RESUMEN
Powdered infant formula (PIF) is rich in nutrients that support the survival and growth of bacteria that trigger food safety disorders in deprived infants through life-threatening illnesses. The study aims to examine and identify the incidence of pathogenic bacteria of concern in PIF upon reconstitution in lukewarm water. A total of 172 samples consisting of 38 brands of PIF available in the Lagos metropolis were sampled, suspended in water (10 g in 100 mL), and bacteria strains were isolated using combinational enrichment and selective culture techniques. Pure bacterial strains were characterized and identified based on their physiology and 16S rRNA gene sequence homology. While 85 bacterial strains were isolated from the enriched culture system, 20 strains were selectively isolated based on tolerance to sodium deoxycholate. Approximately 13% of the selected bacteria were identified as Cronobacter spp., exhibiting virulence traits including extracellular protease production, coagulation and proteolysis of casein, haem-agglutination, and ß-haemolysis of human blood. Approximately 82% of the Cronobacter strains tolerated NaCl (10%) and bile salt; and exhibited resistance to cefotaxime, ceftriaxone, gentamicin, and Amoxicillin-clavulanic acid antibiotics. The presence of Cronobacter spp. in 13% of the PIF brands available to infants calls for concern about the safety of deprived infants that might be fed with such PIF. Consequently, PIF safety alerts need to be activate while further studies on critical points at which the pathogens get introduced to the PIFs need to be identified.
Asunto(s)
Cronobacter , Fórmulas Infantiles , Nigeria , Fórmulas Infantiles/microbiología , Cronobacter/genética , Cronobacter/aislamiento & purificación , Cronobacter/clasificación , Cronobacter/efectos de los fármacos , Humanos , Lactante , Microbiología de Alimentos , Antibacterianos/farmacología , ARN Ribosómico 16S/genética , Pruebas de Sensibilidad Microbiana , Inocuidad de los Alimentos , Farmacorresistencia BacterianaRESUMEN
Ensuring food safety, particularly for vulnerable groups, like infants and young children, requires identifying and prioritizing potential hazards in food chains. We previously developed a web-based decision support system (DSS) to identify specific microbiological hazards (MHs) in infant and toddler foods through a structured five-step process. This study takes the framework further by introducing systematic risk ranking (RR) steps to rank MH risks with seven criteria: process survival, recontamination, growth opportunity, meal preparation, hazard-food association evidence, food consumption habits of infants and toddlers in the EU, and MH severity. Each criterion is given a semi-quantitative or quantitative score or risk value, contributing to the final MH risk calculation via three aggregation methods: semi-quantitative risk scoring, semi-quantitative risk value, and outranking multi-criteria decision analysis (MCDA). To validate the criteria and ranking approaches, we conducted a case study to rank MH risks in infant formula, compared the results of the three risk ranking methods, and additionally evaluated the ranking results against expert opinions to ensure their accuracy. The results showed strong agreement among the three methods, consistently ranking Salmonella non-Typhi and Cronobacter spp. and Shiga-toxin-producing Escherichia coli as the top MH risks in infant formulae, with minor deviations. When MHs were ranked after an initial hazard identification step, all three methods produced nearly identical MH rankings, reinforcing the reliability of the ranking steps and the selected criteria. Notably, the risk value and MCDA methods provided more informative MH rankings compared to the risk scoring method. The risk value and risk scoring methods were implemented into an online tool, called the MIcrobiological hazards risk RAnking decision support system (Mira-DSS), available at https://foodmicrobiologywur.shinyapps.io/MIcrobial_hazards_RAnking/. In conclusion, our framework enables the ranking of MH risks, facilitating intervention comparisons and resource allocations to mitigate MH risks in infant foods, with potential applicability to broader food categories.
