[Molecular characterization of diarrheagenic Escherichia coli from an outpatient pediatric population with diarrhea attended in two hospitals from Buenos Aires, Argentina]. / Caracterización molecular de Escherichia coli diarreogénica proveniente de población pediátrica ambulatoria con diarrea, atendida en dos hospitales de Buenos Aires, Argentina.

Diarrheagenic Escherichia coli comprises a heterogeneous group of pathotypes or pathogenic variants that share phenotypic characteristics with marked differences in virulence genes, colonization sites, pathogenesis, clinical presentation, and epidemiology of infection. The most studied pathotypes are Shiga toxin-producing E.coli (STEC), enterotoxigenic E.coli (ETEC), enteropathogenic E.coli (EPEC), enteroaggregative E.coli (EAEC), and enteroinvasive E.coli (EIEC). The objective of the study was to characterize the isolates of diarrheagenic E.coli from an outpatient pediatric population with diarrhea attended in two public hospitals from Buenos Aires, Argentina. Diarrheagenic E.coli pathotypes were investigated by amplifying characteristic virulence gene fragments: intimin (eae), heat-labile toxin (lt), heat-stable toxins (stp, sth), invasion plasmid antigen H (ipaH), transcriptional activator R (aggR) and Shiga toxins (stx1, stx2). Molecular subtyping of isolates was performed using PFGE (XbaI). Diarrheagenic E.coli was detected in 14% (84/601) of cases. The EAEC pathotype was prevalent, while ETEC, STEC, EPEC and EIEC were found in a lower proportion. EAEC isolates exhibited a high degree of genetic diversity. All pathotypes were found in children under 5years of age, while only EAEC, EIEC and ETEC were detected in the older population. Future studies that include the characterization of isolates from a greater number of genes and populations from other geographical areas will be necessary to determine the relevance of diarrheagenic E.coli in Argentina.

Occurrence and genetic characterization of Shiga toxin-producing Escherichia coli on bovine and pork carcasses and the environment from transport trucks.

Shiga toxin-producing Escherichia coli (STEC) are foodborne pathogens causing severe diseases. The ability of STEC to produce disease is associated with Shiga toxin (Stx) production. We investigated the occurrence of STEC on bovine and pork carcasses and walls of trucks where they were transported, and we characterized virulence genes and serotypes of STEC strains. We compared the whole genomic sequencing of a STEC O157:H7 strain isolated from a bovine carcass in this work and a STEC O157:H7 strain isolated from a child with HUS, both isolated in 2019. We studied the relationship between these isolates and others collected in the database. The results show a 40% of STEC and two different serogroups were identified (O130 and O157). STEC O157:H7 were isolated from bovine carcasses and harbored stx2, eae, ehxA, katP, espP, stcE, ECSP_0242/1773/2687/2870/2872/3286/3620 and were classified as lineage I/II. In STEC non-O157 isolates, three isolates were isolated from bovine carcasses and harbored the serogroup O130 and one strain isolated from pork carcasses was O-non-typeable. All STEC non-O157 harbored sxt1 gene. The analysis from the whole genome showed that both STEC O157:H7 strains belonged to the hypervirulent clade 8, ST11, phylogroup E, carried the allele tir 255 T > A T, and they were not clonal. The analysis of information allows us to conclude that the STEC strains circulate in pork and bovine carcasses arriving in transport. This situation represents a risk for the consumers and the need to implement an integrated STEC control in the food chain.

Antinematodic effect of Enterococcus faecalis CECT7121 using Trichinella spiralis as a model of nematode infection in mice.

Nematode infections affect a significant percentage of the human population worldwide, especially in developing countries. There are a small number of drugs available to treat these infections, with variable outcomes. Therefore, the potential use of probiotics to help control parasitic infections has emerged as a suitable option. The main goal of this work was to assess the antinematodic effect of the probiotic Enterococcus faecalis CECT7121 (EFCECT7121) in vitro and in vivo, using Trichinella spiralis as a nematode model of infection. The in vitro assay showed a reduction in T. spiralis larvae viability of 31.6% when compared with the control group (6.3%) after 48 h incubation with EFCECT7121. Nevertheless, the isolated antimicrobial peptide AP7121 when inoculated at different concentrations did not reveal any larvicidal effect. Different EFCECT7121 treatment schemes in mice were evaluated, and the reduction of the enteral and parenteral burden of T. spiralis was determined. In addition, the protective effect of EFCECT7121 combined with the conventional anthelmintic albendazole (ABZ, 5 mg/kg) was also assessed. The oral administration of EFCECT7121 previous T. spiralis infection produced a reduction in the larvae per gram (LPG) of mice muscle tissue ranging from 32.8 to 47.9% on the 28th day post-infection. ABZ alone and the combination EFCECT7121 + ABZ produced a reduction of the LPG of muscle tissue of 62 and 60.7%, respectively. Results obtained in the current work support the hypothesis that probiotics such as EFCECT7121 have an antinematodic effect, and their combination with conventional anthelmintic drugs may result useful for improving clinical and parasitological outcomes.

