Heterogeneous structure of O-antigenic part of lipopolysaccharide of <Emphasis Type="Italic">Salmonella</Emphasis> telaviv (Serogroup O:28) containing 3-acetamido-3,6-dideoxy-D-glucopyranose

The O-polysaccharide of Salmonella Telaviv was obtained by mild acid degradation of the lipopolysaccharide and studied by chemical methods (sugar and methylation analyses, Smith degradation, de-O-acetylation) and NMR spectroscopy. The structure of the O-polysaccharide was established. The repeating units that are proximal to the lipopolysaccharide core region mostly have a digalactose side chain and lack glucose, whereas those at the other end of the chain mostly do bear glucose but are devoid of the disaccharide side chain. This is the first structure established for the O-polysaccharide of a Salmonella serogroup O:28 (formerly M) strain characterized by subfactors O28₁ and O28₂. Knowledge of this structure and the structure of the O-polysaccharide of Salmonella Dakar (O28₁, O28₃) established earlier is crucial for determination of the exact structures associated with subfactors O28₁, O28₂, and O28₃ and elucidation of the genetic basis of the close relationship between Escherichia coli O71 and S. enterica O:28 O-antigens.     

Simultaneous detection of Salmonella spp and Escherichia coli O157:H7 by multiplex PCR

Contamination of foods with pathogens such as Escherichia coli O157:H7 and Salmonella is a major concern worldwide and rapid, sensitive, and reliable methods are needed for detection of these organisms. Since these pathogens can contaminate similar foods and other types of samples, a multiplex polymerase chain reduction (PCR) was designed to allow simultaneous detection of both E. coli O157:H7 and Salmonella spp directly from enrichment cultures. Samples of apple cider, beef carcass wash water, ground beef, and bovine feces were inoculated with both E. coli O157:H7 and S. typhimurium at various bacterial levels. Following enrichment culturing for 20–24 h at 37°C in modified EC broth or buffered peptone water both containing novobiocin, the samples were subjected to a DNA extraction technique or to immunomagnetic separation then tested by the multiplex PCR assay. Four pairs of primers were employed in the PCR: primers for amplification of E. coli O157:H7 eaeA, stx _1/2 and plasmid sequences and for amplification of a portion of the Salmonella invA gene. Four fragments of the expected sizes were amplified in a single reaction and visualized following agarose gel electrophoresis in all the samples inoculated with ≤ 1 CFU g⁻¹ or ml⁻¹. Results can be obtained in approximately 30 h. The multiplex PCR is a potentially powerful technique for rapid and sensitive co-detection of both pathogens in foods and other types of samples. Received 28 December 1997/ Accepted in revised form 19 March 1998     

Murine Monoclonal Antibodies Specific for Lipopolysaccharide of Escherichia coli O26 and O111

Monoclonal antibody (MAb) 12F5 reacted with 35 Escherichia coli O26 isolates and cross-reacted with 1 of 365 non-E. coli O26 isolates. MAb 15C4 reacted with 30 E. coli O111 strains and 8 Salmonella O35 strains (possessing identical O antigen) but not with 362 other bacterial strains. Lipopolysaccharide immunoblots confirmed MAb O-antigen specificity.     

Effects of Ractopamine HCl on Escherichia coli O157:H7 and Salmonella In Vitro and on Intestinal Populations and Fecal Shedding in Experimentally Infected Sheep and Pigs

The effects of the β-agonist ractopamine, approved for use in finishing swine and cattle to improve carcass quality and performance, were examined on two important foodborne pathogens, Escherichia coli O157:H7 and Salmonella. Ractopamine, administered to sheep before and after oral inoculation with E. coli O157:H7, increased (P < 0.01) fecal shedding and tended to increase (P = 0.08) cecal populations of the challenge strain. Pigs receiving ractopamine in the diet and then experimentally infected with Salmonella Typhimurium, had decreased (P < 0.05) fecal shedding and fewer (P = 0.05) liver samples positive for the challenge strain of Salmonella. Pure cultures of E. coli O157:H7 (used in the present sheep study), E. coli O157:H19 (isolated from pigs with postweaning diarrhea), Salmonella Typhimurium (used in the present pig study), and Salmonella Choleraesuis were incubated with varying concentrations of ractopamine to determine if ractopamine has a direct effect on bacterial growth. No differences in growth rate were observed for either strain of E. coli or for Salmonella Typhimurium when incubated with increasing concentrations of ractopamine. The growth rate for Salmonella Choleraesuis was increased with the addition of 2.0 μg ractopamine/ml compared with the other concentrations examined. Collectively, these results indicate that ractopamine may influence gut populations and fecal shedding of E. coli O157:H7 and Salmonella. Because ractopamine is currently approved to be fed to finishing cattle and swine immediately before slaughter, any potential for decreasing foodborne pathogens has exciting food safety implications. Mention of trade names, proprietary products, or specific equipment does not constitute a guarantee or warranty by the United States Department of Agriculture and does not imply its approval to the exclusion of other products that may be suitable.     

