Analysis of the culturable gut yeast microbiota of dogs with digestive disorders.

Despite the increasing interest in studying the gut mycobiota of dogs, the association between fungal colonization and the development of digestive disorders in this species remains largely understudied. On the other hand, the high prevalence of antifungal-resistant yeasts detected in previous studies in samples from animals represents a major threat to public health. We analyzed the presence of culturable yeasts in 112 rectal swab samples obtained from dogs with digestive disorders attended in a veterinary teaching hospital. Our results revealed that Malassezia pachydermatis was frequently isolated from the studied dog population (33.9% of samples), and that the isolation of this yeast was significantly associated to the age of animals, but not to their sex, disease group, or the presence of vomits and/or diarrhea. In contrast, other yeast species were less prevalent (17.9% of samples in total), and their isolation was not significantly associated to any variable included in the analysis. Additionally, we observed that 97.5% of the studied M. pachydermatis isolates (n = 158, 1-6 per positive episode) displayed a minimum inhibitory concentration (MIC) value >4 µg/ml to nystatin, 31.6% had a MIC ≥32 µg/ml to fluconazole, and 27.2% had a MIC >4 µg/ml to amphotericin B. The antifungal susceptibility profiles of non-Malassezia (n = 43, 1-7 per episode) were more variable and included elevated MIC values for some antifungal-species combinations. These results confirm that the intestine of dogs is a reservoir of opportunistic pathogenic yeasts and suggest that the prevalence of M. pachydermatis colonization depends more on the age of animals than on any specific digestive disorder.

Floral nectar and honeydew microbial diversity and their role in biocontrol of insect pests and pollination.

Sugar-rich plant-related secretions, such as floral nectar and honeydew, that are commonly used as nutrient sources by insects and other animals, are also the ecological niche for diverse microbial communities. Recent research has highlighted the great potential of nectar and honeydew microbiomes in biological pest control and improved pollination, but the exploitation of these microbiomes requires a deep understanding of their community dynamics and plant-microbe-insect interactions. Additionally, the successful application of microbes in crop fields is conditioned by diverse ecological, legal, and ethical challenges that should be taken into account. In this article, we provide an overview of the nectar and honeydew microbiomes and discuss their potential applications in sustainable agricultural practices.

In vitro activity of fidaxomicin and combinations of fidaxomicin with other antibiotics against Clostridium perfringens strains isolated from dogs and cats.

BACKGROUND: Previous studies have demonstrated that fidaxomicin, a macrocyclic lactone antibiotic used to treat recurrent Clostridioides difficile-associated diarrhea, also displays potent in vitro bactericidal activity against Clostridium perfringens strains isolated from humans. However, to date, there is no data on the susceptibility to fidaxomicin of C. perfringens strains of animal origin. On the other hand, although combination therapy has become popular in human and veterinary medicine, limited data are available on the effects of antibiotic combinations on C. perfringens. We studied the in vitro response of 21 C. perfringens strains obtained from dogs and cats to fidaxomicin and combinations of fidaxomicin with six other antibiotics. RESULTS: When tested by an agar dilution method, fidaxomicin minimum inhibitory concentrations (MICs) ranged between 0.004 and 0.032 µg/ml. Moreover, the results of Etest-based combination assays revealed that the incorporation of fidaxomicin into the test medium at a concentration equivalent to half the MIC significantly increased the susceptibility of isolates to metronidazole and erythromycin in 71.4% and 61.9% of the strains, respectively, and the susceptibility to clindamycin, imipenem, levofloxacin, and vancomycin in 42.9-52.4% of the strains. In contrast, » × MIC concentrations of fidaxomicin did not have any effect on levofloxacin and vancomycin MICs and only enhanced the effects of clindamycin, erythromycin, imipenem, and metronidazole in ≤ 23.8% of the tested strains. CONCLUSIONS: The results of this study demonstrate that fidaxomicin is highly effective against C. perfringens strains of canine and feline origin. Although fidaxomicin is currently considered a critically important antimicrobial that has not yet been licensed for veterinary use, we consider that the results reported in this paper provide useful baseline data to track the possible emergence of fidaxomicin resistant strains of C. perfringens in the veterinary setting.

Presence of Aspergillus fumigatus with the TR34/L98H Cyp51A mutation and other azole-resistant aspergilli in the air of a zoological park.

