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Nasal swabs of 423 healthy humans who showed different levels of contact with animals (frequent, 168; sporadic, 94; no contact, 161) were obtained in Tunisia (2008-2009), and 99 of them presented other associated risk factors. Methicillin-resistant Staphylococcus aureus (MRSA) was detected in one of these 423 samples (0.24%), retrieved from a veterinarian. The MRSA isolate was mecA-positive, typed as ST80-t203-SCC mecIVc- agrIII, and contained tet(K), ant(6)-Ia, and aph(3′)-IIIa genes encoding tetracycline, streptomycin, and kanamycin resistance, respectively. This MRSA isolate also contained the lukF/lukS virulence gene encoding Panton-Valentine leukocidin. Fifty-four (12.8%) additional nasal samples contained methicillin-susceptible S. aureus (MSSA) and one isolate/sample was characterized. A high diversity of spa types ( n = 43; 4 new) and pulsed-field gel electrophoresis (PFGE) types ( n = 37) was detected among the 55 recovered S. aureus strains. The percentages of antimicrobial resistance/detected resistance genes were as follows: tetracycline [22%/ tet(K)- tet(L)- tet(M)], erythromycin [5%/ msrA], ciprofloxacin [14.5%], trimethoprim-sulfamethoxazole [2%/ dfrA], streptomycin [11%/ ant(6)-Ia], kanamycin [7%/ aph(3′)-IIIa], amikacin [5%], and chloramphenicol [2%]. Four and two isolates carried the lukF/lukS and eta and/or etb genes, respectively, and always in individuals with contact with animals. Eleven isolates carried the tst gene and were recovered from individuals with different levels of contact with animals. [ABSTRACT FROM AUTHOR]

In the last few years, different surveillances have been published in Africa, especially in northern countries, regarding antimicrobial resistance among husbandry animals. Information is still scarce, but the available data show a worrying picture. Although the highest resistance rates have been described against tetracycline, penicillins and sulphonamides, prevalence of plasmid-mediated quinolone resistance genes and extended spectrum β-lactamase (ESBL) are being increasingly reported. Among ESBLs, the CTX-M-1 group was dominant in most African surveys. Within this group, CTX-M-15 was the main variant both in animals and humans, except in Tunisia where CTX-M-1 was more frequently detected among Escherichia coli from poultry. Certain bla CTX -harbouring clones (ST131/B2 or ST405/D) are mainly identified in humans, but they have also been reported in livestock species from Tanzania, Nigeria or Tunisia. Moreover, several reports suggest an inter-host circulation of specific plasmids (e.g. bla -M-15 -carrying IncI1/ST3 in Tunisia, IncY- and Inc-untypeable replicons co-harbouring qnrS1 and bla CTX -M-1 -carrying IncF-type plasmids). International trade of poultry meat seems to have contributed to the spread of other ESBL variants, such as CTX-M-14, and clones. Furthermore, first descriptions of OXA-48- and OXA-181-producing E. coli have been recently documented in cattle from Egypt, and the emergent plasmid-mediated colistin resistance mcr-1 gene has been also identified in chickens from Algeria, Tunisia and South Africa. These data reflect the urgent need of a larger regulation in the use of veterinary drugs and the implementation of surveillance programmes in order to decelerate the advance of antimicrobial resistance in this continent.CTX -M-15 in Tanzania and the worldwide distributed bla CTX -M-15 -carrying IncF-type plasmids). International trade of poultry meat seems to have contributed to the spread of other ESBL variants, such as CTX-M-14, and clones. Furthermore, first descriptions of OXA-48- and OXA-181-producing E. coli have been recently documented in cattle from Egypt, and the emergent plasmid-mediated colistin resistance mcr-1 gene has been also identified in chickens from Algeria, Tunisia and South Africa. These data reflect the urgent need of a larger regulation in the use of veterinary drugs and the implementation of surveillance programmes in order to decelerate the advance of antimicrobial resistance in this continent., (© 2017 The Society for Applied Microbiology.)

