Detection and characterization of methicillin-resistant and susceptible coagulase-negative staphylococci in milk from cows with clinical mastitis in Tunisia.

International audienceOBJECTIVES: This study investigated prevalence of methicillin-resistant (MR) and methicillin-susceptible (MS) coagulase-negative staphylococci (CNS) and the implicated mechanisms of resistance and virulence in milk of mastitis cows. In addition, the presence of SCCmec type was analyzed in MR Staphylococcus epidermidis (MRSE).RESULTS: Three hundred milk samples from cows with clinical mastitis were obtained from 30 dairy farms in different regions of Tunisia. Sixty-eight of the 300 tested samples contained CNS strains. Various CNS species were identified, with Staphylococcus xylosus being the most frequently found (40%) followed by Staphylococcus warneri (12%). The mecA gene was present in 14 of 20 MR-CNS isolates. All of them were lacking the mecC gene. The SCCmecIVa was identified in four MRSE isolates. Most of CNS isolates showed penicillin resistance (70.6%) and 58.3% of them carried the blaZ gene. MR-CNS isolates (n = 20) showed resistance to erythromycin, tetracycline and trimethoprim-sulfametoxazole harboring different resistance genes such us erm(B), erm(T), erm(C), mph(C) or msr(A), tet(K) and dfr(A). However, a lower percentage of resistance was observed among 48 MS-CNS isolates: erythromycin (8.3%), tetracycline (6.2%), streptomycin (6.2%), clindamycin (6.2%), and trimethoprim-sulfametoxazole (2%). The Inu(B) gene was detected in one Staphylococcus xylosus strain that showed clindamycin resistance. The virulence gene tsst-1 was observed in one MR-CNS strain.DISCUSSION: Coagulase-negative staphylococci containing a diversity of antimicrobial resistance genes are frequently detected in milk of mastitis cows. This fact emphasizes the importance of identifying CNS when an intramammary infection is present because of the potential risk of lateral transfer of resistant genes among staphylococcal species and other pathogenic bacteria

Molecular characterisation of antimicrobial resistance and virulence genes in Escherichia coli strains isolated from diarrhoeic and healthy rabbits in Tunisia

