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Andrea Serraino - One of the best experts on this subject based on the ideXlab platform.
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multilocus sequence typing of Arcobacter Butzleri isolates collected from dairy plants and their products and comparison with their pfge types
Journal of Applied Microbiology, 2016Co-Authors: A De Cesare, Federica Giacometti, Andrea Serraino, A Parisi, Silvia Piva, Marta Caruso, E P L De Santis, Gerardo ManfredaAbstract:Aims The present study aimed to determine, by multilocus sequence type (MLST), the heterogeneity level of Arcobacter Butzleri isolates and to compare MLST and pulsed-field gel electrophoresis (PFGE) in terms of discriminatory power (DI) as well as unidirectional and bi-directional concordance. Methods and Results Arcobacter Butzleri isolates (N = 133) from dairy products and environmental samples, collected from dairy plants, were characterized by MLST and PFGE with SacII and classified in 29 sequence types (STs), 47 PFGE and 62 type strains (TS). Among the 119 alleles, 19 were previously unreported and the same for all the STs but two. A significant linkage disequilibrium was detected when the complete ST data set was analysed The DIs of MLST, PFGE and their combination were 0·937, 0·953 and 0·965 respectively. The adjusted Wallace coefficients between MLST and PFGE as well as PFGE and MLST were 0·535 and 0·720 respectively; the adjusted Rand coefficient was 0·612. Conclusions The A. Butzleri studied population showed recombination to some degree. PFGE showed a DI higher than MLST. Both methods presented good concordance. The TS analysis seems to show persistence of the same strain on time and possible cross-contaminations between food and environmental sites. Significance and Impact of the Study This study provides insights in the A. Butzleri population found in raw milk, cheese, and dairy production plants. The data suggest that MLST and PFGE genotypes correlate reasonably well, although their combination results in optimal resolution.
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Short communication: Arcobacter Butzleri and Arcobacter cryaerophilus survival and growth in artisanal and industrial ricotta cheese
Journal of dairy science, 2015Co-Authors: Federica Giacometti, Paolo Daminelli, Elena Cosciani-cunico, Elena Dalzini, M.n. Losio, Andrea SerrainoAbstract:Ricotta cheese is a ready-to-eat product with properties (pH >6.0, aw >0.98–0.99) and moisture content (75–80%) that may pose a risk to public health due to postprocess contamination by several bacterial pathogens, including Arcobacters. The objective of the study was to evaluate the behavior of Arcobacter Butzleri and Arcobacter cryaerophilus in ricotta cheese during its shelf life assuming postprocessing contamination. Two types of ricotta cheese, artisanal water buffalo (WB) and industrial cow milk ricotta cheese, were experimentally contaminated with A. Butzleri and A. cryaerophilus and the count was monitored at 2 different temperatures (6°C and 12°C) during shelf life of 5 d for WB cheese and 22 d for industrial ricotta cheese. In WB ricotta cheese the A. Butzleri count remained stable during the 5 d of storage at 6°C, whereas a moderate but significant decrease was observed in A. cryaerophilus count. The counts of both species increased when WB ricotta cheese was stored at 12°C. In industrial ricotta cheese stored at 6°C, a significant reduction was observed both in A. Butzleri and A. cryaerophilus counts during the 22-d storage period; at 12°C storage, a count increase was observed for both Arcobacter species up to d 14 of storage after which the log cfu/g count resulted constant until d 22 of storage. The ability of A. Butzleri and A. cryaerophilus to survive at 6°C and to grow at 12°C in ricotta cheese has significant food safety implications.