Effect of Ractopamine HCl Supplementation on Fecal Shedding of Escherichia coli O157:H7 and Salmonella in Feedlot Cattle

The effects of the β-agonist ractopamine, recently approved for use in feedlot cattle to improve carcass quality and performance, on fecal shedding Escherichia coli O157:H7 and Salmonella in feedlot cattle was examined. In the first study, 20 feedlot steers and heifers were randomly assigned to receive ractopamine or no ractopamine (control) by way of oral bolus for 28 days. Fecal samples were collected daily, and shedding of E. coli O157:H7 determined. When examined during the entire 28-day experimental period, ractopamine decreased (P = 0.0006) the percentage of cattle shedding E. coli O157:H7 (58% vs. 42% for control and ractopamine treatments, respectively). A second study was conducted in a commercial feedlot facility in the southwestern United States. Eighteen pens of cross-bred beef heifers (approximately 100 head/pen and 9 pens/treatment) were randomly assigned to receive either 0 (control) or 200 mg ractopamine/head·d^–1. Fresh fecal samples (30/pen) were collected off the pen floor before ractopamine supplementation and again after approximately 28 days of ractopamine supplementation (within a few days of slaughter); the samples were cultured for E. coli O157:H7 and Salmonella. The percentage of animals shedding E. coli O157:H7 was decreased when data were pooled across replicates (P = 0.05) in ractopamine-treated cattle compared with controls. The percentage of animals shedding Salmonella tended to be higher (P = 0.08) with the ractopamine treatment when data were pooled across replicates. Although further research is required to confirm these results, the potential food safety implications of this research are intriguing. Mention of trade name, proprietary product, or specific equipment does not constitute a guarantee or warranty by the United States Drug Administration and does not imply its approval to the exclusion of other products that may be suitable.     

Molecular Analysis of the rfb O Antigen Gene Cluster of Salmonella enterica Serogroup O:6,14 and Development of a Serogroup-Specific PCR Assay

The Kauffmann-White scheme for serotyping Salmonella recognizes 46 somatic (O) antigen groups, which together with detection of the flagellar (H) antigens form the basis for serotype identification. Although serotyping has become an invaluable typing method for epidemiological investigations of Salmonella, it does have some practical limitations. We have been characterizing the genes required for O and H antigen biosynthesis with the goal of developing a DNA-based system for the determination of serotype in Salmonella. The majority of the enzymes involved in O antigen biosynthesis are encoded by the rfb gene cluster. We report the sequencing of the rfb region from S. enterica serotype Sundsvall (serogroup O:6,14). The S. enterica serotype Sundsvall rfb region is 8.4 kb in length and comprises six open reading frames. When compared with other previously characterized rfb regions, the serogroup O:6,14 sequence is most related to serogroup C₁. On the basis of DNA sequence similarity, we identified two genes from the mannose biosynthetic pathway, two mannosyl transferase genes, the O unit flippase gene and, possibly, the O antigen polymerase. The whole cluster is derived from a low-G+C-content organism. Comparative sequencing of an additional serogroup O:6,14 isolate (S. enterica serotype Carrau) revealed a highly homologous sequence, suggesting that O antigen factors O:24 and O:25 (additional O factors associated with serogroup O:6,14) are encoded outside the rfb gene cluster. We developed a serogroup O:6,14-specific PCR assay based on a region of the putative wzx (O antigen flippase) gene. This provides the basis for a sensitive and specific test for the rapid identification of Salmonella serogroup O:6,14.     