[EN] The purpose of this study was to identify Escherichia coli isolates in diarrhoeic and healthy rabbits in Tunisia and characterise their virulence and antibiotic resistance genes. In the 2014-2015 period, 60 faecal samples from diarrhoeic and healthy rabbits were collected from different breeding farms in Tunisia. Susceptibility to 14 antimicrobial agents was tested by disc diffusion method and the mechanisms of gene resistance were evaluated using polymerase chain reaction and sequencing methods. Forty E. coli isolates were recovered in selective media. High frequency of resistance to tetracycline (95%) was detected, followed by different levels of resistance to sulphonamide (72.5%), streptomycin (62.5%), trimethoprim-sulfamethoxazole (60%), nalidixic acid (32.5%), ampicillin (37.5%) and ticarcillin (35%). E. coli strains were susceptible to cefotaxime, ceftazidime and imipenem. Different variants of blaTEM, tet, sul genes were detected in most of the strains resistant to ampicillin, tetracycline and sulphonamide, respectively. The presence of class 1 integron was studied in 29 sulphonamide-resistant E. coli strains from which 15 harboured class 1 integron with four different arrangements of gene cassettes, dfrA17+aadA5 (n=9), dfrA1 + aadA1 (n=4), dfrA12 + addA2 (n=1), dfrA12+orf+addA2 (n=1). The qnrB gene was detected in six strains out of 13 quinolone-resistant E. coli strains. Seventeen E. coli isolates from diarrhoeic rabbits harboured the enteropathogenic eae genes associated with different virulence genes tested (fimA, cnf1, aer), and affiliated to B2 (n=8) and D (n=9) phylogroups. Isolated E. coli strains from healthy rabbit were harbouring fim A and/or cnf1 genes and affiliated to A and B1 phylogroups. This study showed that E. coli strains from the intestinal tract of rabbits are resistant to the widely prescribed antibiotics in medicine. Therefore, they constitute a reservoir of antimicrobial-resistant genes, which may play a significant role in the spread of antimicrobial resistance. In addition, the eae virulence gene seemed to be implicated in diarrhoea in breeder rabbits in Tunisia.The work was supported by Tunisian Ministry of Higher Education, Scientific Research and Technology (LR16IP03). Many thanks go to the members of the Department of Animal Production, National Institute of Agronomy of Tunisia for their help in collecting the samples.Ben Rhouma, R.; Jouini, A.; Klibi, A.; Hamrouni, S.; Boubaker, A.; Kmiha, S.; Maaroufi, A. (2020). Molecular characterisation of antimicrobial resistance and virulence genes in Escherichia coli strains isolated from diarrhoeic and healthy rabbits in Tunisia. World Rabbit Science. 28(2):81-91. https://doi.org/10.4995/wrs.2020.10879OJS8191282Allen H.K., Donato J., Wang H.H, Cloud-Hansen K.A., Davies J. 2010. Call of the wild: antibiotic resistance genes in natural environments. Nat. Rev. Microbiol., 8: 251-259. https://doi.org/10.1038/nrmicro2312Allocatti N., Masulli M., Alexeyev M.F., Di Ilio C. 2013. Escherichia coli in Europe: An overview. Int. J. Environ. Res. Public Health., 10: 6235-6254. https://doi.org/10.3390/ijerph10126235Alonso C.A., Zarazaga M., Ben Sallem R., Jouini A., Ben Slama K., Torres C. 2017. Antibiotic resistance in Escherichia coli in Husbandry animals. The African perspective. Lett. Appl. Microbiol., 64: 318-334. https://doi.org/10.1111/lam.12724Barbosa T.M., Levy S.B. 2000. The impact of antibiotic use on resistance development and persistence. Drug Resist. Updates, 5: 303-311. https://doi.org/10.1054/drup.2000.0167Ben Said L., Jouini A., Klibi N., Dziri R., Alonso C.A., Boudabous A., Ben Slama K., Torres C. 2015. Detection of extended-spectrum beta-lactamase (ESBL)-producing Enterobacteriaceae in vegetables, soil and water of farm environment in Tunisia. Int. J. Food Microbiol., 203: 86-92. https://doi.org/10.1016/j.ijfoodmicro.2015.02.023Ben Sallem R.B., Gharsa H., Slama K.B.., Rojo-Bezares B, Estepa V., Porres-Osante N., Jouini A., Klibi N., Sáenz Y., Boudabous A., Torres C. 2013. First detection of CTX-M-, CMY-2, and QnrB19 resistance mechanisms in fecal Escherichia coli isolates from healthy pets in Tunisia. Vector Borne Zoonotic Dis., 13: 98-102. https://doi.org/10.1089/vbz.2012.1047Ben Slama K., Jouini A., Ben Sallem R., Somalo S., Sáenz Y., Estepa V., Boudabous A., Torres C. 2010. Prevalence of broad-spectrum cephalosporin-resistant Escherichia coli isolates in food samples in Tunisia and characterization of integrons and antimicrobial resistance mechanisms implicated. Int. J. Food Microbiol., 13: 281-286. https://doi.org/10.1016/j.ijfoodmicro.2009.12.003Ben Slama K., Ben Sallem R., Jouini A., Rachid S., Moussa L., Saenz Y., Estepa V., Somalo S., Boudabous A., Torres C. 2011. Diversity of genetic lineages among CTX-M-15 and CTX-M-14 producing Escherichia