Asunto(s)
Microbiología de Alimentos , Inocuidad de los Alimentos , Alimentos Infantiles , Fórmulas Infantiles , Humanos , Lactante , Medición de Riesgo , Alimentos Infantiles/microbiología , Contaminación de Alimentos , Técnicas de Apoyo para la Decisión , Cronobacter/clasificación , Cronobacter/aislamiento & purificaciónRESUMEN
Cronobacter condimenti are environmental commensals that have not been associated with any clinical infections. To date, they are the least understood and described Cronobacter species within the genus. The objective of this study was to use a draft genome sequence (DGS) of the Cronobacter condimenti strain s37 to screen for genes encoding for antibiotic resistance, virulence, response to environmental stress, and biofilm formation. The strain was isolated in Poland from commercial small radish sprouts. This is the second genome of this species available in the GenBank database. The comparative genome analysis (cgMLST) of C. condimenti s37 with other Cronobacter spp. including the pathogenic species C. sakazakii and the plant-associated closely related genera Franconibacter and Siccibacter was also performed. The assembled and annotated genome of the C. condimenti s37 genome was 4,590,991 bp in length, with a total gene number of 4384, and a GC content of 55.7%. The s 37 genome encoded for genes associated with resistance to stressful environmental conditions (metal resistance genes: zinc, copper, osmotic regulation, and desiccation stress), 17 antimicrobial resistance genes encoding resistance to various classes of antibiotics and 50 genes encoding for the virulence factors. The latter were mainly genes associated with adhesion, chemotaxis, hemolysis, and biofilm formation. Cg-MLST analysis (3991 genes) revealed a greater similarity of C. condimenti s37 to S. turicensis, F. pulveris, and C. dublinensis than to other species of the genus Cronobacter. Studies on the diversity, pathogenicity, and virulence of Cronobacter species isolated from different sources are still insufficient and should certainly be continued. Especially the analysis of rare strains such as s37 is very important because it provides new information on the evolution of these bacteria. Comparative cgMLST analysis of s37 with other Cronobacter species, as well as closely related genera Franconibacter and Siccibacter, complements the knowledge on their adaptability to specific environments such as desiccation.
Asunto(s)
Cronobacter , Genoma Bacteriano , Factores de Virulencia , Cronobacter/genética , Cronobacter/patogenicidad , Cronobacter/aislamiento & purificación , Cronobacter/clasificación , Factores de Virulencia/genética , Virulencia/genética , Filogenia , Genómica/métodos , Farmacorresistencia Bacteriana/genética , Antibacterianos/farmacología , Biopelículas/crecimiento & desarrolloRESUMEN
Cronobacter species are potential pathogens that can contaminate powdered infant formula. C. sakazakii and C. malonaticus are the most common species of Cronobacter associated with infections. This study mined new molecular targets for the detection of C. sakazakii and C. malonaticus by using comparative genome approaches. Specific target genes mngB and ompR were obtained and used to detect C. sakazakii and C. malonaticus, respectively. A novel detection method, termed ladder-shape melting temperature isothermal amplification (LMTIA), was developed and evaluated. The detection limit for pure C. sakazakii DNA was 1 pg per reaction and 1 fg per reaction for C. malonaticus. The C. sakazakii, C. malonaticus, and the reference stains were all correctly identified. The amplicons can be successfully visualized and identified by naked eyes when hydroxy naphthol blue dye (HNB dye) was used in the reaction. Therefore, the LMTIA assays developed in this study showed potential application for microorganism identification and detection.
Asunto(s)
Cronobacter sakazakii , Cronobacter , ADN Bacteriano , Técnicas de Amplificación de Ácido Nucleico , Técnicas de Amplificación de Ácido Nucleico/métodos , Cronobacter sakazakii/genética , Cronobacter sakazakii/aislamiento & purificación , Cronobacter sakazakii/clasificación , Cronobacter/genética , Cronobacter/aislamiento & purificación , Cronobacter/clasificación , ADN Bacteriano/genética , Genoma Bacteriano , Fórmulas Infantiles/microbiología , Microbiología de Alimentos , Humanos , Contaminación de Alimentos/análisis , Límite de Detección , Técnicas de Diagnóstico Molecular/métodos , NaftalenosulfonatosRESUMEN
Cronobacter spp. are bacterial pathogens isolated from a wide variety of foods. This study aims at evaluating the occurrence of Cronobacter spp. in low water activity functional food samples, detect the presence of virulence genes, and determine the antibiotic susceptibility of strains. From 105 samples, 38 (36.2%) were contaminated with Cronobacter spp. The species identified by polymerase chain reaction (PCR) and sequencing analyses (rpoB and fusA genes, respectively) were C. sakazakii (60.3%), C. dublinensis (25.4%), C. turincensis (9.5%), and C. malonaticus (4.8%). Nineteen fusA alleles were identified, including four new alleles. The virulence genes were identified by PCR and all isolates were positive for ompX and sodA genes, 60.3% to cpa gene, and 58.7% to hly gene. Using the disk diffusion method, antibiotic susceptibility to twelve antibiotics was assessed twice, separated by a 19-month period. In the first test, the isolates showed diverse antibiotic susceptibility profiles, with nineteen isolates (30.2%) being multi-drug resistant (resistant to three or more antibiotic classes), in the second, the isolates were susceptible to all antibiotics. Cronobacter spp. in functional foods demonstrates the need for continued investigation of this pathogen in foods, and further research is needed to clarify the loss of resistance of Cronobacter strains.
