM, 30 µg), cefazoline (CZ, 30 µg), cefotaxime (CTX, 30 µg), cefpodoxime (CPD, 10 µg), ceftazidime (CAZ, 30 µg), ciprofloxacin (CIP, 5 µg), chloramphenicol (C, 30 µg), doxycycline (DO, 30 µg), enrofloxacin (ENR, 5 µg), fosfomycin (FOS, 200 µg), gentamicin (CN, 10 µg), imipenem (IPM, 10 µg), levofloxacin (LEV, 5 µg), minocycline (MH, 30 µg), netilmicin (NET, 30 µg), norfloxacin (NOR, 10 µg), piperacillin (PRL, 100 µg), sulfadiazine (SUZ, 300 µg), tetracycline (TE, 30 µg), ticarcillin-clavulanic (TTC, 75/10 µg), tobramycin (TOB, 10 µg), and trimethoprim -sulfamethoxazole (SXT, 1.25:23.75 µg). The strains were classified as resistant, susceptible, or intermediate, according to the CLSI guidelines [24].

2.8. Phenotypic Antimicrobial Resistance of Vagococcus fluvialis

The phenotypic antimicrobial resistance of Vagococcus fluvialis was evaluated, using the disk diffusion method, against the following antibiotics: amikacin (AK, 30 µg), ampicillin (AMP, 10 µg), ceftriaxone (CRO, 30 µg), ciprofloxacin (CIP, 5 µg), clindamycin (CMN, 30 µg), chloramphenicol (C, 30 µg), doxycycline (DO,30 µg), enrofloxacin (ENR, 5 µg), erythromycin (ERY, 15 µg), streptomycin (S, 300 µg), fosfomycin (FOS, 200 µg), gentamicin (CN, 10 µg), imipenem (IPM, 10 µg), levofloxacin (LEV, 5 µg), linezolid (LZD, 30 µg), minocycline (MH, 30 µg), nitrofurantoin (F, 300 µg), norfloxacin (NOR, 10 µg), oxacillin (OX, 1 µg), penicillin (P, 10 UI), quinupristin/dalfopristin (QD, 15 µg), rifampicin (RD, 5 µg), sulfadiazine (SUZ, 300 µg), teicoplanin (TEC, 30 µg), tetracycline (TE, 30 µg), tigecycline (TGC, 15 µg), tobramycin (TOB, 10 µg), trimethoprim-sulfamethoxazole (SXT, 1.25:23.75 µg), and vancomycin (VA, 30 µg). The strains were classified as resistant, susceptible, or intermediate, according to the CLSI guidelines [24].

2.9. Statistical Analysis

The analysis of variance was performed using SPSS version 26 software (IBM SPSS S3tatistics). Tukey’s test for comparison of means was carried out using the same program. The level of significance was determined at p < 0.05.

3. Results

Table 2 shows the distribution of microbial counts in horsemeat. The mean counts for mesophiles were 4.89 ± 1.08 log CFU/g. Mesophile counts were below 7 log CFU/g, except in one sample from hypermarket B; counts varied between 2.78 and 7.03 log CFU/g. Pseudomonas populations under 1 log CFU/g were obtained in 9 horsemeat samples (47.37%). The counts in the other 10 samples varied between 1.30 and 2.84 log CFU/g. Enterobacteriaceae populations under 1 log CFU/g were obtained in 12 horsemeat samples (63.16%). The counts in the other 7 samples varied between 1.78 and 4.10 log CFU/g. Staphylococcus spp. populations under 1 log CFU/g were obtained in 15 horsemeat samples (78.95%). The counts in the other 4 samples varied between 1.30 and 3.41 log CFU/g. All the samples from hypermarket B showed Enterococcus spp. counts below 1 log CFU/g, while 9 of 12 samples from hypermarket A shown counts below this value, and the other 3 samples showed counts between 1.30 and 1.60 log CFU/g. No significant differences (p > 0.05) in microbial counts were found between hypermarket A and B.
The counts of Listeria spp. and L. monocytogenes were below 1 log CFU/g in all the samples. Listeria spp. was detected in 6 samples (31.58%), 4 from hypermarket A and 2 from hypermarket B. L. monocytogenes was detected in 4 samples (21.05%), 3 from hypermarket A, and 1 from hypermarket B. L. welshimeri was isolated in 1 sample from hypermarket A, while L. innocua was isolated in 1 sample from hypermarket B. Campylobacter spp. was not found in any sample.
Table 3 shows the bacteria isolated in plate count agar (PCA) in samples taken from hypermarket A and B. In both hypermarkets, the predominant group was lactic acid bacteria (42.86–56.67%). In the hypermarket A samples, C. divergens, Lactobacillus spp., L. lactis, and C. maltaromaticum were identified. On the other hand, in the hypermarket B samples, Lactococcus piscium and Lactobacillus sakei were also found. Brochothrix thermosphacta was found in both hypermarkets A and B, with prevalence of 10.2% and 3.33%, respectively.
Pseudomonas spp. and Enterobacteriaceae accounted for 14.28% and 8.16% of isolates from hypermarket A, respectively. In hypermarket B, Pseudomonas spp. accounted for 20% of the isolates, while Enterobacteriaceae only represented 3.33% of the isolates. P. fragi, P. extremorientalis, and S. proteamaculans were isolated in samples from both hypermarkets. However, the species P. gessardii, P. lundensis, and Buttiauxella gaviniae were only isolated in horsemeat from hypermarket A, while P. fluorescens and P. antarctica were only found in samples from hypermarket B.
Acinetobacter spp., Chryseobacterium spp., Stenotrophomonas spp., Kocuria spp., and Microbaterium spp. were isolated in a lesser extent (2.04–4.08%).
In relation to Pseudomonas spp. isolated from specific chromogenic medium for the isolation of this bacterium, five species were identified in hypermarket A: P. extremorientalis, P. fluorescens, P. fragi, P. rhodesiae, and P. chlororaphis, while in hypermarket B, only P. veronii was found (Figure 1).
In relation to Enterobacteriaceae in horsemeat, in both hypermarkets Serratia liquefaciens and Hafnia alvei were identified. In the horsemeat from hypermarket A, the predominant genus was Serratia, with 50% of the isolates. In the case of horsemeat from hypermarket B, Hafnia alvei was the predominant species, with 60% of the isolates. The species B. warmboldiae, P. agglomerans, P. alcalifaciens, and B. gaviniae were found exclusively in hypermarket A (5.56–22.21%), while R. terrigena was isolated from samples of hypermarket B (20%). E. coli was not isolated in any sample (Figure 2).
When the medium ChromID ESBL was used, E. coli was not isolated. When using ChromID CARBA, Enterobacteriaceae was not isolated, but Stenotrophomonas maltophilia was found in one sample from hypermarket A. This bacterium was also isolated from PCA in the same sample. These two strains of Stenotrophomonas maltophilia showed resistance against ampicillin, aztreonam, cefotaxime, cefpodoxime, cefazolin, imipenem, and piperacillin (Table 4).
When using ChromID colistin, Yersinia enterocolitica was isolated in one sample from hypermarket A. This strain showed resistance against 12 antibiotics: amoxicillin/clavulanic, ampicillin, ampicillin/sulbactan, aztreonan, cefepime, cefotaxime, cefoxitin, cefpodoxime, ceftriaxone, nalidixic acid, nitrofurantoin, and trimethoprim (Table 4).
Figure 3 shows the Staphylococcus spp. identified in horsemeat samples. The species isolated most often from hypermarket A was S. delphini (42.84%), while in hypermarket B, the most often identified species were S. saprophyticus (50%) and S. warneri (50%). S. equorum, S. fleurettii, S. succinus, and S. warneri were also isolated from hypermarket A.
All of the isolated Staphylococcus spp. strains were susceptible to all the antibiotics assayed, except for two. One strain of S. delphini showed resistance against mupirocin and cefoxitin, while one strain of S. saprophyticus showed resistance against doxycycline, penicillin, and tetracycline (Table 4). When using ChromID MRSA, S. aureus was not isolated. However, V. fluvialis was isolated in a sample from hypermarket A. This strain showed resistance against clindamycin, linezolid, nitrofurantoin, oxacillin, penicillin, quinupristin/dalfopristin, teicoplanin, and vancomycin (Table 4).
Enterococcus malodoratus was the only enterococci isolated from horsemeat that was susceptible to all the antibiotics tested (Table 4). None enterococci was isolated from ChromID VRE agar.
All of the isolated L. monocytogenes strains were resistant to ampicillin, cefotaxime, and oxacillin. Resistance against bezilpenicillin, meropenem, and nitrofurantoin was found in 75%, 50%, and 25% of the strains isolated, respectively (Table 4).

