Alkaline phosphatase attenuates LPS-induced liver injury by regulating the miR-146a-related inflammatory pathway.

Lipopolysaccharide (LPS) can remain in dairy products after the sterilization of milk powder and may pose a threat to the health of infants and young children. There is a large amount of alkaline phosphatase (ALP) in raw milk, which can remove the phosphate bond of LPS, thus, detoxifying it. ALP is regarded as an indicator of the success of milk sterilization due to its strong heat resistance. ALP can alleviate the toxicity of LPS in enteritis and nephritis models, but the mechanism by which oral-intake of ALP protects liver tissue from LPS stimulation is unclear. In this study, an in vivo acute mouse liver injury model was induced by C. sakazakii LPS (200 µg/kg) and used to verify the protective mechanism of ALP (200 U/kg) on mice livers. The related pathways were also verified by in vitro cell culture. Enzyme linked immunosorbent assays (ELISAs), quantitative reverse transcription PCR (RT-qPCR) and western blotting were used to detect the levels of inflammatory factors at the protein level and RNA level, and to confirm the inflammation of liver tissue caused by LPS. ALP was found to alleviate acute liver injury in vitro by activating miR-146a. We found that ALP could up-regulate the level of miR146a and subsequently alleviates the expression of TLR4, TNF-α, matured IL-1ß, and NF-κB in mouse liver tissue and hepatocytes; thus, reducing liver inflammation. Herein, we demonstrated for the first time that oral-intake of ALP protected liver tissue by up-regulating the expression of miR-146a and alleviating inflammatory reactions; thus, providing a research basis for the proper processing of milk. This study also suggests that producers should improve the awareness of the protective effects of bioactive proteins in raw milk.

Protective Effect of Recombinant Proteins of Cronobacter Sakazakii During Pregnancy on the Offspring.

Cronobacter sakazakii is a food-borne pathogen carried in milk powder that can cause severe bacteremia, enterocolitis, and meningitis in newborns, which can lead to death of newborns. Preventing infection by this pathogen is significant to the health of newborns. Since infants and young children are the main target group of C. sakazakii, it is considered that maternal immunity can enhance the protection of newborns. Previous studies showed that two proteins of C. sakazakii (GroEL and OmpX) exhibited high expression levels and elicited strong immune reactions, suggesting their potential as vaccine candidates. In this study, GroEL and OmpX were recombinantly expressed in Escherichia coli and purified as immunogens to immunize pregnant rats. Three days after birth, the progeny were challenged with C. sakazakii to determine the protective effect of maternal immunity on the offspring. The results showed that immunization during pregnancy decreased bacterial load in the brain and blood, reduced brain and intestine damage, and significantly increased specific antibody titers in the offspring. Immunization with the recombinant proteins significantly increased cytokine levels in the serum of the progeny. The group whose mothers were immunized with OmpX produced more IL-4, while the group whose mothers were immunized with GroEL produced more IFN-γ, indicating that the immunogens enhanced the Th2 and Th1 responses, respectively. However, although the immune response was induced by both proteins, only the offspring of the pregnant rats immunized with OmpX or OmpX/GroEL mixture showed delayed death, possibly because immunization with OmpX led to a stronger humoral immune response in the offspring, suggesting that OmpX was a better vaccine candidate than GroEL. This study first reported that exposure to C. sakazakii proteins during pregnancy could improve the offspring's ability to resist infection caused by this pathogen.

C. sakazakii activates AIM2 pathway accompanying with excessive ER stress response in mammalian mammary gland epithelium.

