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1.
Methods Mol Biol ; 2291: 19-86, 2021.
Artículo en Inglés | MEDLINE | ID: mdl-33704748

RESUMEN

Cattle and other ruminants are primary reservoirs for Shiga toxin-producing Escherichia coli (STEC) strains which have a highly variable, but unpredictable, pathogenic potential for humans. Domestic swine can carry and shed STEC, but only STEC strains producing the Shiga toxin (Stx) 2e variant and causing edema disease in piglets are considered pathogens of veterinary medical interest. In this chapter, we present general diagnostic workflows for sampling livestock animals to assess STEC prevalence, magnitude, and duration of host colonization. This is followed by detailed method protocols for STEC detection and typing at genetic and phenotypic levels to assess the relative virulence exerted by the strains.


Asunto(s)
Enfermedades de los Bovinos , Infecciones por Escherichia coli , Toxina Shiga II/metabolismo , Escherichia coli Shiga-Toxigénica , Enfermedades de los Porcinos , Animales , Bovinos , Enfermedades de los Bovinos/diagnóstico , Enfermedades de los Bovinos/metabolismo , Enfermedades de los Bovinos/microbiología , Infecciones por Escherichia coli/diagnóstico , Infecciones por Escherichia coli/metabolismo , Infecciones por Escherichia coli/microbiología , Infecciones por Escherichia coli/veterinaria , Escherichia coli Shiga-Toxigénica/clasificación , Escherichia coli Shiga-Toxigénica/aislamiento & purificación , Escherichia coli Shiga-Toxigénica/metabolismo , Escherichia coli Shiga-Toxigénica/patogenicidad , Porcinos , Enfermedades de los Porcinos/diagnóstico , Enfermedades de los Porcinos/metabolismo , Enfermedades de los Porcinos/microbiología
2.
Nat Commun ; 12(1): 765, 2021 02 03.
Artículo en Inglés | MEDLINE | ID: mdl-33536414

RESUMEN

Chickens are the most common birds on Earth and colibacillosis is among the most common diseases affecting them. This major threat to animal welfare and safe sustainable food production is difficult to combat because the etiological agent, avian pathogenic Escherichia coli (APEC), emerges from ubiquitous commensal gut bacteria, with no single virulence gene present in all disease-causing isolates. Here, we address the underlying evolutionary mechanisms of extraintestinal spread and systemic infection in poultry. Combining population scale comparative genomics and pangenome-wide association studies, we compare E. coli from commensal carriage and systemic infections. We identify phylogroup-specific and species-wide genetic elements that are enriched in APEC, including pathogenicity-associated variation in 143 genes that have diverse functions, including genes involved in metabolism, lipopolysaccharide synthesis, heat shock response, antimicrobial resistance and toxicity. We find that horizontal gene transfer spreads pathogenicity elements, allowing divergent clones to cause infection. Finally, a Random Forest model prediction of disease status (carriage vs. disease) identifies pathogenic strains in the emergent ST-117 poultry-associated lineage with 73% accuracy, demonstrating the potential for early identification of emergent APEC in healthy flocks.


Asunto(s)
Infecciones por Escherichia coli/prevención & control , Escherichia coli/genética , Evolución Molecular , Genoma Bacteriano/genética , Enfermedades de las Aves de Corral/prevención & control , Animales , Pollos , Escherichia coli/clasificación , Escherichia coli/patogenicidad , Infecciones por Escherichia coli/diagnóstico , Infecciones por Escherichia coli/microbiología , Genes Bacterianos , Variación Genética , Estudio de Asociación del Genoma Completo/métodos , Genotipo , Humanos , Filogenia , Enfermedades de las Aves de Corral/diagnóstico , Enfermedades de las Aves de Corral/microbiología , Virulencia/genética
3.
Science ; 371(6531)2021 02 19.
Artículo en Inglés | MEDLINE | ID: mdl-33602825

RESUMEN

Although metabolism plays an active role in antibiotic lethality, antibiotic resistance is generally associated with drug target modification, enzymatic inactivation, and/or transport rather than metabolic processes. Evolution experiments of Escherichia coli rely on growth-dependent selection, which may provide a limited view of the antibiotic resistance landscape. We sequenced and analyzed E. coli adapted to representative antibiotics at increasingly heightened metabolic states. This revealed various underappreciated noncanonical genes, such as those related to central carbon and energy metabolism, which are implicated in antibiotic resistance. These metabolic alterations lead to lower basal respiration, which prevents antibiotic-mediated induction of tricarboxylic acid cycle activity, thus avoiding metabolic toxicity and minimizing drug lethality. Several of the identified metabolism-specific mutations are overrepresented in the genomes of >3500 clinical E. coli pathogens, indicating clinical relevance.


Asunto(s)
Antibacterianos/farmacología , Farmacorresistencia Bacteriana/genética , Escherichia coli/efectos de los fármacos , Escherichia coli/genética , Genes Bacterianos , Mutación , Adaptación Fisiológica , Carbenicilina/farmacología , Ciprofloxacino/farmacología , Ciclo del Ácido Cítrico/genética , Evolución Molecular Dirigida , Metabolismo Energético/genética , Escherichia coli/crecimiento & desarrollo , Escherichia coli/metabolismo , Infecciones por Escherichia coli/microbiología , Proteínas de Escherichia coli/genética , Técnicas de Silenciamiento del Gen , Genoma Bacteriano , Complejo Cetoglutarato Deshidrogenasa/genética , Pruebas de Sensibilidad Microbiana , Análisis de Secuencia de ADN , Estreptomicina/farmacología
4.
Appl Environ Microbiol ; 87(6)2021 02 26.
Artículo en Inglés | MEDLINE | ID: mdl-33397699

