Molecular Epidemiology of Streptococcus pneumoniae Serotype 1: A Systematic Review of Circulating Clones and Clonal Clusters

1. Introduction

Streptococcus pneumoniae is a Gram-positive, opportunistic commensal bacterium that colonizes the nasopharynx of the upper respiratory tract of humans [1]. Its ability to spread to other parts of the body and evade host immune response has marked S. pneumoniae as the leading cause of invasive pneumococcal diseases (IPD) in the world, resulting in significant morbidity and mortality, especially among vulnerable populations such as children, older adults, immunocompromised patients [2,3,4,5]. The commonly reported invasive pneumococcal diseases associated with this bacterium are pneumonia, bacteremia, and meningitis. According to the World Health Organization (WHO), S. pneumoniae is responsible for about 300,000 deaths in younger children annually, with the majority of these casualties occurring in developing countries [6]. The epidemiology of S. pneumoniae varies geographically and is influenced by several factors, including carriage patterns, vaccination strategies, antibiotic use, and the emergence of multidrug-resistant strains [7].

To date, one hundred pneumococcal serotypes have been discovered throughout the world [8], with a small number of these serotypes responsible for most invasive diseases. Serotype 1 is among the most prevalent and invasive pneumococcal serotypes globally [9], accounting for a substantial proportion of IPD cases and outbreaks in many regions, including sub-Saharan Africa, Asia, South and North America, and Europe [10]. Although serotype 1 is associated with high invasive potential, it is rarely found in the nasopharyngeal carriage of healthy individuals due to limited carriage duration, capsular characteristics, and genetic factors [9,10]. The global distribution of serotype 1 S. pneumoniae is complex, with different clones circulating in various geographic regions. Understanding the clonal composition of serotype 1 S. pneumoniae is crucial to inform vaccine development, antibiotic treatment, and disease surveillance. Molecular genotyping methods including Multilocus sequence typing (MLST), pulsed-field gel electrophoresis (PFGE), whole genome sequencing (WGS), and a multilocus variable number of tandem repeat analysis (MLVA) have provided a vivid understanding of the genetic structure of S. pneumoniae populations [11]. Studies utilizing these methods have identified several sequence types (STs) associated with serotype 1, highlighting the existence of clonal complexes that demonstrate both virulence potential and antimicrobial resistance profiles.

The introduction of pneumococcal conjugate vaccines (PCVs) has been a significant public health achievement, leading to a substantial reduction in the incidence of invasive pneumococcal disease, especially those caused by vaccine-type serotypes [12,13]. However, the widespread use of these vaccines has been associated with the phenomenon of serotype replacement, where non-vaccine serotypes become more predominant [8]. This has been observed in many countries following the implementation of PCVs, with non-vaccine types such as serotype 1 becoming more prevalent [2]. The emergence of serotype 1 as a major cause of IPD in the post-PCV era, highlights the need for continuous surveillance and monitoring of pneumococcal serotypes, particularly in low-and middle-income countries, where the burden of pneumococcal disease is highest and access to diagnostic tools and vaccines may be limited. Despite the growing body of research on S. pneumoniae serotype 1, a comprehensive synthesis to assess the distribution of its clones is lacking. By consolidating current available data on the continent, this systematic review aims to describe the circulation of serotype 1 clones and their clonal complexes.

2. Results

2.1. Search Results and Selection Process

A total of 1,234 records were identified from all five electronic databases. Two hundred and nine duplicate records were removed, leaving 1,025 unique records to be screened. Of the 1,025 unique records, 813 were excluded based on their title, abstract, and language, and the remaining 212 records were sought for full-text retrieval. After full-text retrieval, 157 articles were excluded following the full-text eligibility assessment. Finally, 43 studies were included in the systematic review (Figure 1).

2.2. Description of Included Studies

Most of the data analyzed in this study were obtained from single-center studies conducted mainly in 19 countries (n = 38 studies). The majority of the studies were found in the United Kingdom (n = 5), Spain (n = 5), the United States (n = 5), and Gambia (n = 5). Five articles were multicenter studies including two intra-continental surveys that grouped data from countries in sub-Saharan Africa and West Africa respectively, one intra-continental survey that described data from the individual countries, and two inter-continental multicenter studies that also reported data from the individual countries. Of the 43 studies, 19 studies (44.2%) focused on participants of all ages (children to adults), 16 studies (37.2) on children only, and 2 studies (4.7%) on adults only. Six studies did not specify the age group of the included participants. Most studies (n = 29, %) described invasive pneumococcal diseases (IPD) among participants while four described pneumococcal carriage. Ten studies reported on two or more isolate types (IPD and NIPD, n = 6; IPD and carriage, n = 3; all three types, n = 1). The PCVs evaluated includes PCV7, PCV9, PCV10, PCV13, PPV23, and PPV24. Studies employed polymerase chain reaction (PCR) or whole genome sequencing (WGS) or a combination thereof, alongside multilocus sequence typing (MLST) analysis for S. pneumoniae to determine their sequence types.