coli strains in a Tunisian hospital. Curr. Microbiol., 62: 1794-1801. https://doi.org/10.1007/s00284-011-9930-4Blanco J.E., Blanco M., Blanco J., Mora A., Balaguer L., Mourino M., JuÁrez A., Jansen W.H. 1996. O serogroups, biotypes, and eae genes in Escherichia coli strains isolated from diarrheic and healthy rabbits. J. Clin. Microbiol., 34: 3101- 3107. https://doi.org/10.1128/JCM.34.12.3101-3107.1996Blanco M., Blanco J.E., Dahbi G., Alonso M.P., Mora A., Corira M.A.A., Madrid C., Juárez A., Bernárdez M.I., González E.A., Blanco J. 2006. Identification of two new intimin types in atypical enteropathogenic Escherichia coli. Int. Microbiol., 9: 103-110.Brinas L., Zarazaga M., Saenz Y., Ruiz-Larrea F., Torres C. 2002. Beta-lactamases in ampicillin-resistant Escherichia coli isolates from foods, humans, and healthy animals. Antimicrob. Agents Chemother., 46: 3156-3163. https://doi.org/10.1128/AAC.46.10.3156-3163.2002Bryan A., Shapir N., Sadowsky M.J. 2004. Frequency and distribution of tetracycline resistance genes in genetically diverse, nonselected and nonclinical Escherichia coli strains isolated from diverse human and animal sources. Appl. Environ. Microbiol., 70: 2503-2507. https://doi.org/10.1128/AEM.70.4.2503-2507.2004Camarda A., Pennelli D., Battista P., Martella V., Greco L., Alloggio I., Mazzolini E. 2004. Virulence genes and antimicrobial resistance patterns of enteropathogenic Escherichia coli from rabbits in southern Italy. In Proc.: 8th World Rabbit Congress., September 7-10, 2004, Puebla, Mexico, 470-476.Clermont O., Bonacorsi S., Bingen E., 2000. Rapid and simple determination of the Escherichia coli phylogenetic group. Appl. Environ. Microb., 66: 4555-4558. https://doi.org/10.1128/AEM.66.10.4555-4558.2000Clermont O., Lavolly M., Vimont S., Deschamps C., Forestier C., Branger C., Denamur E., Arlet G. 2008. The CTX-M15-producing Escherichia coli diffusing clone belongs to a highly virulent B2 phylogenic subgroup. J. Antimicrob. Chemother., 61: 1024-1028. https://doi.org/10.1093/jac/dkn084Clinical and Laboratory Standards Institute. 2015. Performance standards for antimicrobial susceptibility testing. Twenty-fifth informational supplement. Clinical and Laboratory Standards Institute document M100-S25, CLSI, Wayne, PA.Coque T.M., Novais A., Carattoli A., Poirel L., Pitout J., Peixe L., Baquero F., Cantón R., Nordmann P. 2008. Dissemination of clonally related Escherichia coli strains expressing extendedspectrum β-lactamase CTX-M-15. Emerg. Infect. Dis., 14: 195-200. https://doi.org/10.3201/eid1402.070350Da Costa P.M., Loureiro L., Matos A.J.F. 2013. Transfer of Multidrug-Resistant Bacteria between intermingled ecological niches: the interface between Humans, Animals and the environment. Int. J. Environ. Res. Public Health., 10: 278-294. https://doi.org/10.3390/ijerph10010278Dotto G., Giacomelli M., Grilli G., Ferrazzi V., Carattoli A., Fortini D., Piccirillo A. 2014. High prevalence of oqxAB in Escherichia coli isolates from domestic and wild lagomorphs in Italy. Microb. Drug Resist., 20: 118-123. https://doi.org/10.1089/mdr.2013.0141Eager H., Swan G., van Vuuren M. 2015. A survey of antimicrobial usage in animals in South Africa 479 with specific reference to food animals. J. S. Afr. Vet. Assoc., 83: 16. https://doi.org/10.4102/jsava.v83i1.16Guardabassi L., Schwarz S., Lloyd D.H., 2004. Pet animals as reservoirs of antimicrobial-resistant bacteria. J. Antimicrob. Chemother., 54: 321-332. https://doi.org/10.1093/jac/dkh332Hai S.W., Qi R.P., Y. Gao M. et al. 2014. Study on molecular characterization of Class I integron and integron-associated antimicrobial resistance in Escherichia coli from beef cattle. China Animal Husbandry Veterinary Medicine., 41: 63-67.Jacoby G.A., Griffin C., Hooper D.C. 2011. Citrobacter spp. as a source of qnrB alleles. Antimicrob. Agents Chemother., 55: 4979-4984. https://doi.org/10.1128/AAC.05187-11Johnson T.J., Logue C.M., Johnson J.R., Kuskowski M.A., Sherwood J.S., Barnes H.J., DebRoy C., Wannemuehler Y.M., Obata-Yasuoka M., Spanjaard L., Nolan L.K. 2012. Associations between multidrug resistance, plasmid content, and virulence potential among extra-intestinal pathogenic and commensal Escherichia coli from humans and poultry. Foodborne Pathog. Dis., 9: 37-46. https://doi.org/10.1089/fpd.2011.0961Jouini A., Ben Slama K., Saenz Y., Klibi N., Costa D., Vinué L., Zarazaga M., Boudabous A., Torres C. 2009. Detection of Multiple-Antimicrobial Resistance and Characterization of the Implicated Genes in Escherichia coli isolates from Foods of Animal Origin in Tunis. J. Food Protect., 72: 1082-1088. https://doi.org/10.4315/0362-028X-72.5.1082Kaper J.B., Nataro J.P., Mobley H.L. 2004. Pathogenic Escherichia coli. Nat. Rev. Microbiol., 2: 123-140. https://doi.org/10.1038/nrmicro818Lauková A., Strompfová V., Szabóová