Bovine mastitis is a common inflammatory disease caused by various factors. The main factor of mastitis is pathogenic microorganism infection, such as Staphylococcus aureus, Escherichia coli, and Streptococcus. Cronobacter sakazakii (C. sakazakii) is a newly discovered pathogenic bacteria in milk products, which seriously threat human health in recent years. At present, it has not been reported that the pathogenesis of mastitis is caused by C. sakazakii. This study investigated the inflammation of mammary gland epithelium, which was induced by C. sakazakii for the first time. We focused on bacterial isolation, histological observation, AIM2 inflammasome pathways, endoplasmic reticulum stress, and apoptosis. The results showed that C. sakazakii-induced inflammation caused damage of tissue, significantly increased the production of pro-inflammatory cytokines (including TNF-α, IL-1ß, and IL-6), activated the AIM2 inflammasome pathway (increased the expression of AIM2 and cleaved IL-1ß), and induced endoplasmic reticulum stress (increased the expression of ERdj4, Chop, Grp78) and apoptosis (increased the ratio of Bax/Bcl-2, a marker of apoptosis). In conclusion, it is suggested that it maybe inhibite AIM2 inflammasome pathways and alleviate endoplasmic reticulum stress (ER stress) against the C. sakazakii-induced inflammation.

Immunogold Nanoparticles for Rapid Plasmonic Detection of C. sakazakii.

Cronobacter sakazakii is a foodborne pathogen that can cause a rare, septicemia, life-threatening meningitis, and necrotizing enterocolitis in infants. In general, standard methods for pathogen detection rely on culture, plating, colony counting and polymerase chain reaction DNA-sequencing for identification, which are time, equipment and skill demanding. Recently, nanoparticle- and surface-based immunoassays have increasingly been explored for pathogen detection. We investigate the functionalization of gold nanoparticles optimized for irreversible and specific binding to C. sakazakii and their use for spectroscopic detection of the pathogen. We demonstrate how 40-nm gold nanoparticles grafted with a poly(ethylene glycol) brush and functionalized with polyclonal antibodies raised against C. sakazakii can be used to specifically target C. sakazakii. The strong extinction peak of the Au nanoparticle plasmon polariton resonance in the optical range is used as a label for detection of the pathogens. Individual binding of the nanoparticles to the C. sakazakii surface is also verified by transmission electron microscopy. We show that a high degree of surface functionalization with anti-C. sakazakii optimizes the detection and leads to a detection limit as low as 10 CFU/mL within 2 h using a simple cuvette-based UV-Vis spectrometric readout that has great potential for further optimization.

Electrochemical coupled immunosensing platform based on graphene oxide/gold nanocomposite for sensitive detection of Cronobacter sakazakii in powdered infant formula.

A sensitive electrochemical immunosensing platform for the detection of Cronobacter sakazakii was developed using a graphene oxide/gold (GO/Au) composite. Transmission electron microscopy showed that the Au nanoparticles, with an average size of < 30 nm, were well dispersed on the GO surface. For the detection of C. sakazakii, a polyclonal anti-C. sakazakii antibody (IgG) was covalently immobilized to the Au nanoparticles on the surface of the GO/Au composite coated glassy carbon electrode (GCE). The electrochemical sensing performance of immunofunctionalized GCE was characterized by cyclic voltammetry and differential pulse voltammetry. Under optimized conditions, in pure culture there was a linear relationship between electrical signal and C. sakazakii levels over the range 2.0 × 102-2.0 × 107 cfu/mL (R2 = 0.999), with a detection limit of 2.0 × 101 cfu/mL. The total analytical time was 15 min per sample. The C. sakazakii electrochemical immunosensing assay was able to successfully detect 2.0 × 101 cfu/mL of C. sakazakii in artificially contaminated powdered infant formula without any enrichment or pre-enrichment steps. Furthermore, the recovery rates of the C. sakazakii electrochemical immunosensing assay following spiking of powdered infant formula with different concentrations of C. sakazakii (cfu/mL) were 82.58% at 2.0 × 101 cfu/mL, 84.86% at 2.0 × 102 cfu/mL, and 95.40% at 2.0 × 103 cfu/mL. The C. sakazakii electrochemical immunosensing assay had good selectivity, reproducibility, and reactivity compared with other Cronobacter spp. and/or pathogens belonging to other genera, indicating its significant potential in the clinical diagnosis of C. sakazakii.

Detection of Cronobacter sakazakii in powdered infant formula using an immunoliposome-based immunomagnetic concentration and separation assay.