4. Discussion

The mean counts of mesophiles obtained in this work (4.89 ± 1.08 log UFC/g) were higher than those reported by Pavlidis et al. and Gomez and Lorenzo (2.77 ± 0.10 log CFU/g and 4.13 log CFU/g, respectively) [6,25]. Higher pseudomonas counts have been reported by Pavlidis et al. (2021) (2.72 ± 0.17 log10 CFU/g) and Lorenzo and Gomez (4.24 log10 CFU/g) [6,8].
As noted in the current work, other researchers have pointed out that the dominant bacteria in horsemeat from Belgian markets were lactic acid bacteria [7]. In contrast, other authors observed that the dominant bacteria in samples from the European market were pseudomonads, followed by B. thermosphacta [6]. These differences can be explained by the storage temperature and the environmental packaging conditions [8,26]. Also, Geeraerts et al. found that Carnobacterium divergens and Lactobacillus spp. were the dominant lactic acid bacteria in fresh horsemeat [5]. Lactobacillus sakei, Lactococcus piscium, and B. thermosphacta have been also identified by other authors in fresh horsemeat [7]. However, these authors did not isolate C. maltaromaticum from fresh horsemeat [7].
The presence of Pseudomonas spp. is associated with the spoilage of fresh meat, as it is able to develop under non-aerobic conditions [27]. In this study, the species P. fragi, P. extremorientalis, P. gessardii, P. fluorescens, P. antárctica, P. rhodesiae, P. chlororaphis, and P. veronii were identified. These species are frequently found in meat from other animals, as well as on surfaces where meat is handled, some of them being able to form biofilms [28,29].
We also isolated Chryseobacterium spp.; this microorganism has been found in the reproductive system of horses [30].
We only found Enterobacteriaceae counts above 1 log CFU/g in seven samples (36.84%), six from hypermarket A (50%) and one from hypermarket B (14.29%). These results agree with those reported by Geeraerts et al., who only detected Enterobacteriaceae in 11.76% of the samples analysed from the Belgian market, with Hafnia alvei being the only species isolated [7]. In contrast, Furuhata et al. found Enterobacteriaceae in 93.8% of the horsemeat samples from Japan [9]. Furuhata et al. pointed out that the dominant bacterium was Hafnia alvei (19.8%), as we observed in samples from hypermarket B [9]. These authors identified 14 Enterobacteiaceae species, while we only identified 9 different species. As did Furuhata et al., we also isolated Raultella terrigena and Serratia liquefaciens from horsemeat [9]. However, we did not isolate Klebsiella pneuminiae, Enterobacte cloacae, Pantoea spp., Enterobacter ammigenus, Klebsiella oxytoca, Citrobacter yungae, E. coli, or Proteus mirabilis. However, we found other species, such as Serratia liquefaciens, Buttiauxella warmboldiae, Pantoea agglomerans, Providencia alcalifaciens, Buttiauxella gaviniae, Serratia proteamaculans, and Yersinia enterocolitica. We did not detect E. coli in any sample. The differences found could be explained by the varying hygienic conditions, since Enterobacteriaceae are considered as a hygiene indicator associated with faecal contamination [31]. E. coli has been isolated in faecal samples from horses, and faecal meat contamination is possible during slaughter [31].
Other authors have pointed out that contamination of horsemeat with Yersinia enterocolitica could be frequent, since this bacterium is often found in horse faecal samples, and contamination of meat could occur [12]. As in the present work, Seekamela et al. also found Y. enterocolitica strains resistant to ampicillin that were isolated from beef and pork meat in South Africa [32]. However, we did not find resistance against tetracycline, chloramphenicol, aztreonam, imipenem, gentamycin, piperacillin, or amikacin. In agreement with the results of Seekamela et al., we did not find resistance against trimethoprim-sulphamethoxazole [32]. We did observe resistance against amoxicillin/clavulanic, ampicillin/surbactam, cefepime, cefotaxime, cefoxitin, cefpodoxime, ceftriaxone, nalidixic acid, and nitrofurantoin. Terentjeva et al. found resistance to ampicillin, but not to cefotaxime and trimethoprim, in Y. enterocolitica strains isolated from beef and pork in Latvia [33]. Resistance to amoxicillin/clavulanic has also been reported in Y. enterocolitica strains isolated from beef and poultry. It should be note that we found resistance to aztreonam, an antibiotic that is categorized in “Category A: antimicrobial to avoid” for animals [34].
We only isolated Staphylococcus spp. from four samples (21.05%). Other authors have pointed out that the dominant example of Staphylococcus spp. in fresh horsemeat from the Belgian market is S. equorum, instead of S. delphini and S. saprophyticus, as was observed in the present work [7]. Geeraerts et al. isolated S. simulans and S. xylosus, species that were not found in the present work [7]. However, we isolated delphini, S. fleurettii, S. succinus, and S. warneri. S. delphini and S equorum have been isolated from the skin of horses, and in consequence, they could contaminate meat during processing [35,36,37]. We did not isolate S. aureus from horsemeat, although other authors have pointed out that horses are a reservoir of this microorganism, and MRSA strains have even been isolated from horse [13,18,38,39].
Low resistance rates were found in the Staphylococcus spp. strains. We only observed one strain of S. delphini that showed resistance against mupirocin and cefoxitin, while one strain of S. saprophyticus showed resistance against doxycycline, penicillin, and tetracycline. Penicillin is one of the most common antibiotics used in the treatment of horse infections and in consequence, resistance to this antibiotic could be found [30]. The resistance to tetracyclines has also been reported by other authors in staphylococci strains isolated from horses in France and Canada [40,41]. It should be noted that we found resistance to mupirocin, an antibiotic that is categorized in “Category A: antimicrobial to avoid” for animals [34].
Low enterococci levels were found in the present work, and enterococci was identified in only 15.79% of the samples. Higher percentages of samples showing the presence of enterococci have been reported in goat, pork, and poultry meat [20,42]. The only species found in the present work was E. malodoratus, and this isolated strain showed susceptibility to all the antibiotics tested.
A lower prevalence of Listeria spp. and L monocytogenes has been reported by Assis et al. in horsemeat from Brazil (18.2% and 7.4%, respectively, versus 31.58% and 21.05% found in the present work) [11]. Also, a lower prevalence of L monocytogenes has been reported in poultry and pork meat [43,44,45]. In the current study, all the samples showed counts below 2 log CFU/g. High resistance rates of L. monocytogenes against ampicillin, cefotaxime, and oxacillin were observed (100%). Other authors have also found resistance against oxacillin and ampicillin, cefotaxime, meropenem, benzylpenicillin, and nitrofurantoin in L. monocytogenes strains isolated from beef, pork, and poultry meat in Spain and China [43,46]. It should be taken into account that ampicillin and benzylpenicillin are commonly used in the treatment of infections caused by L. monocytogenes [47]. It should also be noted that we found resistance to aztreonam, which is categorized in “Category A: antimicrobial to avoid” for animals [34].
As in the present work Collobert et al. did not isolate Campylobacter spp. from horse carcasses [48]. Several studies indicate that the presence of Campylobacter spp. in horse faeces is low; consequently, the contamination of horsemeat during processing is low compared to that of poultry meat [49].
Stenotrophomonas maltophilia is considered as an emerging Gram-negative multi-resistant bacterium [50,51]. This microorganism has been associated with respiratory and urinary human infections, as well as respiratory infections in horses [51,52]. S. maltophilia has been isolated from goat, rabbit, and poultry meat [53,54,55], but there is no information available regarding its presence in horsemeat. However, S. maltophilia has been detected in horse manure from France and Tunisia, and the majority of strains isolated were resistant to 7–9 antibiotics [56]. The high rates of antibiotic resistance in strains isolated from horse manure has been associated with antibiotic presence, since animals are often treated with antibiotics [56]. In the present study, S. maltophilia showed resistance against seven antibiotics: ampicillin, aztreonam, cefotaxime, cefpodoxime, cefazolin, imipenem, and piperacillin. The high resistance rates of S. maltophilia against ceftazidime and imipenem has also been observed in strains isolated from horse manure in France and Tunisia [56]. It should be highlighted the resistance found against aztreonam, which has been recommended for the treatment of infections caused be this microorganism, is categorized in “Category A: antimicrobial to avoid” for animals [34,57].
V. fluvialis has been isolated from beef, pork, and chicken meat [58,59,60,61,62]. The Vagococcus microorganism is considered as an emerging pathogen in humans [63,64,65]. V. fluvvialis infections could be underestimated, since this bacterium is frequently improperly identified as Enterococcus spp. [65]. The identification by means of MALDI-TOF could help to detect this bacterium. Most of the studies on V. fluvialis antibiotic resistance have been carried out using strains isolated from animals or from human infections. There is not data available on V. fluvialis isolated from meat. In the present work, the V. fluvialis strain showed resistance against clindamycin, linezolid, nitrofurantoin, oxacillin, penicillin, quinupristin/dalfopristin, teicoplanin, and vancomycin. Also, Matajira et al. and Texiera et al. reported that V. fluvialis isolated from pigs in Brazil showed resistance to clindamycin [61,66]. According to Rancero et al., the most active antibiotics against V. fluvialis isolated from human infection were vancomycin, ampicillin, trimethoprim/sulfamethoxazole, linezolid, and teicoplanin, while resistance to fluoroquinolones and tetracyclines was observed in 40 and 80% of the strains, respectively [65]. We observed resistance to vancomycin, teicoplanin, and linezolid. Other studies on strains isolated form human infections reported resistance to trimethoprim/sulfamethoxazole and levofloxacin, while susceptibility to ampicillin, minocycline, vancomycin, and linezolid was observed [63]. Other authors have also reported resistance to clindamycin in strains isolated from human infections [66]. Chen et al. reported susceptibility to tigecycline, vancomycin, quinupristin/dalfopristin, and linezolid in human infection isolates, and moderate sensitivity to ciprofloxacin, levofloxacin, ampicillin/sulbactam, erythromycin, and tetracycline [65]. It should be note that we found resistance to linezolid and vancomycin, antibiotics that are categorized in “Category A: antimicrobial to avoid” for animals [34].

