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Dry-cured ham is a highly appreciated meat product. During the ripening, moulds grow on its surface such as Penicillium nordicum and Aspergillus westerdijkiae producers of ochratoxin A (OTA). This mycotoxin poses a risk to consumers that must be controlled. The aim of this work is to evaluate the effectiveness of Debaryomyces hansenii and Staphylococcus xylosus isolated from dry-cured ham as a combined biocontrol culture (BCA) to reduce the OTA produced by one strain of A. westerdijkiae and two strain of P. nordicum, and to assess the metabolomic changes they cause. Each mould was inoculated alone and in combination with BCA on dry-cured ham for 14 days at 20 °C. OTA and total metabolites were analysed in a mass spectrometer Orbitrap Q- Exactive Plus. The Compound Discoverer software, in-house Python-based software and the Metaboanalyst software were used for metabolite analysis. BCA reduced the OTA of A. westerdijkiae, P. nordicum 15 and P. nordicum 856 by 78 %, 99 % and 65 % respectively. BCA caused changes in their metabolome, mainly in the phenylalanine metabolism pathway altering compounds such as Phenylacetaldehyde, Phenylpyruvate or trans-2-hydroxycinnamate, the synthesis of phenylalanine, tyrosine, and tryptophan altering compounds such as 4-hydroxyphenylpyruvate or L-tryptophan, and in the synthesis of oxylipins derived from the linoleic acid metabolism such as 13-OxoODE, 9(S)-HODE or 9(10)-EpOME., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

In the last few decades, there has been a global increase in the adoption of reptiles as companion animals, mainly turtles and tortoises. Considering the popularity of reptiles as pets in Brazil, and a notable lack of data about potentially pathogenic staphylococci in these animals, this study isolated and evaluate the antimicrobial susceptibility of staphylococcal species from healthy tortoises (Chelonoidis carbonaria) in Brazil. During a 12-month period (February 2019 to February 2020), cloacal swabs from 66 healthy tortoises were collected at the Wild Animals Screening Center in Belo Horizonte, Minas Gerais, Brazil. The swabs were plated onto mannitol salt agar for staphylococci isolation, and species identification was performed using MALDI-TOF MS. Antimicrobial susceptibility was investigated using the disk diffusion method, and the presence of the mecA gene was investigated by PCR to detect methicillin resistance. Of the tested animals, 72.7% were positive for staphylococcal isolation. All isolates were coagulase-negative staphylococci (CoNS), and Staphylococcus sciuri (81.3%), and S. xylosus (12.5%) were the most frequently isolated species. The majority of the isolates (56%) were resistant to at least one antimicrobial agent. A high frequency of resistance was observed for penicillin (35.5%) and tetracycline (29.1 %). All strains were susceptible to cefoxitin, chloramphenicol, ciprofloxacin, erythromycin, and gentamicin. All isolates were negative for the mecA gene. The present work suggests that healthy tortoises are mainly colonized by CoNS, especially S. sciuri. Half of the isolates were resistant to at least one antimicrobial, raising questions regarding the possible role of these animals as reservoirs of antimicrobial resistance genes. [ABSTRACT FROM AUTHOR]

ABSTRACT: In the last few decades, there has been a global increase in the adoption of reptiles as companion animals, mainly turtles and tortoises. Considering the popularity of reptiles as pets in Brazil, and a notable lack of data about potentially pathogenic staphylococci in these animals, this study isolated and evaluate the antimicrobial susceptibility of staphylococcal species from healthy tortoises (Chelonoidis carbonaria) in Brazil. During a 12-month period (February 2019 to February 2020), cloacal swabs from 66 healthy tortoises were collected at the Wild Animals Screening Center in Belo Horizonte, Minas Gerais, Brazil. The swabs were plated onto mannitol salt agar for staphylococci isolation, and species identification was performed using MALDI-TOF MS. Antimicrobial susceptibility was investigated using the disk diffusion method, and the presence of the mecA gene was investigated by PCR to detect methicillin resistance. Of the tested animals, 72.7% were positive for staphylococcal isolation. All isolates were coagulase-negative staphylococci (CoNS), and Staphylococcus sciuri (81.3%), and S. xylosus (12.5%) were the most frequently isolated species. The majority of the isolates (56%) were resistant to at least one antimicrobial agent. A high frequency of resistance was observed for penicillin (35.5%) and tetracycline (29.1 %). All strains were susceptible to cefoxitin, chloramphenicol, ciprofloxacin, erythromycin, and gentamicin. All isolates were negative for the mecA gene. The present work suggests that healthy tortoises are mainly colonized by CoNS, especially S. sciuri. Half of the isolates were resistant to at least one antimicrobial, raising questions regarding the possible role of these animals as reservoirs of antimicrobial resistance genes.

