Human Milk Oligosaccharides Exhibit Biofilm Eradication Activity Against Matured Biofilms Formed by Different Pathogen Species

Human milk oligosaccharides (HMOs) have been shown to exhibit plenty of benefits for infants, such as prebiotic activity shaping the gut microbiota and immunomodulatory and anti-inflammatory activity. For some pathogenic bacteria, antimicrobial activity has been proved, but most studies focus on group B streptococci. In the present study, we investigated the antimicrobial and antibiofilm activities of the total and fractionated HMOs from pooled human milk against four common human pathogenic Gram-negative species (Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Burkholderia cenocepacia) and three Gram-positive species (Staphylococcus aureus, Enterococcus faecium, and Enterococcus faecalis). The activity of HMOs against enterococci and B. cenocepacia are addressed here for the first time. We showed that HMOs exhibit a predominant activity against the Gram-positive species, with E. faecalis being the most sensitive to the HMOs, both in planktonic bacteria and in biofilms. In further tests, we could exclude fucosyllactose as the antibacterial component. The biological significance of these findings may lie in the prevention of skin infections of the mother’s breast as a consequence of breastfeeding-induced skin laceration and/or protection of the infants’ nasopharynx and lung from respiratory pathogens such as staphylococci.

Invasive Group B Streptococcal Disease in Neonates and Infants, Italy, Years 2015–2019

Invasive infections by group B streptococci (iGBS) are the leading cause of sepsis and meningitis in the first three months of life worldwide. The clinical and microbiological characteristics of neonatal and infant iGBS in Italy during the years 2015–2019 were investigated. Voluntary-based surveillance reported 191 cases (67 early-onset (EOD) and 124 late-onset disease (LOD)) and 89 bacterial isolates were received. The main clinical manifestations were sepsis (59.2%) followed by meningitis (21.5%), bacteremia (12.0%) and septic shock (6.3%). Hospitalized preterm babies accounted for one third of iGBS and constituted the most fragile population in terms of mortality (8.2%) and brain damage (16.4%). GBS serotype III was predominant in EOD (56%) and caused almost all LOD (95%). The rate of resistance to clindamycin reached 28.8%. Most of clindamycin-resistant GBS strains (76%) were serotype III-ST17 and possessed the genetic markers of the emerging multidrug resistant (MDR) CC-17 sub-clone. Our data revealed that iGBS is changing since it is increasingly reported as a healthcare-associated infection (22.6%), mainly caused by MDR-CC17. Continuous monitoring of the clinical and microbiological characteristics of iGBS remains of primary importance and it represents, at present, the most effective tool to support prevention strategies and the research on the developing GBS vaccine.

694. Prediction Tool for Infective Endocarditis in Beta-hemolytic Streptococcal Bacteremia

Background Although beta-hemolytic streptococci (BHS) is a rare causative pathogen of infective endocarditis (IE), IE is a serious condition and it is important to predict IE in BHS bacteremia (BHS-IE). The purpose of this study was to develop a predictive score for BHS-IE. Methods We conducted a retrospective study comparing the clinical features of BHS-IE and BHS-non infective endocarditis (BHS-nIE) in adult patients with BHS bacteremia at a 520-bed tertiary hospital in Tokyo, Japan from 2004 to 2020. IE was diagnosed according to modified Duke's criteria, and both “Definite” and “Possible” were included. Univariate and multivariable analyses were conducted using logistic regression. Results Among 250 patients with BHS bacteremia, 47 (19%) were diagnosed with BHS-IE. The median (IQR) patient age was 71 (59, 84) years and 121 (68%) were male. The proportions of A, B, C/G groups were 14%, 38.4%, and 47.6%, respectively. Five predictors, either independently associated with BHS-IE or clinically relevant, were used to develop the prediction score: C-reactive protein ≥ 10 mg/dl (2 points); Group B Streptococci (1 point); Auscultation of heart murmur (1 point); Platelet count < 150 /µl (1 point); and Hypotension (systolic blood pressure < 90 mmHg or on vasopressor) (1 point). In a receiver operating characteristic analysis, the area under the curve was 0.74 (95% confidence interval [CI]: 0.66 - 0.82). The cut-point was 2. A score ≥2 had a sensitivity of 87% (95%CI: 0.743 - 0.952), a specificity of 37% (95%CI: 0.308 - 0.445), a positive predictive value of 24%, and a negative predictive value of 93%, respectively. Conclusion We developed the score to help clinicians rule out IE in BHS bacteremia. Further research is warranted for validation. Disclosures All Authors: No reported disclosures

