High-resolution microbiome analysis reveals exclusionary Klebsiella species competition in preterm infants at risk for necrotizing enterocolitis.

Intestinal colonization with Klebsiella has been linked to necrotizing enterocolitis (NEC), but methods of analysis usually failed to discriminate Klebsiella species or strains. A novel ~ 2500-base amplicon (StrainID) that spans the 16S and 23S rRNA genes was used to generate amplicon sequence variant (ASV) fingerprints for Klebsiella oxytoca and Klebsiella pneumoniae species complexes (KoSC and KpSC, respectively) and co-occurring fecal bacterial strains from 10 preterm infants with NEC and 20 matched controls. Complementary approaches were used to identify cytotoxin-producing isolates of KoSC.

Bacterial Indole as a Multifunctional Regulator of Klebsiella oxytoca Complex Enterotoxicity.

Gastrointestinal microbes respond to biochemical metabolites that coordinate their behaviors. Here, we demonstrate that bacterial indole functions as a multifactorial mitigator of Klebsiella grimontii and Klebsiella oxytoca pathogenicity. These closely related microbes produce the enterotoxins tilimycin and tilivalline; cytotoxin-producing strains are the causative agent of antibiotic-associated hemorrhagic colitis and have been associated with necrotizing enterocolitis of premature infants.

Cytotoxin-producing in the preterm gut and its association with necrotizing enterocolitis.

Necrotizing enterocolitis (NEC) is a devastating intestinal inflammatory disease of premature infants associated with gut bacterial dysbiosis. Using 16S rRNA-based methods, our laboratory identified an unclassified Enterobacteriaceae sequence (NEC_unk_OTU) with high abundance in NEC fecal samples. We aimed to identify this bacterium and determine its potential role in the disease.

Influence of iron and aeration on Staphylococcus aureus growth, metabolism, and transcription.

Staphylococcus aureus is a prominent nosocomial pathogen and a major cause of biomaterial-associated infections. The success of S. aureus as a pathogen is due in part to its ability to adapt to stressful environments.

Staphylococcal response to oxidative stress.

Staphylococci are a versatile genus of bacteria that are capable of causing acute and chronic infections in diverse host species. The success of staphylococci as pathogens is due in part to their ability to mitigate endogenous and exogenous oxidative and nitrosative stress. Endogenous oxidative stress is a consequence of life in an aerobic environment; whereas, exogenous oxidative and nitrosative stress are often due to the bacteria's interaction with host immune systems.

The Staphylococcus aureus two-component regulatory system, GraRS, senses and confers resistance to selected cationic antimicrobial peptides.

The two-component regulatory system, GraRS, appears to be involved in staphylococcal responses to cationic antimicrobial peptides (CAPs). However, the mechanism(s) by which GraRS is induced, regulated, and modulated remain undefined. In this study, we used two well-characterized MRSA strains (Mu50 and COL) and their respective mutants of graR and vraG (encoding the ABC transporter-dependent efflux pump immediately downstream of graRS), and show that (i) the expression of two key determinants of net positive surface charge (mprF and dlt) is dependent on the cotranscription of both graR and vraG, (ii) reduced expression of mprF and dlt in graR mutants was phenotypically associated with reduced surface-positive charge, (iii) this net reduction in surface-positive charge in graR and vraG mutants, in turn, correlated with enhanced killing by a range of CAPs of diverse structure and origin, including those from mammalian platelets (tPMPs) and neutrophils (hNP-1) and from bacteria (polymyxin B), and (iv) the synthesis and translocation of membrane lysyl-phosphatidylglycerol (an mprF-dependent function) was substantially lower in graR and vraG mutants than in parental strains.

CcpA coordinates central metabolism and biofilm formation in Staphylococcus epidermidis.

Staphylococcus epidermidis is an opportunistic bacterium whose infections often involve the formation of a biofilm on implanted biomaterials. In S. epidermidis, the exopolysaccharide facilitating bacterial adherence in a biofilm is polysaccharide intercellular adhesin (PIA), whose synthesis requires the enzymes encoded within the intercellular adhesin operon (icaADBC).

Fur is required for the activation of virulence gene expression through the induction of the sae regulatory system in Staphylococcus aureus.

Our previous studies showed that both Sae and Fur are required for the induction of eap and emp expression in low iron. In this study, we show that expression of sae is also iron-regulated, as sae expression is activated by Fur in low iron. We also demonstrate that both Fur and Sae are required for full induction of the oxidative stress response and expression of non-covalently bound surface proteins in low-iron growth conditions.

Transcriptomic response of Listeria monocytogenes to iron limitation and Fur mutation.

Iron is required by almost all bacteria, but concentrations above physiological levels are toxic. In bacteria, intracellular iron is regulated mostly by the ferric uptake regulator, Fur, or a similar functional protein. Iron limitation results in the regulation of a number of genes, especially those involved in iron uptake.

Staphylococcus aureus cell wall stress stimulon gene-lacZ fusion strains: potential for use in screening for cell wall-active antimicrobials.

lacZ fusion strains were constructed using the promoters of five cell wall stress stimulon genes: pbp2, tcaA, vraSR, sgtB, and lytR. All fusion strains were induced only in the presence of cell wall-active antibiotics, suggesting the potential of these strains for use in high-throughput screening for new cell wall-active agents.

Molecular characterization of the Fur protein of Listeria monocytogenes.

Iron is essential for the survival of almost all organisms, although excess iron can result in the generation of free radicals which are toxic to cells. To avoid the toxic effects of free radicals, the concentration of intracellular iron is generally regulated by the ferric uptake regulator Fur in bacteria. The 150 aa fur ORF from Listeria monocytogenes was cloned into pRSETa, and the His-tagged fusion protein was purified by nickel affinity column chromatography.

Effects of oxacillin and tetracycline on autolysis, autolysin processing and atl transcription in Staphylococcus aureus.

Autolysins are peptidoglycan hydrolases involved in cell growth and cell lysis. Atl is an important autolysin of Staphylococcus aureus and is essential for penicillin-induced autolysis. The objective of our study was to examine the effect of oxacillin, chloramphenicol and tetracycline on autolysis, peptidoglycan hydrolase profiles and transcription of atl encoding the major S.

Regulation of the expression of cell wall stress stimulon member gene msrA1 in methicillin-susceptible or -resistant Staphylococcus aureus.

Genome-wide transcriptional profiling studies of the response of Staphylococcus aureus to cell wall-active antibiotics have led to the discovery of a cell wall stress stimulon of genes induced by these agents. msrA1, encoding methionine sulfoxide reductase, is a highly induced member gene of this stimulon. In the present study we show that msrA1 induction by oxacillin is common to all methicillin-susceptible strains studied but did not occur in two homogeneous and two heterogeneous methicillin-resistant strains.