Asunto(s)
Antibacterianos , Cronobacter , Alimentos Funcionales , Pruebas de Sensibilidad Microbiana , Cronobacter/genética , Cronobacter/efectos de los fármacos , Cronobacter/aislamiento & purificación , Cronobacter/clasificación , Brasil , Antibacterianos/farmacología , Microbiología de Alimentos , Factores de Virulencia/genética , Proteínas Bacterianas/genética , Contaminación de Alimentos/análisis , Agua , Farmacorresistencia Bacteriana/genéticaRESUMEN
This study aimed to evaluate the Cronobacter spp. strains isolated on the American continent and characterized using multi-locus sequence typing (MLST) available in the PubMLST database and current literature. From 465 Cronobacter spp. strains, the majority (n = 267, 57.4%) was from North America, mainly from USA (n = 234) and 198 (42.6%) were from South America, mainly from Brazil (n = 196). A total of 232 (49.9%) were isolated from foods, 102 (21.9%) from environmental, 87 (18.7%) from clinical, 27 (5.8%) from PIF, one from water (0.2%) and 16 (3.5%) from unknown sources. A total of five species were represented: Cronobacter sakazakii (374, 80.4%), Cronobacter malonaticus (41, 8.8%), Cronobacter dublinensis (29, 6.2%), Cronobacter turicensis (16, 3.5%) and Cronobacter muytjensii (5, 1.1%). The strains with complete MLST profile (n = 345) were assigned to 98 STs, a ratio of 3.5 strain by ST found and the calculated Simpson`s index was 0.93. The strains showed a high diversity and after eBURST analysis, 30 STs (n = 189) formed 12 single and/or double-locus variant clonal complexes (CC). A total of 38 STs (38.7%) were associated with clinical cases of infection, including well established C. sakazakii CC 1, 4, 8 and 83; C. malonaticus ST60, 307, 394 and 440; and C. sakazakii ST 12 and 494.
Asunto(s)
Cronobacter/clasificación , Cronobacter/aislamiento & purificación , Infecciones por Enterobacteriaceae/epidemiología , Enfermedades Transmitidas por los Alimentos/microbiología , Fórmulas Infantiles/microbiología , Cronobacter/genética , Cronobacter sakazakii/genética , Cronobacter sakazakii/aislamiento & purificación , Bases de Datos Factuales , Infecciones por Enterobacteriaceae/microbiología , Variación Genética/genética , Humanos , Lactante , Recién Nacido , Tipificación de Secuencias Multilocus , Factor G de Elongación Peptídica/genética , Estados Unidos/epidemiologíaRESUMEN
The emergence of Cronobacter as an important potential pathogen for newborn children and its occurrence in powdered infant formulae has generated a need to develop new management practices for this food group. This includes reduction of the prevalence of Cronobacter in manufacturing environments which can be a source of Cronobacter. This study was performed to assess the suitability of qualitative and quantitative Enterobacteriaceae and coliforms indicator tests for the presence and prevalence of Cronobacter. Environmental swabs (205) from five milk powder factories were examined. The qualitative indicator tests had good sensitivity but they lacked specificity for reliable routine use. Logistic regression analyses revealed a significant relationship between the quantitative indicator tests and Cronobacter prevalence, where the Enterobacteriaceae count was a slightly stronger predictor for Cronobacter than the coliforms count. The optimum test sensitivity (81%) and specificity (66%) was obtained when the indicator count thresholds were set at ≥1 cfu/cm2. However, since 11% of samples were Cronobacter positive when counts of Enterobacteriaceae and coliforms were less than 1 cfu/cm2, specific testing for Cronobacter is advised in addition to Enterobacteriaceae testing to minimise risk of transfer of Cronobacter from the factory environment into powdered infant formulae products.