5. Conclusions

The present work advises that horsemeat could be a source of both emerging (V. fluvialis and S. maltophilia) and recognised foodborne pathogens (L. monocytogenes and Y. enterocolitica). Moreover, horsemeat can be a source of multi-resistant bacteria. The presence of multi-resistant Y. enterocolitica, V. fluvialis, and S. maltophilia in horsemeat is of great concern, and special actions to reduce meat contamination should be adopted in the framework of the One Health approach. Adequate processing, handling, cleaning, and disinfecting procedures could help to avoid cross-contamination. Resistance to critical antibiotics, according to the European Medicine Agency (EMA) standards, including mupirocin, aztreonam, linezolid, and vancomycin, was observed in strains isolated from horsemeat.

Supplementary Materials

The following supporting information can be downloaded at: https://www.mdpi.com/article/10.3390/microorganisms12091775/s1, Table S1: Samples taken by place of purchase.

Author Contributions

Conceptualization, E.G.-F.; methodology, E.G.-F.; formal analysis, E.G.-F. and J.d.S.G.; investigation E.G.-F. and J.d.S.G.; resources, E.G.-F.; data curation E.G.-F. and J.d.S.G.; writing—original draft preparation, E.G.-F.; writing—review and editing, E.G.-F.; supervision, E.G.-F.; project administration and funding acquisition, E.G.-F. All authors have read and agreed to the published version of the manuscript.

Funding

This work was 65% cofinanced by the Interreg V-Spain–France–Andorra (POCTEFA 2014–2020) (EFA 152/16). J.S.G. has received a grant from the EU H2020 research and innovation program, Marie Sklodowska-Curie, No. 801586.

Data Availability Statement

The raw data supporting the conclusions of this article will be made available by the authors on request.

Conflicts of Interest

The authors declare no conflicts of interest.