Primer reporte de Escherichia coli diarreogénica en población pediátrica ambulatoria con diarrea atendida en la ciudad de La Plata, Argentina / First report of diarrheagenic Escherichia coli in pediatric outpatient population with diarrhea in La Plata, Argentina

Resumen Escherichia coli diarreogénica comprende un grupo heterogéneo de cepas que presentan diversos factores de virulencia y causan diferentes síndromes diarreicos. Los patotipos más estudiados son Escherichia coli enteropatogénica (EPEC), Escherichia coli enterotoxigé-nica (ETEC), Escherichia coli enteroagregativa (EAEC), Escherichia coli enteroinvasiva (EIEC) y Escherichia coli productora de toxina Shiga (STEC). El objetivo de este estudio fue estimar la frecuencia de infección de los diversos patotipos de E. coli diarreogénica en una población pediátrica ambulatoria con diarrea, atendida en el Hospital Sor María Ludovica de La Plata, Argentina, durante el período mayo-octubre de 2017. Los patotipos fueron detectados mediante la amplificación molecular de ocho genes de virulencia característicos. Se estudiaron las heces de 211 ninos (76% menores de 5 años). Se detectó infección con E. coli diarreogénica en el 12,3% (n = 26/211) de los niños con diarrea. Los patotipos identificados fueron EAEC, ETEC (todos lt positivos), EPEC y STEC (stx2 y eae positivos). El patotipo EAEC fue prevalente en todos los grupos etarios, mientras que los patotipos ETEC, EPEC y STEC solamente se observaron en niñnos menores de 5 anños. Este estudio constituye el primer reporte de detección por técnicas de amplificación molecular de Escherichia coli diarreogénica en una población pediátrica ambulatoria con diarrea, de la zona de La Plata. Se necesitan estudios más amplios que incluyan la caracterización de los aislamientos abarcando un mayor número de genes, controles asintomáticos, distintas épocas del ano y población de diversas áreas geográficas para esclarecer la relevancia de la infección por E. coli diarreogénica en niños de Argentina.

Abstract Diarrheagenic Escherichia coli is a heterogeneous group of strains that presents various virulence factors and causes different diarrheal syndromes. The most studied pat-hotypes are enteropathogenic Escherichia coli (EPEC), enterotoxigenic Escherichia coli (ETEC), enteroaggregative Escherichia coli (EAEC), enteroinvasive Escherichia coli (EIEC) and Shiga toxin-producing Escherichia coli (STEC). The objective was to estimate the frequency of infec-tion of diarrheagenic E. coli pathotypes in children with diarrhea, attended at the Sor María Ludovica Hospital in La Plata, Argentina, during the period May-October 2017. E. coli pathotypes were detected by molecular amplification of eight characteristic virulence genes. The feces of 211 children (76% under 5 years) were studied. Infection with diarrheagenic E. coli was detected in 12.3% of the samples. The pathotypes were EAEC (10.43%), ETEC (1.42%, all of them positive for thermolabile toxin), EPEC (0.95%) and STEC (0.47%, positive for Shiga toxin 2). The EAEC pathotype was prevalent in children of all age groups, while ETEC, EPEC and STEC were only observed in children under 5 years of age. This study constitutes the first report of diarrheagenic Escherichia coli detection in an outpatient pediatric population with diarrhea from La Plata, using molecular amplification techniques. Broader future studies, including the charac-terization of the isolates with the largest number of genes, asymptomatic controls, different times of the year and population from different geographic areas will be necessary to clarify the relevance of diarrheagenic E. coli infection in children from Argentina.

Características clínicas y microbiológicas de infecciones de piel y partes blandas en pacientes pediátricos de dos hospitales de Buenos Aires / Skin and soft tissue infections in pediatrics