R., Bónai A., Matics Z., Kovács M., Pogány Simonová M. 2019. Enterococci from pannon white rabbits: detection, identification, biofilm and screening for virulence factors. World Rabbit Sci., 27: 31-39. https://doi.org/10.4995/wrs.2019.10875Marinho C., Igrejas G., Gonçalves A., Silva N., Santos T., Monteiro R., Gonçalves D., Rodrigues T., Poeta P. 2014. Azorean wild rabbits as reservoirs of antimicrobial resistant Escherichia coli. Anaerobe, 30: 116-119. https://doi.org/10.1016/j.anaerobe.2014.09.009Panteado A.S., Ugrinivich L.A., Blanco J., Blanco M., Andrade J.R.C., Correa S.S., De Castro A.F.P. 2002. Serobiotypes and virulence genes of Escherichia coli strains isolated from diarrheic and healthy rabbits in Brazil. Vet. Microb., 89: 41-51. https://doi.org/10.1016/S0378-1135(02)00148-7Pohl P.H., Peeters J.E.E., Jacquemin R., Lintermans P.F., Mainil J.G. 1993. Identification of eae sequences in enteropathogenic Escherichia coli strains from rabbits. Infect. Immun., 61: 2203-2206. https://doi.org/10.1128/IAI.61.5.2203-2206.1993Poirel L., Bonnin R.A., Nordmann P. 2012. Genetic support and diversity of acquired extended-spectrum beta-lactamases in Gram-negative rods. Infect. Genet. Evol., 12: 883-893. https://doi.org/10.1016/j.meegid.2012.02.008Qing F.K., Yi G., Ze H.Y., YA J.L., Wei H.M., Zhao B.D., Yu W.Z., Guan W. 2006. Detection of quinolone resistance and plasmid mediated quinolone gene in Escherichia coli isolated from rabbit. Chin. J. Prev. Vet. Med., 38: 944-948.Ruiz J., Simon K., Horcajada J.P., Velasco M., Barranco M., Roig G., Moreno-Martínez A., Martínez J.A., Jiménez de Anta T., Mensa J., Vila J. 2002. Differences in virulence factors among clinical isolates of Escherichia coli causing cystitis and pyelonephritis in women and prostatitis in men. J. Clin. Microbiol., 40: 4445-4449. https://doi.org/10.1128/JCM.40.12.4445-4449.2002Sáenz Y., Zarazaga M., Briñas L., Lantero M., Ruiz-Larrea F., Torres C. 2001. Antibiotic resistance in Escherichia coli isolates obtained from animals, foods and humans in Spain. Int. J. Antimicrob. Agents.,18: 353-358. https://doi.org/10.1016/S0924-8579(01)00422-8Saenz Y., Briñas L., Domínguez E., Ruiz J., Zarazaga M., Vila J., Torres C. 2004. Mechanisms of resistance in multiple-antibio resistant Escherichia coli strains of human, animal, and food origins. Antimicrob. Agents Chemother., 48: 3996-4001. https://doi.org/10.1128/AAC.48.10.3996-4001.2004Santos T., Silva N., Igrejas G, Rodrigues P., Micael J., Rodrigues T., Resendes R., Gonçalves A., Marinho C., Gonçalves D., Cunha R., Poeta P. 2013. Dissemination of antibiotic resistant Enterococcus spp. and Escherichia coli from wild birds of Azores Archipielago. Anaerobe., 24: 25-31. https://doi.org/10.1016/j.anaerobe.2013.09.004Silva N., Igrejas G., Figueiredo N., Gonçalves, A., Radhouani H., Rodrigues J., Poeta P. 2010. Molecular characterization of antimicrobial resistance in enterococci and Escherichia coli isolates from European wild rabbit (Oryctolagus cuniculus). Sci Total Environ., 408: 4871-4876. https://doi.org/10.1016/j.scitotenv.2010.06.046Swennes A.G., Buckley E.M., Parry N.M.A., Madden C.M., Garcia A., Morgan P.B., Astrofsky K.M., Fox J.G. 2012. Enzootic Enteropathogenic Escherichia coli Infection in Laboratory Rabbits. J. Clin. Microbiol., 50: 2353-2358. https://doi.org/10.1128/JCM.00832-12Teshager T., Herrero I.A., Porrero M.C., Garde J., Moreno M.A., Domínguez L. 2000. Surveillance of antimicrobial resistance in Escherichia coli strains isolated from pigs at Spanish slaughterhouses. Int. J. Antimicrob. Agents, 15: 137-142. https://doi.org/10.1016/S0924-8579(00)00153-9Van den Bogaard A.E., London N., Stobberingh E.E. 2000. Antimicrobial resistance in pig faecal samples from The Netherlands (five abattoirs) and Sweden. J. Antimicrobial Chemother., 45: 663-671. https://doi.org/10.1093/jac/45.5.663Wang J., Sang L., Chen Y., Sun S., Chen D., Xie X. 2019. Characterisation of Staphylococcus aureus strain causing severe respiratory disease in rabbits. World Rabbit Sci., 27: 41-48. https://doi.org/10.4995/wrs.2019.10454Werner G., Gfrörer S., Fleige C., Witte W., Klare I. 2008. Tigecycline-resistant Enterococcus faecalis strain isolated from a German ICU patient. J. Antimicrob Chemother., 61: 1182-1183. https://doi.org/10.1093/jac/dkn065Yoon S.H., Jeong H., Kwon S.K., Kim J.F. 2009. Genomics, Biological Features, and Biotechnological Applications of Escherichia coli B: Is B for better. In: Lee S.Y. (eds). Systems Biology and Biotechnology of Escherichia coli., 1-17. https://doi.org/10.1007/978-1-4020-9394-4_1Zhao X., Yang J., Ju Z., Chang W., Sun S. 2018. Molecular characterization of antimicrobial resistance in Escherichia coli from rabbit farms in Tai'an, China. Hindawi, Biomed Research International., 2018: 1-7. https://doi.org/10.1155/2018/860764