RESUMEN

Little is known about the drivers of critically important antibacterial resistance in species with zoonotic potential present on farms (e.g., CTX-M ß-lactamase-positive Escherichia coli). We collected samples monthly between January 2017 and December 2018 on 53 dairy farms in South West England, along with data for 610 variables concerning antibacterial usage, management practices, and meteorological factors. We detected E. coli resistant to amoxicillin, ciprofloxacin, streptomycin, and tetracycline in 2,754/4,145 (66%), 263/4,145 (6%), 1,475/4,145 (36%), and 2,874/4,145 (69%), respectively, of samples from fecally contaminated on-farm and near-farm sites. E. coli positive for bla CTX-M were detected in 224/4,145 (5.4%) of samples. Multilevel, multivariable logistic regression showed antibacterial dry cow therapeutic choice (including use of cefquinome or framycetin) to be associated with higher odds of bla CTX-M positivity. Low average monthly ambient temperature was associated with lower odds of bla CTX-M E. coli positivity in samples and with lower odds of finding E. coli resistant to each of the four test antibacterials. This was in addition to the effect of temperature on total E. coli density. Furthermore, samples collected close to calves had higher odds of having E. coli resistant to each antibacterial, as well as E. coli positive for bla CTX-M Samples collected on pastureland had lower odds of having E. coli resistant to amoxicillin or tetracycline, as well as lower odds of being positive for bla CTX-M IMPORTANCE Antibacterial resistance poses a significant threat to human and animal health and global food security. Surveillance for resistance on farms is important for many reasons, including tracking impacts of interventions aimed at reducing the prevalence of resistance. In this longitudinal survey of dairy farm antibacterial resistance, we showed that local temperature-as it changes over the course of a year-was associated with the prevalence of antibacterial-resistant E. coli We also showed that prevalence of resistant E. coli was lower on pastureland and higher in environments inhabited by young animals. These findings have profound implications for routine surveillance and for surveys carried out for research. They provide important evidence that sampling at a single time point and/or single location on a farm is unlikely to be adequate to accurately determine the status of the farm regarding the presence of samples containing resistant E. coli.


Asunto(s)
Farmacorresistencia Bacteriana , Escherichia coli/genética , beta-Lactamasas/genética , Envejecimiento , Amoxicilina/farmacología , Animales , Antibacterianos/farmacología , Bovinos , Enfermedades de los Bovinos/microbiología , Ciprofloxacino/farmacología , Escherichia coli/efectos de los fármacos , Escherichia coli/aislamiento & purificación , Infecciones por Escherichia coli/microbiología , Granjas , Heces/microbiología , Estreptomicina/farmacología , Temperatura , Tetraciclina/farmacología
5.
Carbohydr Polym ; 255: 117475, 2021 Mar 01.
Artículo en Inglés | MEDLINE | ID: mdl-33436239

RESUMEN

Extraintestinal pathogenic Escherichia coli (ExPEC) has presented a major clinical infection emerged in the past decades. O-polysaccharide (OPS)-based glycoconjugate vaccines produced using the bacterial glycosylation machinery can be utilized to confer protection against such infection. However, constructing a low-cost microbial cell factory for high-efficient production of OPS-based glycoconjugate vaccines remains challenging. Here, we engineered a glyco-optimized chassis strain by reprogramming metabolic network. The yield was enhanced to 38.6 mg L-1, the highest level reported so far. MS analysis showed that designed glycosylation sequon was modified by target polysaccharide with high glycosylation efficiency of 90.7 % and 76.7 % for CTB-O5 and CTB-O7, respectively. The glycoconjugate vaccines purified from this biosystem elicited a marked increase in protection against ExPEC infection in mouse model, compared to a non-optimized system. The glyco-optimized platform established here is broadly suitable for polysaccharide-based conjugate production against ExPEC and other surface-polysaccharide-producing pathogens.


Asunto(s)
Ingeniería Celular/métodos , Infecciones por Escherichia coli/prevención & control , Vacunas contra Escherichia coli/biosíntesis , Escherichia coli Patógena Extraintestinal/inmunología , Glicoconjugados/biosíntesis , Antígenos O/biosíntesis , Secuencia de Aminoácidos , Animales , Animales no Consanguíneos , Anticuerpos Antibacterianos/biosíntesis , Secuencia de Carbohidratos , Infecciones por Escherichia coli/inmunología , Infecciones por Escherichia coli/microbiología , Infecciones por Escherichia coli/mortalidad , Vacunas contra Escherichia coli/administración & dosificación , Vacunas contra Escherichia coli/genética , Vacunas contra Escherichia coli/inmunología , Escherichia coli Patógena Extraintestinal/patogenicidad , Femenino , Glicoconjugados/administración & dosificación , Glicoconjugados/genética , Glicoconjugados/inmunología , Glicosilación , Inmunización , Inmunogenicidad Vacunal , Inmunoglobulina G/biosíntesis , Redes y Vías Metabólicas/genética , Ratones , Antígenos O/genética , Antígenos O/inmunología , Plásmidos/química , Plásmidos/metabolismo , Análisis de Supervivencia , Vacunas Conjugadas
6.
BMJ Case Rep ; 14(1)2021 Jan 26.
Artículo en Inglés | MEDLINE | ID: mdl-33500303

RESUMEN

Rib osteomyelitis is a rare disease, comprising 1% or less of all osteomyelitis. Treatment of rib osteomyelitis includes prolonged antibiotic therapy and surgical intervention. Indications for surgical treatment of rib osteomyelitis remain unclear, however, because of few reported cases. We report the first known case of extended-spectrum ß-lactamase-producing Escherichia coli rib osteomyelitis caused by urosepsis. The 69-year-old male patient remains free of recurrence and symptoms after rib resection and vacuum-assisted closure treatment with antibiotic therapy. Rib osteomyelitis should be considered as differential diagnosis when patients report chest pain after bacteraemic infection. We recommend surgical treatment for patients with drug-resistant bacterial rib osteomyelitis.