This study aimed to optimize the applicability of an immunoliposome-based immunomagnetic concentration and separation assay to facilitate rapid detection of Cronobacter sakazakii in powdered infant formula (PIF). To determine the detection limit, specificity, and pre-enrichment incubation time (0, 4, 6, and 8 h), assay tests were performed with different cell numbers of C. sakazakii (2 × 100 and 2 × 101 CFU/ml) inoculated in 10 g of PIF. The assay was able to detect as few as 2 cells of C. sakazakii/10 g of PIF sample after 6 h of pre-enrichment incubation with an assay time of 2 h 30 min. The assay was assessed for cross-reactivity with other bacterial strains and exhibited strong specificity to C. sakazakii. Moreover, the assay method was applied to the detection of C. sakazakii in PIF without pre-enrichment steps, and the results were compared with INC-ELISA and RT-PCR. The developed method was able to detect C. sakazakii in spiked PIF without pre-enrichment, whereas INC-ELISA failed to detect C. sakazakii. In addition, when compared with the results obtained with RT-PCR, our developed assay required lesser detection time. The developed assay was also not susceptible to any effect of the food matrix or background contaminant microflora.

Immunochromatographic Strip Assay for Detection of Cronobacter sakazakii in Pure Culture.

Cronobacter sakazakii (C. sakazakii) is a foodborne pathogen, posing a high risk of disease to infants and immunocompromised individuals. In order to develop a quick, easy, and sensitive assay for detecting C. sakazakii, a rabbit anti-C. sakazakii immunoglobulin G (IgG) was developed using sonicated cell protein from C. sakazakii. The developed anti-C. sakazakii (IgG) was of good quality and purity, as well as species-specific. The developed rabbit anti-C. sakazakii IgG was attached to the surface of a sulforhodamine B-encapsulated liposome to form an immunoliposome. A test strip was then prepared by coating goat anti-rabbit IgG onto the control line and rabbit anti-C. sakazakii IgG onto the test line, respectively, of a plastic-backed nitrocellulose membrane. A purple color signal both on the test line and the control line indicated the presence of C. sakazakii in the sample, whereas purple color only on the control line indicated the absence of C. sakazakii in the sample. This immunochromatographic strip assay could produce results in 15 min with a limit of detection of 107 CFU/ml in C. sakazakii culture. The immunochromatographic strip assay also showed very good specificity without cross-reactivity with other tested Cronobacter species. Based on these results, the developed immunochromatographic strip assay is efficient for the detection of C. sakazakii and has high potential for on-site detection.

O antigen of FranconibacterpulverisG3872 (O1) is a 4-deoxy-d-arabino-hexose-containing polysaccharide synthesized by the ABC-transporter-dependent pathway.

Franconibacter (Enterobacter, Cronobacter) pulveris bacteria share several typical characteristics with, and hence pose a challenge for the detection of, Cronobacter sakazakii, an emerging opportunistic pathogen, which can cause severe infections in neonates. A structurally variable O-specific polysaccharide (OPS) called O antigen provides the major basis for the typing of Gram-negative bacteria. We investigated the structure and genetics of the O antigen of F. pulveris G3872 (designated O1). An OPS was isolated by mild alkaline degradation of the LPS, whereas the same polysaccharide and its oligosaccharide fragments were obtained by mild acid degradation. Studies by sugar analysis and NMR spectroscopy showed that the OPS contained d-ribose, l-rhamnose (l-Rha) and a rarely occurring monosaccharide 4-deoxy-d-arabino-hexose, and the OPS structure was established. The O-antigen gene cluster of F. pulveris G3872 between JUMPStart and gnd genes includes putative genes for glycosyltransferases, ATP-binding cassette (ABC)-transporter genes wzm and wzt, and genes for the synthesis of l-Rha, but no genes for the synthesis of 4-deoxy-d-arabino-hexose. A mutation test with the wzm gene confirmed that the OPS is synthesized and exported by the ABC-transporter-dependent pathway. A trifunctional transferase was suggested to catalyse formation of two glycosidic linkages and add a methyl group to the non-reducing end of the OPS to terminate the chain elongation. A carbohydrate-binding module that presumably recognizes the terminal methyl-modified monosaccharide was found at the C-terminus of Wzt. Primers specific for F. pulveris G3872 were designed based on the wzm gene, which has potential to be used for identification and detection of the O1 serogroup.