Asunto(s)
Cronobacter/aislamiento & purificación , Enterobacteriaceae/aislamiento & purificación , Contaminación de Alimentos/análisis , Fórmulas Infantiles/microbiología , Leche/microbiología , Animales , Cronobacter/clasificación , Cronobacter/genética , Cronobacter/crecimiento & desarrollo , Enterobacteriaceae/clasificación , Enterobacteriaceae/genética , Enterobacteriaceae/crecimiento & desarrollo , Polvos/análisisRESUMEN
AIMS: The objective of this study was to evaluate the cytotoxic activity of Cronobacter strains isolated from foods (n = 50) and clinical samples (n = 6) in Brazil and genotype selected strains (n = 18) using multi-locus sequence typing (MLST) METHODS AND RESULTS: The cytotoxic activity of C. sakazakii (n = 29), C. dublinensis (n = 13), C. malonaticus (n = 6), C. turicensis (n = 6) and C. muytjensii (n = 2) was screened using Vero, RK13, Hep2c, NCTC clone 929 and BHK-21 cell lines. Selected Cronobacter strains were assigned to C. sakazakii ST 21, C. turicensis ST 252, C. sakazakii ST 647, and three newly assigned STs: C. turicensis STs 738-740. The maximum death caused by non-heat-treated filtrates was 20·4, 86·2, 47·0 and 84·0%, in Vero, RK13, Hep2c and NCTC clone 929 cells, respectively. These were caused by C. sakazakii strains C291 and C292 (ST 494) which had been isolated during neonatal Cronobacter meningitis infection, and C110 (ST 395) isolated from flaxseed flour. Thermal treatment (100°C/20 min) significantly reduced the cytotoxicity activity in NCTC clone 929 and Vero cells (P ≤ 2 × 10-6 ), but not in RK13 (P = 0·12) and Hep2c (P = 0·85), indicating the cytotoxin(s) were probably proteinaceous. Electron microscopy revealed that cytotoxic compounds from C. sakazakii induced several cell death characteristics, including loss of cell-cell contact, microvilli reduction and cellular lysis. Autophagic vacuoles and mitochondrial damage were the most common ultrastructural features observed. CONCLUSIONS: It was concluded that Cronobacter strains, especially C. sakazakii, could produce heat-labile cytotoxic compounds in cell filtrates. SIGNIFICANCE AND IMPACT OF THE STUDY: This study providing insights into the pathogenesis of the Cronobacter genus. Cytotoxins were identified in excreted filtrates of C. sakazakii strains isolated from food and clinical specimens. The presence of Cronobacter strains that can produce cytotoxins in foods can be a potential threat to human health and highlight the need for high levels of hygiene.
Asunto(s)
Cronobacter/clasificación , Cronobacter/patogenicidad , Microbiología de Alimentos , Meningitis Bacterianas/microbiología , Virulencia , Animales , Brasil , Línea Celular , Supervivencia Celular , Chlorocebus aethiops , Cronobacter/genética , ADN Bacteriano , Infecciones por Enterobacteriaceae/microbiología , Genotipo , Interacciones Huésped-Patógeno , Humanos , Tipificación de Secuencias Multilocus , Células VeroRESUMEN
Cronobacter species (Cronobacter spp.) are important foodborne pathogens that can infect and cause serious life-threatening diseases in infants and immunocompromised elderly. This study aimed to acquire data on Cronobacter spp. contamination of aquatic products in China from 2011 to 2016. In total, 800 aquatic products were tested, and the overall contamination rate for Cronobacter spp. was 3.9% (31/800). The average contamination level of the positive samples was 2.05 MPN/g. Four species and nine serotypes were identified among 33 isolates, of which the C. sakazakii serogroup O1 (n = 9) was the primary serotype. The majority of Cronobacter spp. strains harbored highest resistance against cephalothin (84.8%), followed by tetracycline (6.1%), trimethoprim/sulfameth-oxazole (3.0%) and chloramphenicol (3.0%). Two isolates were resistant to three antibiotics. In total, 26 sequence types and 33 CRISPR types (including 6 new STs and 26 new CTs) were identified, which indicates the extremely high diversity of Cronobacter spp. in aquatic products. Pathogenic C. sakazakii ST4, ST1, and C. malonaticus ST7 were also observed. Overall, this large-scale study revealed the relatively low prevalence and high genetic diversity of Cronobacter spp. in aquatic products in China, and the findings provide valuable information that can guide the establishment of effective measures for the control and precaution of Cronobacter spp. in aquatic products during production processes.
Asunto(s)
Repeticiones Palindrómicas Cortas Agrupadas y Regularmente Espaciadas , Cronobacter/clasificación , Cronobacter/aislamiento & purificación , Farmacorresistencia Bacteriana , Alimentos Marinos/microbiología , Antibacterianos/farmacología , Cefalotina/farmacología , China , Cloranfenicol/farmacología , Cronobacter sakazakii/clasificación , Cronobacter sakazakii/aislamiento & purificación , Contaminación de Alimentos , Microbiología de Alimentos , Variación Genética , Tipificación de Secuencias Multilocus , Prevalencia , Serotipificación , Tetraciclina/farmacología , Trimetoprim/farmacologíaRESUMEN
Cronobacter spp. are important foodborne pathogens that infections occur in all age groups, especially cause serious life-threatening diseases in infants. This study aimed to acquire data on Cronobacter spp. contamination of meat and meat products (n = 588) in China during 2011 to 2016, and investigated the use of CRISPR typing technology as an approach for characterizing the genetics of Cronobacter spp. The overall contamination rate for Cronobacter spp. was determined to be 9.18% (54/588). Of the positive samples, 90.74% (49/54) had <10 MPN/g, with duck samples had a relatively high contamination rate (15.69%, 8/51) and highest contamination level (28.90 MPN/g). Four species and nine serotypes were identified among 69 isolates, of which C. sakazakii was the major species (n = 50) and C. sakazakii serogroup O1 and O2 (n = 17) were the primary serotypes. The majority of Cronobacter spp. strains were found to be susceptible to most antibiotics except exhibited high resistance to cephalothin (76.81%, 53/69), and total two multi-drug resistant C. sakazakii strains were isolated from duck. The genetic diversity of Cronobacter spp. was remarkably high, as evidenced by the identification of 40 sequence types (STs) and 60 CRISPR types (CTs). C. sakazakii ST64 (n = 7) was the predominant genotype and was further divided into two sub-lineages based on CRISPR diversity, showing different antibiotic resistance profile. These results demonstrate that CRISPR typing results have a good correspondence with bacterial phenotypes, and it will be a tremendously useful approach for elucidating inter-subtyping during molecular epidemiological investigations while interpreting the divergent evolution of Cronobacter. The presence of Cronobacter spp. in meat and meat product is a potential threat to human public health.