Las infecciones de piel y partes blandas (IPPB) en niños son una de las principales causas de prescripción de antimicrobianos. El objetivo del estudio fue describir las características clínicas y microbiológicas de las IPPB ambulatorias de niños asistidos en dos hospitales zonales. Se realizó un estudio prospectivo entre el 1/11/2017 y el 1/11/2018. Se incluyeron pacientes entre 1 mes y 15 años internados en dos hospitales. Se evaluó: edad, sexo, localidad, factores predisponentes, tipo de IPPB, muestras biológicas realizadas, aislamiento microbiológico, tratamiento empírico indicado y evolución del cuadro. Se realizó antibiograma y determinación genética. Se calculó chi2, IC95, OR; α=5%. N= 94. 58,7% masculinos. 12 pacientes <1 año, 85 >1 año (promedio de edad 4 años, 1-15). El 36% de Tandil y 63,8% de Florencio Varela. El 59,6% corresponden a IPPB purulentas. Se aislaron microorganismos en un 59,6%. Los aislamientos principales: SAMR (40,4%), SAMS (7,4%), S. agalactiae (2,1%) y S. pyogenes (2,1%). El 100% de SAMR son portadores de gen mecA y SCCmec tipo IV, sin multirresistencia. No hubo diferencia estadística entre los factores de riesgo evaluados para el desarrollo de IPPB por SAMR. El 52,1% de los niños recibió tratamiento antibiótico combinado, siendo la más indicada TMS-SMX + CLI en 36 eventos. (38,3%). La evolución fue favorable: no hubo diferencia significativa entre el subgrupo que se aisló SAMR y el que no se aisló SAMR; 91,9% (34/37) y 92,6% (50/54) correspondientemente (chi2: 0,01; p= 0,97 IC95: 0,26-3,88). El principal agente etiológico fue SAMRco, debiendo adecuar los tratamientos a este microorganismo.

Skin and soft tissue infections (SSIs) in children are one of the main causes of antimicrobial prescription. The aim of the study was to describe the clinical and microbiological characteristics of outpatient SSIs in children attended in two hospitals. A prospective study was conducted between 11/1/2017 and 11/1/2018. Patients between 1 month and 15 years old, hospitalized were included. We evaluated: age, sex, locality, predisposing factors, type of IPPB, biological samples taken, microbiological isolation, empirical treatment indicated and evolution of the condition. An antibiogram and genetic determination were performed. Chi2, CI95, OR; α=5% were calculated. N= 94. 58.7% male. 12 patients <1 year, 85 >1 year (mean age 4 years, 1-15). 36% were from Tandil and 63.8% from Florencio Varela. 59.6% corresponded to purulent SSIs. The diagnostic yield was 59.6%. Main isolates: MRSA (40.4%), MSSA (7.4%), S. agalactiae (2.1%) and S. pyogenes (2.1%). 100% of MRSA carried the mecA gene and SCCmec type IV, with no multidrug resistance. There was no statistical difference between the risk factors evaluated. 52.1% of children received combined antibiotic treatment, the most indicated being TMS-SMX + CLI in 36 events. (38,3%). Evolution was favorable: there was no significant difference between the subgroup that isolated MRSA and the subgroup that did not isolate MRSA; 91.9% (34/37) and 92.6% (50/54) respectively (chi2: 0.01; p= 0.97 CI95: 0.26-3.88). The main etiological agent was MRSA, and treatments should be adapted to this microorganism

[First report of diarrheagenic Escherichia coli in pediatric outpatient population with diarrhea in La Plata, Argentina]. / Primer reporte de Escherichia coli diarreogénica en población pediátrica ambulatoria con diarrea atendida en la ciudad de La Plata, Argentina.

Diarrheagenic Escherichia coli is a heterogeneous group of strains that presents various virulence factors and causes different diarrheal syndromes. The most studied pathotypes are enteropathogenic Escherichia coli (EPEC), enterotoxigenic Escherichia coli (ETEC), enteroaggregative Escherichia coli (EAEC), enteroinvasive Escherichia coli (EIEC) and Shiga toxin-producing Escherichia coli (STEC). The objective was to estimate the frequency of infection of diarrheagenic E. coli pathotypes in children with diarrhea, attended at the Sor María Ludovica Hospital in La Plata, Argentina, during the period May-October 2017. E. coli pathotypes were detected by molecular amplification of eight characteristic virulence genes. The feces of 211 children (76% under 5 years) were studied. Infection with diarrheagenic E. coli was detected in 12.3% of the samples. The pathotypes were EAEC (10.43%), ETEC (1.42%, all of them positive for thermolabile toxin), EPEC (0.95%) and STEC (0.47%, positive for Shiga toxin 2). The EAEC pathotype was prevalent in children of all age groups, while ETEC, EPEC and STEC were only observed in children under 5 years of age. This study constitutes the first report of diarrheagenic Escherichia coli detection in an outpatient pediatric population with diarrhea from La Plata, using molecular amplification techniques. Broader future studies, including the characterization of the isolates with the largest number of genes, asymptomatic controls, different times of the year and population from different geographic areas will be necessary to clarify the relevance of diarrheagenic E. coli infection in children from Argentina.

The antimicrobial effect behind Cannabis sativa.