Antifungal-resistant fungi, including Aspergillus fumigatus and other Aspergillus species, pose an urgent threat to human and animal health. Furthermore, the environmental route of azole resistance selection due to the widespread use of azole fungicides in crop protection and other applications is a major public health issue. Although environmental surveillance of fungi is frequently performed in many zoological parks and wildlife rehabilitation centers, the antifungal susceptibility of recovered isolates is only rarely analyzed, which precludes a clear assessment of the threat posed by these fungi to captive animals. In this study, we assessed the presence of airborne azole-resistant Aspergillus spp., including the so-called 'cryptic species' (i.e., species which are phenotypically similar to more well-known aspergilli but clearly constitute different phylogenetic lineages) in a zoological park located in the city of Madrid, Spain. In general, our results revealed a low prevalence A. fumigatus and cryptic aspergilli with decreased susceptibility to azoles. However, we detected an A. fumigatus isolate with the TR34/L98H mutation in the gene encoding the lanosterol 14α-demethylase (Cyp51A), consisting of a tandem repeat of 34 base pairs in the promoter region and a lysine to histidine substitution at codon 98. Notably, this TR34/L98H mutation has been linked to the environmental route of azole resistance selection, thus highlighting the 'One Health' dimension of the emerging problem of antifungal resistance. In this context, continuous environmental surveillance of azole-resistant aspergilli in zoological parks and other similar animal facilities is recommended.

Phylogenomic analysis of the genus Rosenbergiella and description of Rosenbergiella gaditana sp. nov., Rosenbergiella metrosideri sp. nov., Rosenbergiella epipactidis subsp. epipactidis subsp. nov., Rosenbergiella epipactidis subsp. californiensis subsp. nov., Rosenbergiella epipactidis subsp. japonicus subsp. nov., Rosenbergiella nectarea subsp. nectarea subsp. nov. and Rosenbergiella nectarea subsp. apis subsp. nov., isolated from floral nectar and insects.

The genus Rosenbergiella is one of the most frequent bacterial inhabitants of flowers and a usual member of the insect microbiota worldwide. To date, there is only one publicly available Rosenbergiella genome, corresponding to the type strain of Rosenbergiella nectarea (8N4T), which precludes a detailed analysis of intra-genus phylogenetic relationships. In this study, we obtained draft genomes of the type strains of the other Rosenbergiella species validly published to date (R. australiborealis, R. collisarenosi and R. epipactidis) and 23 additional isolates of flower and insect origin. Isolate S61T, retrieved from the nectar of an Antirrhinum sp. flower collected in southern Spain, displayed low average nucleotide identity (ANI) and in silico DNA-DNA hybridization (isDDH) values when compared with other Rosenbergiella members (≤86.5 and ≤29.8 %, respectively). Similarly, isolate JB07T, which was obtained from the floral nectar of Metrosideros polymorpha plants in Hawaii (USA) had ≤95.7 % ANI and ≤64.1 % isDDH with other Rosenbergiella isolates. Therefore, our results support the description of two new Rosenbergiella species for which we propose the names Rosenbergiella gaditana sp. nov. (type strain: S61T=NCCB 100789T=DSM 111181T) and Rosenbergiella metrosideri sp. nov. (JB07T=NCCB 100888T=LMG 32616T). Additionally, some R. epipactidis and R. nectarea isolates showed isDDH values<79 % with other conspecific isolates, which suggests that these species include subspecies for which we propose the names Rosenbergiella epipactidis subsp. epipactidis subsp. nov. (S256T=CECT 8502T=LMG 27956T), Rosenbergiella epipactidis subsp. californiensis subsp. nov. (FR72T=NCCB 100898T=LMG 32786T), Rosenbergiella epipactidis subsp. japonicus subsp. nov. (K24T=NCCB 100924T=LMG 32785T), Rosenbergiella nectarea subsp. nectarea subsp. nov. (8N4T = DSM 24150T = LMG 26121T) and Rosenbergiella nectarea subsp. apis subsp. nov. (B1AT=NCCB 100810T= DSM 111763T), respectively. Finally, we present the first phylogenomic analysis of the genus Rosenbergiella and update the formal description of the species R. australiborealis, R. collisarenosi, R. epipactidis and R. nectarea based on new genomic and phenotypic information.