Staphylococcus xylosus (S. xylosus) has become an emerging opportunistic pathogen due to its strong biofilm formation ability. Simultaneously, the biofilm of bacteria plays an important role in antibiotic resistance and chronic infection. Here, we confirmed that rutin can effectively inhibit biofilm formation in S. xylosus, of which the inhibition mechanism involves its ability to interact with imidazole glycerol phosphate dehydratase (IGPD), a key enzyme in the process of biofilm formation. We designed experiments to target IGPD and inhibited its activities against S. xylosus. Our results indicated that the activity of IGPD and the amount of histidine decreased significantly under the condition of 0.8 mg/ml rutin. Moreover, the expression of IGPD mRNA (hisB) and IGPD protein was significantly down-regulated. Meanwhile, the results from molecular dynamic simulation and Bio-layer interferometry (BLI) technique showed that rutin could bind to IGPD strongly. Additionally, in vivo studies demonstrated that rutin treatment reduced inflammation and protect mice from acute mastitis caused by S. xylosus. In summary, our findings provide new insights into the treatment of biofilm mediated persistent infections and chronic bacterial infections. It could be helpful to design next generation antibiotics to against resistant bacteria.

Staphylococcus xylosus (S. xylosus) has become an emerging opportunistic pathogen due to its strong biofilm formation ability. Simultaneously, the biofilm of bacteria plays an important role in antibiotic resistance and chronic infection. Here, we confirmed that rutin can effectively inhibit biofilm formation in S. xylosus , of which the inhibition mechanism involves its ability to interact with imidazole glycerol phosphate dehydratase (IGPD), a key enzyme in the process of biofilm formation. We designed experiments to target IGPD and inhibited its activities against S. xylosus. Our results indicated that the activity of IGPD and the amount of histidine decreased significantly under the condition of 0.8 mg/ml rutin. Moreover, the expression of IGPD mRNA (hisB) and IGPD protein was significantly down-regulated. Meanwhile, the results from molecular dynamic simulation and Bio-layer interferometry (BLI) technique showed that rutin could bind to IGPD strongly. Additionally, in vivo studies demonstrated that rutin treatment reduced inflammation and protect mice from acute mastitis caused by S. xylosus. In summary, our findings provide new insights into the treatment of biofilm mediated persistent infections and chronic bacterial infections. It could be helpful to design next generation antibiotics to against resistant bacteria. [ABSTRACT FROM AUTHOR]

The effect of taxifolin (TXF) with starter cultures (SC), such as Leuconostoc carnosum, or a mixture of strains Pediococcus pentosaceus and Staphylococcus xylosus, on the TXF stability was evaluated. UPLC analysis demonstrated that after 181 days of storage total TXF content was the highest in samples with TXF and L. carnosum (60%), compared to the 1st day of storage. The sausages with TXF and the mixture of P. pentosaceus and S. xylosus (56%) followed next. The samples treated only with TXF retained 40% of TXF, compared to the 1st day of storage. TXF had no significant effect on the growth of lactic acid bacteria. The accumulation of biogenic amines (BA), including histamine and putrescine, was more effectively reduced in sausages inoculated with the TXF plus P. pentosaceus and S. xylosus mixture. Using this mixture, the rate of lipolysis and processes of lipid oxidation were effectively slowed down. Fatty acid (FA) composition was stable in all cases.