References

  1. Balji, Y.; Knicky, M.; Zamaratskaia, G. Perspectives and safety of horsemeat consumption. Int. J. Food Sci. Technol. 2020, 55, 942–952. [Google Scholar] [CrossRef]
  2. Lorenzo, J.M.; Sarriés, M.V.; Tateo, A.; Polidori, P.; Franco, D.; Lanza, M. Carcass characteristics, meat quality and nutritional value of horsemeat: A review. Meat Sci. 2014, 96, 1478–1488. [Google Scholar] [CrossRef] [PubMed]
  3. Bò, C.D.; Simonetti, P.; Gardana, C.; Riso, P.; Lucchini, G.; Ciappellano, S. Horse meat consumption affects iron status, lipid profile and fatty acid composition of red blood cells in healthy volunteers. Int. J. Food Sci. Nutr. 2013, 64, 147–154. [Google Scholar] [CrossRef] [PubMed]
  4. Food and Agriculture Organization of the United Nations (FAO). FAOSTAT 2022. Available online: https://www.fao.org/faostat/en/#data/QCL (accessed on 31 July 2024).
  5. de Agricultura, M.; Alimentación, P.Y. Anuario de Estadística 2022; MAPA: Madrid, Spain, 2022. [Google Scholar]
  6. Pavlidis, D.E.; Mallouchos, A.; Nychas, G.J. Microbiological assessment of aerobically stored horse fillets through predictive microbiology and metabolomic approach. Meat Sci. 2021, 172, 108323. [Google Scholar] [CrossRef]
  7. Geeraerts, W.; De Vuyst, L.; Leroy, F. Mapping the dominant microbial species diversity at expiration date of raw meat and processed meats from equine origin, an underexplored meat ecosystem, in the Belgian retail. Int. J. Food Microbiol. 2019, 289, 189–199. [Google Scholar] [CrossRef]
  8. Lorenzo, J.M.; Gómez, M. Shelf life of fresh foal meat under MAP, overwrap and vacuum packaging conditions. Meat Sci. 2012, 92, 610–618. [Google Scholar] [CrossRef]
  9. Furuhata, K.; Ishizaki, N.; Sugi, Y.; Fukuyama, M. Isolation and identification of enterobacteriaceae from raw horsemeat intended for human consumption (Basashi). Biocontrol. Sci. 2014, 19, 181–188. [Google Scholar] [CrossRef] [PubMed]
  10. Gill, C.O. Safety and storage stability of horse meat for human consumption. Meat Sci. 2005, 71, 506–513. [Google Scholar] [CrossRef]
  11. Assis, M.A.; Destro, M.T.; Franco, B.D.; Landgraf, M. Incidence of Listeria spp. and Salmonella spp. in horsemeat for human consumption. Int. J. Food Microbiol. 2000, 62, 161–164. [Google Scholar] [CrossRef]
  12. Cattabiani, F.; Ossiprandi, M.C.; Freschi, E. Isolamento di Yersinia enterocolitica da cavalli. L’Igiene Mod. 1995, 103, 675–682. [Google Scholar]
  13. Boyen, F.; Smet, A.; Hermans, K.; Butaye, P.; Martens, A.; Martel, A.; Haesebrouck, F. Methicillin resistant staphylococci and broad-spectrum β-lactamase producing Enterobacteriaceae in horses. Vet. Microbiol. 2013, 167, 67–77. [Google Scholar] [CrossRef]
  14. Colligon, P.J.; McEwen, S.A. One Health. Its importance in helping to better control antimicrobial resistance. Trop. Med. Infect. Dis. 2019, 4, 22. [Google Scholar] [CrossRef] [PubMed]
  15. Dandachi, I.; Chabou, S.; Daoud, Z.; Rolain, J.M. Prevalence and emergence of extended-spectrum cephalosporin, carbapenem and colistin-resistant Gram negative bacteria of animal origin in the Mediterranean basin. Front. Microbiol. 2018, 9, 2299. [Google Scholar] [CrossRef] [PubMed]
  16. David, M.Z.; Daum, R.S. Community-associated methicillin-resistant Staphylococcus aureus: Epidemiology and clinical consequences of an emerging epidemic. Clin. Microbiol. Rev. 2010, 23, 616–687. [Google Scholar] [CrossRef] [PubMed]
  17. Unal, N.; Bal, E.; Kapagoz, A.; ALtun, B.; Kozag, N. Detection of vancomycin-resistant enterococci in samples from broiler flocks and houses. Acta Vet. Hung. 2020, 68, 117–122. [Google Scholar] [CrossRef] [PubMed]
  18. Anwaar, F.; Ijaz, M.; Rasheed, H.; Shah, S.F.A.; Haider, S.A.R.; Sabir, M.J. Evidence and molecular characterization of multidrug resistant Staphylococcus aureus isolated from equines in Pakistan. J. Equine Vet. Sci. 2023, 126, 104498. [Google Scholar] [CrossRef] [PubMed]
  19. Apostolakos, I.; Franz, E.; van Hoek, A.H.A.M.; Florijn, A.; Veenman, C.; Sloet-van Oldruitenborgh-Oosterbaan, M.M.; Dierikx, C.; van Duijkeren, E. Occurrence and molecular characteristics of ESBL/AmpC-producing Escherichia coli in faecal samples from horses in an equine clinic. J. Antimicrob. Chemother. 2017, 72, 1915–1921. [Google Scholar] [CrossRef]
  20. da Silva-Guedes, J.; Martinez-Laorden, A.; Gonzalez-Fandos, E. Effect of the Presence of Antibiotic residues on the microbiological quality and antimicrobial resistance in fresh goat meat. Foods 2022, 11, 3030. [Google Scholar] [CrossRef]
  21. Simón, A.; González-Fandos, E. Influence of modified atmosphere packaging and storage temperature on the sensory and microbiological quality of fresh peeled white asparagus. Food Control. 2011, 22, 369–374. [Google Scholar] [CrossRef]
