Shiga toxin-producing Escherichia coli (STEC) are foodborne pathogens that cause mild or serious diseases and can lead to people death. This study reports the prevalence and characteristics of STEC O157 and non-O157 in commercial ground beef and environmental samples, including meat table, knife, meat mincing machine, and manipulator hands (n = 450) obtained from 90 retail markets over a nine-month period. The STEC isolates were serotyped and virulence genes as stx (Shiga toxin), rfbO157] (O157 lipopolysaccharide), fliCH7 (H7 flagellin), eae (intimin), ehxA (enterohemolysin) and saa (STEC autoagglutinating adhesin), were determined. STEC O157 were identified in 23 (25.5%) beef samples and 16 (4.4%) environmental samples, while STEC non-O157 were present in 47 (52.2%) and 182 (50.5%), respectively. Among 54 strains isolated, 17 were STEC O157:H7 and 37 were STEC non-O157. The prevalent genotype for O157 was stx2/eae/ehxA/fliCH7 (83.4%), and for STEC non-O157 the most frequent ones were stx1/stx2/saa/ehxA (29.7%); stx2 (29.7%); and stx2/saa/ehxA (27%). None of the STEC non-O157 strains were eae-positive. Besides O157:H7, other 20 different serotypes were identified, being O8:H19, O178:H19, and O174:H28 the prevalent. Strains belonging to the same serotype could be isolated from different sources of the same retail market. Also, the same serotype could be detected in different stores. In conclusion, screening techniques are increasingly sensitive, but the isolation of STEC non-O157 is still a challenge. Moreover, with the results obtained from the present work, although more studies are needed, cross-contamination between meat and the environment could be suspected.

The emergence of resistance to antibiotics has been favored by abuse in the application of antimicrobials in human and animal medicine. Essential oils are a great resource to deal with this crisis. Melaleuca armillaris belongs to the family of Myrtaceae, rich in species with essential oils. Plant extracts has shown antimicrobial activity in many investigations. Cloxacillin (CLOX) is an antibiotic widely used in veterinary medicine against Staphylococcus aureus. Our aim was to assess pharmacodynamic interaction established by combining essential oil of M. armillaris (EO) with CLOX in search of a synergistic effect that maximizes the antibacterial activity against S. aureus. The EO was obtained by steam distillation and its composition was analyzed by a GC–FID–MS. The most abundant components in the EO were 1.8 cineole (72.3%), limonene (7.8%). and α-pinene (6%). We worked with wild type S. aureus strains (n = 3) isolated from Holstein cows, and S. aureus ATCC 29213 as the reference strain. The Minimum Inhibitory Concentration (MIC) of CLOX, EO and the combination was determined by microdilution in broth at pH 7.4; 6.5 and 5.0. The checkerboard method was applied to evaluate the interaction between CLOX and EO. The Fractional Inhibitory Concentration index (FIC) was established. From those combinations that yielded the lowest FIC values, we evaluated the index of antibacterial activity (E), established as the difference between the Log10 values of the number of viable bacteria at the initial (nt0) and at the end of the test (nt24). So, time-killing curves with CLOX and EO/CLOX combination at 0.5, 1, 2, 4, and 8 fold the MIC in broth at pH 7.4; 6.5 and 5.0 were prepared. We considered Bacteriostatic effect (E = 0) Bactericidal effect (E = −3) and Effect of virtual eradication of bacteria (E = −4). A clear synergic activity between the EO and the CLOX was demonstrated, which allows reducing the MIC of β-lactam against S. aureus. This interaction was favored by acidification of the medium, where lower concentrations of CLOX achieved a bactericidal effect, close to virtual eradication, in the presence of small amounts of EO.

Staphylococcus aureus is the major subclinical mastitis-causing pathogen in dairy cows. In some European and Latin American countries, rifaximin (RIF) is a commonly used therapy at drying off. Phytotherapeutics are alternatives for the treatment of infectious diseases. Melaleuca armillaris essential oil (EO) has been reported as a good antimicrobial against S. aureus. The aim of this work was to investigate, in vitro, the combined effect of EO and RIF to identify a synergic interaction against S. aureus in order to obtain enough information for subsequent pharmacokinetic/pharmacodynamic studies. The minimum inhibitory concentrations (MIC) for RIF, EO, and combinations of these against S. aureus strains were determined at pH 7.4, 6.5, and 5.0, representing intracellular conditions where S. aureus is usually located. The fractional inhibitory concentration index (FIC) and the index of antibacterial activity (E) were evaluated. The MIC of EO at pH 7.4 was 25-12.5 µL/mL and decreased with the acidity of the medium. RIF presented a high antimicrobial activity (0.032 µg/mL) against S. aureus regardless of the pH conditions. Combining RIF with EO, we found a synergic effect. A mix of 0.004 µg/mL of RIF and 12.5 µL/mL of EO led to a virtual eradication effect against wild-type strains at pH 7.4. Media acidification improves the EO/RIF activity, so EO would be a good adjuvant for RIF to treat staphylococcal infections and decrease antimicrobial resistance.