Immunological detection of Cronobacter and Salmonella in powdered infant formula by plasmonic label-free assay.

UNLABELLED: Cronobacter is an emerging food pathogen, especially in infants and neonates, often associated with the ingestion of contaminated Powdered Infant Formula (PIF). Therefore, regulations require the control of the absence of Cronobacter and of Salmonella, another important food pathogen, in these food products. So far, reference and alternative methods take up to several days, and no validated method exists for the simultaneous detection of these two pathogens. In this work, we propose to address this issue by an innovative and easy-to-operate assay, named Plasmonic Immuno-Assay (PlasmIA), and by producing dedicated polyclonal antibodies. Our approach is based on Surface Plasmon Resonance imaging of antibody-arrays and bacterial growth during a standardized enrichment. Such a single-step assay enables the multiplex detection of both Cronobacter and Salmonella, with concentrations smaller than 30 CFU cells in 25 g PIF samples, in less than 1 day. SIGNIFICANCE AND IMPACT OF THE STUDY: Among bacterial pathogens involved in food contamination, Cronobacter and Salmonella are of particular interest. Nevertheless, all detection methods used so far require several days to assess food safety. In the present paper, we describe the first multiplex immuno-assay ever described for fast and specific detection of these two pathogens in food samples. Such advances were made possible by combining the advantages of protein microarrays with on-biochip culture of contaminated food samples and an easy-to-operate optical detection. By doing so, we managed to detect both viable Cronobacter and Salmonella occurring during the enrichment phase.

Immunoliposome-based immunomagnetic concentration and separation assay for rapid detection of Cronobacter sakazakii.

This study aimed to develop an immunoliposome-based immunomagnetic concentration and separation assay for the rapid detection of Cronobacter sakazakii (C. sakazakii), an acute opportunistic foodborne pathogenic bacterium, in both pure culture and infant formula. To develop the assay, magnetic nanoparticles (diameter 30 nm) were coated with immunoglobulin G (IgG), specifically anti-C. sakazakii IgG, and applied for the sensitive and efficient detection of C. sakazakii using immunoliposomes. The binding efficiency of anti-C. sakazakii IgG to the magnetic nanoparticles was 86.23 ± 0.59%. The assay developed in this study detected as few as 3.3 × 10(3) CFUmL(-1) of C. sakazakii in pure culture within 2h 30 min; in comparison, an indirect non-competitive enzyme-linked immunosorbent assay was able to detect 6.2 × 10(5) CFUmL(-1) of C. sakazakii in pure culture after 17 h. The developed assay did not show any cross-reactivity with other Cronobacter spp. or pathogens belonging to other genera. In addition, the method was able to detect 10(3) CFUmL(-1) of C. sakazakii in infant formula without any pre-incubation. These results confirm that the immunoliposome-based immunomagnetic concentration and separation assay may facilitate highly sensitive, efficient, and rapid detection of C. sakazakii.

Cronobacter sakazakii clinical isolates overcome host barriers and evade the immune response.

Cronobacter sakazakii is the most frequently clinically isolated species of the Cronobacter genus. However the virulence factors of C. sakazakii including their ability to overcome host barriers remains poorly studied. In this study, ten clinical isolates of C. sakazakii were assessed for their ability to invade and translocate through human colonic carcinoma epithelial cells (Caco-2) and human brain microvascular endothelial cells (HBMEC). Their ability to avoid phagocytosis in human macrophages U937 and human brain microglial cells was investigated. Additionally, they were tested for serum sensitivity and the presence of the Cronobacter plasminogen activation gene (cpa) gene, which is reported to confer serum resistance. Our data showed that the clinical C. sakazakii strains invaded and translocated through Caco-2 and HBMEC cell lines and some strains showed significantly higher levels of invasion and translocation. Moreover, C. sakazakii was able to persist and even multiply in phagocytic macrophage and microglial cells. All strains, except one, were able to withstand human serum exposure, the single serum sensitive strain was also the only one which did not encode for the cpa gene. These results demonstrate that C. sakazakii clinical isolates are able to overcome host barriers and evade the host immune response indicating their capacity to cause diseases such as necrotizing enterocolitis (NEC) and meningitis. Our data showed for the first time the ability of C. sakazakii clinical isolates to survive and multiply within human microglial cells. Additionally, it was shown that C. sakazakii clinical strains have the capacity to translocate through the Caco-2 and HBMEC cell lines paracellularly.