The Role of Streptococcal Cell-Envelope Proteases in Bacterial Evasion of the Innate Immune System

Bacteria possess the ability to evolve varied and ingenious strategies to outwit the host immune system, instigating an evolutionary arms race. Proteases are amongst the many weapons employed by bacteria, which specifically cleave and neutralize key signalling molecules required for a coordinated immune response. In this article, we focus on a family of S8 subtilisin-like serine proteases expressed as cell-envelope proteases (CEPs) by group A and group B streptococci. Two of these proteases known as <i>Streptococcus pyogenes</i> CEP (SpyCEP) and C5a peptidase cleave the chemokine CXCL8 and the complement fragment C5a, respectively. Both CXCL8 and C5a are potent neutrophil-recruiting chemokines, and by neutralizing their activity, streptococci evade a key defence mechanism of innate immunity. We review the mechanisms by which CXCL8 and C5a recruit neutrophils and the characterization of SpyCEP and C5a peptidase, including both in vitro and in vivo studies. Recently described structural insights into the function of this CEP family are also discussed. We conclude by examining the progress of prototypic vaccines incorporating SpyCEP and C5a peptidase in their preparation. Since streptococci-producing SpyCEP and C5a peptidase are responsible for a considerable global disease burden, targeting these proteases by vaccination strategies or by small-molecule antagonists should provide protection from and promote the resolution of streptococcal infections.

Russian strains of group B streptococci are different in the content and organization of the PAI-A and PAI-A1 pathogenicity islands

Group B streptococci, or Streptococcus agalactiae, are the major cause of severe diseases in newborns and adults. The PAI-A and PAI-A1 pathogenicity islands containing the sspB1 and sspB1a genes, respectively, were found among group B streptococci mobile genetic elements. The presence of sspB genes correlates with urogenital tract infections. The aim of this study was to determine the frequency of group B streptococci strains with the PAI-A and PAI-A1 pathogenicity islands, circulating in Moscow, in comparison with strains from St. Petersburg. The sspB1 gene, and hence the PAI-A pathogenicity island, was not found in the genomes of strains from Moscow. The frequency of the sspB1a gene and the PAI-A1 pathogenicity island in the genomes of clinical strains was three times higher than in the genomes of colonizing strains. Thus, it can be assumed that the genes of the sspB family are more specific of group B streptococci colonizing pregnant women and newborns.

An in vitro enterocyte cell-like model to study Group B streptococci internalization

We used a 2D in vitro growth model of Caco2 cells to study invasion of Streptococcus agalactiae. This model can be useful for evaluating the virulence of the most relevant enteric pathogens.

Lysine Residues in the MK-Rich Region Are Not Required for Binding of the PbsP Protein From Group B Streptococci to Plasminogen

Binding to plasminogen (Plg) enables bacteria to associate with and invade host tissues. The cell wall protein PbsP significantly contributes to the ability of group B streptococci, a frequent cause of invasive infection, to bind Plg. Here we sought to identify the molecular regions involved in the interactions between Plg and PbsP. The K4 Kringle domain of the Plg molecule was required for binding of Plg to whole PbsP and to a PbsP fragment encompassing a region rich in methionine and lysine (MK-rich domain). These interactions were inhibited by free L-lysine, indicating the involvement of lysine binding sites in the Plg molecule. However, mutation to alanine of all lysine residues in the MK-rich domain did not decrease its ability to bind Plg. Collectively, our data identify a novel bacterial sequence that can interact with lysine binding sites in the Plg molecule. Notably, such binding did not require the presence of lysine or other positively charged amino acids in the bacterial receptor. These data may be useful for developing alternative therapeutic strategies aimed at blocking interactions between group B streptococci and Plg.