ReviewCurrent challenges in the accurate identification of Streptococcus pneumoniae and its serogroups/serotypes in the vaccine era
Introduction
Streptococcus pneumoniae belongs to the mitis group of streptococci that normally (Facklam, 2002) exists as a commensal flora of the upper respiratory tract (URT) (Gray et al., 1980). It is one of the major causes of acute otitis media, pneumonia and in its severe (invasive) form, causes bacteremia and meningitis (Advisory Committee on Immunization Practices, 2000, Musher, 1992) in children leading to high morbidity and mortality, especially in developing countries (CDC, 2013). Among the mitis group, other closely related species such as S. mitis, S. oralis, and S. pseudopneumoniae have also been associated with human disease (Bochud et al., 1994, Douglas et al., 1993, Keith et al., 2006, Rolo et al., 2013). Hence it is very crucial to identify them accurately for an early diagnosis and better treatment (Roberts et al., 1979, Bochud et al., 1994, Elting et al., 1992, Carratalá et al., 1995). Misidentification of causative agents not only affects the true burden of disease surveillance but also falsely increases the pneumococcal antimicrobial resistance rate since other closely related species such as S. mitis are associated with a high rate of penicillin and multidrug resistance (Simões et al., 2010, Wester et al., 2002). In addition, precise identification followed by serotyping is needed for the effective formulation of conjugate vaccine which is based on most prevalent serotypes. Development and implementation of an appropriate vaccine are always driven by proper laboratory identification of the disease causing agents. The current review discusses the different sides of routine culture-based identification of S. pneumoniae which involves insensitive bile solubility and optochin susceptibility testing. This review will also consider the uses and limitations of various molecular assays used for identifying and serotyping of S. pneumoniae.
Section snippets
The role of a conventional method for identification and serotyping
The routine culture-based identification of S. pneumoniae involves bile solubility and optochin susceptibility testing (Lund and Henrichsen, 1978). However, these tests are not completely reliable due to the reports that indicate the presence of bile insoluble (2%) (Richter et al., 2008), optochin resistant strains (10%) of S. pneumoniae (Kellogg et al., 2001). Moreover, the presence of similar biochemical properties in a significant proportion of other closely related Streptococcus spp. makes
Role of molecular methods
In the last decade, different molecular assays were developed using an array of pneumococcal specific targets, like - pneumolysin (ply) (McAvin et al., 2001), autolysin (lytA) (Corless et al., 2001), pneumococcal surface antigen A (psaA) (Morrison et al., 2000), manganese-dependent superoxide dismutase (sodA) (Kawamura et al., 1999), and penicillin binding protein (pbp) (O'Neill et al., 1999) to name a few. But subsequent detection of these pneumococcal targets in viridans group streptococci (
MALDI-TOF
MALDI-TOF Mass Spectrometry (MS) is considered as a rapid tool for identification of bacteria in diagnostic laboratories (Van Veen et al., 2010). MALDI-TOF can identify the streptococcal genus, but could not differentiate the pneumococci and the mitis group. Recent studies have reported selective peaks in MALDI-TOF-MS pattern of S. pneumoniae and S. mitis group using ClinProTools™. Ikryannikova et al. (2013) identified a specific peak of 6949 m/z in S. mitis isolates alone but not seen in S.
Methods for accurate identification of serogroup/serotype of S. pneumoniae from nasopharyngeal specimens
Surveillance of S. pneumoniae serogroup/serotype is important in both disease, carriage and vaccine strategies. Each serotype differs in their disease-causing potential, age group infected, and geographic distribution (Shouval et al., 2006; Scott et al., 1996, Hausdorff et al., 2005). The direct impact of a vaccine can be determined by the reduction in vaccine serotypes causing invasive disease. The first molecular method recommended by CDC for serotyping of S. pneumoniae isolates was
Enhanced capturing of multiple serogroup/serotype of S. pneumoniae from nasopharyngeal sample
Since S. pneumoniae is part of the normal flora of the nasopharynx, carriage studies help to determine the serotypes circulating in the population as well to determine serotype replacement after vaccination. Colonization can be with a single serotype or with more than one serotype. For multiple serotype detection, five colonies from the primary culture have to be serotyped in order to identify the additional serotype which is in 50% proportion. If the second serotype is in 25% proportion 11
Conclusion
The routine use of culture based identification such as bile solubility and optochin susceptibility is not reliable due to the presence of similar properties in closely related Streptococcus spp. of respiratory sites. To overcome insensitivity of phenotypic methods, various different molecular methods (such as real time and conventional PCR targeting ply, lytA, psaA and sodA) have been developed to accurately identify and differentiate S. pneumoniae from other closely related species.
