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Resumen Las cepas de Escherichia coli productoras de toxina Shiga (STEC) son reconocidas como responsables de un alto número de casos de enfermedades de transmisión alimentaria a nivel mundial. Su patogenicidad ha sido vinculada directamente con la actividad de las toxinas (Stx); sin embargo, la habilidad de estas bacterias para colonizar al huésped y otras superficies puede ser esencial para desarrollar su poder patogénico. La gran plasticidad genómica de cepas STEC se infiere de la variabilidad de perfiles de virulencia, con la frecuente emergencia de cepas con nuevos genes, codificados en nuevas islas de patogenicidad vinculadas al metabolismo y la adherencia. La formación de biofilm es un mecanismo espontáneo por el cual las cepas STEC resisten en un ambiente hostil, lo que les permite sobrevivir y, de esa forma, llegar al huésped, a través de los alimentos o de las superficies que están en contacto con ellos. Este mecanismo presenta una alta variabilidad intra e interserotipo y su desarrollo no depende solo de los microorganismos que lo conforman. Factores inherentes al ambiente (pH, temperatura) y la superficie (acero inoxidable, poliestireno) a la que pueden adherirse influyen en la expresión de biofilm. El concepto «una salud¼ implica la interrelación entre los actores de salud pública, animal y ambiental para lograr alimentos inocuos y evitar contaminación cruzada y resistencia a sanitizantes, lo cual pone de manifiesto la necesidad de identificar patógenos emergentes a través de nuevos marcadores moleculares, que detecten cepas STEC portadoras del denominado locus for enterocyte effacement (LEE) o del locus de adherencia y autoagregación (LAA).
Abstract Shiga Toxin-producing Escherichia coli (STEC) is recognized as being responsible for a large number of foodborne illnesses around the world. The pathogenicity of STEC has been related to Stx toxins. However, the ability of STEC to colonize the host and other surfaces can be essential for developing its pathogenicity. Different virulence profiles detected in STEC could cause the emergence of strains carrying new genes codified in new pathogenicity islands linked to metabolism and adherence. Biofilm formation is a spontaneous mechanism whereby STEC strains resist in a hostile environment being able to survive and consequently infect the host through contaminated food and food contact surfaces. Biofilm formation shows intra-and inter-serotype variability, and its formation does not depend only on the microorganisms involved. Other factors related to the environment (such as pH, temperature) and the surface (stainless steel and polystyrene) influence biofilm expression. The «One Health¼ concept implies the interrelation between public, animal, and environmental health actors to ensure food safety, prevent cross-contamination and resistance to sanitizers, highlighting the need to identify emerging pathogens through new molecular markers of rapid detection that involve STEC strains carrying the Locus of Enterocyte Effacement or Locus of Adhesion and Autoaggregation.
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Objective To investigate the antimicrobial resistance ,molecular phenotypes ,virulence gene profiles of Salmonella A gona (S .A gona) isolated from patients with acute diarrhea ,and to better understand its epidemic trend ,prevention and treatment .Methods Clinical data and stool samples of patients with acute diarrhea during April to October in 2013 and 2014 from the Second Hospital of Tianjin Medical University were collected .Enrichment culture ,biochemical identification and serotyping analysis were used to isolate and identify S .A gona strains .The isolated strains were further analyzed with antibiotics susceptibility test ,pulsed field gel electrophoresis (PFGE) ,multiple locus sequence typing (MLST ) , Quinolone resistance determining region (QRDR) .Plasmid-mediated quinolone resistance (PMQR) and β-lactamases genes (TEM ,SHV ,OXA ,and CTX-M) were amplified by polymerase chain reaction (PCR) and sequencing .The representative genes carried by Salmonella pathogenicity islands (SPI) 1 — 6 ,9 — 12 and virulence plasmids were amplified by PCR .And the clinical characteristics of S .Agona infection were analyzed .Results Among 119 non-repetitive (non-typhoidal salmonella ,NTS) isolates during the two years ,eight isolates (6 .7％ ) of S .A gona were identified . The resistance rate of S .A gona strains to streptomycin was 100 .0％ , those to ampicillin and gentamicin were 62 .5％ ,to levofloxacin ,ciprofloxacin and