Asunto(s)
Cronobacter/aislamiento & purificación , Microbiología de Alimentos , Carne/microbiología , Animales , Antibacterianos/farmacología , China/epidemiología , Repeticiones Palindrómicas Cortas Agrupadas y Regularmente Espaciadas , Cronobacter/clasificación , Cronobacter/efectos de los fármacos , Cronobacter/genética , Farmacorresistencia Bacteriana/efectos de los fármacos , Farmacorresistencia Bacteriana/genética , Variación Genética , PrevalenciaRESUMEN
Cronobacter infections of infants are commonly regarded as due to the ingestion of contaminated feed. The aim of this study was to determine the occurrence of Cronobacter, total coliforms and Escherichia coli in different brands of natural mineral waters as sold in 20 l returnable bottles in Rio de Janeiro, Brazil. The quantification of total coliforms and E. coli was performed by Most Probable Number. The detection of Cronobacter was as according to the ISO 22964:2017 and Bacteriological Analytical Manual/FDA. Molecular characterization of Cronobacter isolates was performed by real-time PCR and by multi-locus sequence typing. The antibiotic susceptibility profile was determined and biofilm production was evaluated in polystyrene microplates. Total coliforms and E. coli were detected in 13 (39·4%) and 2 (6·1%) of the 33 lots analysed respectively, and were considered unsatisfactory for human consumption according to Brazilian law. One (3·0%) lot showed contamination by C. malonaticus ST440 (Cronobacter MLST Databases accession no. ID 2646). The strain was susceptible to all (n = 13) antibiotics tested and only formed a weak biofilm. Since there is a high consumption of natural mineral waters by elderly and immunosuppressed persons, epidemiological surveillance agencies should be aware of the risk that these waters may represent for these groups. SIGNIFICANCE AND IMPACT OF THE STUDY: Cronobacter malonaticus ST440 was isolated from 20 l bottled drinking natural mineral waters sold in markets in Rio de Janeiro State, Brazil, and can be a potential threat to human health, particularly for neonates. Thirteen lots (39·4%) were unsatisfactory for human consumption due to the presence of total coliforms and/or Escherichia coli.
Asunto(s)
Cronobacter/aislamiento & purificación , Agua Potable/microbiología , Escherichia coli/aislamiento & purificación , Aguas Minerales/microbiología , Anciano , Antibacterianos/farmacología , Biopelículas , Brasil , Cronobacter/clasificación , Cronobacter/efectos de los fármacos , Infecciones por Enterobacteriaceae/prevención & control , Escherichia coli/efectos de los fármacos , Humanos , Recién Nacido , Pruebas de Sensibilidad Microbiana , Tipificación de Secuencias Multilocus , Reacción en Cadena en Tiempo Real de la PolimerasaRESUMEN
As an emerging food-borne pathogen, Cronobacter species are ubiquitous in the food and environment. In order to know the characteristics of Cronobacter spp. from the environment, we isolated Cronobacter spp. from soil and water, and then studied the molecular typing and antibiotic resistance characteristics of these isolates. In 2016, 141 soil and water samples were collected from farms and Riverside Park in Beijing. Isolates were identified by real-time PCR, 16s rRNA sequencing, and whole-genome sequencing. Molecular subtyping of these isolates was characterized by pulsed-field gel electrophoresis, multilocus sequence typing (MLST), and antibiotic susceptibility tests. Cronobacter species were classified based on fusA sequencing. Twenty-two samples (15.60%) contained Cronobacter spp., and four species were detected, i.e., C. dubliniensis (n = 10), C. sakazakii (n = 6), C. turicensis (n = 4), and C. malonaticus (n = 2). For MLST, 12 types (ST519-ST525, ST533-ST537) were newly identified, indicating high diversity. Most isolates (68.18%) showed resistance to cefazolin. Siccibacter turicensis and Cronobacter both with blue-green colonies on selective media should be respectively identified. Apparently, major Cronobacter species in soil and water samples differed from those in food. Molecular subtyping showed that the environment could not be excluded as a source of Cronobacter infection. The resistance to cefazolin of most isolates indicated natural resistance.
