Group B Streptococcus: molecular epidemiology, pathogenic profiling and control strategies
Group B Streptococcus (GBS) are frequent colonisers of the gastrointestinal tract of healthy adults with worldwide rates of 18% colonisation; however, they are also opportunistic pathogens capable of causing invasive disease, particularly in neonates and adults with underlying conditions. Indeed, GBS is the leading cause of invasive neonatal disease worldwide with a global incidence of 0.49 per 1000 live births and GBS disease in adults is increasing globally, with estimated rates of 10.9 cases per 100,000 people. While GBS remain largely susceptible to penicillin, there have been reports of reduced beta-lactam susceptibility in certain countries and resistance to other antibiotic classes continues to rise. GBS strains are distinguished by their serotype, defined by an expressed capsular polysaccharide, and vaccine efforts have primarily focused on this capsule as a target; however, as of yet, there is no vaccine licensed for use. The aims of this thesis were to identify the predominant serotypes and antimicrobial resistance properties of circulating GBS isolates in Ireland, to investigate pathogenic traits of these isolates and to examine novel therapeutic options for GBS. The molecular epidemiology of n = 253 clinical GBS strains was investigated by determining the serotype and antimicrobial resistance traits in a collection of both colonising (n = 202) and invasive (n = 51) isolates. High levels of erythromycin (22.5%) and clindamycin (21.3%) resistance were observed and the emergence of the unusual L phenotype was detected, which had not been previously reported in Ireland. Resistance to both erythromycin and clindamycin was higher among colonising isolates than colonising isolates (P < 0.05), with mean zones of X mm and X mm for erythromycin and clindamycin respectively. The serotype distribution revealed that overall serotypes Ia (28.1%), III (26.88%), II (13.44%), V (11.86%) and Ib (11.46%) were the most common among GBS strains. Serotype distribution was differed among invasive and colonising isolates, whereby serotype III (31.37%) predominated among invasive isolates while serotypes Ia (29.7%) and III (25.745) were the most common xii in colonising strains. Serotype III predominated among resistant isolates (33.3%). To examine novel antimicrobials for GBS, the bacteriocin nisin was explored, and some level of susceptibility characterised by zones of inhibition was detected in 91% of isolates. Investigation of the genetic basis for resistance to nisin in GBS revealed the presence of a nisin resistance gene (nsr) in 98.4% of isolates, as well as the presence of an ABC transporter, nsrFP, in 84.4% of isolates. A bioengineered derivative of nisin, nisin PV, designed to resist cleavage by the nisin resistance protein (NSR), displayed significantly enhanced activity against isolates (P < 0.05) compared to the wild-type peptide, with an MIC50 of 20.1 μg/ml and 6.3 μg/ml for nisin A and nisin PV respectively. Investigating the potential of novel alternative antimicrobial strategies determined that erythromycin and nisin used in combination exerted a synergistic effect in a proportion of isolates by time-kill curve assays. This combination also reduced TNF-α production (P < 0.05) in U937 macrophages challenged with GBS. The capacity of GBS to form biofilms and the underlying genetic determinants associated with biofilm formation were investigated within the collection. Reducing gastrointestinal colonisation could help limit GBS infections and anti-biofilm agents are an interesting approach to help achieve this. Consuming food products with antimicrobial activity could prevent biofilm formation and therefore the potential for certain food products to inhibit and reduce GBS biofilms was assessed. Biofilm formation was significantly inhibited and reduced by natural food products garlic, honey, feta and goats cheese by the XTT assay for cell viability (P < 0.05). This thesis provides a current assessment of the characteristics of clinical GBS strains circulating in the South of Ireland and has highlighted novel antimicrobial strategies for the control of GBS.