Sugar Concentration, Nitrogen Availability, and Phylogenetic Factors Determine the Ability of Acinetobacter spp. and Rosenbergiella spp. to Grow in Floral Nectar.

The floral nectar of angiosperms harbors a variety of microorganisms that depend predominantly on animal visitors for their dispersal. Although some members of the genus Acinetobacter and all currently known species of Rosenbergiella are thought to be adapted to thrive in nectar, there is limited information about the response of these bacteria to variation in the chemical characteristics of floral nectar. We investigated the growth performance of a diverse collection of Acinetobacter (n = 43) and Rosenbergiella (n = 45) isolates obtained from floral nectar and the digestive tract of flower-visiting bees in a set of 12 artificial nectars differing in sugar content (15% w/v or 50% w/v), nitrogen content (3.48/1.67 ppm or 348/167 ppm of total nitrogen/amino nitrogen), and sugar composition (only sucrose, 1/3 sucrose + 1/3 glucose + 1/3 fructose, or 1/2 glucose + 1/2 fructose). Growth was only observed in four of the 12 artificial nectars. Those containing elevated sugar concentration (50% w/v) and low nitrogen content (3.48/1.67 ppm) were limiting for bacterial growth. Furthermore, phylogenetic analyses revealed that the ability of the bacteria to grow in different types of nectar is highly conserved between closely related isolates and genotypes, but this conservatism rapidly vanishes deeper in phylogeny. Overall, these results demonstrate that the ability of Acinetobacter spp. and Rosenbergiella spp. to grow in floral nectar largely depends on nectar chemistry and bacterial phylogeny.

Nature's Most Fruitful Threesome: The Relationship between Yeasts, Insects, and Angiosperms.

The importance of insects for angiosperm pollination is widely recognized. In fact, approximately 90% of all plant species benefit from animal-mediated pollination. However, only recently, a third part player in this story has been properly acknowledged. Microorganisms inhabiting floral nectar, among which yeasts have a prominent role, can ferment glucose, fructose, sucrose, and/or other carbon sources in this habitat. As a result of their metabolism, nectar yeasts produce diverse volatile organic compounds (VOCs) and other valuable metabolites. Notably, some VOCs of yeast origin can influence insects' foraging behavior, e.g., by attracting them to flowers (although repelling effects have also been reported). Moreover, when insects feed on nectar, they also ingest yeast cells, which provide them with nutrients and protect them from pathogenic microorganisms. In return, insects serve yeasts as transportation and a safer habitat during winter when floral nectar is absent. From the plant's point of view, the result is flowers being pollinated. From humanity's perspective, this ecological relationship may also be highly profitable. Therefore, prospecting nectar-inhabiting yeasts for VOC production is of major biotechnological interest. Substances such as acetaldehyde, ethyl acetate, ethyl butyrate, and isobutanol have been reported in yeast volatomes, and they account for a global market of approximately USD 15 billion. In this scenario, the present review addresses the ecological, environmental, and biotechnological outlooks of this three-party mutualism, aiming to encourage researchers worldwide to dig into this field.

Contrasting effects of nectar yeasts on the reproduction of Mediterranean plant species.

PREMISE: Yeasts are often present in floral nectar and can influence plant fitness directly (independently of pollinators) or indirectly by influencing pollinator visitation and behavior. However, few studies have assessed the effect of nectar yeasts on plant reproductive success or compared effects across different plant species, limiting our understanding of the relative impact of direct vs. indirect effects. METHODS: We inoculated the nectar of six plant species in the field with the cosmopolitan yeast Metschnikowia reukaufii to analyze the direct and indirect effects on female reproductive success over 2 years. The pollinator assemblage for each species was recorded during both flowering years. RESULTS: Direct yeast effects on female fecundity were statistically nonsignificant for all plant species. There were significant indirect, pollinator-mediated effects on fruit production and seed mass for the two species pollinated almost exclusively by bumblebees or hawkmoths, with the direction of the effects differing for the quantity- and quality-related fitness components. There were no consistent effects of the yeast on maternal fecundity for any of the species with diverse pollinator assemblages. CONCLUSIONS: Effects of M. reukaufii on plant reproduction ranged from negative to neutral or positive depending on the plant species. The among-species variation in the indirect effects of nectar yeasts on plant pollination could reflect variation in the pollinator community, the specific microbes colonizing the nectar, and the order of microbial infection (priority effects), determining potential species interactions. Elucidating the nature of these multitrophic plant-pollinator-microbe interactions is important to understand complex processes underlying plant pollination.