  22. González-Fandos, E.; Herrera, B. Efficacy of propionic acid against Listeria monocytogenes attached in poultry skin during refrigerated storage. Food Control. 2013, 34, 601–606. [Google Scholar] [CrossRef]
  23. González-Fandos, E.; Martínez-Laorden, A.; Abad-Fau, A.; Sevilla, E.; Bolea, R.; Serrano, M.J.; Mitjana, O.; Bonastre, C.; Laborda, A.; Falceto, M.V.; et al. Effect of intramuscularly administered oxytetracycline or enrofloxacin on vancomycin-resistant enterococci, extended espectrum beta-lactamase and carbapenemase-producing Enterobacteriaceae in pigs. Animals 2022, 12, 622. [Google Scholar] [CrossRef] [PubMed]
  24. Clinical and Laboratory Standards Institute. Performance Standards for Antimicrobial Susceptibility Testing, 30th ed.; CLSI Document M 100; Clinical and Laboratory Standards Institute: Wayne, PA, USA, 2020. [Google Scholar]
  25. Gómez, M.; Lorenzo, J.M. Effect of packaging conditions on shelf-life of fresh foal meat. Meat Sci. 2012, 91, 513–520. [Google Scholar] [CrossRef] [PubMed]
  26. Cullere, M.; Dalle Zotte, A.; Tasoniero, G.; Giaccone, V.; Szendrő, Z.; Szín, M.; Odermatt, M.; Gerencsér, Z.; Dal Bosco, A.; Matics, Z. Effect of diet and packaging system on the microbial status, pH, color and sensory traits of rabbit meat evaluated during chilled storage. Meat Sci. 2018, 141, 36–43. [Google Scholar] [CrossRef] [PubMed]
  27. Watson, S.C.; Furbeck, R.A.; Fernando, S.C.; Chaves, B.D.; Sullivan, G.A. Spoilage Pseudomonas survive common thermal processing schedules and grow in emulsified meat during extended vacuum storage. J. Food Sci. 2023, 88, 2162–2167. [Google Scholar] [CrossRef] [PubMed]
  28. Emamjomeh, M.; Mohd Hashim, A.; Abdul-Mutalib, N.A.; Khairil Mokhtar, N.F.; Mustapha, N.A.; Maeda, T.; Amin-Nordin, S. Profiling bacterial communities and foodborne pathogens on food-associated surface following contact with raw beef, chicken and pork using 16S amplicon metagenomics. Food Control. 2023, 149, 109698. [Google Scholar] [CrossRef]
  29. Zarei, M.; Rahimi, S.; Fazlara, A.; Anvari, S.E. High biofilm-forming Pseudomonas strains isolated from poultry slaughterhouse surfaces: Their importance in the persistence of Salmonella enteritidis in slaughterhouses. Int. J. Food Microbiol. 2023, 390, 110126. [Google Scholar] [CrossRef]
  30. Tyrnenopoulou, P.; Fthenakis, G.C. Clinical Aspects of Bacterial Distribution and Antibiotic Resistance in the Reproductive System of Equids. Antibiotics 2023, 12, 664. [Google Scholar] [CrossRef]
  31. Barco, L.; Belluco, S.; Roccato, A.; Ricci, A. A systematic review of studies on Escherichia coli and Enterobacteriaceae on beef carcasses at the slaughterhouse. Int. J. Food Microbiol. 2015, 207, 30–39. [Google Scholar] [CrossRef]
  32. Seakamela, E.M.; Diseko, L.; Malatji, D.; Makhado, L.; Motau, M.; Jambwa, K.; Magwedere, K.; Ntushelo, N.; Matle, I. Characterization and antibiotic resistance of Yersinia enterocolitica from various meat categories, South Africa’, Onderstepoort. J. Vet. Res. 2022, 89, a2006. [Google Scholar]
  33. Terentjeva, M.; Ķibilds, J.; Meistere, I.; Gradovska, S.; Alksne, L.; Streikiša, M.; Ošmjana, J.; Valciņa, O. Virulence Determinants and Genetic Diversity of Yersinia Species Isolated from Retail Meat. Pathogens 2022, 11, 37. [Google Scholar] [CrossRef]
  34. European Medicine Agency (EMA). Categorisation of Antibiotics for Use in Animals for Prudent and Responsible Use. 2020. Available online: https://www.ema.europa.eu/en/documents/report/infographic-categorisation-antibiotics-use-animals-prudent-and-responsible-use_en.pdf (accessed on 1 August 2024).
  35. Schleifer, K.H.; Kilpper-Balz, R.; Devriese, L.A. Staphylococcus arlettae sp. nov., S. equorum sp. nov. and S. kloosii sp. nov.: Three new coagulase-negative, novobiocin resistant species from animals. Syst. Appl. Microbiol. 1984, 5, 501–509. [Google Scholar] [CrossRef]
  36. Devriese, L.A.; Nzuambe, D.; Godard, C. Identification and characteristics of staphylococci isolated from lesions and normal skin of horses. Vet. Microbiol. 1985, 10, 269–277. [Google Scholar] [CrossRef]
  37. Stull, J.W. Staphylococcus delphini and Methicillin-Resistant S. pseudintermedius in horses, Canada. Emerg. Infect. Dis. 2014, 20, 485–486. [Google Scholar] [CrossRef]
  38. Islam, M.Z.; Espinosa-Gongora, C.; Damborg, P.; Sieber, R.N.; Munk, R.; Husted, L.; Moodley, A.; Skov, R.; Larsen, J.; Guardabassi, L. Horses in Denmark are a reservoir of diverse clones of methicillin-resistant and -susceptible Staphylococcus aureus. Front. Microbiol. 2017, 8, 543. [Google Scholar] [CrossRef]
  39. Nwobi, O.C.; Anyanwu, M.U.; Jaja, I.F.; Nwankwo, I.O.; Okolo, C.C.; Nwobi, C.A.; Ezenduka, E.V.; Oguttu, J.W. Staphylococcus aureus in horses in Nigeria: Occurrence, antimicrobial, methicillin and heavy metal resistance and virulence potentials. Antibiotics 2023, 12, 242. [Google Scholar] [CrossRef] [PubMed]