Antimicrobial resistance genes and virulence gene encoding intimin in Escherichia coli and Enterococcus isolated from wild rabbits (Oryctolagus cuniculus) in Tunisia

The spread of antimicrobial-resistant bacteria in wildlife must be viewed as a major concern with serious implications for human and animal health. Escherichia coli and enterococcal isolates were recovered from faecal samples of 49 wild rabbits (Oryctolagus cuniculus) on specific media and were characterised using biochemical and molecular tests. For all isolates, antimicrobial susceptibility testing was performed, and resistance genes were detected by PCR. Molecular typing of isolates was carried out by pulsed-field gel-electrophoresis, and E. coli strains were also tested for the presence of intimin (eae) gene characteristic of rabbit enteropathogenic E. coli. A total of 34 E. coli and 36 enterococci [E. hirae (52.8%) and E. faecalis (47.2%)] were obtained. For E. coli, resistance to tetracycline (94%), streptomycin (62%), ciprofloxacin (47%), trimethoprim-sulphamethoxazole (35%) and chloramphenicol (6%) was observed. Resistance to third-generation cephalosporins was detected in one E. coli strain that carried the blaCMY-2 and blaTEM-1 genes. Class 1 integrons were detected in eight isolates. For enterococci, resistance to tetracycline (63.9%), erythromycin (30.5%), streptomycin (18.2%), and chloramphenicol (5.5%) was detected. The tet(M)+tet(L), erm(B) and ant (6)-Ia genes were identified in thirteen, seven and three resistant Enterococcus strains, respectively. Molecular typing showed a high diversity among our strains. Wild rabbits could represent a reservoir of E. coli, and enterococci carrying antimicrobial resistance genes and E. coli additionally carrying the eae gene of enteropathogenic pathotypes could both contaminate the environment. our finding seems to represent the first report of eae-positive E. coli in wild rabbits