2.3. Pneumococcal Serotype 1 Clones Reported Worldwide

Table S1 describes the molecular characteristics of 103 pneumococcal serotype 1 clones identified globally. Based on the BURST analysis, three main clonal complexes (CCs) were identified: CC217, CC306, and CC615.

2.3.1. Africa

Our search identified seventeen (17) studies describing pneumococcal serotype 1 STs from African countries (Table 3). BURST analysis revealed that clonal complex 217 (CC217) is the largest group, comprising five main clusters CC217-ST217, ST303, ST612, ST618, and ST3081. This group was predominantly reported in Africa. The ancestral clone, ST217, was predominantly associated with pneumococcal meningitis in Niger [52], Ghana [32,33], Burkina Faso [49], and Egypt [61]. Additionally, ST217 was widely responsible for several IPD cases described in South Africa [27,39], Kenya [27,40], Ethiopia [59], Mozambique [61], Malawi [56], and Gambia [25,48,61]. Carriage of serotype 1 ST217 was reported in Gambia [25], Kenya [40], Ethiopia [61], and Nigeria [61]. Single locus variants of ST217 such as ST613 and ST614 were identified in Kenya as associated with IPD cases and, to a lesser extent, carriage [27,40]. Other SLVs including ST1316, ST1325, ST2034, ST2206, ST2839, ST3575, ST5012, ST5632, ST8158, ST8314, ST9067, and ST9529, as well as DLVs like ST12150 and ST12739 were identified in countries in sub-Saharan Africa [20,25,33,39,61].

ST303, a triple locus variant of 217 was recovered from IPD cases in Gambia [35,57,61], Ghana [32,33,61], Malawi [61], Niger [61], and Togo [58]. Its SLVs, ST1322, ST1323, and ST4755 were reported in Ghana [33,61] while others including ST11104, ST11736, ST12305, and ST12803 were identified in sub-Saharan African countries [20]. ST3081 (TLV of 217) and its SLVs, ST11779, and ST12197 were identified in Gambia [25,35,61], Senegal [61], and other countries in sub-Saharan Africa [20]. ST612 (TLV of 217) was described in studies from Ghana [33], Gambia [25], and South Africa [27,39,61] while its SLVs including ST1327, ST1328 and ST3570, were found in Ghana [33] and Gambia [25]. Another TLV of 217, ST618 was reported in Burkina Faso [49], Egypt [61], Gambia [25,35,48,57,61], South Africa [39], and Togo [58]. SLV of ST618 such as ST1331 was only reported in Ghana [33], ST2084 in Ghana and Gambia [25,61], and ST3336, ST3573, ST3574, ST3577, and ST3581 in Gambia [25]. Some studies from South Africa reported ST304. ST306, and ST611 [27,39].

2.3.2. Europe

A total of 16 studies from the European region were identified (Table 3). Clonal complex 306 was the second largest group based on the BURST analysis. It was divided into three main clusters: CC306-ST306, ST304, and ST227. This group was widespread and diverse across Europe, with a significant presence in cases of IPD reported in the 16 studies. The ancestral clone, ST306 was the predominant clone in the region frequently associated with IPD cases in the United Kingdom (UK) [36,44,45,55], Spain [23,24,43,46,51], Sweden [61], Portugal [29,60], Poland [27], Norway [27], Netherland [27], Germany [27], France [27], Czech Republic [27,42], and Croatia [61]. Alongside ST306, its double locus variant, ST304 was notably reported in France [27], Netherlands [27], Norway [27], Portugal [60], Slovenia [61], Spain [23,24,43,46,51,61], and the UK [36,44,45,55]. ST227 was reported in Denmark [27] and the UK [27,36,45,53,55] while ST228 was described in Portugal [60], France [27], and Croatia [61]. Both ST227 and ST228 were identified in Spain [23,24,43,46,61] and the Netherlands [27]. Other STs with CC306, including ST1247 (DLV of 306), ST1310 (SLV of 306), and ST2376 (SLV of 306) were reported in the UK [36] in Spain [23,46]. Additionally, ST617 (SLV of 306), ST1239 (SLV of 227), and ST3861 (SLV of 304) were reported in Norway [27], UK [36] and Spain [24] respectively. Clonal complex 217 (CC217) was also identified in European countries, with ST217 widely documented in studies reporting IPD cases in the UK [36], Denmark [27], France [27], Germany [27], and Spain [23,24]. ST618 was only reported in the Netherlands [27] but identified along with ST614 (SLV of 217) in Spain [43]. To a lesser extent ST300 (DLV), ST616 (SLV), and ST3860 (DLV) of ST615 were documented in reports from the UK [27,55], Poland [27], and Spain [24] respectively. Regarding serotype 1 carriage, ST306, ST304, ST350, ST228, and ST217 were identified in nasopharyngeal samples from healthy children attending nursery school in Belgium [28] and Portugal [22].