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occurrence and genetic diversity of Arcobacter Butzleri in an artisanal dairy plant in italy
Applied and Environmental Microbiology, 2013Co-Authors: Federica Giacometti, Gerardo Manfreda, Alex Lucchi, Daniela Florio, Renato Giulio Zanoni, Andrea SerrainoAbstract:The present study aimed to investigate the presence, distribution, and persistence of Arcobacter spp. in an artisanal dairy plant and to test the isolates to determine their different genotypes in the processing plant and in foods. Samples were collected in an artisanal cheese factory on four occasions between October and December 2012. Food samples (raw milk, ricotta cheese, mozzarella cheese, and conditioning liquid), water samples, and environmental samples were analyzed by the culture method; isolates were identified by multiplex PCR and genotyped by pulsed-field gel electrophoresis (PFGE) analysis. Arcobacter Butzleri was isolated from 29 out of 59 samples (46.6%), 22 of which were from environmental samples and 7 of which were from food samples. Cluster analysis divided the strains into 47 PFGE patterns: 14 PFGE clusters and 33 unique types. Our findings indicate that the plant harbored numerous A. Butzleri pulsotypes and that the manual cleaning and sanitation in the studied dairy plant do not effectively remove Arcobacter. The recurrent isolation of A. Butzleri suggests that the environmental conditions in the dairy plant constitute a good ecological niche for the colonization of this microorganism. In some cases, the presence of indistinguishable strains isolated from the same facilities on different sampling days showed that these strains were persistent in the processing environment.
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survival of Arcobacter Butzleri during production and storage of artisan water buffalo mozzarella cheese
Foodborne Pathogens and Disease, 2013Co-Authors: Andrea Serraino, Federica Giacometti, Paolo Daminelli, M.n. Losio, G Finazzi, Giacomo Marchetti, Angelo Vittorio Zambrini, Roberto RosminiAbstract:Abstract Water buffalo mozzarella cheese (WBMC) is a fresh stretched cheese produced from whole chilled buffalo milk. Although pasteurization of milk and the use of defined starter cultures are recommended, traditional technology involving unpasteurized milk and natural whey cultures is still employed for WBMC production in Italy. The purpose of this study was to assess the behavior of Arcobacter Butzleri during WBMC production and storage under different temperature conditions (5, 10, and 20°C). Raw milk was experimentally inoculated with one reference strain and two isolates of A. Butzleri, and the count was monitored during WBMC production and storage. The bacterial count of A. Butzleri decreased during curd ripening (from 7.83 log colony-forming units (CFU)/g to 4.14 log CFU/g in about 4 h) and a further decrease (>4 log CFU/g) was observed at the end of curd stretching. During storage testing, A. Butzleri was never detected by direct plating, whereas it was recovered from 12 of the total 162 WBMC unt...
Kurt Houf - One of the best experts on this subject based on the ideXlab platform.
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Antimicrobial susceptibility testing of Arcobacter Butzleri and Arcobacter cryaerophilus strains isolated from Belgian patients
The Journal of antimicrobial chemotherapy, 2016Co-Authors: Anne-marie Van Den Abeele, Dirk Vogelaers, Elke Vanlaere, Kurt HoufAbstract:OBJECTIVES To evaluate the feasibility of different methods for susceptibility testing of human Arcobacter isolates, to assess susceptibility to antibiotics commonly used to treat diarrhoeal illness and to obtain MIC distribution data. METHODS One-hundred-and-six unique Arcobacter strains were collected during an epidemiological study on pathogens in gastroenteritis. Strains were identified by multiplex PCR and PCR-RFLP, and characterized by PFGE. Susceptibility to ampicillin, erythromycin, tetracycline, doxycycline, gentamicin and ciprofloxacin was determined using gradient strip and disc diffusion methodology. Optimal conditions for growth and incubation were tested. Azithromycin was tested with gradient strip diffusion on a subset of 40 strains. Sequence analysis of the quinolone resistance-determining region of gyrA was performed for a subset of 18 strains. RESULTS Based on gradient diffusion results, most Arcobacter strains were susceptible to gentamicin (99%) and tetracycline (89%). Erythromycin (78%), ciprofloxacin (72%) and doxycycline (76%) retained moderate activity against Arcobacter spp. Only 9% of the strains were susceptible to ampicillin. Most Arcobacter Butzleri strains were susceptible to ciprofloxacin (87%), whereas half of the Arcobacter cryaerophilus isolates (51%) showed high-level resistance (MIC >32 mg/L). MIC50 values were comparable for both macrolide antibiotics. Ciprofloxacin-resistant strains possessed an identical mutation in gyrA. Overall, categorical agreement between gradient and disc diffusion results was 60%. Gradient diffusion showed superior readability. CONCLUSIONS Gradient diffusion methodology is preferred for routine susceptibility testing. Acquired resistance to fluoroquinolones was observed in A. cryaerophilus. Macrolides are not first-choice empirical antibiotics for Arcobacter infections. Tetracyclines can be suggested for treatment of documented Arcobacter-related gastrointestinal infections.
