Molecular determinants underlying the production of siderophores in the human and animal pathogen Staphylococcus aureus and the contribution of siderophore production to the virulence of this bacterium have, until now, remained undefined. Here, we show that S. aureus strains RN6390 and Newman produce siderophore when the cells are starved for iron. We further identified and characterized a nine-gene, iron-regulated operon, designated sbn and situated between sirABC and galE on the S. aureus chromosome, that is involved in the production of a siderophore. Mutation of the sbnE gene, in both RN6390 and Newman, eliminates the ability of these strains to produce a siderophore under iron-limited growth conditions, while introduction of multicopy sbnE into sbnE mutants complemented the inability of the mutants to produce the siderophore. sbnE mutants, in both the RN6390 and Newman backgrounds, displayed a drastic growth deficiency, compared to the wild type, in iron-restricted growth medium, whereas no such deficiency was observed during growth in iron-replete medium. Complemented mutants showed a restored ability to grow under iron restriction. We further showed that an sbnE mutant was compromised in a murine kidney abscess model of S. aureus infection, illustrating the importance of siderophore production to the pathogenicity of S. aureus. sbn genes were present in all S. aureus strains tested (and all S. aureus genome sequences) but were undetectable in any of the 13 coagulase-negative staphylococci tested, including Staphylococcus epidermidis.

Staphylococcus aureus SirA was previously identified as a lipoprotein, and SirB and SirC are thought to encode the transmembrane domains of an ABC transporter. Sir proteins show similarity to iron-siderophore transporters in several bacteria. Here, we show that the iron-regulated sirABC operon is divergently transcribed from the sbn operon that encodes enzymes involved in the synthesis of staphylobactin, a recently described siderophore produced by S. aureus. Mutation of either sirA or sirB increased the resistance of iron-starved S. aureus to streptonigrin and resulted in compromised growth in iron-restricted, but not ironrich, media. We also demonstrated that sirA and sirB mutants are compromised in the ability to transport iron complexed to staphylobactin but are not compromised for uptake of other iron complexes, such as ferric hydroxamates, ferric enterobactin, or ferric citrate. SirA-and SirB-deficient S. aureus, however, retain the ability to produce staphylobactin. Moreover, we found that transcription from the sbn operon was increased, relative to the wild type, in both sirA and sirB knockout strains, likely in response to an increased level of iron starvation in these cells. These results provide evidence of a role for these proteins in iron import in S. aureus and for full fitness of the bacterium in iron-restricted environments and demonstrate a function for S. aureus genes encoding proteins involved in the transport of an endogenously produced siderophore.The ability of bacterial pathogens to acquire iron from host iron-binding glycoproteins, such as transferrin and lactoferrin, is an important attribute that aids in the establishment of many bacterial infections (25,33,34). To access these extracellular iron stores, many bacteria produce small organic molecules called siderophores that have a high affinity for ferric iron (35). Iron-siderophore complexes (ferrisiderophores) are recognized and transported into the bacterial cytoplasm by specific receptor proteins and associated transport systems expressed at the cell surface (11). In gram-negative bacteria, these transport systems include high-affinity outer membrane receptor proteins that capture ferrisiderophores and shuttle them across the outer membrane (4, 9). Once in the periplasm, ferrisiderophores are bound by periplasmic binding proteins (16,31) that direct the ligand to membrane-associated ATP-binding cassette (ABC) transporters (3,19,23). In gram-positive bacteria, ferrisiderophores are initially recognized and bound by lipoproteins, tethered at the external face of the cytoplasmic membrane, that direct the ligand to ABC transporters. One ferrisiderophore import system in gram-positive bacteria that has been studied in our laboratory is the ferric hydroxamate uptake (fhu) system. In Staphylococcus aureus, the fhu system is comprised of FhuC (ATPase), FhuB and FhuG (together they form a membrane-embedded permease), and the lipoproteins FhuD1 and FhuD2 (high-affinity receptors), and together, these proteins function to scavenge hydroxamate ...