Susceptibility testing of Prototheca bovis isolates from cases of bovine mastitis using the CLSI reference broth microdilution method and the Sensititre YeastOne colorimetric panel.

A total of 62 Prototheca bovis isolates from cases of bovine mastitis were tested for susceptibility to different antifungal compounds by the Clinical and Laboratory Standards Institute (CLSI) reference microdilution method and a commercial colorimetric microdilution panel (Sensititre YeastOne). All isolates displayed low susceptibility to echinocandins (MICs > 8 µg/ml for anidulafungin, caspofungin, and micafungin), flucytosine (MIC > 64 µg/ml), and the azoles enilconazole and fluconazole (MICs > 4 and > 64 µg/ml, respectively). Moreover, 45.2, 32.3, and 1.6% of isolates had MICs > 4 µg/ml for ketoconazole, terbinafine, and voriconazole, respectively, when tested by the CLSI method. In contrast, all isolates were more susceptible to the polyene compounds amphotericin B and nystatin, and itraconazole, posaconazole, and ravuconazole (MICs ≤ 2 µg/ml, in all cases). Comparison of the results obtained in the CLSI and Sensititre methods showed excellent essential agreement (EA) for azoles (98.4% for itraconazole and posaconazole, and 100% for voriconazole) and moderate EA for amphotericin B (72.6%), when MICs were read after 48 and 24 h of incubation, respectively. In contrast, much lower EA values were obtained in some cases when the MICs for both techniques were determined after 48 h of incubation (e.g., 9.7% for amphotericin B and 69.4% for posaconazole). Therefore, the CLSI broth microdilution method and the Sensititre YeastOne panel can be used indistinctly for susceptibility testing of P. bovis isolates against azoles but not against amphotericin B until further optimization of the test conditions. LAY SUMMARY: The antifungal susceptibility of Prototheca bovis isolates was analyzed. All tested isolates displayed low susceptibility to echinocandins, flucytosine, and some azoles. Excellent agreement of the results of two different test methods was obtained for azoles, but not for the polyene amphotericin B.

Acinetobacter pollinis sp. nov., Acinetobacter baretiae sp. nov. and Acinetobacter rathckeae sp. nov., isolated from floral nectar and honey bees.

A detailed evaluation of eight bacterial isolates from floral nectar and animal visitors to flowers shows evidence that they represent three novel species in the genus Acinetobacter. Phylogenomic analysis shows the closest relatives of these new isolates are Acinetobacter apis, Acinetobacter boissieri and Acinetobacter nectaris, previously described species associated with floral nectar and bees, but high genome-wide sequence divergence defines these isolates as novel species. Pairwise comparisons of the average nucleotide identity of the new isolates compared to known species is extremely low (<83 %), thus confirming that these samples are representative of three novel Acinetobacter species, for which the names Acinetobacter pollinis sp. nov., Acinetobacter baretiae sp. nov. and Acinetobacter rathckeae sp. nov. are proposed. The respective type strains are SCC477T (=TSD-214T=LMG 31655T), B10AT (=TSD-213T=LMG 31702T) and EC24T (=TSD-215T=LMG 31703T=DSM 111781T).

Nitrogen Assimilation Varies Among Clades of Nectar- and Insect-Associated Acinetobacters.

Floral nectar is commonly colonized by yeasts and bacteria, whose growth largely depends on their capacity to assimilate nutrient resources, withstand high osmotic pressures, and cope with unbalanced carbon-to-nitrogen ratios. Although the basis of the ecological success of these microbes in the harsh environment of nectar is still poorly understood, it is reasonable to assume that they are efficient nitrogen scavengers that can consume a wide range of nitrogen sources in nectar. Furthermore, it can be hypothesized that phylogenetically closely related strains have more similar phenotypic characteristics than distant relatives. We tested these hypotheses by investigating the growth performance on different nitrogen-rich substrates of a collection of 82 acinetobacters isolated from nectar and honeybees, representing members of five species (Acinetobacter nectaris, A. boissieri, A. apis, and the recently described taxa A. bareti and A. pollinis). We also analyzed possible links between growth performance and phylogenetic affiliation of the isolates, while taking into account their geographical origin. Results demonstrated that the studied isolates could utilize a wide variety of nitrogen sources, including common metabolic by-products of yeasts (e.g., ammonium and urea), and that phylogenetic relatedness was associated with the variation in nitrogen assimilation among the studied acinetobacters. Finally, nutrient source and the origin (sample type and country) of isolates also predicted the ability of the acinetobacters to assimilate nitrogen-rich compounds. Overall, these results demonstrate inter-clade variation in the potential of the acinetobacters as nitrogen scavengers and suggest that nutritional dependences might influence interactions between bacteria and yeasts in floral nectar.