Prevalence of verotoxigenic Escherichia coli O157 (VTEC O157) and compliance with microbiological safety standards in bovine carcasses from an industrial beef slaughter plant

The presence of Salmonella spp. and levels of Enterobacteriaceae and aerobic plate count were determined in 300 bovine carcasses randomly collected in an industrial cattle slaughterhouse in Catalonia (Spain) as part of a control programme to validate good slaughter practices according to Commission Regulation No 2073/2005. The verotoxigenic Escherichia coli O157 (VTEC O157), although not currently legislated, was also investigated in the same carcasses due to the importance of bovines as a reservoir for this micro‐organism. Virulence genes (vtx1, vtx2 and eae), the presence of fliC_H7 and antimicrobial susceptibility were studied in E. coli O157 isolates. Levels of Enterobacteriaceae and aerobic colonies and the presence of Salmonella were within the admissible range stipulated by current legislation. However, VTEC O157 was detected in 14·7% of carcasses. Among the VTEC O157 strains tested for antimicrobial susceptibility, 65% were multiresistant. Overall, the results of this study indicate that even with good manufacturing practices, contamination with VTEC O157 can occur and cattle meat can pose a risk to human health. These results confirm the need for a review of the appropriateness of introducing antimicrobial treatments in the processing of cattle carcasses in Europe.

Significance and Impact of the Study

This study describes the prevalence of verotoxigenic and multidrug‐resistant E. coli O157 strains in bovine carcasses. These results suggest that despite the good manufacturing practices used in the slaughterhouse studied (the largest in Catalonia slaughtering over 81 000 cattle per year), the absence of verotoxigenic E. coli O157 in bovine carcasses cannot be guaranteed.     

The growth potential of Escherichia coli O157:H7, Salmonella spp. and Listeria monocytogenes in dairy manure‐based compost in a greenhouse setting under different seasons

Aim: The pathogen growth in dairy compost was studied in a greenhouse setting under different seasons. Methods and Results: The five‐strain mixtures of each Escherichia coli O157:H7, Salmonella spp. and Listeria monocytogenes were inoculated separately into dry compost to yield c. 1 log CFU g^?1. After acclimation at room temperature, the inoculated compost was initially adjusted to moisture levels of 10–50% and then kept in a greenhouse under different seasons. The populations of all three pathogens increased by 2·1–3·9 log CFU g^?1 within 3 days in autoclaved compost with initial moisture content of at least 40%. Listeria monocytogenes multiplied up to 2·4 log CFU g^?1 in compost with initial moisture content of 30% and was detected up to 28 days for all seasons, whereas populations of both E. coli O157:H7 and Salmonella increased by c. 1 log in compost with initial moisture content of 30% during winter months only. No pathogen growth in nonautoclaved compost was detected. Conclusion: Bacterial species, temperature, light intensity and moisture content affected the growth potential and survival of pathogens in compost when the population of background microflora was low. Significance and Impact of the Study: Keeping compost as dry as possible and maintaining certain levels of background microflora may be critical to prevent the growth of pathogens.     

Structure of an abequose-containing O-polysaccharide from Citrobacter freundii O22 strain PCM 1555

The lipopolysaccharide of Citrobacter freundii O22 (strain PCM 1555) was degraded under mild acidic conditions and the O-polysaccharide released was isolated by gel chromatography. Sugar and methylation analyses along with ¹H and ¹³C NMR spectroscopy, including two-dimensional ¹H,¹H ROESY and ¹H,¹³C HMBC experiments, showed that the repeating unit of the O-polysaccharide has the following structure:

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where Abe is abequose (3,6-dideoxy-d-xylo-hexose). SDS–PAGE and immunoblotting revealed that the O-antigen of C. freundii O22 is serologically indistinguishable from those of Salmonella group B serovars (Typhimurium, Brandenburg, Sandiego, Paratyphi B) but not related to other abequose-containing O-antigens tested (Citrobacter werkmanii O38 and Salmonella Kentucky) or colitose (l enantiomer of abequose)-containing O-antigen of Escherichia coli O111.     