Les déterminants de prêts non performants

Cette étude explore une approche empirique pour détecter les déterminants de crédits à problèmes pour un échantillon composé de 135 banques européennes. nous avons focalisé l’intérêt sur les banques appartenant aux pays les plus touchés par la crise financière de 2008 à savoir l’Espagne, la Grèce et l’Italie. En s’inspirant des travaux précurseurs, nous avons choisi des facteurs spécifiques à la banque et des facteurs macro-économiques comme déterminants des créances douteuses. Notre résultat montre que les prêts non performants dépendent positivement de taux de chômage, de taux d’inflation et des provisions pour pertes sur prêts. Ces crédits à problèmes varient négativement de taux de croissance de PIB et de rentabilité des fonds propres

Antibacterial Effect of Eight Essential Oils against Bacteria Implicated in Bovine Mastitis and Characterization of Primary Action Mode of <i>Thymus capitatus</i> Essential Oil

During the current investigation, eight essential oils (EOs) were tested for their antimicrobial activity against six species, belonging to the genus of staphylococcus, multi-resistant to antibiotics (S. epidermidis, S. cohni, S. wareneri, S. scuiri, S. chromogenes, S. pasteuri), three methicillin-resistant Staphylococcus aureus strains (MRSA) and two strains of Escherichia coli, producing extended-spectrum β-lactamase (ESBL) responsible for bovine mastitis. Our results indicated that the antimicrobial activities of eight EOs varied significantly among the types of EOs and bacterial species. Thymus capitatus and Trachyspermum ammi EOs display important antibacterial activity against all tested strains, with the inhibition zone diameters situated between 20 and 45 mm, while EOs of Artemisia absinthium, Eucalyptus globulus, Eucalyptus camaldulensis, Myrtus communis and Mentha pulegium exerted an intermediate activity. For Cymbopogon citratus, this effect depends on bacteria species. In fact, an important effect was observed against S. warneri, S. epidermidis, S. cohenii, S. pasteuri and MRSA (EC 39+) strains. In addition, the important lytic effect was observed against MRSA strains, showing that Gram-positive bacteria were more sensitive to T. capitatus EO than Gram-negative ones. Concerning the characterization of the mode action of T. capitatus, experiments of kill-time, bacteriolytic, loss of salt tolerance and loss of cytoplasmic material showed that the used EO was able to destroy cell walls and membranes followed by the loss of vital intracellular materials. In addition, it inhibits the normal synthesis of DNA, causing the bacterial death of E. coli and MRSA strains. This study shows the potential of using of EOs, particularly T. capitaus, to inhibit the growth of Gram-positive and Gram-negative bacteria multi-resistant to antibiotics causing bovine mastitis

Detection of Extended-Spectrum β-Lactamases (ESBL) Producing Enterobacteriaceae from Fish Trapped in the Lagoon Area of Bizerte, Tunisia

Extended-spectrum β-lactamase and their molecular mechanism in Enterobacteriaceae were analyzed in 126 fish samples of 9 various wild species, living in the lagoon of Bizerte in Tunisia. Fifty-nine (59) Gram-negative strains were isolated and identified as Escherichia coli (n=24), Klebsiella pneumonia (n=21), Citrobacter freundii (n=8), and Shigella boydii (n=6). Forty-seven ESBL producers were identified using the synergic test. β-Lactamase genes detected were blaCTX-M-1 (E. coli/15; K. pneumonia/8; C. freundii/1; Sh. boydii/1), blaCTX-M-1+ blaOXA-1 (E. coli/4; K. pneumonia/3), blaCTX-M-1+ blaTEM-1-a (K. pneumonia/2), blaCTX-M-15+ blaTEM-1-a (K. pneumonia/1; Sh. boydii/1), blaCTX-M-15+ blaOXA-1 (K. pneumonia/1), blaCTX-M-15 (E. coli/3; K. pneumonia/1; Sh. boydii/3), and blaCTX-M-9 (C. freundii/3). Most strains (84.7%) showed a multiresistant phenotype. qnrA and qnrB genes were identified in six E. coli and in ten E. coli+one K. pneumonia isolates, respectively. The resistance to tetracycline and sulfonamide was conferred by the tet and sul genes. Characterization of phylogenic groups in E. coli isolates revealed phylogroups D (n=20 strains), B2 (n=2), and A (n=2). The studied virulence factor showed prevalence of fimA genes in 9 E. coli isolates (37.5%). Similarly, no strain revealed the three other virulence factors tested (eae, aer, and cnf1). Our findings confirmed that the lagoons of Bizerte may be a reservoir of multidrug resistance/ESBL-producing Enterobacteriaceae. This could lead to indisputable impacts on human and animal health, through the food chain