Acknowledgement
The authors would like to thank Dr. R.S. Suresh Kumar Mr. James John and Mr. Francis Yesurajan for their constructive comments on the manuscript.
References (82)
- et al.
Crossing the barrier: evolution and spread of a major class of mosaic pbp2x in Streptococcus pneumoniae, S. mitis and S. oralis
International Journal of Medical Microbiology
(2007) - et al.
Epidemiological differences among pneumococcal serotypes
Lancet Infect. Dis.
(2005) - et al.
Discrimination between Streptococcus pneumoniae and Streptococcus mitis based on sorting of their MALDI mass spectra
Clin. Microbiol. Infect.
(2013) - et al.
Chapter XI laboratory diagnosis, serology and epidemiology of Streptococcus pneumoniae
Methods Microbiol.
(1978) - et al.
Using whole genome sequencing to identify resistance determinants and predict antimicrobial resistance phenotypes for year 2015 invasive pneumococcal disease isolates recovered in the United States
Clin. Microbiol. Infect.
(2016) - et al.
lytA-based identification methods can misidentify Streptococcus pneumoniae
Diagn. Microbiol. Infect. Dis.
(2016) - et al.
Customized sequential multiplex PCR for accurate and early determination of invasive pneumococcal serotypes found in India
J. Microbiol. Methods
(2016) - et al.
Serotype replacement in disease after pneumococcal vaccination
Lancet
(2011) - et al.
Possible overestimation of penicillin resistant Streptococcus pneumoniae colonization rates due to misidentification of oropharyngeal streptococci
Diagn. Microbiol. Infect. Dis.
(2002) Preventing pneumococcal disease among infants and young children. Recommendations of the Advisory Committee on Immunization Practices (ACIP)
Accuracy of phenotypic and genotypic testing for identification of Streptococcus pneumoniae and description of Streptococcus pseudopneumoniae sp. nov
J. Clin. Microbiol.
DNA sequence sampling of the Streptococcus pneumoniae genome to identify novel targets for antibiotic development
Microb. Drug Resist.
Assigning strains to bacterial species via the internet
BMC Biol.
Efficacy, safety and immunogenicity of heptavalent pneumococcal conjugate vaccine in children
Pediatr. Infect. Dis. J.
Bacteremia due to viridans streptococcus in neutropenic patients with cancer: clinical spectrum and risk factors
Clin. Infect. Dis.
Immunoblot method to detect Streptococcus pneumoniae and identify multiple serotypes from nasopharyngeal secretions
J. Clin. Microbiol.
Bacteremia due to viridans streptococci that are highly resistant to penicillin: increase among neutropenic patients with cancer
Clin. Infect. Dis.
Progress in introduction of pneumococcal conjugate vaccine-worldwide, 2000–2012
Simultaneous detection of Neisseria meningitidis, Haemophilus influenzae, and Streptococcus pneumoniae in suspected cases of meningitis and septicemia using real-time PCR
J. Clin. Microbiol.
Revisiting pneumococcal carriage by use of broth enrichment and PCR techniques for enhanced detection of carriage and serotypes
J. Clin. Microbiol.
Potential nonpneumococcal confounding of PCR-based determination of serotype in carriage
J. Clin. Microbiol.
Non-pneumococcal mitis-group streptococci confound detection of pneumococcal capsular serotype-specific loci in upper respiratory tract
PeerJ
Population structure of Streptococcus oralis
Microbiology
Identity of viridans streptococci isolated from cases of infective endocarditis
J. Med. Microbiol.
Evolution of penicillin resistance in Streptococcus pneumoniae; the role of Streptococcus mitis in the formation of a low affinity PBP2B in S. pneumoniae
Mol. Microbiol.
rpoB gene sequence-based identification of aerobic Gram-positive cocci of the genera Streptococcus, Enterococcus, Gemella, Abiotrophia, and Granulicatella
J. Clin. Microbiol.
Identification of clinical Streptococcus pneumoniae isolates among other alpha and nonhemolytic streptococci by use of the Vitek MS matrix-assisted laser desorption ionization–time of flight mass spectrometry system
J. Clin. Microbiol.
Septicemia and shock syndrome due to viridans streptococci: a case-control study of predisposing factors
Clin. Infect. Dis.