Asunto(s)
Absceso/terapia , Antibacterianos/uso terapéutico , Cefmetazol/uso terapéutico , Desbridamiento/métodos , Infecciones por Escherichia coli/terapia , Osteomielitis/terapia , Costillas/cirugía , Absceso/etiología , Absceso/patología , Anciano , Infecciones por Escherichia coli/microbiología , Humanos , Masculino , Terapia de Presión Negativa para Heridas/métodos , Osteomielitis/etiología , Osteomielitis/patología , Costillas/patología , Sepsis/complicaciones , Pared Torácica , Tomografía Computarizada por Rayos X , Infecciones Urinarias/complicaciones , Resistencia betalactámica/fisiología
7.
ACS Appl Mater Interfaces ; 13(2): 2303-2315, 2021 Jan 20.
Artículo en Inglés | MEDLINE | ID: mdl-33395246

RESUMEN

Numerous studies have found that the surface topography affects the material antibacterial properties by reducing the attachment of bacteria on the surfaces without influencing the viability of the adhered cells. For Cu-bearing alloys with excellent contact-killing properties, bacterial adhesion on the surface is also accompanied by short-range interactions which regulate the toxic effects of the material surface against bacterial cells. Thus, the surface topography of Cu-bearing alloys, as an important factor dominating the exposure level of bacteria on the surfaces, should affect the subsequent contact-killing efficiency. In this work, our major focus was on the regulation mechanism of the surface features on the material-bacterial interactions. We correlated the surface properties including different surface roughnesses of Cu-bearing stainless steel (SS) with the bacterial damage pattern and attempted to clarify the role of surface roughness in mediating the contact-killing behavior of Cu-bearing SS. The results of both atomic force microscopy and scanning electron microscopy investigations showed that E. coli cells experienced the most rapid physical and mechanical damages after incubating with the diamond-polished Cu-bearing SS surface. The bacterial cells noticeably stiffened and the adhesion force significantly increased, as evidenced by force-distance curve measurements. Because of the enhanced hydrophobicity and higher surface potential of the diamond-polished surface, which strengthened the Lewis acid-base attractive forces and weakened the electrostatic barrier between the bacteria and the surface, a higher exposure surface for bacteria was generated. Furthermore, the contact-induced charge transfer, manifested by Cu ion burst release, and reactive oxygen species overexpression contribute to an efficient contact-killing process.


Asunto(s)
Antibacterianos/farmacología , Cobre/farmacología , Escherichia coli/efectos de los fármacos , Acero Inoxidable/farmacología , Aleaciones/química , Aleaciones/farmacología , Antibacterianos/química , Adhesión Bacteriana/efectos de los fármacos , Cobre/química , Escherichia coli/fisiología , Infecciones por Escherichia coli/microbiología , Infecciones por Escherichia coli/prevención & control , Humanos , Acero Inoxidable/química , Propiedades de Superficie
8.
Gene ; 773: 145415, 2021 Mar 20.
Artículo en Inglés | MEDLINE | ID: mdl-33444678

RESUMEN

Heat shock protein 27 (HSP27) plays an important role in protecting cells from various stress factors. This study aimed to investigate the function of HSP27 gene and its regulatory mechanism as infected by Escherichia coli (E. coli) at the tissue and cellular levels. Real-time PCR was used to detect the differential expression of HSP27 gene in F18 resistant and sensitive Sutai pigs and the differential expression upon E. coli F18ab, F18ac, K88ac bacterial supernatant, thallus infection and LPS induction in IPEC-J2. In addition, the HSP27 gene overexpression vector was constructed to detect the effect of the HSP27 gene overexpression on the adhesion of E. coli F18 to IPEC-J2, secretion of pro-inflammatory factors, and the expression of the upstream key genes in Mitogen-activated protein kinase (MAPK) pathway. Ribosomal S6 kinase (RSK2) is an important protein in the MAPK pathway. Therefore, the RSK2 gene overexpression vector was constructed and the number of colonies was counted after co-transfection of HSP27 and RSK2 gene. Results revealed that the expression level of HSP27 gene in resistant individuals in 11 tissues was higher than sensitive type. At the cellular level, the relative expression levels of HSP27 gene were increased after F18ab, F18ac bacterial supernatant, F18ab thallus infection, and LPS induction for 4 h (P < 0.01). The adhesion ability of E. coli F18ab to IPEC-J2 was significantly reduced after HSP27 gene overexpression (P < 0.01), and the concentration of pro-inflammatory factors in the HSP27 gene overexpression group was significantly reduced compared with the control group after F18ab infection (P < 0.05). Furthermore, the expression of RSK2 was significantly increased in HSP27 overexpression group upon F18ab infection (P < 0.01). The colonies quantitative results also showed that the number of colonies was significantly reduced after co-transfection of HSP27 and RSK2 gene. We indicated that the high expression of HSP27 gene may resist the inflammatory response caused by exogenous stress and enhance the ability of IPEC-J2 to resist E. coli F18 infection. RSK2 gene in the MAPK pathway may cooperate with HSP27 gene to participate in the immune response of the organism, which provides a theoretical basis for the study of the mechanism of anti-E. coli infection in piglets.