Methicillin-Resistant <i>Staphylococcus</i> <i>aureus</i> Strains Isolated from Burned Patients in a Tunisian Hospital: Molecular Typing, Virulence Genes, and Antimicrobial Resistance

Methicillin-resistant Staphylococcus aureus (MRSA) is one of the major causes of a variety of infections in hospitals and the community. Their spread poses a serious public health problem worldwide. Nevertheless, in Tunisia and other African countries, very little molecular typing data on MRSA strains is currently available. In our study, a total of 64 MRSA isolates were isolated from clinical samples collected from burned patients hospitalized in the Traumatology and Burns Center of Ben Arous in Tunisia. The identification of the collection was based on conventional methods (phenotypic and molecular characterization). The characterization of the genetic support for methicillin resistance was performed by amplification of the mecA gene by polymerase chain reaction (PCR), which revealed that 78.12% of S. aureus harbors the gene. The resistance of all the collection to different antibiotic families was studied. Indeed, the analysis of strain antibiotic susceptibility confirmed their multi-resistant phenotype, with high resistance to ciprofloxacin, gentamicin, penicillin, erythromycin, and tetracycline. The resistance to the last three antibiotics was conferred by the blaZ gene (73.43%), the erm(C) gene (1.56%), the msr(A) gene (6.25%), and tet(M) gene (7.81%), respectively. The clonal diversity of these strains was studied by molecular typing of the accessory gene regulator (agr) system, characterization of the SCCmec type, and spa-typing. The results revealed the prevalence of agr types II and III groups, the SCCmec type III and II cassettes, and the dominance of spa type t233. The characterization of the eight enterotoxins genes, the Panton-Valentine leukocidin and the toxic shock syndrome toxin, was determined by PCR. The percentage of virulence genes detected was for enterotoxins (55%), tst (71.88%), leukocidin E/D (79.69%), and pvl (1.56%) factors. Furthermore, our results revealed that the majority of the strains harbor IEC complex genes (94%) with different types. Our findings highlighted the emergence of MRSA strains with a wide variety of toxins, leukocidin associated with resistance genes, and specific genetic determinants, which could constitute a risk of their spread in hospitals and the environment and complicate infection treatment

Lineages, Virulence Gene Associated and Integrons among Extended Spectrum &beta;-Lactamase (ESBL) and CMY-2 Producing Enterobacteriaceae from Bovine Mastitis, in Tunisia

Extended Spectrum Beta-Lactamase (ESBL) Enterobacteriaceae are becoming widespread enzymes in food-producing animals worldwide. Escherichia coli and Klebseilla pneumoniae are two of the most significant pathogens causing mastitis. Our study focused on the characterization of the genetic support of ESBL/pAmpC and antibiotic resistance mechanisms in cefotaxime-resistant (CTXR) and susceptible (CTXS) Enterobacteriaceae isolates, recovered from bovine mastitis in Tunisia, as well as the analyses of their clonal lineage and virulence-associated genes. The study was carried out on 17 ESBL/pAmpC E. coli and K. pneumoniae and 50 CTXS&nbsp;E. coli. Detection of resistance genes and clonal diversity was performed by PCR amplification and sequencing. The following &beta;-lactamase genes were detected: blaCTX-M-15 (n = 6), blaCTX-M-15 + blaOXA-1 (2), bla CTX-M-15 + blaOXA-1 + blaTEM-1b (2), blaCTX-M-15 + blaTEM-1b (4), blaCMY-2 (3). The MLST showed the following STs: ST405 (n = 4 strains); ST58 (n = 3); ST155 (n = 3); ST471 (n = 2); and ST101 (n = 2). ST399 (n = 1) and ST617 (n = 1) were identified in p(AmpC) E. coli producer strains. The phylogroups A and B1 were the most detected ones, followed by the pathogenic phylogroup B2 that harbored the shigatoxin genes stx1/stx2, associated with the cnf, fimA, and aer virulence factors. The qnrA/qnrB, aac(6&prime;)-Ib-cr genes and integrons class 1 with different gene cassettes were detected amongst these CTXR/S isolated strains. The presence of different genetic lineages, associated with resistance and virulence genes in pathogenic bacteria in dairy farms, may complicate antibiotic therapies and pose a potential risk to public health