Fate of manure-borne pathogen surrogates in static composting piles of chicken litter and peanut hulls

The fate of manure-borne pathogen surrogates (gfp-labeled Escherichia coli O157:H7 and Listeria innocua and avirulent Salmonella Typhimurium) in the field was monitored at both sub-surface (30 cm from surface) and surface sites of static composting piles (3.5-m base diameter) composed of chicken litter and peanut hulls. Despite exposure to elevated temperatures, Salmonella was detected by enrichment culture in sub-surface samples following 14 days of composting. In surface samples, pathogen surrogates were detected in the summer after 4 days of composting by enrichment culture only, whereas E. coli O157:H7 and L. innocua remained detectable by direct plating (>2log₁₀ cfu/g) up to 28 days in piles composted during the fall and winter. All three types of bacteria remained detectable by enrichment culture in surface samples composted for 56 days during the winter.     

Amplification of rfbE and fliC Genes by Polymerase Chain Reaction for Identification and Detection of Salmonella Serovar Enteritidis,Dublin and Gallinarum-Pullorum

Polymerase chain reaction (PCR) primers for O9 antigen (rfbE) and phase 1 flagellin antigen (fliC) were designed for the rapid identification and detection of Salmonella serovar Enteritidis and Dublin. The rfbE primer pairs selectively amplified the rfbE region of group O9 Salmonella serovars. The fliC primer pairs amplified the DNAs of g,m and g,p-type flagellar antigen; Salmonella serovar Enteritidis, Dublin, and Essen. However, DNA from flagellar-negative Salmonella serovar Gallinarum-Pullorum was also amplified. The sensitivity of PCR primer pairs was 10 CFU/assay by boiled DNA preparation and 10² CFU/assay by proteinase K-treated DNA preparation.     

Ultrasensitive chemiluminescent immunoassay of Salmonella with silver enhancement of nanogold labels

In this paper, silver enhancement of nanogold labels coupled with chemiluminescence detection was developed for ultrasensitive immunoassay of Salmonella based upon antigen–antibody immunoreaction. Polyclonal rabbit anti‐Salmonella sp. antibodies (pAb) were employed to establish the analytical protocol. The pAb coated onto ELISA microwell plates and Au nanoparticles (Au NPs) conjugated pAb capture target Salmonella to form a sandwich‐type complex. Silver then was in situ deposited around the Au NPs core and resulted in the signal amplification. In consequence, silver was dissolved to form Ag⁺ and a sensitive chemiluminescence based on the Ag⁺–K₂S₂O₈–Mn²⁺–luminol system was coupled for further signal amplification. Under the optimized conditions, the chemiluminescent intensity is proportional to target Salmonella over the range of 5–1038 cfu mL^?1 with a detection limit of 5 cfu mL^?1. The relative standard deviation for 11 measurements of about 50–100 cfu/mL target Salmonella is 4.7%. The proposed method was successfully applied to measure Salmonella in food samples and the results are identical to those of the offical standard method of China. These offer us a more powerful tool for ultrasensitive assay of foodborne pathogens. Copyright © 2010 John Wiley & Son, Ltd.     

Sequence Analysis of the rfb Loci, Encoding Proteins Involved in the Biosynthesis of the Salmonella enterica O17 and O18 Antigens: Serogroup-Specific Identification by PCR

We report sequencing of the O antigen encoded by the rfb gene cluster of Salmonella enterica serotype Jangwani (O17) and Salmonella serotype Cerro (O18). We developed serogroup O17- and O18-specific PCR assays based on rfb gene targets and found them to be sensitive and specific for rapid identification of Salmonella serogroups O17 and O18.     

Physical Covering for Control of Escherichia coli O157:H7 and Salmonella spp. in Static and Windrow Composting Processes

This study investigated the effect of a 30-cm covering of finished compost (FC) on survival of Escherichia coli O157:H7 and Salmonella spp. in active static and windrow composting systems. Feedstocks inoculated with E. coli O157:H7 (7.41 log CFU/g) and Salmonella (6.46 log CFU/g) were placed in biosentry tubes (7.5-cm diameter, 30-cm height) at three locations: (i and ii) two opposing sides at the interface between the FC cover layer (where present) and the feedstock material (each positioned approximately 10 cm below the pile''s surface) and (iii) an internal location (top) (approximately 30 cm below the surface). On specific sampling days, surviving populations of inoculated E. coli O157:H7 and Salmonella, generic E. coli, and coliforms in compost samples were determined. Salmonella spp. were reduced significantly within 24 h in windrow piles and were below the detection limit after 3 and 7 days at internal locations of windrow and static piles containing FC covering, respectively. Likewise, E. coli O157:H7 was undetectable after 1 day in windrow piles covered with finished compost. Use of FC as a covering layer significantly increased the number of days that temperatures in the windrows remained ≥55°C at all locations and in static piles at internal locations. These time-temperature exposures resulted in rapid reduction of inoculated pathogens, and the rate of bacterial reduction was rapid in windrow piles. The sample location significantly influenced the survival of these pathogens at internal locations compared to that at interface locations of piles. Finished compost covering of compost piles aids in the reduction of pathogens during the composting process.     