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Inhibitory effect of organic acids on Arcobacters in culture and their use for control of Arcobacter Butzleri on chicken skin.
International journal of food microbiology, 2010Co-Authors: E. Skřivanová, Kurt Houf, Zuzana Molatova, Michaela Matěnová, Milan MarounekAbstract:Abstract The inhibitory effects of 17 organic acids (C2–C16 fatty acids, sorbic, benzoic, phenylacetic, fumaric, succinic, lactic, malic and citric) on Arcobacter Butzleri, Arcobacter cryaerophilus and Arcobacter skirrowii were investigated by determining their IC50 values, defined as the concentration of acid at which the target DNA sequence was expressed at 50% of the positive control level in cultures incubated at 30 °C for 24 h. DNA was analysed by real-time PCR. The Arcobacter strains tested were inhibited by all the organic acids, with the sensitivities in the order A. skirrowii > A. cryaerophilus > A. Butzleri. Eight acids with IC50 values of
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Antimicrobial susceptibility patterns of Arcobacter Butzleri and Arcobacter cryaerophilus strains isolated from humans and broilers.
Microbial drug resistance (Larchmont N.Y.), 2004Co-Authors: Kurt Houf, Luc Devriese, Freddy Haesebrouck, Olivier Vandenberg, Jean-paul Butzler, Jan Van HoofAbstract:The MICs of five antimicrobial agents were determined by the agar dilution method for 98 Arcobacter Butzleri and 28 Arcobacter cryaerophilus strains from humans, and poultry. With gentamicin, a MIC of 16 microg/ml was recorded for one A. Butzleri strain isolated from poultry, whereas for the other strains MICs ranged from 0.25 to 4 microg/ml. With ciprofloxacin, a bimodal distribution of susceptibility levels was seen for human A. Butzleri isolates (0.015-0.03 versus 0.12-0.25), whereas MICs for 65 of the 68 A. Butzleri poultry strains ranged from 0.12 to 0.5 microg/ml and three strains from three different broilers were resistant with a MIC of 16 microg/ml. One A. cryaerophilus strain from poultry was resistant to erythromycin at a MIC of 128 microg/ml, whereas MICs for the other Arcobacter strains ranged from 2 to 32 microg/ml. No difference in susceptibility or resistance among the human and poultry strains tested was observed with doxycycline and nalidixic acid. The presence of acquired resistance to erythromycin and ciprofloxacin among poultry isolates is a matter of concern, because the two antimicrobials are generally prescribed as first-line drugs for the treatment of infections with Campylobacteraceae in humans.
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Development of a multiplex PCR assay for the simultaneous detection and identification of Arcobacter Butzleri, Arcobacter cryaerophilus and Arcobacter skirrowii
FEMS microbiology letters, 2000Co-Authors: Kurt Houf, Ann Tutenel, Lieven De Zutter, Jan Van HoofAbstract:A multiplex PCR assay with five primers targeting the 16S and 23S rRNA genes was developed for the simultaneous detection and identification of Arcobacter Butzleri, Arcobacter cryaerophilus and Arcobacter skirrowii. The selected primers amplify a 257-bp fragment from A. cryaerophilus, a 401-bp fragment from A. Butzleri and a 641-bp fragment from A. skirrowii. No PCR product was generated for closely related bacteria including Campylobacter and Helicobacter species. The assay was useful to identify cultures after in vitro cultivation and to detect and identify A. butlzeri and A. cryaerophilus from poultry samples present in 24-h old enrichment in Arcobacter broth with cefoperazone, amphotericin and teicoplanin (CAT)-supplement.
