Enterococcus faecalis is among the predominant causes of nosocomial infections. Surface molecules like d -alanine lipoteichoic acid (LTA) perform several functions in gram-positive bacteria, such as maintenance of cationic homeostasis and modulation of autolytic activities. The aim of the present study was to evaluate the effect of d -alanine esters of teichoic acids on biofilm production and adhesion, autolysis, antimicrobial peptide sensitivity, and opsonic killing. A deletion mutant of the dltA gene was created in a clinical E. faecalis isolate. The absence of d -alanine in the LTA of the dltA deletion mutant was confirmed by nuclear magnetic resonance spectroscopy. The wild-type strain and the deletion mutant did not show any significant differences in growth curve, morphology, or autolysis. However, the mutant produced significantly less biofilm when grown in the presence of 1% glucose (51.1% compared to that of the wild type); adhesion to eukaryotic cells was diminished. The mutant absorbed 71.1% of the opsonic antibodies, while absorption with the wild type resulted in a 93.2% reduction in killing. Sensitivity to several cationic antimicrobial peptides (polymyxin B, colistin, and nisin) was considerably increased in the mutant strain, confirming similar results from other studies of gram-positive bacteria. Our data suggest that the absence of d -alanine in LTA plays a role in environmental interactions, probably by modulating the net negative charge of the bacterial cell surface, and therefore it may be involved in the pathogenesis of this organism.

Group B streptococcus (GBS) is the most important cause of neonatal sepsis, which is mediated in part by TLR2. However, GBS components that potently induce cytokines via TLR2 are largely unknown. We found that GBS strains of the same serotype differ in released factors that activate TLR2. Several lines of genetic and biochemical evidence indicated that lipoteichoic acid (LTA), the most widely studied TLR2 agonist in Gram-positive bacteria, was not essential for TLR2 activation. We thus examined the role of GBS lipoproteins in this process by inactivating two genes essential for bacterial lipoprotein (BLP) maturation: the prolipoprotein diacylglyceryl transferase gene (lgt) and the lipoprotein signal peptidase gene (lsp). We found that Lgt modification of the N-terminal sequence called lipobox was not critical for Lsp cleavage of BLPs. In the absence of lgt and lsp, lipoprotein signal peptides were processed by the type I signal peptidase. Importantly, both the Δlgt and the Δlsp mutant were impaired in TLR2 activation. In contrast to released factors, fixed Δlgt and Δlsp GBS cells exhibited normal inflammatory activity indicating that extracellular toxins and cell wall components activate phagocytes through independent pathways. In addition, the Δlgt mutant exhibited increased lethality in a model of neonatal GBS sepsis. Notably, LTA comprised little, if any, inflammatory potency when extracted from Δlgt GBS. In conclusion, mature BLPs, and not LTA, are the major TLR2 activating factors from GBS and significantly contribute to GBS sepsis.

Background. Recent population-based cohort studies have questioned the role of pneumococci as the most frequent pathogen causing severe infection in patients after splenectomy. The aim of the study was to define the causative pathogens and clinical presentation of patients with overwhelming postsplenectomy infection (OPSI). Methods. In a prospective cohort study in 173 German intensive care units, we searched for patients with and without asplenia and community-acquired severe sepsis/septic shock. Clinical and laboratory variables and survival of patients were assessed. Results. Fifty-two patients with severe sepsis or septic shock with asplenia and 52 without asplenia were included. OPSI patients more often had a history of malignancy (38% vs 17%; P = .016) and had a lower body mass index (24 kg/m² vs 28 kg/m²; P = .004). Streptococcus pneumoniae was detected more frequently in OPSI patients (42% vs 12% without asplenia; P < .001) and more frequently manifested as bloodstream infection (31% vs 6%; P = .002). Gram-negative infection was similar in both groups (12% vs 19%; P = .157). Pneumococcal vaccine coverage of OPSI patients was low overall (42% vs 8% among patients without asplenia; P < .001). Purpura fulminans was a frequent complication, developing in 19% of OPSI patients vs 5% of patients without asplenia (P = .038). The interval between splenectomy and OPSI was 6 years (range, 1 month–50 years). On multivariable Poisson regression, asplenia was the only predictive variable independently associated with pneumococcal sepsis (adjusted relative risk, 2.53 [95% confidence interval, 1.06–6.08]). Conclusions. Pneumococcal infections remain the most important cause of severe sepsis and septic shock following splenectomy.