Immunoproteomic identification of immunogenic proteins in Cronobacter sakazakii strain BAA-894.

Cronobacter spp. are emerging opportunistic pathogens. Cronobacter sakazakii is considered as the predominant species in all infections. So far, our understanding of the species' immunogens and potential virulence factors of Cronobacter spp. remains limited. In this study, an immunoproteomic approach was used to investigate soluble and insoluble proteins from the genome-sequenced strain C. sakazakii ATCC BAA-894. Proteins were separated using two-dimensional electrophoresis, detected by Western blotting with polyclonal antibodies of C. sakazakii BAA-894, and identified using tandem mass spectrometry (MALDI-MS and MALDI-MS/MS, MS/MSMS). A total of 11 immunoreactive proteins were initially identified in C. sakazakii BAA-894, including two outer membrane proteins, four periplasmic proteins, and five cytoplasmic proteins. In silico functional analysis of the 11 identified proteins indicated three proteins that were initially described as immunogens of pathogenic bacteria. For the remaining eight proteins, one protein was categorized as a potential virulence factor involved in protection against reactive oxygen species, and seven proteins were considered to play potential roles in adhesion, invasion, and biofilm formation. To our knowledge, this is the first time that immunogenic proteins of C. sakazakii BAA-894 have been identified as immunogens and potential virulence factors by an immunoproteomics approach. Future studies should investigate the roles of these proteins in bacterial pathogenesis and modulation of host immune responses during infection to identify their potential as molecular therapeutic targets.

Development of sandwich enzyme-linked immunosorbent assay for the detection of Cronobacter muytjensii (formerly called Enterobacter sakazakii).

This study aimed to produce a polyclonal antibody against Cronobacter muytjensii (C. muytjensii, formerly called Enterobacter sakazakii) and to develop an immunoassay for its detection. The optimum production of rabbit anti-C. muytjensii immunoglobulin G (IgG) and chicken anti-C. muytjensii IgY was reached in weeks 8 and 9, respectively. Purification of rabbit anti-C. muytjensii IgG from immunized rabbit sera was accomplished using the caprylic acid and ammonium sulfate precipitation method. As a result, sodium dodecyl sulfate-polyacrylamide gel electrophoresis produced two bands around 25 and 50 kDa, corresponding to a light and a heavy chain, respectively. The optimized conditions for sandwich enzyme-linked immunosorbent assay were using rabbit anti-C. muytjensii IgG (1 µg/mL) as a detection antibody and chicken anti-C. muytjensii IgY (10 µg/mL) as a capture antibody. In this assay, no cross-reactivity was observed with the other genera of pathogenic bacteria tested, which included Escherichia coli O157:H7, Salmonella typhimurium, Staphylococcus aureus, Bacillus cereus and Listeria monocytogenes. The developed assay did not show cross-reactivity with other tested species of Cronobacter and Enterobacter genera such as C. turicensis, C. sakazakii, E. aerogenes, E. pulveris and E. helveticus. The detection limit of sandwich ELISA for C. muytjensii was found to be 2.0 × 10(4) colony forming units (CFU)/mL. In addition, detection of C. muytjensii in infant formula powder showed a low matrix effect on the detection curve of sandwich ELISA for C. muytjensii, the detection limit being found to be 6.3 × 10(4) CFU/mL. These findings demonstrate that the developed method is able to detect all strains of C. muytjensii. Hence, this ELISA technique has potent application for the rapid and accurate detection of C. muytjensii in dietary foods.

Genetic analysis of the Cronobacter sakazakii O4 to O7 O-antigen gene clusters and development of a PCR assay for identification of all C. sakazakii O serotypes.