Livestock-associated Staphylococcus aureus isolates, and especially those belonging to ST398, have been increasingly described in colonized and infected animals and humans, and also in food samples in several countries. The purpose of this study was to determine the frequency of S. aureus and methicillin-resistant S. aureus (MRSA) isolates in raw meat samples destined for food consumption in Tunisia, and to characterize the recovered isolates. One hundred sixty-nine food samples of animal origin were collected. Samples were inoculated onto selective mediums for S. aureus and MRSA recovery. Different molecular typing methods were implemented (pulsed-field gel electrophoresis [PFGE], multilocus sequence typing, spa-, agr-, and SCCmec typing). MRSA was detected in 2 of these 169 samples (1.2%), both of which were of chicken origin. The two MRSA isolates (one/sample) were typed as ST30-CC30-t012-agrIII-SCCmecV and ST398-CC398-t4358-agrI-SCCmecIVa. The MRSA ST398 strain presented resistance, in addition to β-lactams, to tetracycline (tet[M]) and erythromycin (erm[C]) and harbored the sen, hla, hlg, and hlgv virulence genes. Methicillin-susceptible S. aureus (MSSA) isolates were recovered from 42 of the 169 tested samples (24.8%). A high diversity of spa types (n=21) and SmaI-PFGE patterns (27 different profiles; 4 nontypeable) were detected among MSSA isolates. Four MSSA isolates were typed as ST398/CC398. The percentage of antimicrobial resistance and detected genes in MSSA isolates were as follows: tetracycline (28.6% tet[K] and tet[L]), kanamycin (9.5%, aph[3']-IIIa), tobramycin (2.4%, ant[4']-Ia), erythromycin (14.3%, erm[A], erm[C], msr[A]), and penicillin (95%). The genes lukS-lukF were detected in two MSSA isolates (4.5%), the gene tst in one isolate, and the gene eta in five isolates. To our knowledge, this is the first detection of MRSA and MSSA ST398 in food in an African country. The risk of transmission of S. aureus and MRSA carrying different antimicrobial resistance and virulence genes through the food chain cannot be ignored.

Reasons for Performing Study: Staphylococcus intermedius group (SIG) bacteria can colonise the nares of some animals but are also emerging pathogens in humans and animals., Objectives: To analyse SIG nasal carriage in healthy donkeys destined for food consumption in Tunisia and to characterise recovered isolates., Methods: Nasal swabs from 100 healthy donkeys were tested for SIG recovery, and isolates were identified by biochemical and molecular methods. Antimicrobial susceptibility of isolates was tested and detection of antimicrobial resistance and virulence genes was performed. Isolates were typed at the clonal level by multilocus sequence typing and SmaI pulsed-field gel electrophoresis., Results: Staphylococcus delphini and Staphylococcus pseudintermedius (included in SIG) were obtained in 19% and 2% of the tested samples, respectively, and one isolate per sample was characterised. All isolates were meticillin susceptible and mecA negative. Most S. delphini and S. pseudintermedius isolates showed susceptibility to all antimicrobials tested, with the exception of 2 isolates resistant to tetracycline (tet(M) gene) or fusidic acid. The following toxin genes were identified (percentage of isolates): lukS-I (100%), lukF-I (9.5%), siet (100%), se-int (90%), seccanine (19%) and expA (9.5%). Thirteen different pulsed-field gel electrophoresis profiles were identified among the 21 SIG isolates. Additionally, the following 9 different sequence types (STs) were detected by multilocus sequence typing, 6 of them new: ST219 (6 isolates), ST12 (5 isolates), ST220 (3 isolates), ST13, ST50, ST193, ST196, ST218 and ST221 (one isolate each)., Conclusions: Staphylococcus delphini and S. pseudintermedius are common nasal colonisers of donkeys, generally susceptible to the antimicrobials tested; nevertheless, these SIG isolates contain virulence genes, including the recently described exfoliative gene (expA) and several enterotoxin genes, with potential implications for public health. This is the first description of S. delphini in Tunisia. The Summary is available in Chinese - see Supporting information., (© 2014 EVJ Ltd.)

A total of 261 healthy farm and pet animals (75 cattle, 52 goats, 100 dogs, and 34 cats) from different regions of Tunisia were screened for Staphylococcus aureus nasal carriage. Molecular typing of isolates (by spa- and multilocus sequence-typing) was performed, and their antimicrobial resistance and virulence genotypes were determined by PCR and sequencing. S. aureus isolates were detected in 17 of 261 tested samples (6.5%). All S. aureus isolates recovered were methicillin-susceptible (MSSA), and one isolate/sample was further studied. Eight different spa types were detected (t189, t279, t582, t701, t1166, t1268, t1534, and t1773), and eight different sequence types were identified (ST6, ST15, ST45, ST133, ST188, ST700 [clonal complex CC130], ST2057, and a new ST2121). MSSA from pets (six isolates) showed resistance to (number of isolates, resistance gene): penicillin (six, blaZ), tetracycline (one, tet[M]), erythromycin one, erm[A]), streptomycin (one, ant[6]-Ia), and ciprofloxacin (one). All isolates from farm animals showed susceptibility to the tested antimicrobials, except for two penicillin-resistant isolates. Five S. aureus isolates from goats and cats harbored the lukF/lukS-PV genes, encoding the Panton-Valentine leukocidin, and six isolates from goats harbored the tst virulence gene. In addition, diverse combinations of enterotoxin genes were detected, including two variants of the egc cluster. Goats and cats could represent a reservoir of important toxin genes, with potential implications in animal and human health.