2.3.3. Asia

Pneumococcal serotype 1 clones reported in five studies conducted in Asia (Table 3) revealed a predominant clustering of member clones of CC217. This group encompasses subgroups such as CC303, CC618, and CC3081. In India, clones belonging to this group including ST217 and its variants ST2839, and ST5002, were identified along with ST5044 related to ST618 and ST5672 (SLV of 303) [61]. Following the introduction of PCV7 in Singapore, serotype 1 strains causing meningitis and sepsis were primarily ST217, ST303, ST5044, and ST5254 [31]. In other Asian countries, such as the Philippines, Bangladesh, Nepal, and Thailand, STs belonging to CC217 including ST217, ST303, and ST3081 were detected [61]. Notably, ST615, part of the CC615 group, was also identified in clinical isolates in Thailand [61]. In Qatar, clinical isolates from blood specimens were represented by ST217, ST303, and ST3081, as well as ST306 (CC306) [61] (Figure 4). Porat et al. found that serotype 1 ST217 was the predominant clone in the southern region of Israel, accounting for cases of acute otitis media (AOM) and invasive pneumococcal disease (IPD) in children under 5 years old [54]. Notably, ST304 was also identified in children with IPD in this study. In China, ST2296 was predominant along with singletons such as ST81, ST242, ST342, ST1263, ST3397, and ST13362 identified in isolates from invasive and non-invasive pneumococcal disease [38].

2.3.4. Americas

Six studies reported pneumococcal serotype 1 clones circulating in North American countries (Table 3). Related clones of clonal complex 306 were widely reported in this region. Studies conducted in the United States from 1990 to 2002 consistently reported ST227 (DLV of 306) as the dominant ST (Figure 5). For instance, a study assessing the clonal compositions of invasive pneumococcal serotypes in the United States in 1999, 2001, and 2002 found that ST227 was the prevalent serotype 1 isolates [47]. This was consistent with the findings of Brueggemann and Spratt [27] and Gonzalez et al. [26] confirming ST227 as the most prevalent serotype 1 clone associated with cases of IPD. However, from 2003 onwards, multiple STs closely related to ST227, including ST304, 306, 2126, 4290 (SLV 306), 4288, and 4289, were reported in cases of Pediatric Pneumococcal Empyema (PPE) in Utah [19] (Figure 5). Like in the United States, ST227 was the predominant ST in Canada [27]. In contrast, in Mexico, cases of serotype 1 IPD were predominantly associated with ST304 [42].

Three studies reported pneumococcal serotype 1 clones in six countries from South America (Table). These studies documented the widespread of member clones of CC306 and CC615. In Argentina and Uruguay [42,61], the dominant clones identified in isolates of IPD cases were ST615 and ST304. ST615 clone was reported in Chile [27] and Peru [61] in isolates from blood and pleural fluid. According to Zemlicková et al. [42], serotype 1 IPD in Brazil and Colombia were caused by ST304 and ST306 respectively.

2.3.5. Oceania

Our search identified six studies from Oceania that reported clones of pneumococcal serotype 1 (Table 3). In this region, most reported STs belonged to clonal complex 306. In Australia, ST304 and ST227 were widely reported in serotype 1 IPD cases and carriage in the north according to Smith-Vaughan et al. [30]. An outbreak of serotype 1 IPD in Australia between 2008 and 2012 was predominantly caused by ST306 clones [34,37]. The proportion of ST304 isolates reported in Australia significantly decreased during this period, attributed to the clonal expansion of CC306, characterized by an increase in ST306 isolates (Figure 6). In New Caledonia (NC), ST306 emerged as the most predominant serotype 1 clone responsible for outbreaks reported [21]. Genomic characterization of serotype 1 isolates from outbreaks in NC discovered ST306 strains (62/67, 92.53%) and their single locus variants, ST3717 (5/67, 7.46%) [21]. Additionally, ST306 serotype 1 isolates were reported in cases of IPD 61 in New Zealand. ,