Asunto(s)
Resistencia a la Enfermedad/genética , Infecciones por Escherichia coli/genética , Escherichia coli/genética , Proteínas de Choque Térmico HSP27/genética , Animales , Adhesión Bacteriana/genética , Diarrea/genética , Diarrea/microbiología , Diarrea/veterinaria , Escherichia coli/patogenicidad , Infecciones por Escherichia coli/microbiología , Infecciones por Escherichia coli/patología , Proteínas de Escherichia coli/genética , Regulación de la Expresión Génica/genética , Porcinos/genética , Porcinos/microbiología , Enfermedades de los Porcinos/genética , Enfermedades de los Porcinos/microbiología
9.
Biochim Biophys Acta Gen Subj ; 1865(1): 129762, 2021 01.
Artículo en Inglés | MEDLINE | ID: mdl-33053413

RESUMEN

BACKGROUND: Previous studies have demonstrated the formation of stable complexes between inorganic pyrophosphatase (PPase) and three other Escherichia coli enzymes - cupin-type phosphoglucose isomerase (cPGI), class I fructose-1,6-bisphosphate aldolase (FbaB) and l-glutamate decarboxylase (GadA). METHODS: Here, we determined by activity measurements how complex formation between these enzymes affects their activities and oligomeric structure. RESULTS: cPGI activity was modulated by all partner proteins, but none was reciprocally affected by cPGI. PPase activity was down-regulated upon complex formation, whereas all other enzymes were up-regulated. For cPGI, the activation was partially counteracted by a shift in dimer ⇆ hexamer equilibrium to inactive hexamer. Complex stoichiometry appeared to be 1:1 in most cases, but FbaB formed both 1:1 and 1:2 complexes with both GadA and PPase, FbaB activation was only observed in the 1:2 complexes. FbaB and GadA induced functional asymmetry (negative kinetic cooperativity) in hexameric PPase, presumably by favoring partial dissociation to trimers. CONCLUSIONS: These four enzymes form all six possible binary complexes in vitro, resulting in modulated activity of at least one of the constituent enzymes. In five complexes, the effects on activity were unidirectional, and in one complex (FbaB⋅PPase), the effects were reciprocal. The effects of potential physiological significance include inhibition of PPase by FbaB and GadA and activation of FbaB and cPGI by PPase. Together, they provide a mechanism for feedback regulation of FbaB and GadA biosynthesis. GENERAL SIGNIFICANCE: These findings indicate the complexity of functionally significant interactions between cellular enzymes, which classical enzymology treats as individual entities, and demonstrate their moonlighting activities as regulators.


Asunto(s)
Proteínas de Escherichia coli/metabolismo , Escherichia coli/metabolismo , Fructosa-Bifosfato Aldolasa/metabolismo , Glucosa-6-Fosfato Isomerasa/metabolismo , Glutamato Descarboxilasa/metabolismo , Pirofosfatasa Inorgánica/metabolismo , Proteínas de la Membrana/metabolismo , Escherichia coli/química , Infecciones por Escherichia coli/microbiología , Proteínas de Escherichia coli/química , Fructosa-Bifosfato Aldolasa/química , Glucosa-6-Fosfato Isomerasa/química , Glutamato Descarboxilasa/química , Humanos , Pirofosfatasa Inorgánica/química , Cinética , Proteínas de la Membrana/química , Multimerización de Proteína
10.
Nat Rev Microbiol ; 19(1): 37-54, 2021 01.
Artículo en Inglés | MEDLINE | ID: mdl-32826992

RESUMEN

Escherichia coli is a commensal of the vertebrate gut that is increasingly involved in various intestinal and extra-intestinal infections as an opportunistic pathogen. Numerous pathotypes that represent groups of strains with specific pathogenic characteristics have been described based on heterogeneous and complex criteria. The democratization of whole-genome sequencing has led to an accumulation of genomic data that render possible a population phylogenomic approach to the emergence of virulence. Few lineages are responsible for the pathologies compared with the diversity of commensal strains. These lineages emerged multiple times during E. coli evolution, mainly by acquiring virulence genes located on mobile elements, but in a specific chromosomal phylogenetic background. This repeated emergence of stable and cosmopolitan lineages argues for an optimization of strain fitness through epistatic interactions between the virulence determinants and the remaining genome.


Asunto(s)
Infecciones por Escherichia coli/microbiología , Escherichia coli/genética , Genoma Bacteriano , Genómica , Escherichia coli/clasificación , Evolución Molecular , Genómica/métodos , Humanos , Fenotipo , Filogenia , Virulencia , Factores de Virulencia
11.
Int J Food Microbiol ; 339: 109029, 2021 Feb 02.
Artículo en Inglés | MEDLINE | ID: mdl-33360585