Yeast-nectar interactions: metacommunities and effects on pollinators.

About 90% of all flowering plant species are pollinated by animals. Animals are attracted to flowers because they often provide food in the form of nectar and pollen. While floral nectar is assumed to be initially sterile, it commonly becomes colonized by yeasts after animals have visited the flowers. Although yeast communities in floral nectar appear simple, community assembly depends on a complex interaction between multiple factors. Yeast colonization has a significant effect on the scent of floral nectar, foraging behavior of insects and nectar consumption. Consumption of nectar colonized by yeasts has been shown to improve bee fitness, but effects largely depended on yeast species. Altogether, these results indicate that dispersal, colonization history and nectar chemistry strongly interact and have pronounced effects on yeast metacommunities and, as a result, on bee foraging behavior and fitness. Future research directions to better understand the dynamics of plant-microbe-pollinator interactions are discussed.

Antimicrobial Resistance of Coagulase-Positive Staphylococcus Isolates Recovered in a Veterinary University Hospital.

The Staphylococcus pseudintermedius group (SIG) is an emerging threat in veterinary medicine, particularly methicillin-resistant (MRSP) isolates, which are frequently associated with multidrug resistance. Reliable identification of SIG members is critical to establish correct antimicrobial treatments. However, information on the molecular epidemiology and antimicrobial resistance patterns of MRSP in some regions is still limited. This study aimed to assess the antimicrobial resistance of SIG isolates recovered from animals at the Veterinary Teaching Hospital of Complutense University of Madrid (Spain) during a 10-year period (2007-2016). A total of 139 selected Staphylococcus isolates were subjected to species-level identification by different bioanalytical techniques (PCR, VITEK, MALDI-TOF) and subsequent antimicrobial susceptibility testing. Methicillin-resistant isolates (n = 20) were subjected to whole genome sequencing for further characterization of their antibiotic resistance determinants. Our results showed that there was a good correlation between PCR and MALDI-TOF identification, whereas VITEK showed very divergent results, thus confirming MALDI-TOF as a good alternative for species-level identification of coagulase-positive staphylococci. Notably, S. pseudintermedius, including the epidemic MRSP genotype ST71, was the only SIG species found among canine isolates. In addition, we found a high prevalence of multidrug resistance and resistance to fluoroquinolones, cephalosporins and macrolides. Finally, diverse genes associated with antibiotic resistance were detected among MRSP isolates, although the genetic basis of some of the resistant phenotypes (particularly to fluoroquinolones) could not be determined. In conclusion, our study reveals the circulation of MRSP in the veterinary setting in Spain, thus highlighting the emerging threat posed by this bacterial group and the need for further epidemiological surveillance.

Candida metrosideri pro tempore sp. nov. and Candida ohialehuae pro tempore sp. nov., two antifungal-resistant yeasts associated with Metrosideros polymorpha flowers in Hawaii.

Flowers produce an array of nutrient-rich exudates in which microbes can thrive, making them hotspots for microbial abundance and diversity. During a diversity study of yeasts inhabiting the flowers of Metrosideros polymorpha (Myrtaceae) in the Hawai'i Volcanoes National Park (HI, USA), five isolates were found to represent two novel species. Morphological and physiological characterization, and sequence analysis of the small subunit ribosomal RNA (rRNA) genes, the D1/D2 domains of the large subunit rRNA genes, the internal transcribed spacer (ITS) regions, and the genes encoding the largest and second largest subunits of the RNA polymerase II (RPB1 and RPB2, respectively), classified both species in the family Metschnikowiaceae, and we propose the names Candida metrosideri pro tempore sp. nov. (JK22T = CBS 16091 = MUCL 57821) and Candida ohialehuae pro tempore sp. nov. (JK58.2T = CBS 16092 = MUCL 57822) for such new taxa. Both novel Candida species form a well-supported subclade in the Metschnikowiaceae containing species associated with insects, flowers, and a few species of clinical importance. The ascosporic state of the novel species was not observed. The two novel yeast species showed elevated minimum inhibitory concentrations to the antifungal drug amphotericin B (>4 µg/mL). The ecology and phylogenetic relationships of C. metrosideri and C. ohialehuae are also discussed.