Characterization of Primary Action Mode of Eight Essential Oils and Evaluation of Their Antibacterial Effect against Extended-Spectrum β-Lactamase (ESBL)-Producing <i>Escherichia coli</i> Inoculated in Turkey Meat

The current study aims to evaluate the antimicrobial activity of eight essential oils (EOs) against multidrug-resistant Escherichia coli strains, producing extended-spectrum β-lactamase (ESBL) enzymes and isolated from foods. Disc-diffusion assay showed that the inhibition diameters generated by EOs varied significantly among the tested EOs and strains. In fact, EOs extracted from Thymus capitaus, Eucalyptus camaldulensis, Trachyspermum ammi and Mentha pulegium exerted an important antimicrobial effect against tested strains, with the diameters of inhibition zones varied between 20 and 27 mm. Moreover, minimal inhibition and bactericidal concentration (MIC and MBC) values demonstrated that T. capitatus EOs generate the most important inhibitory effect against E. coli strains, with MIC values ranging from 0.02 to 0.78%. Concerning the mode of action of T. capitatus EO, the obtained data showed that treatment with this EO at its MIC reduced the viability of E. coli strains, their tolerance to NaCl and promoted the loss of 260-nm-absorbing material. In addition, in the presence of T. capitatus EO, cells became disproportionately sensitive to subsequent autolysis. Moreover, the inhibitory effect of T. capitatus was evaluated against two E. coli strains, experimentally inoculated (105 CFU/g) in minced turkey meat, in the presence of two different concentrations of EO (MIC and 2 × MIC), and stored for 15 days. In both samples, EO exerted a bacteriostatic effect in the presence of concentrations equal to MIC. Interestingly, at 2 × CMI concentration, the bactericidal activity was pronounced after 15 days of storage. Our results highlighted that the use of essential oils, specially of T. capitatus, to inhibit or prevent the growth of extended-spectrum β-lactamase (ESBL)-producing E. coli in food, may be a promising alternative to chemicals

First Detection of CTX-M-1 in Extended-Spectrum β-Lactamase-Producing Escherichia coli in Seafood from Tunisia.

International audienceThe purpose of this study was to determine the carriage rate of Escherichia coli isolates in seafood, to analyze the phenotype and genotype of antimicrobial resistance in the recovered isolates, and to characterize extended-spectrum β-lactamase (ESBL) E. coli producers. E. coli isolates were recovered from 24 (34.3%) of the 70 seafood samples analyzed, and one isolate per sample was further characterized. Antibiotic resistance was determined by the disk diffusion method in the 24 isolates, with the following results (number of resistant isolates): tetracycline (8), streptomycin (7), ampicillin (6), trimethoprim-sulfamethoxazole (4), chloramphenicol (4), ciprofloxacin (3), cefotaxime (2), and ceftazidime (2). Six isolates showed a multiresistant phenotype (including at least three families of antibiotics). Among tetracycline-resistant E. coli isolates, tet(A) was detected in five isolates and tet(B) in two isolates. The qnr(A) or aac(6')-1b-cr genes were detected in two ciprofloxacin-resistant E. coli isolates, and the sul2 gene in two trimethoprim-sulfamethoxazole-resistant isolates. ESBL-containing E. coli isolates, carrying the blaCTX-M-1 gene, were detected in 2 of the 70 seafood samples, obtained from gilt-head bream aquaculture. The ESBL isolates were typed phylogenetically and by multilocus sequence typing, and they were ascribed to lineage ST48/A and to the new ST3497/B1; these isolates carried the fimA, aer, and papGIII virulence genes. One of the ESBL-producing E. coli isolates carried an unusual class 1 integron (with the array dfr32-ereA-aadA1). Seafood could be a source of multiresistant E. coli isolates for the aquatic environment, and these could enter the food chain