The Gram-negative bacterium Cronobacter sakazakii is an emerging food-borne pathogen that causes severe invasive infections in neonates. Variation in the O-antigen lipopolysaccharide in the outer membrane provides the basis for Gram-negative bacteria serotyping. The O-antigen serotyping scheme for C. sakazakii, which includes seven serotypes (O1 to O7), has been recently established, and the O-antigen gene clusters and specific primers for three C. sakazakii serotypes (O1, O2, and O3) have been characterized. In this study, the C. sakazakii O4, O5, O6, and O7 O-antigen gene clusters were sequenced, and gene functions were predicted on the basis of homology. C. sakazakii O4 shared a similar O-antigen gene cluster with Escherichia coli O103. The general features and anomalies of all seven C. sakazakii O-antigen gene clusters were evaluated and the relationship between O-antigen structures and their gene clusters were investigated. Serotype-specific genes for O4 to O7 were identified, and a molecular serotyping method for all C. sakazakii O serotypes, a multiplex PCR assay, was developed by screening against 136 strains of C. sakazakii and closely related species. The sensitivity of PCR-based serotyping method was determined to be 0.01 ng of genomic DNA and 10(3) CFU of each strain/ml. This study completes the elucidation of C. sakazakii O-antigen genetics and provides a molecular method suitable for the identification of C. sakazakii O1 to O7 strains.

Cpa, the outer membrane protease of Cronobacter sakazakii, activates plasminogen and mediates resistance to serum bactericidal activity.

Cronobacter spp. are emerging neonatal pathogens in humans, associated with outbreaks of meningitis and sepsis. To cause disease, they must survive in blood and invade the central nervous system by penetrating the blood-brain barrier. C. sakazakii BAA-894 possesses an ~131-kb plasmid (pESA3) that encodes an outer membrane protease (Cpa) that has significant identity to proteins that belong to the Pla subfamily of omptins. Members of this subfamily of proteins degrade a number of serum proteins, including circulating complement, providing protection from the complement-dependent serum killing. Moreover, proteins of the Pla subfamily can cause uncontrolled plasmin activity by converting plasminogen to plasmin and inactivating the plasmin inhibitor α2-antiplasmin (α2-AP). These reactions enhance the spread and invasion of bacteria in the host. In this study, we found that an isogenic cpa mutant showed reduced resistance to serum in comparison to its parent C. sakazakii BAA-894 strain. Overexpression of Cpa in C. sakazakii or Escherichia coli DH5α showed that Cpa proteolytically cleaved complement components C3, C3a, and C4b. Furthermore, a strain of C. sakazakii overexpressing Cpa caused a rapid activation of plasminogen and inactivation of α2-AP. These results strongly suggest that Cpa may be an important virulence factor involved in serum resistance, as well as in the spread and invasion of C. sakazakii.

Enterobacter sakazakii targets DC-SIGN to induce immunosuppressive responses in dendritic cells by modulating MAPKs.

Enterobacter sakazakii (ES) is an emerging pathogen that causes meningitis and necrotizing enterocolitis in infants. Dendritic cells (DCs) are professional phagocytic cells that play an essential role in host defense against invading pathogens; however, the interaction of ES with DCs is not known. In this study, we demonstrate that ES targets DC-specific ICAM nonintegrin (DC-SIGN) to survive in myeloid DCs for which outer membrane protein A (OmpA) expression in ES is critical, although it is not required for uptake. In addition, DC-SIGN expression was sufficient to cause a significant invasion by ES in HeLa cells and intestinal epithelial cells, which are normally not invaded by ES. OmpA(+) ES prevented the maturation of DCs by triggering the production of high levels of IL-10 and TGF-beta and by suppressing the activation of MAPKs. Pretreatment of DCs with Abs to IL-10 and TGF-beta or of bacteria with anti-OmpA Abs significantly enhanced the maturation markers on DCs. Furthermore, DCs pretreated with various inhibitors of MAPKs prohibited the increased production of proinflammatory cytokines stimulated by LPS or OmpA(-) ES. LPS pretreatment followed by OmpA(+) ES infection of DCs failed to induce maturation of DCs, indicating that OmpA(+) ES renders the cells in immunosuppressive state to external stimuli. Similarly, OmpA(+) ES-infected DCs failed to present Ag to T cells as indicated by the inability of T cells to proliferate in MLR. We conclude that ES interacts with DC-SIGN to subvert the host immune responses by disarming MAPK pathway in DCs.