Heriberto Fernández - One of the best experts on this subject based on the ideXlab platform.
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intra amoebic localization of Arcobacter Butzleri as an endocytobiont of acanthamoeba castellanii
Archives of Microbiology, 2019Co-Authors: Gustavo Medina, P Leyan, Viera C Da Silva, S Floresmartin, Carolina Manosalva, Heriberto FernándezAbstract:Acanthamoeba castellanii is a free-living amoeba found mainly in humid environments and Arcobacter Butzleri is an emerging zoonotic pathogen, both can establish in vitro endosymbiotic relationships in the absence of bacterial replication. We analyzed the localization of A. Butzleri within A. castellanii establishing their association with endoplasmic reticulum vesicles and mitochondria. Through confocal microscopy, we observed that during the early stages of endosymbiosis, there is not colocalization between amoebic vacuoles containing A. Butzleri and mitochondria or ER vesicles of A. castellanii. Considering that energy production of this bacterium occurs via metabolism of amino acids or the tricarboxylic acid cycle, these results contribute to explain the absence of bacterial replication, since A. Butzleri would not have access to the nutrients found in endoplasmic reticulum vesicles and mitochondria. In addition, we observe that A. Butzleri induces significantly the actin polymerization of A. castellanii during the early stages of endosymbiosis.
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transcriptional analysis of flagellar and putative virulence genes of Arcobacter Butzleri as an endocytobiont of acanthamoeba castellanii
Archives of Microbiology, 2019Co-Authors: Gustavo Medina, Carola Otth, S Floresmartin, Carolina Manosalva, Patricia R Neves, Marcela Andaur, Nilton Lincopan, Heriberto FernándezAbstract:Arcobacter Butzleri is an emerging foodborne zoonotic pathogen that has been isolated from environmental water sources. This pathogen establishes in vitro endosymbiotic relationships with Acanthamoeba castellanii, a free-living amoeba found in environmental matrices such as soil and water. The principal aim of this study was to analyse the transcriptional pattern of flagellar (flaA-flaB-flgH-motA) and other putative virulence genes (ciaB-cadF-mviN-pldA) of A. Butzleri during its interaction with A. castellanii by quantitative real-time PCR. The transcriptional analysis showed up-regulation of all genes analysed before A. Butzleri became established as an endocytobiont of A. castellanii. In contrast, while A. Butzleri remains an endocytobiont, a significant and sustained decrease in the transcription of all analysed genes was observed. Our findings suggest that A. Butzleri requires a biphasic transcriptional pattern of flagellar and other putative virulence genes to establish an endosymbiotic relationship with A. castellanii.
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Arcobacter Butzleri survives within trophozoite of acanthamoeba castellanii
Revista Argentina De Microbiologia, 2016Co-Authors: Maria Paz Villanueva, Gustavo Medina, Heriberto FernándezAbstract:The survival of three Arcobacter Butzleri strains inside Acanthamoeba castellanii was assessed using axenic cultures of A. castellanii that were inoculated with the tested strains and incubated at 26°C under aerobic conditions for 240h. The behavior of bacteria in contact with amoebae was monitored using phase contrast microscopy. The bacterial survival rate within amoebae was assessed through counting colony forming units, using the gentamicin protection assay. All A. Butzleri strains were able to survive during 240h within the amoebae, thus suggesting that (i) A. Butzleri resists the amoebic digestion processes at least for the analyzed time; (ii) that A. castellanii could serve as an environmental reservoir for this bacterium, probably acting as a transmission vehicle for A. Butzleri.