The presence of determinants of resistance to antibiotics can constitute a possible safety hazard in coagulase-negative staphylococci (CNS), which are widely present in food of animal origin. Among CNS, S. xylosus is a species frequently isolated from fermented meat products. Resistance to tetracycline was found to be one of the most distributed resistances occurring in S. xylosus strains isolated from fermented sausages. We evaluated the transfer of tetracycline resistance in vitro and in situ between S. xylosus strains. We selected three strains isolated from dry fermented sausages, resistant to tetracycline but not to minocycline, their resistance occurring by a mechanism of active efflux encoded by the tetK gene. Only one strain was able to transfer its tetracycline resistance to a recipient strain initially susceptible and plasmid-free using a filter mating procedure. Transfer of tetracycline resistance was observed at very low frequencies of 3.4 × 10−9 per recipient. To further investigate the transferability of this tetracycline resistance, the donor and recipient strains were tested in pilot-scale fermented sausage production. This transfer was possible but at a low rate, 1.4 × 10−7, and only under conditions of a high inoculation level of 108 CFU/g of meat. The tetK gene is located on a small mobilizable plasmid close to Staphylococcus aureus pT181 plasmid. In conclusion, the transfer of tetracycline resistance between strains of S. xylosus is possible, but at a really low frequency in vitro and in situ in fermented sausages. Even if this represents a very moderate risk, it should be taken into account as required by the European approach of Qualified Presumption of Safety (QPS) and AFSSA safety recommendations, advising that strains used as starter cultures should not carry any transferable antibiotic resistance.

The effect of taxifolin (TXF) with starter cultures (SC), such as Leuconostoc carnosum, or a mixture of strains Pediococcus pentosaceus and Staphylococcus xylosus, on the TXF stability was evaluated. UPLC analysis demonstrated that after 181 days of storage total TXF content was the highest in samples with TXF and L. carnosum (60%), compared to the 1st day of storage. The sausages with TXF and the mixture of P. pentosaceus and S. xylosus (56%) followed next. The samples treated only with TXF retained 40% of TXF, compared to the 1st day of storage. TXF had no significant effect on the growth of lactic acid bacteria. The accumulation of biogenic amines (BA), including histamine and putrescine, was more effectively reduced in sausages inoculated with the TXF plus P. pentosaceus and S. xylosus mixture. Using this mixture, the rate of lipolysis and processes of lipid oxidation were effectively slowed down. Fatty acid (FA) composition was stable in all cases. [ABSTRACT FROM AUTHOR]

The presence of determinants of resistance to antibiotics can constitute a possible safety hazard in coagulase-negative staphylococci (CNS), which are widely present in food of animal origin. Among CNS, S. xylosus is a species frequently isolated from fermented meat products. Resistance to tetracycline was found to be one of the most distributed resistances occurring in S. xylosus strains isolated from fermented sausages. We evaluated the transfer of tetracycline resistance in vitro and in situ between S. xylosus strains. We selected three strains isolated from dry fermented sausages, resistant to tetracycline but not to minocycline, their resistance occurring by a mechanism of active efflux encoded by the tetK gene. Only one strain was able to transfer its tetracycline resistance to a recipient strain initially susceptible and plasmid-free using a filter mating procedure. Transfer of tetracycline resistance was observed at very low frequencies of 3.4 × 10−9 per recipient. To further investigate the transferability of this tetracycline resistance, the donor and recipient strains were tested in pilot-scale fermented sausage production. This transfer was possible but at a low rate, 1.4 × 10−7, and only under conditions of a high inoculation level of 108 CFU/g of meat. The tetK gene is located on a small mobilizable plasmid close to Staphylococcus aureus pT181 plasmid. In conclusion, the transfer of tetracycline resistance between strains of S. xylosus is possible, but at a really low frequency in vitro and in situ in fermented sausages. Even if this represents a very moderate risk, it should be taken into account as required by the European approach of Qualified Presumption of Safety (QPS) and AFSSA safety recommendations, advising that strains used as starter cultures should not carry any transferable antibiotic resistance. [ABSTRACT FROM AUTHOR]