  40. Roudaud, M.; Allano, M.; Fairbrother, J.-H.; Sauvé, F. A retrospective study on methicillin-resistant Staphylococcus spp. isolated from horses admitted to a Canadian veterinary teaching hospital between 2008 and 2018. Can Vet. J. 2020, 61, 1197–12022. [Google Scholar]
  41. Léon, A.; Castagnet, S.; Maillard, K.; Paillot, R.; Giard, J.C. Evolution of in vitro antimicrobial susceptibility of equine clinical isolates in France between 2016 and 2019. Anim. Open Access J. 2020, 10, 812. [Google Scholar] [CrossRef] [PubMed]
  42. Martinez-Laorden, A.; Arraíz-Fernandez, C.; Gonzalez-Fandos, E. Identification and characterisation of antimicrobial resistance of Enterococcus spp. Isolated from pork and poultry meat. Int. J. Food Sci. Technol. 2023, 58, 4455–4463. [Google Scholar] [CrossRef]
  43. Martinez-Laorden, A.; Arraíz-Fernandez, C.; Cantalejo, M.J.; Gonzalez-Fandos, E. Prevalence, identification and antimicrobial resistance of Listeria monocytogenes and Listeria spp. isolated from poultry and pork meat. Int. J. Food Sci. Technol. 2024, 59, 2667–2675. [Google Scholar] [CrossRef]
  44. Cufaoglu, G.; Ambarcioglu, P.; Ayaz, N.D. Meta-analysis of the prevalence of Listeria spp. and antibiotic resistant L. monocytogenes isolates from foods in Turkey. Food Sci. Technol. 2021, 144, 111210. [Google Scholar] [CrossRef]
  45. Liu, Y.; Sun, W.; Sun, T.; Gorris, L.G.; Wang, X.; Liu, B.; Dong, Q. The prevalence of Listeria monocytogenes in meat products in China: A systematic literature eview and novel meta-analysis approach. Int. J. Food Microbiol. 2020, 312, 108358. [Google Scholar] [CrossRef]
  46. Shen, J.; Zhang, G.; Yang, J.; Zhao, L.; Jiang, Y.; Guo, D.; Wang, X.; Zhi, S.; Xu, X.; Dong, Q.; et al. Prevalence, antibiotic resistance, and molecular epidemiology of Listeria monocytogenes isolated from imported foods in China during 2018 to 2020. Int. J. Food Microbiol. 2023, 382, 109916. [Google Scholar] [CrossRef] [PubMed]
  47. Kayode, A.J.; Okoh, A.I. Antibiotic resistance profile of Listeria monocytogenes recovered from ready-to-eat foods surveyed in South Africa. J. Food Prot. 2022, 85, 1807–1814. [Google Scholar] [CrossRef] [PubMed]
  48. Collobert, J.F.; Guyon, R.; Dieuleveux, V.; Dorey, F. Etude de la contamination de carcasses de chevaux par Salmonella spp. Campylobacter spp. et Escherichia coli O157:H7. Bull. Soc. Vet. Prat. Fr. 2001, 85, 186–191. [Google Scholar]
  49. Moriarty, E.M.; MDowning, M.; Bellamy, J.; Gilpin, B.J. Concentrations of faecal coliforms, Escherichia coli, enterococci and Campylobacter spp. in equine faeces. N. Z. Vet. J. 2015, 63, 104–109. [Google Scholar] [CrossRef] [PubMed]
  50. Brooke, J.S. Stenotrophomonas maltophilia: An emerging global opportunistic pathogen. Clin. Microbiol. 2012, 25, 2–41. [Google Scholar] [CrossRef] [PubMed]
  51. Brooke, J.S. Advances in the microbiology of Stenotrophomonas maltophilia. Clin. Microbiol. Rev. 2021, 34, e00030-19. [Google Scholar] [CrossRef] [PubMed]
  52. Winther, L.; Andersen, R.M.; Baptiste, K.E.; Aalbæk, B.; Guardabassi, L. Association of Stenotrophomonas maltophilia infection with lower airway disease in the horse: A retrospective case series. Vet. J. 2010, 186, 358–363. [Google Scholar] [CrossRef]
  53. Singh, F.; Hirpurkar, S.D.; Rawat, N.; Shakya, S.; Kumar, R.; Rajput, P.K.; Kumar, S. Occurrence of the genes encoding carbapenemases, ESBLs and class 1 integron-integrase among fermenting and non-fermenting bacteria from retail goat meat. Lett. Appl. Microbiol. 2020, 71, 611–619. [Google Scholar] [CrossRef]
  54. Yamamoto, S.; Nakayama, T.; Asakura, H. Draft genome sequence of Stenotrophomonas maltophilia CRB139-1, isolated from poultry meat in Japan. Microbiol. Resour. Announc. 2020, 12, e00075. [Google Scholar] [CrossRef]
  55. da Silva Guedes, J.; Velilla-Rodriguez, D.; González-Fandos, E. Microbiological quality and safety of fresh rabbit meat with special reference to methicillin-resistant S. aureus (MRSA) and ESBL-producing E. coli. Antibiotics 2024, 13, 256. [Google Scholar] [CrossRef] [PubMed]
  56. Deredjian, A.; Alliot, N.; Blanchard, L.; Brothier, E.; Anane, M.; Cambier, P.; Jolivet, C.; Khelil, M.N.; Nazaret, S.; Saby, N.; et al. Occurrence of Stenotrophomonas maltophilia in agricultural soils and antibiotic resistance properties. Res. Microbiol. 2016, 167, 313–324. [Google Scholar] [CrossRef] [PubMed]
  57. Tamma, P.D.; Aitken, S.L.; Bonomo, R.A.; Mathers, A.J.; Van Duin, D.; Clancy, C.J. Infectious diseases Society of America Guidance on the treatment of AmpC β-lactamase-producing Enterobacterales, carbapenem-resistant Acinetobacter baumannii and Stenotrophomonas maltophilia Infections. Clin. Infect. Dis. 2021, 74, 2089–2114. [Google Scholar] [CrossRef] [PubMed]
  58. Yang, X.; Zhu, L.; Zhang, Y.; Liang, R.; Luo, X. Microbial community dynamics analysis by high-throughput sequencing in chilled beef longissimus steaks packaged under modified atmospheres. Meat Sci. 2018, 141, 94–102. [Google Scholar] [CrossRef] [PubMed]
