Removal of Group B Streptococci Colonizing the Vagina and Oropharynx of Mice with a Bacteriophage Lytic Enzyme

Abstract

Group B streptococci (GBS) are the leading cause of neonatal meningitis and sepsis worldwide. The current treatment strategy is limited to intrapartum antibiotic prophylaxis in pregnant women to prevent early-onset neonatal diseases, but considering the potential for antibiotic resistance, the risk of losing control over the disease is high. To approach this problem, we have developed a bacteriophage (phage) lytic enzyme to remove colonizing GBS. Bacteriophage muralytic enzymes, termed lysins, are highly evolved molecules designed to degrade the cell wall of host bacteria to release phage particles from the bacterial cytoplasm. Several different lysins have been developed to specifically kill bacterial pathogens both on mucosal surfaces and in blood and represent a novel approach to control infection. A lysin cloned from a phage infecting GBS was found to contain two putative catalytic domains and one putative binding domain, which is similar to the domain organization of some staphylococcal phage lysins. The lysin (named PlyGBS) was recombinantly expressed in Escherichia coli, and purified PlyGBS efficiently killed all tested GBS serotypes in vitro. In a mouse model, a single dose of PlyGBS significantly reduced bacterial colonization in both the vagina and oropharynx. As an alternative strategy for intrapartum antibiotic prophylaxis, this approach may be used to reduce vaginal GBS colonization in pregnant women before delivery or to decontaminate newborns, thus reducing the incidence of GBSassociated neonatal meningitis and sepsis.Group B streptococci (GBS), or Streptococcus agalactiae, are the leading cause of neonatal sepsis and meningitis in the United States and Europe (1). First identified as a pathogen in bovine mastitis, GBS were later found to colonize the human genital and lower gastrointestinal tract, where the organism can be transmitted perinatally to the fetus. In addition to newborn infections, GBS can also cause substantial morbidity and mortality in adults, particularly those who are immunocompromised or elderly (6). GBS are classified into nine serotypes based on differences in the polysaccharide capsule (24). Historically, the most commonly reported serotype associated with diseases is type III; however, recent epidemiological studies found that serotype V now accounts for approximately 30% of nonpregnant adults (11) and 14 to 23% of pregnant women and neonates (30).The efficacy of antibiotic administration during various stages of pregnancy in order to reduce the rate of early-onset GBS infection in neonates has been tested. Hall et al. (10) demonstrated that treatment during pregnancy (antepartum prophylaxis) reduced maternal GBS colonization transiently, but colonization recurred at delivery. For postpartum prophylaxis, universal administration of penicillin to neonates shortly after birth reduced the early-onset GBS infection by 68%, but it was also associated with a 40% increase in overall mortality and therefore is not recommended (2). The most successful approach to block vertical tr...

Antimicrobial bacteriophage-derived proteins and therapeutic applications

“…Washing a mouse vagina with the S. agalactiae phage endolysin PlyGBS could reduce the bacterial burden of a streptomycin-resistant S. agalactiae strain by 3-logs. 58 Notably, PlyGBS had an optimal lytic activity around pH 5 coinciding with the normal pH range of healthy human vaginal tracts. However, Cheng et al considered PlyGBS to have a relatively low streptolytic activity in vitro compared to other endolysins described in the literature.…”

“…GBS is particularly problematic for causing sepsis and acute inflammation of the brain and spinal cord (meningitis) among newborns. 58,66 Mouse S. agalactiae bacteremia is typically fatal within 3 d. Treatment with the S. dysgalactiae prophage endolysin PlySK1249, which also lyses S. agalactiae, could prolong the survival of infected mice after a single intraperitoneal treatment of 22.5 mg/kg. PlySK1249 treatment became curative when increased to 3 intraperitoneal injection treatments of 45 mg/kg over the course of the first day after GBS challenge.…”

“…[58][59][60] The most problematic streptococci are the b-hemolytic group A streptococci (GAS) and group B streptococci (GBS), and the a-hemolytic streptococci Streptococcus pneumoniae and Streptococcus viridans.…”

“…In addition, neither PlyGBS nor its hyperactive mutant harmed other common beneficial vaginal colonizers, such as Lactobacillus acidophilus. 58,66 S. pneumoniae is an a-hemolytic bacterium that colonizes the nasopharynx of most adults and children. It is also a common cause of bacterial pneumonia.…”

Plastid production of protein antibiotics against pneumonia via a new strategy for high-level expression of antimicrobial proteins

“…Cell lysis is mediated by the action of a single enzyme encoded in the phage genome, a cell wall hydrolase also referred to as lysin. Purified phage lysin proteins have been demonstrated to rapidly kill pathogenic bacteria and effectively prevent bacterial colonization of mucosal surfaces (5,6), thus qualifying as excellent candidates for next-generation antibiotics. We have recently shown that a lysin (PlyGBS) targeted against group A and group B streptococci can accumulate to high levels in plant cells if encapsulated inside the chloroplast (3).…”