Towards a better understanding of the role of nectar-inhabiting yeasts in plant-animal interactions.

Flowers offer a wide variety of substrates suitable for fungal growth. However, the mycological study of flowers has only recently begun to be systematically addressed from an ecological point of view. Most research on the topic carried out during the last decade has focused on studying the prevalence and diversity of flower-inhabiting yeasts, describing new species retrieved from floral parts and animal pollinators, and the use of select nectar yeasts as model systems to test ecological hypotheses. In this primer article, we summarize the current state of the art in floral nectar mycology and provide an overview of some research areas that, in our view, still require further attention, such as the influence of fungal volatile organic compounds on the foraging behavior of pollinators and other floral visitors, the analysis of the direct and indirect effects of nectar-inhabiting fungi on the fitness of plants and animals, and the nature and consequences of fungal-bacterial interactions taking place within flowers.

Yeast-Bacterium Interactions: The Next Frontier in Nectar Research.

Beyond its role as a reward for pollinators, floral nectar also provides a habitat for specialized and opportunistic yeasts and bacteria. These microbes modify nectar chemistry, often altering mutualistic relationships between plants and pollinators in ways that we are only beginning to understand. Many studies on this multi-partite system have focused on either yeasts or bacteria without consideration of yeast-bacterium interactions, but recent evidence suggests that such interactions drive the assembly of nectar microbial communities and its consequences for pollination. Unexplored potential mechanisms of yeast-bacterium interactions include the formation of physical complexes, nutritional interactions, antibiosis, signaling-based interactions, and horizontal gene transfer. We argue that studying these mechanisms can elucidate how nectar microbial communities are established and affect plant fitness via pollinators.

In vitro activity of amphotericin B-azole combinations against Malassezia pachydermatis strains.

Combination therapy has become popular in clinical practice, but limited data on the effects of combinations of antifungal agents is still available for most fungal pathogens. We studied the in vitro response of 30 genetically diverse clinical strains of the basidiomycetous lipophilic yeast Malassezia pachydermatis obtained from cases of canine otitis to several amphotericin B (AMB)-azole combinations. Broth microdilution checkerboard tests revealed that AMB antagonized the effects of itraconazole (ITC) and voriconazole (VRC) in 50% and 6.7% of the strains, respectively, but did not interact with fluconazole or posaconazole (fractional inhibitory concentration index (FICI) values were <4 in all cases). Subsequent Etest-based assays performed for a subset of strains did not confirm the antagonism between AMB and ITC or AMB and VRC. In summary, the results of this study suggest that antagonistic combination effects between AMB and azoles might occur when tested against M. pachydermatis. Nevertheless, as observed for other fungi, different in vitro analyses yielded contrasting results, and the response to AMB-azole combinations was compound- and strain-dependant.

Distribution and tracking of Clostridium difficile and Clostridium perfringens in a free-range pig abattoir and processing plant.

The presence and genetic diversity of Clostridium difficile and C. perfringens along the slaughtering process of pigs reared in a free-range system was assessed. A total of 270 samples from trucks, lairage, slaughter line and quartering were analyzed, and recovered isolates were toxinotyped and genotyped. C. difficile and C. perfringens were retrieved from 14.4% and 12.6% of samples, respectively. The highest percentage of positive samples for C. difficile was detected in trucks (80%) whereas C. perfringens was more prevalent in cecal and colonic samples obtained in the slaughter line (85% and 45%, respectively). C. difficile isolates (n = 105) were classified into 17 PCR ribotypes (including 010, 078, and 126) and 95 AFLP genotypes. C. perfringens isolates (n = 85) belonged to toxinotypes A (94.1%) and C (5.9%) and were classified into 80 AFLP genotypes. The same genotypes of C. difficile and C. perfringens were isolated from different pigs and occasionally from environmental samples, suggesting a risk of contaminated meat products.