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Biotyping of Arcobacter Butzleri isolated from poultry products: rescuing a phenotypic method
'EDUFU - Editora da Universidade Federal de Uberlandia', 2016Co-Authors: Heriberto Fernández, Karen Ringler, María Laura AriasAbstract:Arcobacter Butzleri is an emergent zoonotic foodborne pathogen associated to enteritis and occasionally to bacteremia in human beings. Biotyping of this bacterium is important in order to establish the circulating strains and its dissemination routes. The purpose of this work was to determine the circulating A. Butzleri biotypes in poultry products for human consumption in Southern Chile using the method proposed by Lior and Woodward, in order to explore the possibility of introducing this biotyping scheme as a routine laboratory tool. From the 60 strains studied the prevalent biotypes were 8A, 8B, 7A, 4A and 4B. The most frequently isolated biotype, independently of the sample of origin, was 8A with (44 strains, 73.3%). The less frequently isolated biotype was 4B (two strains 3.3%). The biotyping method used results to be simple, easy to handle and yields stable results. Therefore, it might be rescued to be used as a phenotypic tool for epidemiological marking of A. Butzleri
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Mechanisms associated with phagocytosis of Arcobacter Butzleri by Acanthamoeba castellanii
Parasitology research, 2014Co-Authors: Gustavo Medina, S Flores-martin, Belchiolina Fonseca, Carola Otth, Heriberto FernándezAbstract:Acanthamoeba castellanii is a free-living amoeba widely found in environmental matrices such as soil and water. Arcobacter Butzleri is an emerging potential zoonotic pathogen that can be isolated from environmental water sources, where they can establish endosymbiotic relationships with amoebas. The aim of this study was to describe the implication of mannose-binding proteins and membrane-associated receptors of glucose and galactose present in the amoebic membrane, during the attachment of Arcobacter Butzleri by blocking with different saccharides. Another objective was to describe the signaling pathways involved in phagocytosis of these bacteria using specific inhibitors and analyze the implication of phagolysosome formation on the survival of Arcobacter Butzleri inside the amoeba. We infer that the attachment of Arcobacter Butzleri to the amoeba is a process which involves the participation of mannose-binding proteins and membrane-associated receptors of glucose and galactose present in the amoeba. We also demonstrated an active role of protozoan actin polymerization in the phagocytosis of Arcobacter Butzleri and a critical involvement of PI3K and RhoA pathways. Further, we demonstrated that the tyrosine kinase-induced actin polymerization signal is essential in Acanthamoeba-mediated bacterial uptake. Through phagolysosomal formation analysis, we conclude that the survival of Arcobacter Butzleri inside the amoeba could be related with the ability to remain inside vacuoles not fused with lysosomes, or with the ability to retard the fusion between these structures. All these results help the understanding of the bacterial uptake mechanisms used by Acanthamoeba castellanii and contribute to evidence of the survival mechanisms of Arcobacter Butzleri.
Federica Giacometti - One of the best experts on this subject based on the ideXlab platform.
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multilocus sequence typing of Arcobacter Butzleri isolates collected from dairy plants and their products and comparison with their pfge types
Journal of Applied Microbiology, 2016Co-Authors: A De Cesare, Federica Giacometti, Andrea Serraino, A Parisi, Silvia Piva, Marta Caruso, E P L De Santis, Gerardo ManfredaAbstract:Aims The present study aimed to determine, by multilocus sequence type (MLST), the heterogeneity level of Arcobacter Butzleri isolates and to compare MLST and pulsed-field gel electrophoresis (PFGE) in terms of discriminatory power (DI) as well as unidirectional and bi-directional concordance. Methods and Results Arcobacter Butzleri isolates (N = 133) from dairy products and environmental samples, collected from dairy plants, were characterized by MLST and PFGE with SacII and classified in 29 sequence types (STs), 47 PFGE and 62 type strains (TS). Among the 119 alleles, 19 were previously unreported and the same for all the STs but two. A significant linkage disequilibrium was detected when the complete ST data set was analysed The DIs of MLST, PFGE and their combination were 0·937, 0·953 and 0·965 respectively. The adjusted Wallace coefficients between MLST and PFGE as well as PFGE and MLST were 0·535 and 0·720 respectively; the adjusted Rand coefficient was 0·612. Conclusions The A. Butzleri studied population showed recombination to some degree. PFGE showed a DI higher than MLST. Both methods presented good concordance. The TS analysis seems to show persistence of the same strain on time and possible cross-contaminations between food and environmental sites. Significance and Impact of the Study This study provides insights in the A. Butzleri population found in raw milk, cheese, and dairy production plants. The data suggest that MLST and PFGE genotypes correlate reasonably well, although their combination results in optimal resolution.