The development of multidrug-resistant bacteria has revealed the need for new antimicrobial compounds. Cannabis sativa preparations have a long history of medical applications, including the treatment of infectious diseases. This review collects the information about the activity of C. sativa extracts and its main components (cannabinoids and terpenes) against pathogenic bacteria and fungus, to assess its potential using as antimicrobial agents.

Bacteriemia por Staphylococcus aureus con sensibilidad disminuida heterogénea a vancomicina: reporte de un caso / Staphylococcus aureus bacteremia with heterogeneous intermediate-level vancomycin resistance: a case report

Staphylococcus aureusmeticilina-resistente (SAMR)es una causa frecuente de bacteriemias intrahospitalarias. Para su tratamiento se utiliza vancomicina y han emergido cepas con sensibilidad disminuida heterogénea (h-VISA) que albergan subpoblaciones con sensibi-lidad reducida a vancomicina. Se comunica un caso de bacteriemia intra-tratamiento con vancomicina por SAMR h-VISA. El aislamiento muestra sensibilidad a vancomicina (CIMvan: 1 µg/mL), sin embargo E-test GRD sugiere h-VISA (CIMvan: 2 µg/mL y CIMtei: 8 µg/mL). El análisis del perfil poblacional - área bajo la curva (PAP-AUC) valida este hallazgo. Se rota a linezolid con resolución clínica.

Methicillin-resistant Staphylococcus aureus (SAMR) is a common cause of nosocomial bacteremia. Vancomycin, a glycopeptide, is widely employed for the therapy of SAMR infections. In recent years, heterogeneous vancomycin-intermediate strains (h-VISA) have emerged. We report a case of intra-treatment bacteremia caused by SAMR h-VISA. The isolate shows susceptibility to vancomycin (MICvan: 1 µg/mL). But the GRD E-test suggests h-VISA (MICvan: 2 µg/mL and MICtei: 8 µg/mL). The population analysis profile - area under the curve (PAP-AUC) validates SAMR h-VISA. Rotation of antibiotic therapy with linezolid is done, with good clinical outcome.

Gram-positive bacteriocins: usage as antimicrobial agents in veterinary medicine.

Antimicrobial resistance is a worldwide spread phenomenon that affects both human and veterinary medicine. This issue has led to a "One Health" approach in order to coordinate efforts and set back the development of drug-resistant microbes. In the search for alternatives therapies, bacteriocins or antimicrobial peptides have proven to be effective both in vitro and in vivo for multiples pathogens, even those resistant to many classic antibiotics. Gram-positive bacteriocins have been the most studied to the present. The use of bacteriocins as therapeutically active molecules is limited mainly due to difficulties in production, purification, delivery systems and regulatory approvals. To overcome some of these limitations, biotechnological and nanotechnological approaches are evaluated. Bacteriocins proved to be a good complement for conventional antibiotics therapy. Antimicrobial peptides are nowadays included in the veterinary products such as udder disinfectant for dairy cattle and dermatological medicated wipe for topical use on dogs, cats, and horses. But there are other potential uses to explore in the veterinary field for both companion and production animals.

In vitro activity of the antimicrobial peptide AP7121 against the human methicillin-resistant biofilm producers Staphylococcus aureus and Staphylococcus epidermidis.

In vitro activity against methicillin-resistant Staphylococcus aureus (MRSA) and Staphylococcus epidermidis biofilm producers from blood cultures of patients with prosthetic hip infections was evaluated. The Minimum Inhibitory Concentration (MIC) for AP7121 was determined and the bactericidal activity of AP7121 (MICx1, MICx4) against planktonic cells was studied at 4, 8 and 24 h. The biofilms formed were incubated with AP7121 (MICx1, MICx4) for 1 and 24 h. The anti-adhesion effect of an AP7121-treated inert surface over the highest MIC isolate was studied with scanning electron microscopy (SEM). The bactericidal activity of AP7121 against all the planktonic staphylococcal cells was observed at 4 h at both peptide concentrations. Dose-dependent anti-biofilm activity was detected. AP7121 (MICx4) showed bactericidal activity at 24 h in all isolates. SEM confirmed prevention of biofilm formation. This research showed the in vitro anti-biofilm activity of AP7121 against MRSA and S. epidermidis and the prevention of biofilm formation by them on an abiotic surface.