  59. Kim, H.E.; Lee, J.J.; Lee, M.J.; Kim, B.S. Analysis of microbiome in raw chicken meat from butcher shops and packaged products in South Korea to detect the potential risk of foodborne illness. Food Res. Int. 2019, 122, 517–527. [Google Scholar] [CrossRef]
  60. Peruzy, M.F.; Houf, K.; Joossens, M.; Yu, Z.; Proroga, Y.T.R.; Murru, N. Evaluation of microbial contamination of different pork carcass areas through culture-dependent and independent methods in small-scale slaughterhouses. Int. J. Food Microbiol. 2021, 336, 108902. [Google Scholar] [CrossRef]
  61. Matajira, C.E.C.; Poor, A.P.; Moreno, L.Z.; Monteiro, M.S.; Dalmutt, A.C.; Gomes, V.T.M.; Dutra, M.C.; Barbosa, M.R.F.; Sato, M.I.Z.; Moreno, A.M. Vagococcus sp. a porcine pathogen: Molecular and phenotypic characterization of strains isolated from diseased pigs in Brazil. J. Infect. Develop. Ctries 2020, 14, 1314–1319. [Google Scholar] [CrossRef]
  62. Pot, B.; Devriese, L.A.; Hommez, J. Characterization and identification of Vagococcus fluvialis strains isolated from domestic animals. J. Appl. Microbiol. 1994, 77, 362–369. [Google Scholar] [CrossRef]
  63. Brunswick, J.; Spiro, J.; Wisniewski, P. Vagococcus: An under-recognized and emerging cause of antibiotic-resistant infection. IDCases 2024, 36, e01995. [Google Scholar] [CrossRef]
  64. Chen, Q.; Tan, S.; Long, S.; Wang, K.; Liu, Q. Vagococcus fluvialis isolation from the urine of a bladder cancer patient: A case report. BMC Infect. Dis. 2024, 24, 261. [Google Scholar] [CrossRef]
  65. Racero, L.; Barberis, C.; Traglia, G.; Loza, M.S.; Vay, C.; Almuzara, M. Infections due to Vagococcus spp. Microbiological and clinical aspects and literature review. Enfermedades Infecc. Y Microbiol. Clin. (Engl. Ed.) 2021, 39, 335–339. [Google Scholar] [CrossRef]
  66. Teixeira, L.M.; Carvalho, M.D.G.; Merquior, V.L.; Steigerwalt, A. Phenotypic and genotypic characterization of Pediococcus strains isolated from human clinical sources. J. Clin. Microbiol. 1997, 39, 1241–1246. [Google Scholar]
Microorganisms 12 01775 g001
Figure 1. Identification of Pseudomonas spp. isolated from fresh horsemeat by place of purchase, in percentages (recovered from medium specific for Pseudomonas).
Figure 1. Identification of Pseudomonas spp. isolated from fresh horsemeat by place of purchase, in percentages (recovered from medium specific for Pseudomonas).
Microorganisms 12 01775 g001
Microorganisms 12 01775 g002
Figure 2. Identification of Enterobacteriaceae isolated from fresh horsemeat by place of purchase, in percentages (recovered from McConkey agar).
Figure 2. Identification of Enterobacteriaceae isolated from fresh horsemeat by place of purchase, in percentages (recovered from McConkey agar).
Microorganisms 12 01775 g002
Microorganisms 12 01775 g003
Figure 3. Identification of Staphylococcus spp. isolated from fresh horsemeat according to place of purchase, in percentages (recovered from MSA).
Figure 3. Identification of Staphylococcus spp. isolated from fresh horsemeat according to place of purchase, in percentages (recovered from MSA).
Microorganisms 12 01775 g003
Table 1. Media and conditions used for microbiological evaluation.
Table 1. Media and conditions used for microbiological evaluation.
Culture MediaMicrobial GroupIncubation Time (h)Incubation Temperature (°C)
Plate Count AgarMesophiles4830
Chromogenic Agar for PseudomonasPseudomonas7230
MacConkey AgarEnterobacteriaceae2437
Mannitol Salt AgarStaphylococci3635
Kanamycin Esculin Azide AgarEnterococci4837
ALOA AgarListeria spp. and L. monocytogenes2430
Brilliance Campy Count Agar 1Campylobacter spp.4842
ChromID ESBL AgarESBL-producing E. coli2437
ChromID CARBA AgarEnterobacteriaceae carbapenemase-producers2437
ChromID Colistin AgarColistin-resistant Enterobacteriaceae2437
ChromID VRE AgarVancomycin-resistant enterococci2437
ChromID MRSA AgarMethicillin-resistant S. aureus2437
1 incubated under microaerobic conditions.
Table 2. Distribution of microbial counts in fresh horsemeat (log CFU/g).
Table 2. Distribution of microbial counts in fresh horsemeat (log CFU/g).
Microbial GroupPlace of PurchaseDistribution of Microbial Counts 2Minimum CountsMaximum CountsMeans± Standard Deviation 4
Mesophiles <1 31–7 3>7 3
A (n 1 = 12)01203.056.304.92 ± 1.06 A 5
B (n = 7)0612.787.034.84 ± 1.13 A
A + B (n = 19)01812.787.034.89 ± 1.08
Pseudomonas spp. <1 31–6 3>6 3
A (n = 12)5701.302.842.18 ± 0.00 A
B (n = 7)4302.002.472.26 ± 0.70 A
A + B (n = 19)91001.302.842.20 ± 0.40
Enterobacteriaceae <1 31–4 3>4 3
A (n = 12)6601.783.832.31 ± 0.79 A
B (n = 7)6014.104.104.10 ± 0.00 A
A + B (n = 19)12611.784.102.56 ± 0.97
Staphylococcus spp. <1 31–4 3>4 3
A (n = 12)9301.303.412.32 ± 0.30 A
B (n = 7)6101.601.601.60 ± 0.00 A
A + B (n = 19)15401.303.412.14 ± 0.69