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Short communication: Arcobacter Butzleri and Arcobacter cryaerophilus survival and growth in artisanal and industrial ricotta cheese
Journal of dairy science, 2015Co-Authors: Federica Giacometti, Paolo Daminelli, Elena Cosciani-cunico, Elena Dalzini, M.n. Losio, Andrea SerrainoAbstract:Ricotta cheese is a ready-to-eat product with properties (pH >6.0, aw >0.98–0.99) and moisture content (75–80%) that may pose a risk to public health due to postprocess contamination by several bacterial pathogens, including Arcobacters. The objective of the study was to evaluate the behavior of Arcobacter Butzleri and Arcobacter cryaerophilus in ricotta cheese during its shelf life assuming postprocessing contamination. Two types of ricotta cheese, artisanal water buffalo (WB) and industrial cow milk ricotta cheese, were experimentally contaminated with A. Butzleri and A. cryaerophilus and the count was monitored at 2 different temperatures (6°C and 12°C) during shelf life of 5 d for WB cheese and 22 d for industrial ricotta cheese. In WB ricotta cheese the A. Butzleri count remained stable during the 5 d of storage at 6°C, whereas a moderate but significant decrease was observed in A. cryaerophilus count. The counts of both species increased when WB ricotta cheese was stored at 12°C. In industrial ricotta cheese stored at 6°C, a significant reduction was observed both in A. Butzleri and A. cryaerophilus counts during the 22-d storage period; at 12°C storage, a count increase was observed for both Arcobacter species up to d 14 of storage after which the log cfu/g count resulted constant until d 22 of storage. The ability of A. Butzleri and A. cryaerophilus to survive at 6°C and to grow at 12°C in ricotta cheese has significant food safety implications.
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occurrence and genetic diversity of Arcobacter Butzleri in an artisanal dairy plant in italy
Applied and Environmental Microbiology, 2013Co-Authors: Federica Giacometti, Gerardo Manfreda, Alex Lucchi, Daniela Florio, Renato Giulio Zanoni, Andrea SerrainoAbstract:The present study aimed to investigate the presence, distribution, and persistence of Arcobacter spp. in an artisanal dairy plant and to test the isolates to determine their different genotypes in the processing plant and in foods. Samples were collected in an artisanal cheese factory on four occasions between October and December 2012. Food samples (raw milk, ricotta cheese, mozzarella cheese, and conditioning liquid), water samples, and environmental samples were analyzed by the culture method; isolates were identified by multiplex PCR and genotyped by pulsed-field gel electrophoresis (PFGE) analysis. Arcobacter Butzleri was isolated from 29 out of 59 samples (46.6%), 22 of which were from environmental samples and 7 of which were from food samples. Cluster analysis divided the strains into 47 PFGE patterns: 14 PFGE clusters and 33 unique types. Our findings indicate that the plant harbored numerous A. Butzleri pulsotypes and that the manual cleaning and sanitation in the studied dairy plant do not effectively remove Arcobacter. The recurrent isolation of A. Butzleri suggests that the environmental conditions in the dairy plant constitute a good ecological niche for the colonization of this microorganism. In some cases, the presence of indistinguishable strains isolated from the same facilities on different sampling days showed that these strains were persistent in the processing environment.