In Staphylococcus aureus, fhuCBG encodes an ATP-binding cassette (ABC) transporter that is required for the transport of iron(III)-hydroxamates; mutation of either fhuB or fhuG eliminates transport. In this paper, we describe construction and characterization of an S. aureus fhuCBG deletion strain. The ⌬fhuCBG::ermC mutation not only resulted in a strain that was incapable of growth on iron(III)-hydroxamates as a sole source of iron but also resulted in a strain which had a profound growth defect in iron-restricted laboratory media. The growth defect was not a result of the inability to transport iron(III)-hydroxamates since S. aureus fhuG::Tn917 and S. aureus fhuD1::Km fhuD2::Tet mutants, which are also unable to transport iron(III)-hydroxamates, do not have similar iron-restricted growth defects. Complementation experiments demonstrated that the growth defect of the ⌬fhuCBG::ermC mutant was the result of the inability to express FhuC and that this was the result of an inability to transport iron complexed to the S. aureus siderophore staphylobactin. Iron is an essential micronutrient for virtually all microorganisms owing to its wide range of redox potentials. Either alone or incorporated into heme or iron-sulfur clusters, iron serves as the catalytic center of enzymes involved in critical cellular processes such as DNA synthesis and electron transport. However, despite the fact that iron is plentiful on Earth, the amount of free iron in biological systems is low due to its tendency to form insoluble oxyhydroxides under aerobic conditions at a neutral pH. The amount of iron available in the host environment is further reduced by sequestration into proteins such as transferrin and lactoferrin; virtually no free iron exists in living organisms (7). Transport of iron(III)-staphylobactin is dependent uponIn order to overcome iron limitation, most successful pathogens employ several strategies to acquire the quantities of iron necessary to cause infection (49). One such strategy is production of low-molecular-weight iron-chelating molecules termed siderophores which, together with their cognate cell surface binding proteins and transporters, provide an efficient system for iron acquisition (for a recent review, see reference 25). Indeed, there is considerable evidence demonstrating the importance of siderophore-mediated iron acquisition systems for the virulence of many disease-causing bacteria, including gram-negative bacteria such as Escherichia coli (50) and Yersinia enterocolitica (22) and gram-positive bacteria such as Staphylococcus aureus (14) and Bacillus anthracis (9).S. aureus is an important human pathogen in both community and hospital settings; it is a leading cause of nosocomially acquired infection. This organism is capable of causing infections ranging from minor (e.g., impetigo and food poisoning) to more severe (e.g., bacteremia, necrotizing pneumonia, endocarditis, and osteomyelitis) (13). Moreover, S. aureus is a major threat to the health care system since it is becoming increasingly resistant ...

https://emergency.cdc.gov/han/2022/pdf/CDC_HAN_462.pdf † Since April 2022, providers have been encouraged to report to public health authorities persons under investigation for acute hepatitis meeting the following definition: children aged <10 years with elevated aspartate aminotransferase or alanine aminotransferase levels (>500 U/L) with an unknown etiology for their hepatitis since October 1, 2021.

Heteroresistance is a form of antibiotic resistance where a bacterial strain is comprised of a minor resistant subpopulation and a majority susceptible subpopulation. We showed previously that colistin heteroresistance can mediate the failure of colistin therapy in an in vivo infection model, even for isolates designated susceptible by clinical diagnostics. We sought to characterize the extent of colistin heteroresistance among the highly drug-resistant carbapenem-resistant Enterobacterales (CRE). We screened 408 isolates for colistin heteroresistance. These isolates were collected between 2012 and 2015 in eight U.S. states as part of active surveillance for CRE. Colistin heteroresistance was detected in 10.1% (41/408) of isolates, and it was more common than conventional homogenous resistance (7.1%, 29/408). Most (93.2%, 38/41) of these heteroresistant isolates were classified as colistin susceptible by standard clinical diagnostic testing. The frequency of colistin heteroresistance was greatest in 2015, the last year of the study. This was especially true among Enterobacter isolates, of which specific species had the highest rates of heteroresistance. Among Klebsiella pneumoniae isolates, which were the majority of isolates tested, there was a closely related cluster of colistin-heteroresistant ST-258 isolates found mostly in Georgia. However, cladistic analysis revealed that, overall, there was significant diversity in the genetic backgrounds of heteroresistant K. pneumoniae isolates. These findings suggest that due to being largely undetected in the clinic, colistin heteroresistance among CRE is underappreciated in the United States. IMPORTANCE Heteroresistance is an underappreciated phenomenon that may be the cause of some unexplained antibiotic treatment failures. Misclassification of heteroresistant isolates as susceptible may lead to inappropriate therapy. Heteroresistance to colistin was more common than conventional resistance and was overwhelmingly misclassified as susceptibility by clinical diagnostic testing. Higher proportions of colistin heteroresistance observed in certain Enterobacter species and clustering among heteroresistant Klebsiella pneumoniae strains may inform colistin treatment recommendations. Overall, the rate of colistin nonsusceptibility was more than double the level detected by clinical diagnostics, suggesting that the prevalence of colistin nonsusceptibility among CRE may be higher than currently appreciated in the United States.