Asunto(s)
Cronobacter/clasificación , Microbiología del Suelo , Microbiología del Agua , Antibacterianos/farmacología , Beijing , Biodiversidad , Cronobacter/efectos de los fármacos , Cronobacter/genética , Cronobacter/aislamiento & purificación , Electroforesis en Gel de Campo Pulsado , Genoma Bacteriano/genética , Pruebas de Sensibilidad Microbiana , Tipificación de Secuencias Multilocus , Filogenia , ARN Ribosómico 16S/genéticaRESUMEN
Cronobacter spp. have been recognized as causative agents of various severe infections in pre-term or full-term infants as well as elderly adults suffering from serious underlying disease or malignancy. A surveillance study was designed to identify antibiotic resistance among clinical Cronobacter spp. strains, which were isolated from patients of two hospitals between May 2007 and August 2013. Altogether, 52 Cronobacter spp. isolates were analyzed. Although MALDI-TOF mass spectrometry recognized all Cronobacter sakazakii and Cronobacter malonaticus strains, it could not identify Cronobacter muytjensii strain. Nevertheless, all strains were identified as Cronobacter spp. using multilocus sequence typing (MLST). Strains were tested against 17 types of antibiotics, using the standard microdilution method according to the 2018 European Committee on Antimicrobial Susceptibility Testing criteria. Three Cronobacter species were identified as C. sakazakii (n = 33), C. malonaticus (n = 18), and C. muytjensii (n = 1); all isolates were susceptible to all tested antibiotics. All strains were PCR-negative for bla TEM, bla SHV, and bla CTX-M ß-lactamase genes, as well. Even though the results of this study showed that Cronobacter spp. isolates were pan-susceptible, continued antibiotic resistance surveillance is warranted.Cronobacter spp. have been recognized as causative agents of various severe infections in pre-term or full-term infants as well as elderly adults suffering from serious underlying disease or malignancy. A surveillance study was designed to identify antibiotic resistance among clinical Cronobacter spp. strains, which were isolated from patients of two hospitals between May 2007 and August 2013. Altogether, 52 Cronobacter spp. isolates were analyzed. Although MALDI-TOF mass spectrometry recognized all Cronobacter sakazakii and Cronobacter malonaticus strains, it could not identify Cronobacter muytjensii strain. Nevertheless, all strains were identified as Cronobacter spp. using multilocus sequence typing (MLST). Strains were tested against 17 types of antibiotics, using the standard microdilution method according to the 2018 European Committee on Antimicrobial Susceptibility Testing criteria. Three Cronobacter species were identified as C. sakazakii (n = 33), C. malonaticus (n = 18), and C. muytjensii (n = 1); all isolates were susceptible to all tested antibiotics. All strains were PCR-negative for bla TEM, bla SHV, and bla CTX-M ß-lactamase genes, as well. Even though the results of this study showed that Cronobacter spp. isolates were pan-susceptible, continued antibiotic resistance surveillance is warranted.
Asunto(s)
Antibacterianos/farmacología , Cronobacter/clasificación , Cronobacter/efectos de los fármacos , Técnicas de Tipificación Bacteriana , Cronobacter sakazakii/clasificación , Cronobacter sakazakii/efectos de los fármacos , Cronobacter sakazakii/genética , Farmacorresistencia Bacteriana Múltiple , Genotipo , Infecciones por Bacterias Gramnegativas/microbiología , Humanos , Pruebas de Sensibilidad Microbiana , Tipificación de Secuencias Multilocus , Polonia , Reacción en Cadena de la PolimerasaRESUMEN
Cronobacter spp. are associated with serious infections in neonates with the clinical presentations of necrotizing enterocolitis, bacteraemia and meningitis. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) was used to identify 203 Cronobacter isolates from imported food during 2006-2015 with an optimized in-house database. The isolates were predominantly C. sakazakii (88.18%), followed by C. malonaticus (8.37%), C. muytjensii (1.48%), C. turicensis (0.99%) and C. dublinensis (0.99%). The result was totally consistent with that of fusA allele sequencing. 12.32% (25/203) of isolates gave inconsistent spectra following separate protein extractions. Sixty C. sakazakii isolates and 24 isolates from the other four species were chosen for multi-locus sequence type analyses (MLST) and PCR-serotyping. Thirty-one sequence types were identified. The common sequence types were ST1 (19/60) and ST4 (13/60) for C. sakazakii and ST7 (12/17) for C. malonaticus. The primary serotypes were Csak O:1 (30/60), Csak O:2 (25/60) and Cmal O:2 (16/17) for C. sakazakii and C. malonaticus isolates, respectively. In conclusion, appropriate in-house database could make MALDI-TOF MS method identifying Cronobacter spp. isolates to the species level. But the spectra data were not sufficiently consistent for subtyping, unlike MLST. The Cronobacter spp. isolates have a high diversity including recognized pathovars.