RESUMEN

Shiga toxin-producing Escherichia coli (STEC) O145 is a major serotype associated with severe human disease. Production of Shiga toxins (Stxs), especially Stx2a, is thought to be correlated with STEC virulence. Since stx genes are located in prophages genomes, induction of prophages is required for effective Stxs production. Here, we investigated the production of Stxs in 12 environmental STEC O145:H28 strains under stresses STEC encounter in natural habitats and performed comparative analysis with two O145:H28 clinical strains, one linked to a 2010 U.S. lettuce-associated outbreak (RM13514) and the other linked to a 2007 Belgium ice cream-associated outbreak (RM13516). Similar to the outbreak strains, all environmental strains belong to Sequence Type (ST)-78 using the EcMLST typing scheme. Although all Stx1a-prophages were grouped together, variations in Stx1a production were observed prior to or following the inductions. Among all stx2a positive environmental strains, only the Stx2a-prophage in cattle isolate RM9154-C1 was clustered with the Stx2a-prophages in RM13514, the Stx2a-phage induced from a STEC O104:H4 strain linked to the 2011 outbreak of enterohemorrhagic infection in Germany, and the Stx2a-prophage in STEC O157:H7 strain EDL933, a prototype of enterohemorrhagic E. coli. Furthermore, the Stx2a-prophage in RM9154-C1 shared the same chromosomal insertion site and carried the same antiterminator Q gene and the late promoter PR' as the Stx2a-prophage in RM13514. Following mitomycin C or enrofloxacin treatment, the production of Stx2a in RM9154-C1 was the highest among all environmental strains tested. In contrast, following acid challenge and recovery, the production of Stx2a in RM9154-C1 was the lowest among all the environmental strains tested, at a level comparable to the clinical strains. A significant increase in Stx2a production was detected in all strains when exposed to H2O2, although the induction fold was much lower than those by other inducers. This low-efficiency induction of Stx-prophages by H2O2, a natural inducer of Stx-prophages, supports the hypothesis of bacterial altruism in controlling Stxs production, a strategy that assures the survival of the STEC population as a whole by sacrificing a small fraction of cells for Stxs production and release. Differential induction of Stxs among strains carrying nearly identical Stx-prophages suggests a role of host bacteria in regulating Stxs production. Our study revealed diverse Stx-prophages in STEC O145:H28 strains that were genotypically indistinguishable. Identification of a cattle isolate harboring a Stx2a-prophage associated with high virulence supports the premise that cattle, a natural reservoir of STEC, serve as a source of hypervirulent STEC strains.


Asunto(s)
Toxina Shiga/metabolismo , Escherichia coli Shiga-Toxigénica/genética , Animales , Bacteriófagos/genética , Bélgica , Bovinos , Brotes de Enfermedades , Escherichia coli Enterohemorrágica , Microbiología Ambiental , Infecciones por Escherichia coli/microbiología , Genoma , Genotipo , Alemania , Humanos , Peróxido de Hidrógeno , Profagos/genética , Serogrupo , Toxina Shiga/genética , Toxina Shiga II/genética , Virulencia
12.
PLoS One ; 15(12): e0235583, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-33320853

RESUMEN

BACKGROUND: Escherichia coli O157 is an emerging foodborne pathogen of great public health concern. It has been associated with bloody diarrhoea, haemorrhagic colitis and haemolytic uremic syndrome in humans. Most human infections have been traced to cattle and the consumption of contaminated cattle products. In order to understand the risk associated with the consumption of cattle products, this study sought to investigate the prevalence and identify virulence genes in E. coli O157 from cattle in Cameroon. METHOD: A total of 512 rectal samples were obtained and analysed using conventional bacteriological methods (enrichment on modified Tryptone Soy Broth and selective plating on Cefixime-Tellurite Sorbitol Mac-Conkey Agar) for the isolation of E. coli O157. Presumptive E. coli O157 isolates were confirmed serologically using E. COLIPROTM O157 latex agglutination test and molecularly using PCR targeting the rfb gene in the isolates. Characterisation of the confirmed E. coli O157 strains was done by amplification of stx1, stx2, eaeA and hlyA virulence genes using both singleplex and multiplex PCR. RESULTS: E. coli O157 was detected in 56 (10.9%) of the 512 samples examined. The presence of the virulence genes stx2, eaeA and hylA was demonstrated in 96.4% (54/56) of the isolates and stx1 in 40 (71.4%) of the 54. The isolates exhibited three genetic profiles (I-III) with I (stx1, stx2, eaeA and hlyA) being the most prevalent (40/56; 71.4%) while two isolates had none of the virulence genes tested. CONCLUSION: A proportion of cattle slaughtered in abattoirs in Buea are infected with pathogenic E. coli O157 and could be a potential source of human infections. We recommend proper animal food processing measures and proper hygiene be prescribed and implemented to reduce the risk of beef contamination.


Asunto(s)
Infecciones por Escherichia coli/microbiología , Escherichia coli O157/genética , Factores de Virulencia/genética , Virulencia/genética , Animales , Camerún , Bovinos , Proteínas de Escherichia coli/genética , Microbiología de Alimentos/métodos , Perfil Genético , Carne/microbiología , Reacción en Cadena de la Polimerasa/métodos , Prevalencia
13.
J Vis Exp ; (166)2020 12 04.
Artículo en Inglés | MEDLINE | ID: mdl-33346201

RESUMEN

Recurrent urinary tract infections (rUTI) caused by uropathogenic Escherichia coli (UPEC) are common and costly. Previous articles describing models of UTI in male and female mice have illustrated the procedures for bacterial inoculation and enumeration in urine and tissues. During an initial bladder infection in C57BL/6 mice, UPEC establish latent reservoirs inside bladder epithelial cells that persist following clearance of UPEC bacteriuria. This model builds on these studies to examine rUTI caused by the emergence of UPEC from within latent bladder reservoirs. The urogenital bacterium Gardnerella vaginalis is used as the trigger of rUTI in this model because it is frequently present in the urogenital tracts of women, especially in the context of vaginal dysbiosis that has been associated with UTI. In addition, a method for in situ bladder fixation followed by scanning electron microscopy (SEM) analysis of bladder tissue is also described, with potential application to other studies involving the bladder.