Structural variation in the O-specific polysaccharides of Klebsiella pneumoniae serotype O1 and O8 lipopolysaccharide: evidence for clonal diversity in rfb genes

The O-polysaccharide fraction of the lipopolysaccharide from Klebsiella pneumoniae serotype O8 was found to comprise two galactose-containing homopolymers. Structural analysis, using chemical and high-field nuclear magnetic resonance (NMR) techniques, established that the K. pneumoniae O8 polysaccharides are composed of the linear, disaccharide repeating units OAc 1 2/6 →3)-β-d -Galf-(1 →3)-α- d -Galp-(1→d -Galactan I-OAc →3)-α-d -Galp-(1 →3)-β-d -Galp-(1→d -Galactan II. K. pneumoniae O8 mutant RFK-1 was isolated by resistance to phage KO1-2; strain RFK-1 expressed only d -galactan I-OAc. The ¹H- and ¹³C-NMR resonances from this O-polysaccharide indicate that all of the O-acetyl groups within the K. pneumoniae O8 polysaccharide are carried on d -galactan I and O-acetylation occurs only on the β- d -galactofuranose residues; 60% of the available β- d -galactofuranose residues are non-acetylated. The O-acetylation of the remaining residues is equally distributed between the O-2 and O-6 positions. The carbohydrate backbone structures in the O8 polysaccharide are identical to d -galactan I and II expressed by K. pneumoniae O1, accounting for the antigenic cross-reaction between strains belonging to serotypes O1 and O8. However, the O1 polysaccharides are not acetylated and the O-acetyl groups present in the K. pneumoniae serotype O8 polysaccharides provide a structural basis for their recognition as distinct serotypes. The rfb (O-polysaccharide biosynthesis) gene cluster of K. pneumoniae serotype O1 determines the synthesis of d -galactan I. rfb_Kpo1-specific gene probes were used to examine conservation in the rfb gene clusters of other K. pneumoniae serotypes which produce d -galactan I. Six O1 strains were examined and all showed hybridization with rfb_KpO1 probes under conditions of high stringency. Three serotype O2 strains produce d -galactan I and these strains also contained DNA sequences recognized by rfb_KpO1 probes under high stringency. The physical maps of these homologous rfb chromosomal regions showed some polymorphism. Surprisingly, the rfb_KpO8 region from K. pneumoniae serotype O8 was only recognized by rfb_KpO1 probes under low-stringency hybridization conditions, providing evidence for two substantially different clonal groups of rfb genes from K. pneumoniae strains with structurally related O-antigens.     

Enterotoxigenicity and invasiveness of Salmonella species

A large number of enterotoxigenic strains was encountered in a group 56 Salmonella cultures belonging to 8 species viz., S. alachua, S. anatum, S. dublin, S. enteritidis, S. hindmarsh, S. newport, S. typhimurium, S. weltevreden, and 5 serotypes of S. arizona (16:z₄:-; 48:1,v:z₅₆; 53:z₅₂:z₅₃; 60:r:z; 61:i:z₅₃). These cultures were isolated mainly from humans and animals suffering from gasteroenteritis. The enterotoxigenic (diarrhoeagenic) Salmonella cultures possess capacities for both skin permeation and epithelial penetration (invasiveness). Preliminary characterization revealed that Salmonella enterotoxin is a heatlabile protein of high molecular weight. It is suggested that enterotoxigenic and invasive propeties play a vital role in the pathogenesis of Salmonella diarrhoea.     