Asunto(s)
Cronobacter/clasificación , Contaminación de Alimentos , Microbiología de Alimentos , Variación Genética , Técnicas de Tipificación Bacteriana , Beijing , Cronobacter/aislamiento & purificación , Tipificación de Secuencias Multilocus , Reacción en Cadena de la Polimerasa , Serogrupo , Serotipificación , Espectrometría de Masa por Láser de Matriz Asistida de Ionización DesorciónRESUMEN
The aim of this study was to determine the prevalence Cronobacter from 30 samples of oats and 30 of linseeds commercially available in Brazil. The detection of Cronobacter was as according to the ISO 22964:2017. The isolates were characterized according to their phenotypically using Vitek 2.0 and antibiotic susceptibility profile. Molecular characterization was accomplished by real-time PCR targeting dnaG gene, PCR targeting rpoB gene, multiplex-PCR targeting cgcA gene and fusA allele sequencing. A total of 34 samples (56.7%) contained Cronobacter; 19 (63.3%) of linseeds and 15 (50.0%) of oats. The isolates were identified as C. sakazakii (n = 18, 52.9%), C. dublinensis (n = 7, 20.6%), C. turicensis (n = 6, 17.7%) and C. malonaticus (n = 3, 8.8%). Thirty-four Cronobacter isolates were assigned to 11 different fusA alleles of which 3 were new (169, 170 and 171). The PCR targeting rpoB gene and cgcA gene failed to identify 19 isolates. Seven (20.6%) strains showed resistance or intermediate/resistance to tetracycline, and one (2.9%) strain had intermediate resistance to piperacilin-tazobactam. The presence of Cronobacter in oats and linseeds indicate that these foods can be a potential threat to human health, particularly when preparing food for elderly or immunosuppressed persons. The incorrect use of this foods for feeding of neonates (<6 months) by careers should also be avoided.
Asunto(s)
Avena/microbiología , Cronobacter/efectos de los fármacos , Cronobacter/genética , Lino/microbiología , Microbiología de Alimentos , Antibacterianos/farmacología , Técnicas de Tipificación Bacteriana , Brasil , Cronobacter/clasificación , Genes Bacterianos/genética , Humanos , Pruebas de Sensibilidad Microbiana , Reacción en Cadena en Tiempo Real de la PolimerasaRESUMEN
Cronobacter are opportunistic bacterial pathogens of both infants and adults. We investigated the incidence and distribution of Cronobacter in 1245 samples of cereal and related environments. 39.1% (101/258) rice-related and 46.9% (98/209) wheat-related samples tested positive for Cronobacter, and the positive rate differed notably according to processing method. Cronobacter was found in rice and wheat plants at the tillering, filling and mature stages. Soil, water and swab samples from nearby milling plants were assayed, and results revealed that 6.3% (7/122) of water from paddy fields, 49.1% (28/57) and 62.1% (41/67) of swab samples from rice and wheat flour milling plants were Cronobacter positive. Pulsed-field gel electrophoresis (PFGE) subtyping indicated that some strains had a common profile, which suggested their persistence in the environment, potential transmission routes and cross-contamination in processing. Finally, we surveyed 18 families to evaluate potential risks. None of the families who primarily ate rice cooked with water tested positive for Cronobacter, though of 66.7% families (6/9) whose food staples were produced from wheat flour tested positive. Taken together, our results are important for understanding Cronobacter transmission and will aid in the development of additional control measures to reduce the risk of infection by these opportunistic pathogenic bacteria.