Asunto(s)
Infecciones por Escherichia coli/microbiología , Gardnerella vaginalis/fisiología , Vejiga Urinaria/microbiología , Vejiga Urinaria/patología , Infecciones Urinarias/microbiología , Animales , Modelos Animales de Enfermedad , Reservorios de Enfermedades/microbiología , Infecciones por Escherichia coli/patología , Infecciones por Escherichia coli/orina , Femenino , Ratones Endogámicos C57BL , Recurrencia , Espectrofotometría , Vejiga Urinaria/ultraestructura , Infecciones Urinarias/patología , Infecciones Urinarias/orina , Orina/citología , Escherichia coli Uropatógena/fisiología
14.
PLoS One ; 15(12): e0244713, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-33382795

RESUMEN

The prevalence of Shiga toxin (Stx)-producing Escherichia coli (STEC) was determined by evaluating its presence in faecal samples from 155 heifers, and 254 dairy cows in 21 farms at North of Portugal sampled between December 2017 and June 2019. The prevalence of STEC in heifers (45%) was significantly higher than in lactating cows (16%) (p<0.05, Fisher exact test statistic value is <0.00001). A total of 133 STEC were isolated, 24 (13.8%) carried Shiga-toxin 1 (stx1) genes, 69 (39.7%) carried Shiga-toxin 2 (stx2) genes, and 40 (23%) carried both stx1 and stx2. Intimin (eae) virulence gene was detected in 29 (21.8%) of the isolates. STEC isolates belonged to 72 different O:H serotypes, comprising 40 O serogroups and 23 H types. The most frequent serotypes were O29:H12 (15%) and O113:H21 (5.2%), found in a large number of farms. Two isolates belonged to the highly virulent serotypes associated with human disease O157:H7 and O26:H11. Many other bovine STEC serotypes founded in this work belonged to serotypes previously described as pathogenic to humans. Thus, this study highlights the need for control strategies that can reduce STEC prevalence at the farm level and, thus, prevent food and environmental contamination.


Asunto(s)
Adhesinas Bacterianas/genética , Infecciones por Escherichia coli/veterinaria , Proteínas de Escherichia coli/genética , Heces/microbiología , Escherichia coli Shiga-Toxigénica/aislamiento & purificación , Animales , Bovinos , Infecciones por Escherichia coli/microbiología , Femenino , Portugal , Serogrupo , Serotipificación , Escherichia coli Shiga-Toxigénica/genética , Virulencia
15.
Int J Mol Sci ; 21(24)2020 Dec 15.
Artículo en Inglés | MEDLINE | ID: mdl-33334000

RESUMEN

Enteroaggregative Escherichia coli (EAEC) is an emerging pathogen frequently associated with acute diarrhea in children and travelers to endemic regions. EAEC was found the most prevalent bacterial diarrheal pathogen from hospitalized Bolivian children less than five years of age with acute diarrhea from 2007 to 2010. Here, we further characterized the epidemiology of EAEC infection, virulence genes, and antimicrobial susceptibility of EAEC isolated from 414 diarrheal and 74 non-diarrheal cases. EAEC isolates were collected and subjected to a PCR-based virulence gene screening of seven virulence genes and a phenotypic resistance test to nine different antimicrobials. Our results showed that atypical EAEC (a-EAEC, AggR-negative) was significantly associated with diarrhea (OR, 1.62, 95% CI, 1.25 to 2.09, p < 0.001) in contrast to typical EAEC (t-EAEC, AggR-positive). EAEC infection was most prevalent among children between 7-12 months of age. The number of cases exhibited a biannual cycle with a major peak during the transition from warm to cold (April-June). Both typical and a-EAEC infections were graded as equally severe; however, t-EAEC harbored more virulence genes. aap, irp2 and pic were the most prevalent genes. Surprisingly, we detected 60% and 52.6% of multidrug resistance (MDR) EAEC among diarrheal and non-diarrheal cases. Resistance to ampicillin, sulfonamides, and tetracyclines was most common, being the corresponding antibiotics, the ones that are frequently used in Bolivia. Our work is the first study that provides comprehensive information on the high heterogenicity of virulence genes in t-EAEC and a- EAEC and the large prevalence of MDR EAEC in Bolivia.


Asunto(s)
Infección Hospitalaria/epidemiología , Farmacorresistencia Bacteriana Múltiple , Infecciones por Escherichia coli/epidemiología , Infecciones por Escherichia coli/microbiología , Escherichia coli/efectos de los fármacos , Escherichia coli/genética , Antibacterianos/farmacología , Bolivia/epidemiología , Preescolar , Diarrea/epidemiología , Diarrea/microbiología , Femenino , Genes Bacterianos , Humanos , Lactante , Pruebas de Sensibilidad Microbiana , Epidemiología Molecular , Filogenia , Prevalencia , Vigilancia en Salud Pública , Índice de Severidad de la Enfermedad , Virulencia/genética , Factores de Virulencia/genética
16.
Epidemiol Infect ; 149: e12, 2020 12 17.
Artículo en Inglés | MEDLINE | ID: mdl-33327984

RESUMEN

The prevalence of extended-spectrum beta-lactamase (ESBL)-producing Escherichia coli and Klebsiella pneumoniae urinary tract infections (UTIs) is increasing worldwide. We investigated the prevalence, clinical findings, impact and risk factors of ESBL E. coli/K. pneumoniae UTI through a retrospective review of the medical records of children with UTI aged <15 years admitted to Prince of Songkla University Hospital, Thailand over 10 years (2004-2013). Thirty-seven boys and 46 girls had ESBL-positive isolates in 102 UTI episodes, compared with 85 boys and 103 girls with non-ESBL isolates in 222 UTI episodes. The age of presentation and gender were not significantly different between the two groups. The prevalence of ESBL rose between 2004 and 2008 before plateauing at around 30-40% per year, with a significant difference between first and recurrent UTI episodes of 27.3% and 46.5%, respectively (P = 0.003). Fever prior to UTI diagnosis was found in 78.4% of episodes in the non-ESBL group and 61.8% of episodes in the ESBL group (P = 0.003). Multivariate analysis indicated that children without fever (odds ratio (OR) 2.14, 95% confidence interval (CI) 1.23-3.74) and those with recurrent UTI (OR 2.67, 95% CI 1.37-5.19) were more likely to yield ESBL on culture. Congenital anomalies of the kidney and urinary tract were not linked to the presence of ESBL UTI. In conclusion, ESBL producers represented one-third of E. coli/K. pneumoniae UTI episodes but neither clinical condition nor imaging studies were predictive of ESBL infections. Recurrent UTI was the sole independent risk factor identified.