Doxycycline (DOX), tetracycline of second generation, is mainly active against Grampositive and Gram-negative bacteria, aerobic and anaerobic. Although there are few pharmacokinetic studies in chickens, it is frequently used for the colibacillosis treatment, salmonellosis, staphylococcal infections, avian mycoplasmosis and chlamydia. The objective of this study was to evaluate the withdrawal time (WT) of DOX formulation at 25 % in edible tissues, after its oral (PO) use in broilers. Forty healthy chicks (30-35 days of age) were used. DOX was administered with drinking water for 5 days at 10 mg kg-1 (N = 36); four untreated animals were reserved (control). Six animals per group were euthanized by cervical dislocation after desensitization by passage of an electric current through the head, after 24 hours until 9 d post treatment and control animals also. Muscle, liver, kidney and skin/fat samples were obtained. DOX was determined by HPLC with UV detection. DOX concentrations were determined in all tissues examined; generally falling below the MRL at 7 d after administration is terminated. It was estimated 6.58, 8.18, 8.69 and 6.96 d of WT for muscle, liver, kidney and skin/fat, respectively. After DOX administration at a rate of 10 mg kg-1 for 5 days in drinking water, a WT of 9 d is suggested in poultry destined for human consumption.

Resumen: La problemática actual de la resistencia bacteriana, promovida básicamente por el abuso en la utilización de antibióticos, lleva a la necesidad de encontrar alternativas terapéuticas más eficaces para enfrentar las infecciones bacterianas. Los aceites esenciales representan una gran herramienta para enfrentar esta crisis. Nuestro objetivo fue evaluar la interacción farmacodinámica establecida al combinar aceite esencial de Melaleuca armillaris con cloxacilina en busca de un efecto sinérgico que maximice la actividad del antibiótico frente a Staphylococcus aureus. Cepas de referencia y salvajes de dicha especie fueron enfrentadas al extracto vegetal y al antimicrobiano en forma individual y combinada mediante microdilución en caldo, modificando las condiciones del pH para emular el ambiente subcelular. Se determinó la interacción mediante la técnica de tablero de damas o checkerboard, se calculó el índice de concentración fraccionaria inhibitoria y, finalmente, se estableció el índice de actividad antibacteriana de la combinación. Nuestros resultados demuestran una clara actividad antimicrobiana del aceite esencial, que permite reducir las concentraciones mí-nimas inhibitorias del antibiótico betalactámico para el S. aureus. Esta interacción se ve favorecida por la acidificación del medio, lo que permite que menores concentraciones de cloxacilina logren un efecto bactericida, cercano a la erradicación virtual, en presencia de pequeñas cantidades de aceite esencial. Estos hallazgos resultan prometedores para enfrentar infecciones estafilocócicas de difícil resolución. Palabras clave: Melaleuca armillaris, cloxacilina, Staphylococcus aureus, resistencia, sinergismoAbstract: The current problem of bacterial resistance, largely promoted by indiscriminate use of antibiotics, leads to find more effective therapeutic alternatives to combat bacterial infections. Essential oils are a potential tool to deal with this issue. Our objective was to evaluate the pharmacodynamic interaction of Melaleuca armillaris essential oil and cloxacillin in search of a synergistic effect that maximizes the activity of the antibiotic against Staphylococcus aureus. Reference and wild strains of S. aureus were confronted to the essential oil and the antibiotic individually and in combination by microdilution in broth, modifying the pH conditions to emulate the subcellular environment. The interaction was determined by the checkerboard technique, the index of inhibitory fractional concentration was calculated and the index of antibacterial activity of the combination was finally established. Our results demonstrate a clear antimicrobial activity of the essential oil, which allows reducing the minimum inhibitory concentrations of cloxacillin for S. aureus. This interaction is favored by medium acidification, where lower concentrations of cloxacillin achieved a bactericidal effect, close to virtual eradication, in the presence of small amounts of essential oil. These findings are promising for dealing with staphylococcal infections with...