The structural characterization of the O-polysaccharide antigen of the lipopolysaccharide of Escherichiacoli serotype O118 and its relation to the O-antigens of Escherichiacoli O151 and Salmonellaenterica O47

Mild acid hydrolysis of the lipopolysaccharide produced by Escherichiacoli O118:H16 standard strain (NRCC 6613) afforded an O-polysaccharide (O-PS) composed of d-galactose, 2-acetamidoylamino-2,6-dideoxy-l-galactose , 2-acetamido-2-deoxy-d-glucose, ribitol, and phosphate (1:1:1:1:1). From DOC-PAGE, sugar and methylation analyses, one- and two-dimensional NMR spectroscopy, capillary electrophoresis-mass spectrometry, hydrolysis, and sequential Smith-type periodate oxidation studies, the O-PS was determined to be an unbranched linear polymer having the structure:[6)-α-d-Galp-(1→3)-α-l-FucpNAm-(1→3)-β-d-GlcpNAc-(1→3)-Rib-ol-5-P-(O→]_nThe structure of the O-PS is consistent with the reported DNA data on the O-antigen gene-cluster of E. coli O118 and interestingly, the O-PS is similar to the structures of the O-antigens of Salmonellaenterica O47 and E. coli O151:H10 reference strain 880-67, as predicted from the results of DNA sequencing of their respective O-antigen gene-clusters.     

Examination of indigenous microbiota and survival of Escherichia coli 0157 and Salmonella in a paper milling environment

Aims: A public beach was frequently cited for health advisories because of high Escherichia coli levels, the source suspected to be a paper mill located upstream. This investigation sought to confirm whether or not the paper mill was the pollution source, and to characterize the risk to recreational bathers imposed by the source. Methods and Results: Quantification of E. coli in river water collected at incremental distances showed that paper mill effluent caused elevated E. coli levels in beach samples. Samples collected throughout the mill were variably positive for heterotrophic bacteria, total coliforms and E. coli, but negative for pathogenic E. coli O157 and Salmonella. Escherichia coli O157 or Salmonella spiked into mill samples (4·2 log₁₀ or 5·6 log₁₀ CFU per 100 ml, respectively) fell below detection levels within 14–24 h in raw (unaltered) samples, while in heat‐sterilized replicates, the counts remained at initial levels or increased over 36 h. Conclusions: Pathogenic E. coli O157 and Salmonella were not isolated from paper mill samples. The absence of native bacteria allowed the survival of pathogens, while their presence accelerated pathogen decline. Significance and Impact of the Study: The co‐existence of paper mill and swimming beach may be reasonable for now in spite of the limitations of an E. coli‐based assay for beach water.     

Conflicting roles for a cell surface modification in Salmonella

Chemical modifications of components of the bacterial cell envelope can enhance resistance to antimicrobial agents. Why then are such modifications produced only under specific conditions? Here, we address this question by examining the role of regulated variations in O‐antigen length in the lipopolysaccharide (LPS), a glycolipid that forms most of the outer leaflet of the outer membrane in Gram‐negative bacteria. We determined that activation of the PmrA/PmrB two‐component system, which is the major regulator of LPS alterations in Salmonella enterica serovar Typhimurium, impaired growth of Salmonella in bile. This growth defect required the PmrA‐activated gene wzz_st, which encodes the protein that determines long O‐antigen chain length and confers resistance to complement‐mediated killing. By contrast, this growth defect did not require the wzz_fepE gene, which controls production of very long O‐antigen, or other PmrA‐activated genes that mediate modifications of lipid A or core regions of the LPS. Additionally, we establish that long O‐antigen inhibits growth in bile only in the presence of enterobacterial common antigen, an outer‐membrane glycolipid that contributes to bile resistance. Our results suggest that Salmonella regulates the proportion of long O‐antigen in its LPS to respond to the different conditions it faces during infection.     

Salmonella exploits host Rho GTPase signalling pathways through the phosphatase activity of SopB

Salmonella uses Type 3 secretion systems (T3SSs) to deliver virulence factors, called effectors, into host cells during infection. The T3SS effectors promote invasion into host cells and the generation of a replicative niche. SopB is a T3SS effector that plays an important role in Salmonella pathogenesis through its lipid phosphatase activity. Here, we show that SopB mediates the recruitment of Rho GTPases (RhoB, RhoD, RhoH, and RhoJ) to bacterial invasion sites. RhoJ contributes to Salmonella invasion, and RhoB and RhoH play an important role in Akt activation. R‐Ras1 also contributes to SopB‐dependent Akt activation by promoting the localised production of PI(3,4)P₂/PI(3,4,5)P₃. Our studies reveal new signalling factors involved in SopB‐dependent Salmonella infection.