A novel coagulase negative staphylococci strain as flavor agent for the processing of fermented meat products, Staphylococcus xylosus P2, was screened from Chinese bacon. It was identified on the basis of morphology and 16S rDNA sequence analysis. Its proteolytic, lipolytic and nitrate reductive activities were evaluated. And its growth ability at different temperatures, pH, sodium chloride, sodium nitrite concentrations, biogenic amine and pigment were investigated or detected. Furthermore, the fermentation evaluation with S. xylosus P2 as starter culture was carried out. The result showed that the fermented beef jerky was more attractive on the color, odor, texture, tasty and overall acceptability. It indicated that S. xylosus P2 would be a good starter culture for fermented meat products (jerky). [ABSTRACT FROM AUTHOR]

Toxigenic moulds can develop on the surface of dry-cured meat products during ripening due to their ecological conditions, which constitutes a risk for consumers. A promising strategy to control this hazard is the use of antifungal microorganisms usually found in these foods. However, to date, the effectiveness of gram-positive catalase-positive cocci (GCC+) has not been explored. The aim of this work was to select GCC+ isolates with antifungal activity to study its effectiveness in a dry-cured ham model system at the environmental conditions reached during the ripening. Forty-five strains of GCC+ were evaluated and the isolate Staphylococcus xylosus Sx8 was selected to assess its efficacy at two different concentrations (106 and 104 cfu/mL) against Penicillium nordicum, Aspergillus flavus, Aspergillus parasiticus, and Penicillium griseofulvum at 15, 20, and 25 °C. The results showed that the inoculation of 106 cfu/mL of S. xylosus completely inhibited the growth of most fungi. In addition, in the presence of this strain at 104 cfu/mL, a significant reduction in fungal growth and mycotoxins production was observed at the three temperatures studied. In conclusion, S. xylosus Sx8 possesses great potential as a biological agent to control toxigenic moulds in dry-cured meat products.

Staphylococcus xylosus (S. xylosus) is an AT-rich and coagulase-negative Staphylococcus (CNS). It is normally regarded as non-pathogenic, however, recent studies have demonstrated that it is related to human opportunistic infections and bovine mastitis. In addition, S. xylosus strains have the ability to form biofilm. Biofilms are also involved in chronic infections and antibiotic resistance, there are only a few reports about cefquinome inhibiting S. xylosus biofilm formation and the protein targets of cefquinome. In our study, we found that sub-MICs of cefquinome were sufficient to inhibit biofilm formation. To investigate the potential protein targets of cefquinome, we used iTRAQ for the analyses of cells at two different conditions: 1/2-MIC (0.125 μg/mL) cefquinome treatment and no treatment. Using iTRAQ technique and KEGG database analysis, we found that proteins differently expression in histidine metabolism pathway may play a role in the process by which 1/2-MIC (0.125 μg/mL) cefquinome inhibits S. xylosus biofilm formation. Interestingly, we found a sharply down-regulated enzyme [A0A068E9J3 imidazoleglycerol-phosphate dehydratase (IGPD)] involved in histidine metabolism pathway in cefquinome-treated cells. We demonstrated the important role of IGPD in sub-MICs cefquinome inhibiting biofilm formation of S. xylosus by gene (hisB) knockout, IGPD enzyme activity and histidine content assays. Thus, our data sheds light on important role of histidine metabolism in S. xylosus biofilm formation; especially, IGPD involved in histidine metabolism might play a crucial role in sub-MICs cefquinome inhibition of biofilm formation of S. xylosus, and we propose IGPD as an attractive protein target of cefquinome.

Staphylococcus xylosus (S. xylosus) is an AT-rich and coagulase-negative Staphylococcus (CNS). It is normally regarded as non-pathogenic, however, recent studies have demonstrated that it is related to human opportunistic infections and bovine mastitis. In addition, S. xylosus strains have the ability to form biofilm. Biofilms are also involved in chronic infections and antibiotic resistance, there are only a few reports about cefquinome inhibiting S. xylosus biofilm formation and the protein targets of cefquinome. In our study, we found that sub-MICs of cefquinome were sufficient to inhibit biofilm formation. To investigate the potential protein targets of cefquinome, we used iTRAQ for the analyses of cells at two different conditions: 1/2-MIC (0.125 mg/mL) cefquinome treatment and no treatment. Using iTRAQ technique and KEGG database analysis, we found that proteins differently expression in histidine metabolism pathway may play a role in the process by which 1/2-MIC (0.125 mg/mL) cefquinome inhibits S. xylosus biofilm formation. Interestingly, we found a sharply down-regulated enzyme [A0A068E9J3 imidazoleglycerol-phosphate dehydratase (IGPD)] involved in histidine metabolism pathway in cefquinome-treated cells. We demonstrated the important role of IGPD in sub-MICs cefquinome inhibiting biofilm formation of S. xylosus by gene (hisB) knockout, IGPD enzyme activity and histidine content assays. Thus, our data sheds light on important role of histidine metabolism in S. xylosus biofilm formation; especially, IGPD involved in histidine metabolism might play a crucial role in sub-MICs cefquinome inhibition of biofilm formation of S. xylosus, and we propose IGPD as an attractive protein target of cefquinome. [ABSTRACT FROM AUTHOR]