nalicixic acid were 25 .0％ ,to chloramphenicol ,amoxillin/clavulanic acid and piperacillin tazobactam were 12 .5％ .The strains were susceptible to other drugs .All 8 isolates had the identical ST13 genotype .PFGE showed 5 clones ,and 4 out of 5 isolates had the exact same patterns of PFGE and drug susceptibility .Two (fluoroquinolones ,FQ) resistant strains carried gyrA mutation leading to amino acid substitutions at position 87 in GyrA ,and no PMQR genes was detected ,while one of which was sensitive to ciprofloxacin by K-B method .All five ampicillin-resistant isolates were positive for TEM-1b gene and one isolate of them was resistant to β-lactam/β-lactamase inhibitor complex .The representative genes carried by SPI 1 — 6 , 9 ,11 ,12 (hilA ,sseL ,mgtC ,siiE ,sopB ,pagN ,bapA ,pagC and sspH2) were 100 .0％ positive ,while the genes carried by SPI10 (sef A ) virulence plasmids (spvB , prot6E) were negative . Two patients with FQ resistant strains infection were clinically diagnosed with bacillary dysentery ,and the remaining six cases with FQ susceptible strains infection were clinically diagnosed with acute gastroenteritis .Conclusions FQs-resistant and multi-drug resistant S .A gonaisolates have emerged in clinical settings .These isolates carry a variety of virulence genes .Resistance to FQ of S .Agonamay cause more severe illness .ST13 might be the dominant genotype of S . A gona in China ,and we should try to prevent the infection outbreak of S .A gona .
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Phenomenon of pathogenicity is the polyfunctional biological potency of germs that are realized by factors (determinants) of pathogenicity [1]. Bacterial pathogenicity is an ability of bacteria to induce and develop infectious diseases in multi-cellular organisms (human, animals and plants). Virulence is a degree of pathogenicity measured by the in vivo (LD50) and in vitro (ID50) tests (highly virulent, weakly virulent and non-virulent strains). Pathogenic factors (determinants) are the bio-molecules produced by pathogen and are responsible for interaction with the host tissue cells. “Pathogenicity Islands” are the bacterial genome mobile elements that carry genes encoding the pathogenicity factors production.
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Para la comprensión de las bases genéticas de los mecanismos de patogenicidad de Salmonella se han descrito diversas metodologías para manipular el ADN genómico y generar mutantes con características particulares. En este estudio se reporta la construcción de mutantes a partir de varios serotipos de S. enterica, por sustitución e inactivación de los genes invG/invE en SPI-1 y de los genes ssaJ/ssaK en SPI-2 mediante la técnica de recombinasa Red del fago λ descrita por Datsenko y Wanner (2000). Los genes delecionados en las SPI-1 y SPI-2 codifican para las proteínas que participan en la formación de los sistemas de secreción tipo III, responsables de la invasión y supervivencia intracelular de S. enterica en las células hospedadoras. Los resultados de este trabajo permitirán realizar estudios futuros in vivo para evaluar la posible atenuación de la virulencia de las cepas mutantes, así como aportar nuevos conocimientos sobre los mecanismos genéticos involucrados en la fisiopatogenia de las enfermedades producidas por los serovares estudiados. Además, esta técnica se recomienda para generar de manera eficiente mutantes de diferentes serotipos de S. enterica con la finalidad de estudiar los genes cromosómicos y sus productos.
To understand the genetic basis of Salmonella pathogenicity mechanisms, various methods have been described to manipulate and generate mutant genomic DNA with specific characteristics. In this study we report the construction of mutants from several serotypes of S. enterica, substitution and inactivation of genes invG/invE in SPI-1 gene and ssaJ/ssaK in SPI-2 by the technique of phage λ Red recombinase, as described by Datsenko and Wanner (2000). The gene deletion in SPI-1 and SPI-2 encodes proteins involved in the formation of type III secretion systems responsible for the invasion and intracellular survival of S. enterica in the host cells. The results of this work will allow in vivo studies to evaluate the possible attenuation of virulence of the mutant strains, as well as to provide new insights into the genetic mechanisms involved in the pathogenesis of diseases caused by these bacteria. Moreover, this technique is recommended to efficiently generate mutants of different serotypes of S. enterica in order to study the chromosomal genes and their products.