Background: The objective of this study was to determine the genetic lineages and the incidence of antibiotic resistance and virulence determinants of nasal Staphylococcus aureus isolates of healthy donkeys destined to food consumption in Tunisia., Results: Nasal swabs of 100 donkeys obtained in a large slaughterhouse in 2010 were inoculated in specific media for S. aureus and methicillin-resistant S. aureus (MRSA) recovery. S. aureus was obtained in 50% of the samples, being all of isolates methicillin-susceptible (MSSA). Genetic lineages, toxin gene profile, and antibiotic resistance mechanisms were determined in recovered isolates. Twenty-five different spa-types were detected among the 50 MSSA with 9 novel spa-types. S. aureus isolates were ascribed to agr type I (37 isolates), III (7), II (4), and IV (2). Sixteen different sequence-types (STs) were revealed by MLST, with seven new ones. STs belonging to clonal clomplex CC133 were majority. The gene tst was detected in 6 isolates and the gene etb in one isolate. Different combinations of enterotoxin, leukocidin and haemolysin genes were identified among S. aureus isolates. The egc-cluster-like and an incomplete egc-cluster-like were detected. Isolates resistant to penicillin, erythromycin, fusidic acid, streptomycin, ciprofloxacin, clindamycin, tetracycline, or chloramphenicol were found and the genes blaZ, erm(A), erm(C), tet(M), fusC were identified., Conclusions: The nares of donkeys frequently harbor MSSA. They could be reservoirs of the ruminant-associated CC133 lineage and of toxin genes encoding TSST-1 and other virulence traits with potential implications in public health. CC133 seems to have a broader host distribution than expected.

Beta-lactamase characterization was carried out in a collection of 18 extended-spectrum beta-lactamase (ESBL)-positive Escherichia coli isolates from blood (n=8) and urine (n=10) obtained in 2007 in a tunisian Hospital. All isolates were clonally unrelated according to PFGE analysis. Seventeen strains presented the bla(CTX-M-)₁₅ gene associated with bla (OXA-)₁ and four of these strains with the (TEM-)₁(b) gene. The remaining ESBL-positive strain contained the bla (CTX-M-)₉ gene associated with the bla (OXA-)₁ and bla (TEM-)₁(b) genes. The orf477 sequence was identified downstream of the bla(CTX-M-)₁₅ gene in all 17 bla(CTX-M-)₁₅-positive strains, and ISEcp1 upstream in 15 of them (in eight cases truncated by IS26). The presence of a class 1 integron was demonstrated in 4 of the 18 ESBL-positive strains (22.2%), with dfrA17 + aadA5 (3 strains) and dfrA12 + orfF + aadA2 (1 strain) being the gene cassettes identified. The variant aac(6´)-Ib-cr was found in 15 bla(CTX-M-)₁₅-containing strains. All 18 ESBL-positive strains were typed as phylogroup B2 and contained at least three of the eight tested virulence genes (fimA, papGIII, hlyA, cnf1, papC, aer, eae and bfp). Six bla(CTX-M-)₁₅-positive strains were included in the serotype O25b and one of them was typed as ST131. Another bla(CTX-M-)₁₅-positive strain serotype-O25 was typed as ST638. The bla(CTX-M-)₁₅, aac(6')- Ib-cr, and aac(3)-II genes were co-transferred by conjugation from 7 donor strains to E. coli CSH26 recipient strain. The bla(CTXM-)₁₅ gene is prevalent among ESBL-positive E. coli strains in the studied hospital, that is frequently found together with aac(6')- Ib-cr, and aac(3)-II genes. The detection of the clone O25b-St131 in a bla(CTX-M-)₁₅ strain corroborates its worldwide dissemination.

Fifty-five Escherichia coli isolates were acquired from chicken and turkey meat obtained from two slaughterhouses in Tunis. Eighty-nine percent, 80%, 78%, 67%, 45%, 27%, 7%, 4%, and 2% of these isolates showed resistance to tetracycline, trimethoprim/sulfamethoxazole, streptomycin, nalidixic acid, ampicillin, chloramphenicol, ciprofloxacin, colistine, and gentamicin, respectively. No resistance was detected to cefotaxime, ceftazidime, or amikacin. bla(TEM) gene was found in 22 of 25 ampicillin-resistant isolates, and 1 isolate harbored bla(OXA-1) gene. Tetracycline resistance was predominately mediated by the tetA gene. The sul1, sul2, and sul3 genes, alone or combined, were detected in 46 of 48 sulfonamide-resistant isolates, and sul1 and sul3 were included in class 1 integrons in some cases. Sixty percent of isolates harbored integrons (class 1, 30 isolates; class 2, 5 isolates). Class 2 integrons contained in all cases the dfrA1-sat1-aadA1-orfX gene cassette arrangement. Nine gene cassette arrangements have been detected among class 1 integrons, containing different alleles of dfrA (five alleles) and aadA (2 alleles) genes, which encode trimethoprim and streptomycin resistance, respectively. An uncommon gene cassette array (sat-psp-aadA2-cmlA1-aadA1-qacH-IS440-sul3) has been identified in three class 1 integron-positive isolates, and one additional isolate had this same structure with the insertion of IS26 inside the aadA1 gene (included in GenBank with accession no. FJ160769). The 55 studied isolates belong to the four phylogenic groups of E. coli, and phylogroups A and D were the most prevalent ones. At least one virulence-associated gene (fimA, papC, or aer) was detected in 44 of the 55 (80%) studied isolates. E. coli isolates of poultry origin could be a reservoir of antimicrobial-resistance genes and of integrons, and its evolution should be tracked in the future.

