Population structure of ocular Streptococcus pneumoniae is highly diverse and formed by lineages that escape current vaccines

Abstract

Streptococcus pneumoniae is a leading cause of ocular infections including serious and sight-threatening conditions. The use of pneumococcal conjugate vaccines (PCV) has substantially reduced the incidence of pneumonia and invasive pneumococcal diseases, but has had limited impact on ocular infections. Additionally, widespread vaccine use has resulted in ongoing selective pressure and serotype replacement in carriage and disease. To gain insight into the population structure of pneumococcal isolates causing ocular infections in a post-PCV-13 time period, we investigated the genomic epidemiology of ocular S. pneumoniae isolates (n=45) collected at Massachusetts Eye and Ear between 2014 and 2017. By performing a series of molecular typing methods from draft genomes, we found that the population structure of ocular S. pneumoniae is highly diverse with 27 sequence types (grouped into 18 clonal complexes) and 17 serotypes being identified. Distribution of these lineages diverged according to the site of isolation, with conjunctivitis being commonly caused by isolates grouped in the Epidemic Conjunctivitis Cluster-ECC (60 %), and ST448 (53.3 %) being most frequently identified. Conversely, S. pneumoniae keratitis cases were caused by a highly diverse population of isolates grouping within 15 different clonal complexes. Serotyping inference demonstrated that 95.5 % of the isolates were non-PCV-13 vaccine types. Most of the conjunctivitis isolates (80 %) were unencapsulated, with the remaining belonging to serotypes 15B, 3 and 23B. On the other hand, S. pneumoniae causing keratitis were predominantly encapsulated (95.2 %) with 13 different serotypes identified, mostly being non-vaccine types. Carriage of macrolide resistance genes was common in our ocular S. pneumoniae population (42.2 %), and usually associated with the mefA +msrD genotype (n=15). These genes were located in the Macrolide Efflux Genetic Assembly cassette and were associated with low-level in vitro resistance to 14- and 15-membered macrolides. Less frequently, macrolide-resistant isolates carried an ermB gene (n=4), which was co-located with the tetM gene in a Tn-916-like transposon. Our study demonstrates that the population structure of ocular S. pneumoniae is highly diverse, mainly composed by isolates that escape the PCV-13 vaccine, with patterns of tissue/niche segregation, adaptation and specialization. These findings suggest that the population structure of ocular pneumococcus may be shaped by multiple factors including PCV-13 selective pressure, microbial-related and niche-specific host-associated features.

Sulfated motifs in heparan sulfate inhibit Streptococcus pneumoniae adhesion onto fibronectin and attenuate corneal infection

“…S. pneumoniae consists of many serotypes, which differ in their tropism and infectivity of different tissues. Interestingly, an epidemiologic study at Massachusetts Eye and Ear Institute found that 95% of S. pneumoniae keratitis isolates are encapsulated, whereas 80% of conjunctivitis isolates are unencapsulated 37 . While polysaccharide capsule may not be an essential virulence factor for S. pneumoniae keratitis 58,59 the ongoing selective pressure of widely used vaccines targeting a small number of capsule serotypes has resulted in the emergence of novel serotypes not covered by these vaccines, which are becoming common colonizers of the nasopharynx and seem to be well adapted to cause keratitis 37 …”

“…37 While polysaccharide capsule may not be an essential virulence factor for S. pneumoniae keratitis 58,59 the ongoing selective pressure of widely used vaccines targeting a small number of capsule serotypes has resulted in the emergence of novel serotypes not covered by these vaccines, which are becoming common colonizers of the nasopharynx and seem to be well adapted to cause keratitis. 37 We, therefore, examined if 2-O-sulfated heparan compounds also inhibit corneal infection by other serotypes of S. pneumoniae, including the emerging serotype 35B. Injured mouse corneas were topically infected with IDI30/44 (serotype 35B), a clinical keratitis isolate, or with L82016 (serotype 6B) and NDS-HP, 2ODS-HP, H, NS-H, or NS2OS-H and the corneal bacterial burden was enumerated at 6 h pi.…”

“…We 80% of conjunctivitis isolates are unencapsulated. 37 While polysaccharide capsule may not be an essential virulence factor for S. pneumoniae keratitis 58,59 the ongoing selective pressure of widely used vaccines targeting a small number of capsule serotypes has resulted in the emergence of novel serotypes not covered by these vaccines, which are becoming common colonizers of the nasopharynx and seem to be well adapted to cause keratitis. 37 We, therefore, examined if 2-O-sulfated heparan compounds also inhibit corneal infection by other serotypes of S. pneumoniae, including the emerging serotype 35B.…”

“…Fluorophore-conjugated secondary antibodies were purchased from Invitrogen/Thermo Fisher. Bacteria S. pneumoniae strains TIGR4 (serotype 4) and L82016 (serotype 6B), 36 and clinical keratitis isolate IDI30/44 (sequence type 558, serotype 35B) 37 were grown on 5% sheep blood TSA plates or in 4 mL of Todd-Hewitt broth supplemented with 0.5% yeast extract (THY broth) in glass tubes without agitation to mid-log growth phase. The bacterial concentration was approximated by measuring absorbance at OD600 nm.…”

Targeted Killing of Ocular Streptococcus pneumoniae by the Phage Endolysin MSlys

“…The antibacterial activity of MSlys was tested against ocular S. pneumoniae isolates (n=31) molecularly characterized in our previous studies 5,6 . Protocols for J o u r n a l P r e -p r o o f obtaining discarded isolates were approved by the Mass General Brigham Institutional Review Board, and the study was conducted in accordance with the Declaration of Helsinki.…”