ENCAPSULACIÓN DEL PÉPTIDO ANTIMICROBIANO AP7121 PARA SU ADMINISTRACIÓN POR VÍA ORAL / Encapsulation of antimicrobial peptide AP7121 for oral administration

El péptido antimicrobiano AP7121, producido por Enterococcus faecalis CECT7121, presenta actividad bactericida sobre patógenos Gram positivos. Sin embargo, su vía de administración oral está inhibida por la acidez gástrica y enzimas proteolíticas digestivas. El objetivo del presente trabajo fue analizar la eficacia de la encapsulación de AP7121 para su administración por vía oral. Para ello, se realizó la encapsulación del péptido antimicrobiano mediante la formación de gotas de alginato de sodio estéril 2,2% conteniendo AP7121 (30,0 mg/L) y circulando en dispositivo extrusor. Se evaluó la actividad inhibitoria de cápsulas obtenidas mediante la determinación de la Concentración Inhibitoria Mínima de AP7121 (CIMAP7121), con Listeria monocytogenes ATCC 19111 (LM, CIMAP7121: 0,8 mg/L). Asimismo, se investigó la estabilidad del péptido antimicrobiano frente a las enzimas proteolíticas tripsina, α-quimotripsina, proteinasa K y pronasa E (1 mg/mL) y se evaluó el efecto del pH utilizando una solución de HCl, pH=2,0. Las cápsulas obtenidas fueron uniformes y se obtuvo una concentración final de AP7121 de 29,7±0,3 mg/L con una CIMAP7121: 0,8 mg/L para LM. Frente a enzimas proteolíticas, no se observó descenso de actividad, permaneciendo inalterable en las cápsulas (CIMAP7121 de LM: 0,8 mg/L). Luego de la exposición a pH=2,0, se observó pérdida significativa de actividad a las 4 h de exposición. Los resultados obtenidos habilitarían la utilización de AP7121 en cápsulas para su administración por vía oral, dada su resistencia al pH ácido estomacal y enzimas proteolíticas, factores limitantes para su uso sin protección de su actividad.

The antimicrobial peptide AP7121, produced by Enterococcus faecalis CECT7121, presents bactericidal activity on Gram-positive pathogens. However, its administration via oral route is inhibited by gastric acidity and proteolytic digestive enzymes. The objective of this work was to analyze the effectiveness of the encapsulation of AP7121 for oral administration. In order to do this, the encapsulation of the antimicrobial peptide was performed by forming sterile 2.2% sodium alginate drops containing AP7121 (30.0 mg / L) and circulating in an extruder device. The inhibitory activity of the obtained capsules was evaluated by determining the Minimum Inhibitory Concentration of AP7121 (MICAP7121), with Listeria monocytogenes ATCC 19111 (LM, MICAP7121: 0.8 mg / L). Likewise, the stability of the antimicrobial peptide was investigated against trypsin, α-chymotrypsin, proteinase K and pronase E (1 mg / mL) proteolytic enzymes, and the effect of pH was evaluated using an HCl, pH = 2.0 solution. The capsules obtained were uniform and a final AP7121 concentration of 29.7 ± 0.3 mg / L with a MIC AP7121: 0.8 mg / L for ML was obtained. Against proteolytic enzymes, no decrease in activity was observed, remaining unchanged in the capsules (MICAP7121 of LM: 0.8 mg / L). After exposure to pH = 2.0, a significant loss of activity was observed after 4 h of exposure. The results obtained would enable the use of AP7121 in capsules for oral administration given its resistance to stomach acid pH and proteolytic enzymes, factors that limit the use without protection of its activity.

Detection of ß-Lactamase-Producing Enterococcus faecalis and Vancomycin-Resistant Enterococcus faecium Isolates in Human Invasive Infections in the Public Hospital of Tandil, Argentina.

The study's aim was to analyze the population structure of enterococci causing human invasive infections in a medium-sized Argentinian Hospital coincidental with a 5 year-period of increased recovery of antibiotic resistant enterococci (2010-2014). Species identification (biochemical testing/MALDI-TOF-MS), antimicrobial susceptibility (disk-diffusion) and clonal relatedness (PFGE/MLST/BAPS) were determined according to standard guidelines. ß-lactamase production was determined by a nitrocefin test and confirmed by PCR/sequencing. The isolates were identified as Enterococcus faecalis and Enterococcus faecium at a 2:1 ratio. Most of the E. faecalis isolates, grouped in 25 PFGE-types (ST9/ST179/ST236/ST281/ST388/ST604/ST720), were resistant to high-levels (HLR) of gentamicin/streptomycin. A ST9 clone (bla+/HLR-gentamicin) was detected in patients of different wards during 2014. E. faecium isolates were grouped in 10 PFGE-types (ST25/ST18/ST19/ST52/ST792), with a low rate of ampicillin resistance. Five vancomycin-resistant E. faecium, three vanA (ST792/ST25) and two vanB (ST25) were detected. The ST25 clone carried either vanA or vanB. The recovery of a bla+-ST9-E. faecalis clone similar to that described in the late 1980s in Argentina suggests the possibility of a local hidden reservoir. These results reflect the relevance of local epidemiology in understanding the population structure of enterococci as well as the emergence and spread of antimicrobial resistance in predominant enterococcal clonal lineages.