Asunto(s)
Cronobacter/aislamiento & purificación , Grano Comestible/microbiología , Manipulación de Alimentos/estadística & datos numéricos , Microbiología de Alimentos/estadística & datos numéricos , Técnicas de Tipificación Bacteriana , Análisis por Conglomerados , Cronobacter/clasificación , Cronobacter/genética , Grano Comestible/crecimiento & desarrollo , Electroforesis en Gel de Campo Pulsado , Microbiología Ambiental , Conducta Alimentaria , Harina/microbiología , Manipulación de Alimentos/métodos , Abastecimiento de Alimentos , Humanos , Oryza/crecimiento & desarrollo , Oryza/microbiología , Triticum/crecimiento & desarrollo , Triticum/microbiologíaRESUMEN
Identification of Cronobacter represent a major challenge for laboratories testing powdered infant formula (PIF). In the present study, two biochemical galleries and three molecular methods have been applied to confirm 276 Cronobacter spp. and non-Cronobacter isolates from different sources. Using the latest database of API 20â¯E and ID 32â¯E biochemical miniaturized kits, 53% and 78% of the isolates were identified respectively. From the available results, total accuracy for Cronobacter detection was in 97% (API 20â¯E) and 99% (ID 32â¯E). The three molecular methods were based on rRNA based lateral flow, Real Time PCR combined with either a hybridization or hydrolysis probe. For all three methods total accuracy was more than 99%. A pilot trial using Next Generation Sequencing (NGS) correctly identified 58 out of 66 isolates (88%) in DNA mixtures. The results indicate that the commercially available approaches such as ID 32â¯E, rRNA based lateral flow and Real Time PCR are all suitable for Cronobacter identification at the genus level. The NGS method may become a suitable alternative in the future, provided that the sequence database is improved.
Asunto(s)
Bioquímica/métodos , Cronobacter/genética , ADN Bacteriano/genética , Fórmulas Infantiles/microbiología , ARN Ribosómico 16S/genética , Reacción en Cadena en Tiempo Real de la Polimerasa/métodos , Técnicas de Tipificación Bacteriana , Bioquímica/instrumentación , Cronobacter/clasificación , Cronobacter/aislamiento & purificación , Exactitud de los Datos , Contaminación de Alimentos/análisis , Microbiología de Alimentos/métodos , Humanos , Lactante , Recién Nacido , Miniaturización , Papel , Filogenia , Juego de Reactivos para Diagnóstico , Tiras Reactivas/análisisRESUMEN
In Bogotá, Colombia, a large number of babies are fed with breast milk substitutes made from corn and plantain starch. We found 34.3% of tested samples to be contaminated with Cronobacter spp.; C. sakazakii was the most recovered species. Our findings underscore the risk for contamination of breast milk substitutes.
Asunto(s)
Cronobacter , Contaminación de Alimentos , Microbiología de Alimentos , Enfermedades Transmitidas por los Alimentos/epidemiología , Enfermedades Transmitidas por los Alimentos/microbiología , Sustitutos de la Leche , Colombia/epidemiología , Cronobacter/clasificación , Cronobacter/genética , Cronobacter/aislamiento & purificación , Humanos , Vigilancia en Salud PúblicaRESUMEN
Cronobacter spp. (previously Enterobacter sakazakii), an opportunistic food-borne pathogen, involved in fatal infections in neonates and infants. Contaminated powdered infant formula (PIF) was considered to be the uppermost infective resource. In the present study, a thermophilic helicase-dependent isothermal amplification (tHDA) based method combined with silica-coated magnetic nanoparticles (Si-MNPs) as a DNA separator for rapid and sensitive detection of Cronobacter has been developed. The whole detection process can be completed in <3â¯h, with a detection limit of 100 and 101â¯CFU/ml Cronobacter in pure culture and artificially contaminated PIF, respectively, with a great sensitivity (94%) and specificity (100%). Hence, this protocol might be useful for screening and monitoring the contamination of Cronobacter spp. in food industry, and helpful for avoiding the economic loss by retard of feedback of the contamination of pathogens in dairy products.
Asunto(s)
Cronobacter/genética , ADN Helicasas/química , Microbiología de Alimentos/métodos , Fórmulas Infantiles/microbiología , Nanopartículas de Magnetita/química , Técnicas de Amplificación de Ácido Nucleico/métodos , Cronobacter/clasificación , Humanos , Lactante , Recién Nacido , Dióxido de Silicio/químicaRESUMEN
Crononbacter spp. is an opportunistic foodborne pathogen that causes infections in neonates, infants, and immunocompromised adults. Although the contamination of spices with Cronobacter has been previously reported in some countries, there have been no studies on Cronobacter contamination in China. Therefore, this study aimed to investigate the prevalence of Cronobacter spp. in Chinese retail spices. Fifty-six packaged Chinese spices were collected from different markets, and 32 of these were found to be contaminated with Cronobacter. Five species were identified from the 54 isolates of the 32 positive samples: Cronobacter sakazakii (n = 35), Cronobacter muytjensii (n = 8), Cronobacter malonaticus (n = 6), Cronobacter turicensis (n = 3), and Cronobacter dublinensis (n = 2). Pulsed-field gel electrophoresis demonstrated high genetic diversity, as 53 PFGE profiles were revealed for the 54 isolates. Multilocus sequence typing analysis revealed 46 sequence types, and of these, 26 were newly identified. Most of the isolates were sensitive to antibiotics (n = 15), with the exception of cefazolin. This study revealed that the contamination of Chinese retail spices by Cronobacter spp. poses a potential risk to the consumer.