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survival of Arcobacter Butzleri during production and storage of artisan water buffalo mozzarella cheese
Foodborne Pathogens and Disease, 2013Co-Authors: Andrea Serraino, Federica Giacometti, Paolo Daminelli, M.n. Losio, G Finazzi, Giacomo Marchetti, Angelo Vittorio Zambrini, Roberto RosminiAbstract:Abstract Water buffalo mozzarella cheese (WBMC) is a fresh stretched cheese produced from whole chilled buffalo milk. Although pasteurization of milk and the use of defined starter cultures are recommended, traditional technology involving unpasteurized milk and natural whey cultures is still employed for WBMC production in Italy. The purpose of this study was to assess the behavior of Arcobacter Butzleri during WBMC production and storage under different temperature conditions (5, 10, and 20°C). Raw milk was experimentally inoculated with one reference strain and two isolates of A. Butzleri, and the count was monitored during WBMC production and storage. The bacterial count of A. Butzleri decreased during curd ripening (from 7.83 log colony-forming units (CFU)/g to 4.14 log CFU/g in about 4 h) and a further decrease (>4 log CFU/g) was observed at the end of curd stretching. During storage testing, A. Butzleri was never detected by direct plating, whereas it was recovered from 12 of the total 162 WBMC unt...
Fernanda C. Domingues - One of the best experts on this subject based on the ideXlab platform.
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current insights on Arcobacter Butzleri in food chain
Current opinion in food science, 2019Co-Authors: Susana Ferreira, Mónica Oleastro, Fernanda C. DominguesAbstract:Arcobacter Butzleri is an emerging pathogen with a global distribution, frequently found in the food chain. Its wide spread and association with human illness may point this bacterium as a possible causative agent of foodborne diseases usually associated with unknown etiology. Thus, the true pathogenic potential of A. Butzleri and the associated risk of disease development must be clarified. Since its dispersion throughout the food chain is clear, this points toward a potential public health problem due to contaminated food and water, which is strengthened by the ability of this microorganism to survive in food products and water and by its resistance to stress created during food storage and processing.
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Arcobacter Butzleri ciprofloxacin resistance point mutations in dna gyrase a and role on fitness cost
Microbial Drug Resistance, 2018Co-Authors: Susana Ferreira, Mónica Oleastro, Daniela R Correia, Fernanda C. DominguesAbstract:Arcobacter Butzleri is a widely distributed emerging pathogen resistant to various classes of antimicrobial agents, namely fluoroquinolones. A. Butzleri resistance to fluoroquinolones is conferred by point mutations at the antibiotic target. The aim of this study was to evaluate mutations at gyrA associated with ciprofloxacin resistance and evaluate whether acquisition of resistance impacts on fitness and stress tolerance of A. Butzleri. A. Butzleri ciprofloxacin mutants were generated by laboratory induction. Identification of mutations associated with ciprofloxacin resistance was performed by gyrA sequencing. Growth kinetics, cost of fitness, biofilm formation ability, and stress tolerance were assessed. Two amino acid substitutions in the quinolone resistance-determining region of GyrA were identified in the mutant strains, one previously described (Thr-85-Ile) and a new substitution (Asp-89-Tyr). No differences in growth kinetics were recorded between parental and mutant strains; however, fitness cost...
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genotypic and phenotypic features of Arcobacter Butzleri pathogenicity
Microbial Pathogenesis, 2014Co-Authors: Susana Ferreira, João A. Queiroz, Mónica Oleastro, Fernanda C. DominguesAbstract:Abstract Even though Arcobacter Butzleri has been implicated in some human disease as diarrhoea and bacteraemia, much of its pathogenesis and virulence factors remain unclear. In this work we have compared pathogenic and genotypic properties of six A. Butzleri isolates from human and non-human sources. The tested isolates showed to be susceptible to tetracyclines and aminoglycosides, however non-human isolates were all resistant to quinolones. The ability to form biofilms was variable among the tested strains, and all of them showed a weak haemolytic activity. The presence of nine putative virulence genes was determined, with cadF , ciaB , cj1349 , mviN , pldA , tlyA being detected in all strains, while irgA (3/6), hecA (5/6), hecB (4/6) were detected only in some strains. High levels of adhesion were observed for A. Butzleri on Caco-2 cells, with pre-existing inflammation showing no significant effect on the adherence ability; yet variable levels of invasion were observed. A. Butzleri isolates were able to survive intracellularly in Caco-2 cells and to induce a significant up-regulation of interleukin-8 secretion and structural cell rearrangements. These data brings new insights on A. Butzleri virulence and highlights its pathogenic potential.