3. Discussion

Streptococcus pneumonaie serotype 1 has been a leading cause of severe pneumococcal disease worldwide, affecting both children and adults. Notably, this serotype is seldom found in healthy individuals, suggesting that while it may not be easily transmitted, it has a high tendency to progress to invasive disease once carried. This implies that serotype 1 is remarkably hypervirulent, corroborating its frequent association with outbreaks of severe pneumococcal infection. Furthermore, several clones, including ST217 and ST306, have been linked to outbreaks of IPD globally. In light of these findings, we conducted a systematic review to elucidate the molecular epidemiology of S. pneumoniae serotype 1 clones circulating worldwide. This comprehensive analysis seeks to identify high-risk clones with the potential to trigger outbreaks in various countries or continents, ultimately informing targeted evidence-based strategies for disease prevention and control.

The multilocus sequence typing (MLST) technique is a widely accepted method for bacterial genotyping, leveraging seven housekeeping genes (aroE, gdh, gki, recP, spi, xpt, and ddl) to characterize S. pneumoniae isolates. As the golden standard for S. pneumoniae genotyping, MLST was mainly used in most studies to elucidate the genetic diversity of this pathogen. We identified one hundred and three (103) different MLST sequence types associated with serotype 1. Most of these STs shared genetic relationships with ST217 followed by ST306.

The S. pneumoniae ST217 clonal complex is a hypervirulent lineage of significant public health concern. In our study, this lineage was mostly reported in studies from Africa and Asia. The ancestral clone, ST217 was predominantly associated with pneumococcal meningitis across several countries, including, Nigeria, Niger, Israel, India, Thailand, Singapore, and the Philippines. ST217 was also linked to numerous IPD cases in Kenya, South Africa, Ethiopia, Mozambique, and Malawi. Although not the most common clone, ST217 was associated with a serotype 1 pneumococcal meningitis outbreak in northern Ghana [33]. This finding is not surprising, since the predominance of ST217 in sub-Saharan African countries has been previously reported in several studies [62]. This widespread presence of ST217 may be attributed to intra and intercontinental movements of people between countries, leading to the transfer of clones. Other clones within CC217, such as ST618, were prevalent among serotype 1 isolates from both invasive and carriage isolates in Gambia. Also, ST618 was responsible for pneumococcal meningitis in patients from Burkina Faso [49] and Egypt [61]. Additionally, studies from neighboring countries like Ghana and Togo, as well as Nepal and Bangladesh, mostly described ST303 as associated with bacterial meningitis. Several other STs within CC217 were also identified in Africa and Asia (Table 3) indicating an extensive clonal expansion and diversity of the ST217 clonal complex, particularly in Africa. The propensity of ST217 and its clonal complex 217 to cause pneumococcal meningitis poses a significant threat to these regions, emphasizing the importance of implementing mitigation strategies to curb their spread. Fortunately, many isolates of CC217 are sensitive to antibiotics, making their treatment easy and effective [33]. While the ST217 clonal complex is predominantly reported in Africa and Asia, its presence has also been noted in other parts of the world. For example, studies have reported STs from the clonal complex in regions like Australia, the US, and specific countries in Europe.

ST306 clonal complex is another lineage of serotype 1 of public health importance. This group emerged as the prominent group in Europe, Oceania, North America, and some countries in South America characterized by its division into three main subgroups: ST306, ST304, and ST227. ST306 (ancestral clone), was the most predominant ST among this group widely dispersed in most European countries. ST304 was common in Mexico, Brazil, Uruguay, and Slovenia. STs of CC306 were more associated with bacteremia than other pneumococcal diseases. Our analysis revealed that ST227 was the predominant clone in Canada and the US, supporting the findings of Brueggemann & Spratt [27]. However, our results diverged from theirs regarding the UK, where they reported ST227 as the dominant clone. This discrepancy suggests that a clonal shift may have occurred in the UK over time, potentially replacing ST227 with ST306 as discovered in our study. Like CC217, the genetic variation of ST306 clonal complex is also illustrated by the identification of several other related STs.