Phenotypic and genotypic characterization of antimicrobial resistance was conducted for 98 Escherichia coli isolates recovered from 40 food samples of animal origin (poultry, sheep, beef, fish, and others) obtained in supermarkets and local butcheries in Tunis during 2004 and 2005. Susceptibility to 15 antimicrobial agents was tested by disk diffusion and agar dilution methods, the mechanisms of resistance were evaluated using PCR and sequencing methods, and the clonal relationship among isolates was evaluated using pulsed-field gel electrophoresis. High resistance was detected to tetracycline, sulphonamides, nalidixic acid, ampicillin, streptomycin, and trimethoprim-sulfamethoxazole (29 to 43% of isolates), but all isolates were susceptible to cefotaxime, ceftazidime, cefoxitin, azthreonam, and amikacin. One-third of the isolates had multiresistant phenotypes (resistance to at least five different families of antimicrobial agents). Different variants of blaTEM, tet, sul, dfrA, aadA, and aac(3) genes were detected in most of the strains resistant to ampicillin, tetracycline, sulphonamide, trimethoprim, streptomycin, and gentamicin, respectively. The presence of class 1 and class 2 integrons was studied in 15 sulphonamide-resistant unrelated E. coli strains, and 14 of these strains harbored class 1 integrons with five different arrangements of gene cassettes, and a class 2 integron with the dfrA 1 + sat + aadA 1 arrangement was found in one strain. This study revealed the high diversity of antimicrobial resistance genes, some of them included in integrons, in E. coli isolates of food origin.

The polymorphism in pbp5 gene was investigated in nine unrelated clinical gentamicin-resistant Enterococcus faecium strains with different minimal-inhibitory-concentration values for ampicillin (six ampicillin-resistant and three ampicillin-susceptible). Five alleles were detected when the pbp5 C-terminal region was analysed, two of them in the ampicillin-resistant strains showed a new allele characterised by the Thr416Ala and Val462Ala substitutions. Two different alleles were identified when the pbp5 N-terminal region was studied; one of them in the unique strain (E. faecium 83) that presented very low ampicillin MIC (<0.125 microg/ml) and a nucleotidic mutation implicating a stop codon at 451 position. RT-PCR experiments carried out on five E.faecium positive results indication the expression of this gene. Specific mutations in pbp5 gene could be responsible of the high MIC values of some of the E. faecium strains.

Susceptibility testing for 15 antibiotics was performed in a series of 191 clinical enterococci recovered in a Tunisian Hospital during 2000-2003. Species detected were the following ones (number of isolates): E. faecalis (139), E. faecium (41), E. casseliflavus (5), E. gallinarum (3), E. avium (2) and E. hirae (1). The percentages of antibiotic resistance detected were as follows (E. faecalis/ E. faecium/ other species) : penicillin (0/ 73/ 9%), tetracycline (78/ 44/ 54%), chloramphenicol (52/ 29/ 27%), erythromycin (66/ 100/ 82%), spiramycin (84/ 83/ 64%), pristinamycin (100/ 0/ 73%), trimethoprim-sulfamethoxazole (88/ 78/ 91%), rifampicin (72/ 41/ 0%), vancomycin (0/ 0/ 36%), teicoplanin (0/ 0/ 0%), high-level-resistance for gentamicin (24/ 29/ 45%), streptomycin (34/ 56/ 55%) and kanamycin (41/ 68/ 55%). Increased vancomycin minimum inhibitory concentrations (MICs) were only detected in E. casseliflavus and E. gallinarum isolates (MIC range 8-24 microg/ml). The erm(B), catA, tet(M), aac(6')-aph(2''), aph(3')-IIIa, and ant(6)-Ia genes were detected in 91%, 32%, 86%, 98%, 100%, and 72% of the E. faecium and E. faecalis isolates resistant to erythromycin, chloramphenicol, tetracycline and high-level-resistant to gentamicin, kanamycin and streptomycin, respectively. A total of 20 unrelated pulsed-field-gel-electrophoresis patterns were found in the series of 46 high-level gentamicin-resistant E. faecalis and E. faecium isolates of this study.