Caracterización fenotípica de variantes pequeñas de Staphylococcus aureus: un nuevo desafío / Phenotypic characterization of small colony variants of Staphylococcus aureus: a new challenge / Caracterizaçao fenotípica de pequenas variantes de Staphylococcus aureus: um novo desafio

En infecciones crónicas y recurrentes por Staphylococcus aureus se han descripto subpoblaciones de colonias pequeñas (VCPSa). El objetivo de este trabajo fue reconocer las características fenotípicas de VCPSa para optimizar su detección y caracterización a partir de materiales clínicos provenientes de infecciones crónicas. Se analizaron n=3 VCPSa de pacientes adultos con infecciones crónicas de tejidos blandos. Las muestras se inocularon en agar nutritivo, agar sangre, agar chocolate y agar Schaedler suplementado. Se realizaron tinción de Gram, catalasa, coagulasa libre, pruebas de dependencia para hemina, menadiona y timidina y, desarrollo/ataque del manitol en agar manitol salado. La sensibilidad antibiótica se efectuó en agar Mueller Hinton suplementado, según las pruebas de dependencia. Se investigó la presencia de proteína ligadora de penicilina anómala (PBP2´) por aglutinación con látex. Las VCPSa se detectaron en los medios de cultivo enriquecidos. Estas bacterias dieron positivas las pruebas de catalasa y coagulasa, y eran dependientes de menadiona y hemina. En los tres aislamientos se observó resistencia a cefoxitina y se detectó la PBP2´.

In chronic and recurrent infections, small colonies of Staphylococcus aureus subpopulations (SCVSa) have been observed. The objective of the present study was to recognize the phenotypic characteristics of SCVSa isolated from patients with chronic infections to optimize their detection. SCVSa of adult patients n=3 with chronic soft tissue infections were analyzed. Samples were inoculated on nutritive agar, blood-agar, chocolate agar and supplemented Schaedler agar. Subsequently, Gram stain, catalase, free coagulase, dependence tests for hemin, menadione and thymidine, and growth/fermentation of mannitol on salt mannitol agar were performed. Antibiotic susceptibility tests were performed by the agar diffusion method on supplemented Mueller Hinton agar, according to dependence assays results. Anomalous penicillin binding protein (PBP2') was investigated by latex agglutination. SCVSa were detected in all enriched culture media. They showed catalase and coagulase activities, and menadione and hemin dependence. By the agar diffusion test, cefoxitin resistance was found in all isolates; PBP2' was detected as well.

Nas infecções crônicas e recorrentes por Staphylococcus aureus, subpopulações de pequenas colônias (VCPSa) foram descritas. O objetivo desse trabalho foi reconhecer as características fenotípicas de VCPSa para otimizar sua detecção e caracterização a partir de materiais clínicos provenientes de infecções crônicas. Foram analisados n=3 VCPSa de pacientes adultos com infecções crônicas de tecidos moles. As amostras foram inoculadas em agar nutritivo, agar sangue; agar chocolate e agar Schaedler enriquecido. Foram realizados testes de coloração de Gram, catalase, coagulase livre, testes de dependência para hemina, menadiona e timidina, e desenvolvimento/fermentação do manitol em agar manitol salgado. A sensibilidade antibiótica foi realizada em agar Mueller Hinton suplementado, de acordo com os testes de dependência. Foi investigada a presença de proteína ligante de penicilina anômala (PBP2´) por aglutinação com látex. Os VCPSa foram detectados em meios de cultura enriquecidos. Estas bactérias deram positivas nos testes de catalase e coagulase positivos e eram dependentes de menadiona e hemina. A resistência à cefoxitina foi detectada nos três isolados e detectou-se a PBP2'.

Draft Genome Sequence of Enterococcus faecalis Strain CECT7121, a Corn Silage Isolate with Antibacterial Activity against Gram-Positive Pathogens.

Enterococcus faecalis CECT7121 is a corn silage probiotic bacterium that shows antibacterial activity against Gram-positive pathogens from different origins. Its genome sequence is 2.9 Mb long with a G+C content of 37.3%. Genome annotation identified three bacteriocin gene clusters in the genome.

Impact on Public Health of the Spread of High-Level Resistance to Gentamicin and Vancomycin in Enterococci.