One key finding of our study is the geographic structure among major serotype 1 lineages, revealing distinctive regional differences in circulating clones. This observation may be attributed to the rarity of serotype 1 carriage. Unlike other serotypes that can easily be carried and transferred between populations via human movement, the rarity of serotype 1 carriage restricts its geographic spread and may contribute to unique clonal distributions. Also, this unique clonal distribution may be attributed to geographic weather conditions in the various regions. For instance, it might be that the ST217 clonal complex thrives in humid and tropical areas which may explain why they are widely found in Africa and Asia. This possibly applies to ST306 and ST615 clonal complexes that are common in temperate climate areas.

While our study provides valuable insight into the global circulation of serotype 1 S. pneumoniae, it is not without limitations. One notable constraint is the geographical representation of the included studies, predominantly clustered in forty-eight (48) countries. The uneven distribution leaves a significant knowledge gap in many countries, highlighting the need for further research to understand better the epidemiological landscape of pneumococcal serotype 1 clones in these underrepresented regions.

4. Materials and Methods

This systematic review adhered to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines ¹⁴.

4.1. Literature Search Strategy and Selection Process

An extensive literature search was conducted in Google Scholar, PubMed, Scopus, ScienceDirect, and Web of Science using keywords related to Streptococcus pneumoniae serotype 1. The search was conducted until October 2024 and restricted to free full-text or open-access research articles published in English (Table 1). Retrieved records were exported in RIS and CSV formats.

Citations of the retrieved records were imported to the Rayyan application [15] (a web-based tool for systematic reviews) for screening. Duplicate records were first detected and removed, followed by title and abstract screening of the remaining unique records, where all unrelated records were excluded. After title and abstract screening, the remaining articles were assessed for full-text eligibility based on the abovementioned inclusion criteria. Included studies were imported to Zotero 7.0.8 for data extraction. O.K.N. and E.S.D. assessed the eligibility of the retrieved records.

4.2. Eligibility Criteria

This study included English-published peer-reviewed reports that utilized at least one molecular genotyping tool to describe S. pneumoniae serotype 1 clones based on their sequence types (STs). Multicenter studies describing serotype 1 clones in specific geographic regions (e.g., West Africa, sub-Saharan Africa) or those reporting clone data for individual countries were eligible for inclusion. Exclusions included non-peer-reviewed sources (Theses, dissertations, editorials, comments, preprints, etc.), studies not reporting serotype 1 clones, studies that selected specific serotype 1 clones for genetic analysis, and studies that focused or reported on other serotypes or their clones.

4.3. Data Extraction and Analysis

The relevant data were extracted from each included study as described in Table 2 by O.K.N. and E.S.D. The sequence types (STs) of S. pneumoniae serotype 1 clones were extracted and stratified by country/region. Microsoft Excel 365 was used to extract and manage the extracted data.

4.4. BURST Cluster Analysis

The allelic profiles of identified STs were downloaded from the Streptococcus pneumoniae database on the MLST website (https://pubmlst.org/organisms/streptococcus-pneumoniae). STs were grouped into clonal complexes according to their allelic profiles using the BURST algorithm [16]. Furthermore, the geoBURST algorithm in the Phyloviz online application (https://online.phyloviz.net/index) was used to construct the minimum spanning tree, providing a visual representation of the genetic relationship among the STs.

4.5. Risk of Bias Assessment

Two investigators (O.K.N. and E.S.D) assessed the risk of bias of the included studies using the STROBE checklist for reporting observational studies [17], as adapted according to Sanderson et al. [18] The assessment tool comprised of five key domains, method of selecting study participants, method of measuring exposure and outcome variables, method of control confounding, design-specific source of bias, and statistical methods (Supplementary table S2). Studies were graded into three categories: High risk of bias (≥1 of any criteria classified as H, or ≥2 major criteria as M or U); Moderate risk of bias (≥2 of any criteria classified as M or U (<2 major criteria)); Low risk of bias (all major criteria classified as L and <2 minor criteria as M or U.

5. Conclusion

This systematic review highlights the complex genetic diversity of S. pneumoniae serotype 1, characterized by distinct geographic structures among major lineages. A striking continental distribution emerged, with the ST217 clonal complex predominantly clustered in Africa and Asia, whereas the ST306 clonal complex was widespread across Europe, the Americas, and Oceania. Notably, invasive pneumococcal disease (IPD) was primarily linked to clones from these clonal complexes, with few instances of carriage. The findings of this study emphasize the need for enhanced surveillance and targeted interventions to mitigate the spread of hypervirulent clones, ultimately informing evidence-based strategies for disease prevention and control. To address existing knowledge gaps, further research is warranted, particularly in countries with limited or no data on serotype 1 S. pneumoniae clones.