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Resveratrol against Arcobacter Butzleri and Arcobacter cryaerophilus: activity and effect on cellular functions.
International journal of food microbiology, 2014Co-Authors: Susana Ferreira, Filomena Silva, João A. Queiroz, Mónica Oleastro, Fernanda C. DominguesAbstract:The frequent isolation of Arcobacter Butzleri and Arcobacter cryaerophilus from food samples makes it imperative to search for potential compounds able to inhibit the development of these bacteria. Taking this into consideration, this study focuses on the antimicrobial activity of resveratrol and its mechanism of action against A. Butzleri and A. cryaerophilus. The activity of resveratrol was assessed by a microdilution method and time-kill curves. Resveratrol effect on cellular functions was assessed by flow cytometry evaluating intracellular DNA content and metabolic activity. Ethidium bromide (EtBr) accumulation in the presence of resveratrol was also evaluated, as well as the susceptibility to resveratrol in the presence of phenylalanine-arginine β-naphthylamide (PAβN). Scanning electron microscopy (SEM) was used to further evaluate cell damage caused by resveratrol. Resveratrol presented MIC values of 100 and 50μg/mL to A. Butzleri and A. cryaerophilus, respectively. Based on the time-kill curves, resveratrol exhibited bactericidal activity, leading to a ≥3log10CFU/mL reduction of initial inoculums, for A. Butzleri exponential phase cells incubated for 6h with 1× MIC or with 2× MIC after 24h for stationary phase cells. For A. cryaerophilus cells in exponential growth phase, 99.9% killing was achieved after 24h incubation with 2× MIC, whereas, for stationary phase cells, bactericidal activity was only detected after incubation with 4× MIC. Incubation with resveratrol led to a decrease in both intracellular DNA content and metabolic activity. An increase in the accumulation of EtBr was observed in the presence of resveratrol, and the efflux pump inhibitor PAβN reduced the MIC of resveratrol. SEM analysis revealed disintegration of A. Butzleri cells treated with resveratrol, whereas no morphological alteration was observed for A. cryaerophilus cells. Resveratrol has a good anti-Arcobacter activity, and the results obtained suggest that this compound could act through several different mechanisms in the inhibition of this microorganism. The results encourage the use of this compound for the development of potential strategies to control Arcobacter in food products.
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genetic diversity antibiotic resistance and biofilm forming ability of Arcobacter Butzleri isolated from poultry and environment from a portuguese slaughterhouse
International Journal of Food Microbiology, 2013Co-Authors: Susana Ferreira, João A. Queiroz, Fernanda C. Domingues, Maria Joao Fraqueza, Mónica OleastroAbstract:The genus Arcobacter is an emerging pathogen associated with several clinical symptoms. This genus is widely distributed and has been isolated from environmental, animal, food and human samples, where poultry is considered the major source. In this study, forty three Arcobacter Butzleri strains isolated from poultry and environment of a Portuguese slaughterhouse, were characterized by pulsed field gel electrophoresis (PFGE) and assessed for antimicrobial susceptibility and ability to form biofilms. PFGE patterns obtained using restriction enzymes SmaI and SacII revealed high genetic diversity, with 32 distinct PFGE patterns. Most of A. Butzleri isolates presented multiple antimicrobial resistance, exhibiting four different resistance profiles. All 43 isolates were susceptible to gentamicin and 2.3% were resistant to chloramphenicol, in contrast to twenty four (55.8%) that were resistant to ciprofloxacin. Among 36 selected isolates, 26 strains presented biofilm-forming ability, which was dependent on the atmosphere and initial inoculum density. Overall, the results showed that A. Butzleri displays a high genetic diversity, and presents resistance to several antibiotics, which together with its biofilm formation ability may represent a potential hazard for foodborne infections and a considerable risk for human health.