Antibiotic resistance has turned into a global public health issue. Enterococci are intrinsically resistant to many antimicrobials groups. These bacteria colonize dairy and meat products and integrate the autochthonous microbiota of mammal's gastrointestinal tract. Over the last decades, detection of vanA genotype in Enterococcus faecium from animals and from food of animal origin has been reported. Vancomycin-resistant E. faecium has become a prevalent nosocomial pathogen. Hospitalized patients are frequently treated with broad-spectrum antimicrobials and this leads to an increase in the presence of VanA or VanB vancomycin-resistant enterococci in patients' gastrointestinal tract and the risk of invasive infections. In humans, E. faecium is the main reservoir of VanA and VanB phenotypes. Acquisition of high-level aminoglycoside resistance is a significant therapeutic problem for patients with severe infections because it negates the synergistic effect between aminoglycosides and a cell-wall-active agent. The aac(6')-Ie-aph (2″)-Ia gene is widely spread in E. faecalis and has been detected in strains of human origin and in the food of animal origin. Enzyme AAC(6')-Ie-APH(2″)-Ia confers resistance to available aminoglycosides, except to streptomycin. Due to the fast dissemination of this genetic determinant, the impact of its horizontal transferability among enterococcal species from different origin has been considered. The extensive use of antibiotics in food-producing animals contributes to an increase in drug-resistant animal bacteria that can be transmitted to humans. Innovation is needed for the development of new antibacterial drugs and for the design of combination therapies with conventional antibiotics. Nowadays, semi-purified bacteriocins and probiotics are becoming an attractive alternative to the antibiotic in animal production. Therefore, a better understanding of a complex and relevant issue for Public Health such as high-level vancomycin and gentamicin resistance in enterococci and their impact is needed. Hence, it is necessary to consider the spread of vanA E. faecium and high-level gentamicin resistant E. faecalis strains of different origin in the environment, and also highlight the potential horizontal transferability of these resistance determinants to other bacteria.

Bactericidal Activity and Synergy Studies of Peptide AP-CECT7121 Against Multi-resistant Bacteria Isolated from Human and Animal Soft Tissue Infections.

AP-CECT7121 is an antimicrobial peptide, produced by Enterococcus faecalis CECT7121, with bactericidal activity against Gram-positive bacteria. The aim of this study was to evaluate the bactericidal activity of AP-CECT7121, alone and with gentamicin, against multi-resistant bacteria isolated from human and animals with soft tissue infections. During the period 2014-2015, bacterial strains producing human and animal soft tissue infections were studied. Samples from patients attended at a general hospital and cattle from four dairies in the Province of Buenos Aires (Argentina) were included. Twenty-two methicillin-resistant Staphylococcus aureus (11, human blood samples; 11, cow milk) and five vancomycin-resistant Ent. faecium strains isolated from four mastitic dairy cows were tested. AP-CECT7121 (12 mg/L) potency was assessed by time-kill curves alone or with sub-inhibitory concentrations of gentamicin. All staphylococcal strains were susceptible to gentamicin; enterococci did not show high-level gentamicin resistance. Colony counts were carried out at 0, 2, 4, 8, and 24 h of incubation. AP-CECT7121 showed bactericidal activity against all the enterococcal strains. In addition, AP-CECT7121 had a bactericidal effect on most staphylococci (16/22). Early AP-CECT7121/gentamicin synergy (4-8 h) for all staphylococci was detected. At 24 h, synergy (19/22) and indifference (3/22) were observed. Synergy with gentamicin was detected for staphylococci. AP-CECT7121 constitutes an attractive candidate for its use as a natural therapeutic tool for the treatment of infections produced by multi-resistant Staph. aureus and vancomycin-resistant Ent. faecium isolated from humans and animals.

Throat Carriage Rate and Antimicrobial Resistance of Streptococcus pyogenes In Rural Children in Argentina.

OBJECTIVES: The aim of this study was to determine the prevalence of asymptomatic carriers of group A ß-hemolytic streptococci (GAS) in children living in a rural community and to investigate the association between episodes of acute pharyngitis and carrier status. METHODS: Throat swabs were collected from September to November 2013 among children 5-13 years of age from a rural community (Maria Ignacia-Vela, Argentina). The phenotypic characterization of isolates was performed by conventional tests. Antimicrobial susceptibility was assayed for penicillin, tetracycline, chloramphenicol, erythromycin, and clindamycin (disk diffusion). The minimum inhibitory concentration was determined for penicillin, cefotaxime, tetracycline, and erythromycin. RESULTS: The carriage of ß-hemolytic streptococci was detected in 18.1% of participants, with Streptococcus pyogenes in 18 participants followed by S. dysgalactiae ssp. equisimilis in 5. The highest proportion of GAS was found in 8 to 10-year-old children. No significant association between the number of episodes of acute pharyngitis suffered in the last year and the carrier state was detected (p>0.05). Tetracycline resistance (55.5%) and macrolide-resistant phenotypes (11.1%) were observed. Resistance to penicillin, cefotaxime, or chloramphenicol was not expressed in any streptococcal isolate. CONCLUSIONS: The present study demonstrated significant throat carriage of GAS and the presence of group C streptococci (S. dysgalactiae ssp. equisimilis) in an Argentinian rural population. These results point out the need for continuous surveillance of GAS and non-GAS carriage as well as of antimicrobial resistance in highly susceptible populations, such as school-aged rural children. An extended surveillance program including school-aged children from different cities should be considered to estimate the prevalence of GAS carriage in Argentina.