The aim of the present study was to determine virulence factors and antibiotic susceptibility of Staphylococcus xylosus isolated from subclinical mastitis in cows. The material consisted of 42 isolates of S. xylosus obtained from 276 samples of milk collected from cows with subclinical mastitis. The isolates were obtained from the udder secretions of 33 cows from farms in the Lublin region (Poland). S. xylosus was found in 15.2% of tested milk samples. The study did not reveal any macroscopic changes in the milk or symptoms in the cow's body. The number of somatic cells in milk samples ranged from 245,000 to 416,000/ml of milk (on average 268,000/ml of milk). The ability to produce slime was observed in 42.9% of S. xylosus isolates. None of the tested isolates demonstrated the ability to produce protease or cause haemolysis. Five isolates of S. xylosus (11.9%) were classified to the methicillin-resistant group. The mecA gene was not found in any of these isolates. The enzyme β-lactamase was detected in 28.6% of S. xylosus isolates. The highest efficacy against S. xylosus was demonstrated for cephalosporin antibiotics: cefacetrile and cefoperazone (80.1% and 76.2% of susceptible isolates of S. xylosus, respectively). A significant quantity of isolates was resistant to streptomycin, linkomycin, penicillin and neomycin (approximately 10% of susceptible isolates of S. xylosus). [ABSTRACT FROM AUTHOR]

The purpose of this study was to analyze and compare genes encoding superantigens (SAgs) in Staphylococcus xylosus and Staphylococcus aureus isolates collected simultaneously from milk of the same cows with clinical mastitis. Genes encoding staphylococcal enterotoxins and enterotoxin-like proteins ( sea- selu), toxic shock syndrome toxin 1 ( tst-1) and exfoliative toxins ( eta and etd) were investigated. It was found that among 30 isolates of S. xylosus, 16 (53.3%) harbored from 1 to 10 SAg genes. In total, in 16 SAg positive S. xylosus, 11 different enterotoxin genes were detected: sec, sed, seg, seh, sei, selm, seln, selo, selp, ser, selu and one etd gene encoding exfoliative toxin D. The most prevalent genes were ser, selu, and selo. Among all the positive isolates of S. xylosus, a total of 14 different SAg gene combinations were detected. One combination was repeated in 3 isolates, whereas the rest were detected only once. However, in the case of S. aureus all the 30 isolates harbored the same combination of SAg genes: seg, sei, selm, seln, selo and on the basis of PFGE analysis all belonged to the same clonal type. Also noteworthy was the observation that SAg genes detected in S. aureus have also been found in S. xylosus. The findings of this study further extend previous observations that SAg genes are present not only in S. aureus but also in coagulase-negative staphylococci, including S. xylosus. Therefore, taking into account that the SAg genes are encoded on mobile genetic elements it is possible that these genes can be transferred between different species of coexisting staphylococci. [ABSTRACT FROM AUTHOR]