Asunto(s)
Infecciones por Escherichia coli/microbiología , Escherichia coli/efectos de los fármacos , Infecciones por Klebsiella/microbiología , Klebsiella pneumoniae/efectos de los fármacos , Infecciones Urinarias/microbiología , beta-Lactamasas/metabolismo , Adolescente , Antibacterianos/farmacología , Niño , Preescolar , Farmacorresistencia Bacteriana Múltiple , Escherichia coli/enzimología , Femenino , Humanos , Lactante , Klebsiella pneumoniae/enzimología , Masculino , Estudios Retrospectivos , beta-Lactamasas/genética
17.
PLoS One ; 15(12): e0244358, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-33362261

RESUMEN

Escherichia coli are one of the commonest bacteria causing bloodstream infection (BSI). The aim of the research was to identify the serotypes, MLST (Multi Locus Sequence Type), virulence genes, and antimicrobial resistance of E. coli isolated from bloodstream infection hospitalized patients in Cipto Mangunkusumo National Hospital Jakarta. We used whole genome sequencing methods rather than the conventional one, to characterized the serotypes, MLST (Multi Locus Sequence Type), virulence genes, and antimicrobial resistance (AMR) of E. coli. The composition of E. coli sequence types (ST) was as follows: ST131 (n = 5), ST38 (n = 3), ST405 (n = 3), ST69 (n = 3), and other STs (ST1057, ST127, ST167, ST3033, ST349, ST40, ST58, ST6630). Enteroaggregative E. coli (EAEC) and Extra-intestinal pathogenic E. coli (ExPEC) groups were found dominant in our samples. Twenty isolates carried virulence genes for host cells adherence and 15 for genes that encourage E. coli immune evasion by enhancing survival in serum. ESBL-genes were present in 17 E. coli isolates. Other AMR genes also encoded resistance against aminoglycosides, quinolones, chloramphenicol, macrolides and trimethoprim. The phylogeny analysis showed that phylogroup D is dominated and followed by phylogroup B2. The E. coli isolated from 22 patients in Cipto Mangunkusumo National Hospital Jakarta showed high diversity in serotypes, sequence types, virulence genes, and AMR genes. Based on this finding, routinely screening all bacterial isolates in health care facilities can improve clinical significance. By using Whole Genome Sequencing for laboratory-based surveillance can be a valuable early warning system for emerging pathogens and resistance mechanisms.


Asunto(s)
Bacteriemia/microbiología , Infecciones por Escherichia coli/microbiología , Escherichia coli/clasificación , Secuenciación de Nucleótidos de Alto Rendimiento/métodos , Farmacorresistencia Bacteriana Múltiple , Escherichia coli/genética , Escherichia coli/patogenicidad , Escherichia coli Patógena Extraintestinal/aislamiento & purificación , Genoma Bacteriano , Humanos , Evasión Inmune , Tipificación de Secuencias Multilocus , Filogenia , Factores de Virulencia/genética , Secuenciación Completa del Genoma
18.
PLoS One ; 15(12): e0243630, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-33332370

RESUMEN

Enterobacterales resistant to carbapenems, a class of last-resort antimicrobials, are ranked as an "urgent" and "critical" public health hazard by CDC and WHO. IMP-type carbapenemase-containing Enterobacterales are endemic in Japan, and blaIMP-6 is one of the notable carbapenemase genes responsible for the resistance. The gene is plasmid-encoded and confers resistance to meropenem, but not to imipenem. Therefore, IMP-6-producing Enterobacterales isolates are occasionally overlooked in clinical laboratories and are referred to as 'stealth-type'. Since previous reports in Japan were confined only to some geographical regions, their distribution across prefectures and the factors affecting the distribution remain unclear. Here, we revealed the dynamics of the geographical distribution of Enterobacterales with IMP-6 phenotype associated with antimicrobial use in Japan. We utilized comprehensive national surveillance data of all routine bacteriological test results from more than 1,400 hospitals in 2015 and 2016 to enumerate Escherichia coli and Klebsiella pneumoniae isolates with the antimicrobial susceptibility pattern (phenotype) characteristic of IMP-6 (imipenem susceptible, meropenem resistant), and to tabulate the frequency of isolates with the phenotype for each prefecture. Isolates were detected in approximately half of all prefectures, and combined analysis with the national data of antimicrobial usage revealed a statistically significant association between the frequency and usage of not carbapenems but third-generation cephalosporins (p = 0.006, logistic mixed-effect regression) and a weaker association between the frequency and usage of fluoroquinolones (p = 0.043). The usage of third-generation cephalosporins and fluoroquinolones may select the strains with the IMP-6 phenotype, and contribute to their occasional spread. We expect the findings will promote antimicrobial stewardship to reduce the spread of the notable carbapenemase gene.