Effects of Enterococcus faecalis CECT 7121 on Cryptosporidium parvum infection in mice.

Cryptosporidium is an opportunistic protozoan parasite of humans and animals worldwide and causes diarrheal disease that is typically self-limiting in immunocompetent hosts but often life threatening to immunocompromised individuals. However, there is a lack of completely efficient therapy available. Probiotics have attracted the attention as potential antiparasite compounds against protozoa involved in intestinal infections. This study investigated the effects of administration of probiotic Enterococcus faecalis CECT 7121 on Cryptosporidium parvum infection in immunosuppressed mice. Effects on C. parvum infection at the intestinal mucosa were studied and scored at each portion of the gut. It was demonstrated that Ef CECT 7121 interfered with C. parvum infection when both probiotic and parasite were present in the same intestinal location suggesting that Ef CECT 7121 supplementation can alleviate the negative effects of C. parvum infection.

Clinical and microbiological features of bacteremia caused by Enterococcus faecalis.

INTRODUCTION: Enterococcus faecalis is a frequent etiologic agent of invasive infections in hospitalized patients. The aim of this study was to analyze clinical and microbiological features of bacteremia caused by E. faecalis. METHODOLOGY: Between 2011 and 2013, significant bacteremia caused by E. faecalis in hospitalized patients was studied. Patient characteristics, comorbid conditions, and 14-day mortality were recorded. Virulence genes esp, gelE, and cylA; opsonophagocytosis resistance; resistance to bactericidal effect of normal serum; beta lactamase production; and susceptibility to ampicillin, vancomycin, teicoplanin, gentamicin, and streptomycin were investigated. RESULTS: E. faecalis strains were recovered from 33 bacteremic patients. Polymicrobial bacteremia was diagnosed in 2 patients; 10 patients died. Virulence genes were found in strains from both deceased patients and survivors. Sources of bacteremia included urinary tract infections (36.4%), vascular catheters (15.1%), abscesses (9.1%), and unknown (48.5%). Underlying diseases included cancer (30.3%), diabetes (36.4%), cirrhosis (6.1%), renal (36.4%), and chronic obstructive pulmonary disease (2.0%). Co-morbidities included alcohol use (26.1%); glucocorticoid therapy (19.0%); prior antibiotic therapy (60.6%); and central venous (21.2%), arterial (12.1%), and urinary (63.6%) catheters. Also, 57.6% of patients came from the intensive care unit (ICU); 33.3% had mechanical ventilation. Significant mortality-associated conditions included polymicrobial bacteremia, oncological disease, APACHE II score ≤ 20, ICU stay, renal disease, central venous catheter, and mechanical ventilation. CONCLUSIONS: Outcome of patients was associated with their status and not with the presence of virulence genes in E. faecalis strains. A significant percentage of bacteremia had undetermined origin. An alternate origin may be the gastrointestinal tract, through translocation.

Immunomodulatory properties of cell wall extract from Enterococcus faecalis CECT7121.

The aim of this study was to investigate the immunomodulatory properties of cell wall extract from Enterococcus faecalis CECT7121, measuring the induction of cytokines TNF-α, IL-6, IL-10 and IL-12 from human peripheral blood mononuclear cells (PBMCs). Cell wall extract was prepared from their growth in brain heart infusion broth (18h, 35°C). Subsequently, toxicity of the obtained cell wall extract was tested in Balb-C mice. PBMCs were isolated from buffy coats at the Blood Transfusion Service of Hospital Ramón Santamarina (Tandil, Argentina). PBMCs were purified using standard Ficoll-Paque gradient centrifugation. Aliquots of purified leukocytes were incubated at 37°C for 24h with heat-killed E. faecalis CECT7121 and cell wall extract. Concentrations of IL-6, TNFα, IL-10 and IL-12 (p70) were measured by solid phase sandwich ELISA. Changes in appearance and behavior of mice were evidenced only in the group with the maximal concentration of wall cell extract used (10,000µg). Cell wall extract and heat-killed E. faecalis CECT7121 induced the production of significantly higher amounts of Il-12, IL-6, TNF-α and IL-10 cytokines compared to the nonstimulated PBMCs. These findings provide helpful information on immunomodulation activity by cell wall extract in sight of the application of this compound in controlling certain infectious diseases.