Enterococcus spp. <1 31–3 3>3 3
A (n = 12)9301.301.601.40 ± 0.13
B (n = 7)700---
A + B (n = 19)16301.301.601.40 ± 0.13
1 n, number of samples. 2 Data in the table indicate the number of samples presenting the counts indicated (log CFU/g). 3 range of counts in log CFU/g). 4 means ± standard deviation of samples with counts. 5 For each microbial group, means in the same column, not followed by the same letter (subscript), are significantly different (p < 0.05).
Table 3. Bacteria identified in horsemeat isolated from plate count agar, according to the place of purchase.
Table 3. Bacteria identified in horsemeat isolated from plate count agar, according to the place of purchase.
Place of PurchaseMicrobial GroupPercentage (%)SpeciesPercentage (%)
Brochothrix sp.10.20Brochothrix thermosphacta10.20
ALactic acid bacteria42.86Carnobacterium divergens18.37
Lactobacillus sp.16.33
Lactococcus lactis4.08
Carnobacterium maltaromaticum4.08
Pseudomonas spp.14.28P. fragi8.16
P. extremorientalis2.04
P. gessardii2.04
P. lundensis2.04
Enterobacteriaceae8.16Serratia proteamaculans6.12
Buttiauxella gaviniae2.04
Micrococcaceae2.04Staphylococcus saprophyticus2.04
Other Gram-negative bacteria16.33Acinetobacter guillouiae4.08
Chryseobacterium scophthalmum4.08
Acinetobacter harbinensis2.04
Chryseobacterium indologenes2.04
Stenotrophomonas rhizophila2.04
Stenotrophomonas maltophilia2.04
BBrochothrix sp.3.33Brochothrix thermosphacta3.33
Lactic acid bacteria56.67Carnobacterium divergens16.67
Lactobacillus sp.16.67
Carnobacterium maltaromaticum10.00
Lactococcus lactis6.67
Lactococcus piscium3.33
Lactobacillus sakei3.33
Pseudomonas spp.20.00P. fragi6.67
P. fluorescens6.67
P. antarctica3.33
P. extremorientalis3.33
Enterobacteriaceae3.33Serratia proteamaculans3.33
Micrococcaceae6.66Staphylococcus warneri3.33
Kocuria rhizophila3.33
Other Gram-negative bacteria9.99Chryseobacterium indologenes3.33
Chryseobacterium shigense3.33
Microbacterium liquefaciens3.33
Table 4. Antimicrobial resistance phenotypes of bacteria isolated from horsemeat.
Table 4. Antimicrobial resistance phenotypes of bacteria isolated from horsemeat.
SpeciesAntibiotic Resistance Phenotype 1Sample 2Place of Purchase
S. maltophiliaAMP-ATM-CTX-CPD-CZ-IPM-PRLH11(PCA)A
H11 (CARB)A
Y. enterocoliticaAUG-AMP-SAM-ATM-FEP-CTX-FOX-CPD-CRO-NA-F-WH07 (COL)A
S. delphiniPUM-FOXH02 (MSA)A
susceptible to all antibiotics testedH03 (MSA)A
S. saprophyticusDO-P-TEH06 (MSA)B
PH11 (PCA)A
S. warnerisusceptible to all antibiotics testedH03 (MSA)A
H06(PCA)B
H06 (MSA)B
S. fleurettisusceptible to all antibiotics testedH02 (MSA)A
S. succinussusceptible to all antibiotics testedH06 (MSA)A
S. equorumsusceptible to all antibiotics testedH05 (MSA)A
E. maldoratussusceptible to all antibiotics testedH13 (KN)A
V. fluvialisCMN-LZD-F-OX-P-QD-TEC-VAH14 (MRSA)A
L. monocytogenesAMP-PNG-CTX-MEM-OXH13 (ALOA)A
AMP-PNG-CTX-MEM-OXH14 (ALOA)A
CTX-F-OXH11 (ALOA)B
AMP-PNG-CTX-OXH16 (ALOA)A
1 AMP: ampicillin, ATM: aztreonan, CTX: cefotaxime, CPD: cefpodoxime, CZ: cefazoline, IPM: imipenem, PRL: piperacillin, AUG: amoxicillin-clavulanate, SAM: ampicillin/surbactam, FEP: cefepime, FOX: cefoxitin, CRO: ceftriaxone, NA: nalidixic acid, F: nitrofurantoin, W: trimethoprim, DO: doxycycline, PUM: mupirocin, P: penicillin, TE: tetracycline, CMN: clindamycin, LZD: linezolid, OX: oxacillin, QD: quinupristin/dalfopristin, TEC: teicoplanin, VA: vancomycin, PNG: bezilpenicillia, MEM: meropenem. 2 isolation medium. PCA: plate count agar, CARB: ChromID CARBA agar, COL: ChromID Colistin agar, MSA: Mannitol salt agar, KN: Kanamycin Esculin Azide agar, MRSA: ChromID MRSA Agar, ALOA: ALOA agar.
Disclaimer/Publisher’s Note: The statements, opinions and data contained in all publications are solely those of the individual author(s) and contributor(s) and not of MDPI and/or the editor(s). MDPI and/or the editor(s) disclaim responsibility for any injury to people or property resulting from any ideas, methods, instructions or products referred to in the content.

Share and Cite

MDPI and ACS Style

Gonzalez-Fandos, E.; da Silva Guedes, J. Microbiological Quality and Antibiotic Resistance of Relevant Bacteria from Horsemeat. Microorganisms 2024, 12, 1775. https://doi.org/10.3390/microorganisms12091775

AMA Style

Gonzalez-Fandos E, da Silva Guedes J. Microbiological Quality and Antibiotic Resistance of Relevant Bacteria from Horsemeat. Microorganisms. 2024; 12(9):1775. https://doi.org/10.3390/microorganisms12091775

Chicago/Turabian Style

Gonzalez-Fandos, Elena, and Jessica da Silva Guedes. 2024. "Microbiological Quality and Antibiotic Resistance of Relevant Bacteria from Horsemeat" Microorganisms 12, no. 9: 1775. https://doi.org/10.3390/microorganisms12091775

Note that from the first issue of 2016, this journal uses article numbers instead of page numbers. See further details here.

Article Metrics

Back to TopTop