Asunto(s)
Antibacterianos/farmacología , Proteínas Bacterianas/genética , Carbapenémicos/farmacología , Farmacorresistencia Bacteriana , Escherichia coli/genética , Klebsiella pneumoniae/genética , beta-Lactamasas/genética , Antibacterianos/uso terapéutico , Carbapenémicos/uso terapéutico , Enterobacteriaceae/efectos de los fármacos , Enterobacteriaceae/genética , Infecciones por Enterobacteriaceae/tratamiento farmacológico , Infecciones por Enterobacteriaceae/epidemiología , Infecciones por Enterobacteriaceae/microbiología , Escherichia coli/efectos de los fármacos , Infecciones por Escherichia coli/tratamiento farmacológico , Infecciones por Escherichia coli/epidemiología , Infecciones por Escherichia coli/microbiología , Humanos , Imipenem/farmacología , Imipenem/uso terapéutico , Japón/epidemiología , Infecciones por Klebsiella/tratamiento farmacológico , Infecciones por Klebsiella/epidemiología , Infecciones por Klebsiella/microbiología , Klebsiella pneumoniae/efectos de los fármacos , Meropenem/farmacología , Meropenem/uso terapéutico , Fenotipo
19.
BMC Infect Dis ; 20(1): 813, 2020 Nov 09.
Artículo en Inglés | MEDLINE | ID: mdl-33167875

RESUMEN

BACKGROUND: Uncomplicated urinary tract infections (UTIs) in women are usually managed in primary care with antibiotics. However, many women seem to prefer to handle UTI symptoms with nonsteroidal anti-inflammatory drugs (NSAIDs) and other remedies. The aim of this study was to compare UTI management as recommended by physicians with the patients' management at home. METHODS: This prospective cohort study in German primary care is based on clinical data from local practices and patient questionnaires. Participating women completed a baseline data sheet in the practice; their urine sample was tested by a dipstick in the practice and cultured by a laboratory. The women reported treatment and symptom-related impairment on an eight-item symptom questionnaire daily for 7 days. Using growth curve models, we analysed the influence of time on the total severity score to examine how symptoms changed across days. We then examined whether symptom severity and symptom course differed between patients who took antibiotics or NSAIDs. RESULTS: A total of 120 women (mean age of 43.3 ± 16.6 years) were enrolled. The urine dipstick was positive for leucocytes in 92%, erythrocytes in 87%, and nitrites in 23%. Physicians prescribed antibiotics for 102 (87%) women and recommended NSAIDs in 14 cases. According to the women's reports, only 60% (72/120) took antibiotics, while the remainder took NSAIDs and other remedies. Symptoms declined from day 0 to day 6, irrespective of whether women decided to take an antibiotic, NSAIDs, none or both, as confirmed by a significant curvilinear time effect (B = 0.06, SE = 0.005, p < .001). The symptom course, however, was moderated by taking antibiotics so that the change in symptom severity was somewhat more pronounced in women taking antibiotics (B = 0.06) than in the remainder (B = 0.04). CONCLUSION: A substantial proportion of women did not follow their physicians' treatment recommendations, and many used NSAIDs. All women had a good chance of recovery irrespective of whether they decided to take antibiotics. A sensitive listening to patient preferences in the consultation may encourage physicians to recommend and prescribe symptomatic treatment with NSAID more often than antibiotic medicines.


Asunto(s)
Antibacterianos/uso terapéutico , Antiinflamatorios no Esteroideos/uso terapéutico , Infecciones por Escherichia coli/tratamiento farmacológico , Escherichia coli/efectos de los fármacos , Atención Primaria de Salud , Derivación y Consulta , Infecciones Urinarias/tratamiento farmacológico , Adulto , Farmacorresistencia Bacteriana/efectos de los fármacos , Infecciones por Escherichia coli/microbiología , Femenino , Humanos , Persona de Mediana Edad , Cooperación del Paciente , Médicos/psicología , Estudios Prospectivos , Encuestas y Cuestionarios , Infecciones Urinarias/microbiología
20.
Nat Commun ; 11(1): 4915, 2020 10 01.
Artículo en Inglés | MEDLINE | ID: mdl-33004811

RESUMEN

A phenotype of Escherichia coli and Klebsiella pneumoniae, resistant to piperacillin/tazobactam (TZP) but susceptible to carbapenems and 3rd generation cephalosporins, has emerged. The resistance mechanism associated with this phenotype has been identified as hyperproduction of the ß-lactamase TEM. However, the mechanism of hyperproduction due to gene amplification is not well understood. Here, we report a mechanism of gene amplification due to a translocatable unit (TU) excising from an IS26-flanked pseudo-compound transposon, PTn6762, which harbours blaTEM-1B. The TU re-inserts into the chromosome adjacent to IS26 and forms a tandem array of TUs, which increases the copy number of blaTEM-1B, leading to TEM-1B hyperproduction and TZP resistance. Despite a significant increase in blaTEM-1B copy number, the TZP-resistant isolate does not incur a fitness cost compared to the TZP-susceptible ancestor. This mechanism of amplification of blaTEM-1B is an important consideration when using genomic data to predict susceptibility to TZP.


Asunto(s)
Antibacterianos/farmacología , Farmacorresistencia Bacteriana Múltiple/genética , Infecciones por Escherichia coli/tratamiento farmacológico , Proteínas de Escherichia coli/genética , Escherichia coli/genética , beta-Lactamasas/genética , Antibacterianos/uso terapéutico , Cromosomas Bacterianos/genética , Elementos Transponibles de ADN/genética , ADN Bacteriano/genética , Quimioterapia Combinada/métodos , Escherichia coli/efectos de los fármacos , Escherichia coli/aislamiento & purificación , Infecciones por Escherichia coli/microbiología , Amplificación de Genes , Regulación Bacteriana de la Expresión Génica , Genoma Bacteriano/genética , Humanos , Pruebas de Sensibilidad Microbiana , Piperacilina/farmacología , Piperacilina/uso terapéutico , Polimorfismo de Longitud del Fragmento de Restricción , ARN Ribosómico 16S/genética , Tazobactam/farmacología , Tazobactam/uso terapéutico , Secuenciación Completa del Genoma
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