The effect of starter culture, containing the strains Lactobacillus plantarum GM77 and Staphylococcus xylosus GM92, isolated from traditional sucuk, on the quality characteristics of dry fermented sausage (sucuk) during ripening period was studied. The microbiological (lactic acid bacteria, Micrococcus/Staphylococcus, Enterobacteriaceae) and physicochemical (pH, aw, NPN, TBARS) properties and volatile compounds, extracted by solid phase microextraction and analyzed by gas chromatography-mass spectrometry, were determined in sucuk samples with starter culture and in the control group (without starter culture). Both starter culture and ripening period had significant effects on lactic acid bacteria, Micrococcus/Staphylococcus counts, and pH and aw values ( P < 0.01). The highest value for nonprotein nitrogen (NPN) was observed to occur in samples with starter culture ( P < 0.05). TBARS value increased during ripening, the highest value was determined at 14th day in samples with starter culture. Among volatile compounds, terpenes were the major group in sucuk. Other volatile compounds (aldehydes, ketones, sulfur compounds, acids, esters, aliphatic hydrocarbons, aromatic hydrocarbons, and alcohols) can be found in fewer amounts. While the use of starter culture was affecting a few compounds, ripening period had affected most of the compounds. [ABSTRACT FROM AUTHOR]

The aim of the study was to identify lactic acid bacteria and Gram-positive catalase-positive cocci isolated from Turkish dry fermented sausage (sucuk) produced by 7 different manufacturers without using starter culture. A total of 129 isolates of lactic acid bacteria were identified phenotypically. Lactobacillus plantarum was the dominant species (45.7%) followed by L. curvatus (10.9%) and L. fermentum (9.3%). Pediococcus isolates were identified as P. pentosaceus and P. acidilactici. All the isolates of gram-positive and catalase-positive cocci (123 isolates) were classified as Staphylococcus except for 1 isolate assigned to Kocuria rosea. The species isolated most often were S. xylosus (41.5%) and S. saprophyticus (28.5%). Four isolates were identified as S. equorum (3.3%), 1 isolate was assigned to S. carnosus (0.8%). [ABSTRACT FROM AUTHOR]

s.c. morot-bizot, r. talon and s. leroy. 2004.To develop a multiplex PCR that allows the identification of bacteria belonging to theStaphylococcusgenus and in particular to the speciesStaphylococcus xylosus,S. saprophyticus,S. epidermidisandS. aureusisolated from food manufacturing plants.Five primer pairs were used in the multiplex PCR, one specific to theStaphylococcusgenus and four specific toS. xylosus,S. saprophyticus,S. epidermidisandS. aureusspecies. All the 31Staphylococcusreference strains yielded a specific PCR product with the genus-specific primers.Staphylococcus xylosus,S. saprophyticus,S. epidermidisandS. aureusgave a specific PCR fragment with the corresponding species-specific primers. No amplification with theKocuria,MacrococcusandMicrococcusstrains was observed in our conditions. This multiplex PCR was performed on 30 strains of Gram-positive cocci isolated from different workshops and fermented sausages. Among them, 28 belonged to theStaphylococcusgenus and 14 were identified toS. saprophyticus, four toS. xylosus, two toS. aureusand one toS. epidermidis.This multiplex PCR provided reliable and repeatable PCR results. It allowed the identification of a major part of the isolates, highlighting the predominance of theS. saprophyticusspecies in the workshops studied.This tool is a useful way to screen the strains isolated from foodstuff and food environment and to monitor these species during the food processing. [ABSTRACT FROM AUTHOR]

Toxigenic moulds can develop on the surface of dry-cured meat products during ripening due to their ecological conditions, which constitutes a risk for consumers. A promising strategy to control this hazard is the use of antifungal microorganisms usually found in these foods. However, to date, the effectiveness of gram-positive catalase-positive cocci (GCC+) has not been explored. The aim of this work was to select GCC+ isolates with antifungal activity to study its effectiveness in a dry-cured ham model system at the environmental conditions reached during the ripening. Forty-five strains of GCC+ were evaluated and the isolate Staphylococcus xylosus Sx8 was selected to assess its efficacy at two different concentrations (106 and 104 cfu/mL) against Penicillium nordicum, Aspergillus flavus, Aspergillus parasiticus, and Penicillium griseofulvum at 15, 20, and 25 °C. The results showed that the inoculation of 106 cfu/mL of S. xylosus completely inhibited the growth of most fungi. In addition, in the presence of this strain at 104 cfu/mL, a significant reduction in fungal growth and mycotoxins production was observed at the three temperatures studied. In conclusion, S. xylosus Sx8 possesses great potential as a biological agent to control toxigenic moulds in dry-cured meat products. [ABSTRACT FROM AUTHOR]