Structure of the antigenic repeating pentasaccharide unit of the LPS O-polysaccharide of Cronobacter sakazakii implicated in the Tennessee outbreak.

Strains of the Gram-negative bacterium Cronobacter (Enterobacter) sakazakii have been identified as emerging opportunistic pathogens that can cause enterocolitis, bacteraemia, meningitis, and brain abscess, and have been particularly associated with meningitis in neonates where infant-milk formulae has been epidemiologically linked to the disease. A study of the lipopolysaccharides produced by clinical isolates using chemical, 2D 1H and 13C NMR, and MS methods revealed that the O-polysaccharide produced by C. sakazakii (3290), a clinical strain from the Tennessee outbreak, was a branched polymer of repeating pentasaccharide units composed of 2-acetamido-2-deoxy-D-galactose, 3-(N-acetyl-L-alanylamido)-3-deoxy-D-quinovose, D-glucuronic acid, and D-glucose present in the molar ratio 1:1:1:2 and had the structure:

Enterobacter sakazakii enhances epithelial cell injury by inducing apoptosis in a rat model of necrotizing enterocolitis.

Necrotizing enterocolitis (NEC) is an inflammatory intestinal disorder that affects 2%-5% of all premature infants. Enterobacter sakazakii, a common contaminant of milk-based powdered infant formula, has been implicated as a causative agent of sepsis, meningitis, and NEC in newborn infants, with high mortality rates. However, the role played by E. sakazakii in the pathogenesis of NEC is, to date, not known. Here, we demonstrate for the first time that E. sakazakii can induce clinical and histological NEC in newborn rats. E. sakazakii was found to bind to enterocytes in rat pups at the tips of villi and to intestinal epithelial cells (IEC-6) in culture, with no significant invasion. Exposure to E. sakazakii induced apoptosis and increased the production of interleukin-6 in IEC-6 cells and in the animal model. These data suggest that E. sakazakii could be a potential pathogen that induces NEC and triggers intestinal disease by modulating enterocyte intracellular signaling pathways.

Preparación y manejo de las fórmulas infantiles en polvo. Reflexiones en torno a las recomendaciones del Comité de Nutrición de la ESPGHAN / Preparation and handling of powdered infant formulae: comments on the recommendations of the European Society for Paediatric Gastroenterology, Hepatology and Nutrition Committee on Nutrition

Las fórmulas infantiles en polvo no son productos estériles. Se han descrito infecciones graves en recién nacidos y lactantes vulnerables relacionadas con la contaminación de las fórmulas infantiles en polvo. De entre todos los gérmenes contaminantes, adquiere especial relevancia el Enterobacter sakazakii. Por este motivo, la OMS/FAO y la Sociedad Europea de Gastroenterología, Hepatología y Nutrición Pediátricas han elaborado una serie de recomendaciones sobre la preparación y la manipulación de estas fórmulas en polvo, dirigidas tanto a los fabricantes de fórmulas infantiles, como a las instituciones en las que se utilicen estos productos y a los padres que preparan los biberones en casa. Estas recomendaciones y algunas consideraciones sobre el uso de las fórmulas infantiles en polvo constituyen el objetivo de este trabajo de revisión

Powdered infant formulas are not sterile and can contain small numbers of bacteria. Serious infections, especially in newborns and infants at risk, have been attributed to the contamination of these products. Of all the contaminating microorganisms, Enterobacter sakazakii, a gram-negative rod-shaped bacterium, appears to pose particularly serious risks. For this reason, in recent years, the World Health Organization, the Food and Agriculture Organization and the European Society for Pediatric Gastroenterology, Hepatology and Nutrition (ESPGHAN) have become concerned with this problem and have drawn up a series of recommendations for the preparation and handling of powdered infant formulae, in the attempt to minimize the risk of infections. These recommendations are intended not only for the manufacturers of these products, but for the institutional and home settings, as well. The purpose of this review is to discuss these recommendations and a number of considerations concerning the use of powdered infant formulas