Simple genetic method to identify viridans group streptococci by colorimetric dot hybridization and fluorometric hybridization in microdilution wells
J. Clin. Microbiol.
What happened to the streptococci: overview of taxonomic and nomenclature changes
Clin. Microbiol. Rev.
GyrB polymorphisms accurately assign invasive viridans group streptococcal species
J. Clin. Microbiol.
Partial sequence comparison of the rpoB, sodA, groEL and gyrB genes within the genus Streptococcus
Int. J. Syst. Evol. Microbiol.
Epidemiologic studies of Streptococcus pneumoniae in infants: acquisition, carriage, and infection during the first 24 months of life
J. Infect. Dis.
Accuracy of using the lytA gene to distinguish Streptococcus pneumoniae from related species
J. Med. Microbiol.
Mosaic genes and mosaic chromosomes: intra-and interspecies genomic variation of Streptococcus pneumoniae
Infect. Immun.
A homologue of aliB is found in the capsule region of nonencapsulated Streptococcus pneumoniae
J. Bacteriol.
Incidence of pneumococcal disease due to non-pneumococcal conjugate vaccine (PCV7) serotypes in the United States during the era of widespread PCV7 vaccination, 1998–2004
J Infect Dis
Differences in genotype and virulence among four multidrug-resistant Streptococcus pneumoniae isolates belonging to the PMEN1 clone
PLoS One
Diversity of PspA: mosaic genes and evidence for past recombination in Streptococcus pneumoniae
Infect. Immun.
Lack of utility of serotyping multiple colonies for detection of simultaneous nasopharyngeal carriage of different pneumococcal serotypes
Pediatr. Infect. Dis. J.
Characterization of nontypeable and atypical Streptococcus pneumoniae pediatric isolates from 1994 to 2010
J. Clin. Microbiol.
Cited by (36)
A phase 3 clinical trial of MINHAI PCV13 in Chinese children aged from 7 months to 5 years old
2021, VaccineCitation Excerpt :Pneumonia can be induced by various kinds of pathogens such as bacteria, viruses, and fungi. Among which, Streptococcus pneumoniae, also known as pneumococcus, remain to be the leading cause [4,5]. Besides, pneumococcus may also induce diverse invasive diseases such as sinusitis, otitis media, meningitis, septicemia, and bacteremia [6–8].
Streptococcus pneumoniae serotypes with meningeal tropism in cases of invasive pneumococcal disease. Community of Madrid, 2007-2018
2020, Enfermedades Infecciosas y Microbiologia ClinicaHigh-throughput proteotyping of bacterial isolates by double barrel chromatography-tandem mass spectrometry based on microplate paramagnetic beads and phylopeptidomics
2020, Journal of ProteomicsCitation Excerpt :While whole-cell MALDI-TOF mass spectrometry has quickly become a standard approach for identifying human pathogens because of its simplicity to implement and low cost [35,36], some limitations have been highlighted [1]. In particular, several microorganisms are not well identified at the species level; for example, Streptococcus pneumoniae and Streptococcus mitis cannot be easily discriminated with this methodology [37]. Tandem mass spectrometry proteotyping is based on many more experimental features than whole-cell MALDI-TOF mass spectrometry: thousands of peptide masses recorded with high accuracy (error below 5 ppm) versus less than one hundred protein molecular weights with low accuracy (error above 400 ppm), respectively.
Development of a novel MALDI-TOF MS-based bile solubility test for rapid discrimination of Streptococcus pneumoniae
2020, International Journal of Medical MicrobiologyCitation Excerpt :Discrimination of Streptococcus pneumoniae from other viridans streptococci is clinically important due to their different pathogenic potential (Spellerberg et al., 2019). However, reliable differentiation of pneumococci from other S. mitis group streptococci (SMGS) remains challenging despite the introduction of matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) (Burckhardt et al., 2017; Ikryannikova et al., 2013; Slotved et al., 2017; Spellerberg et al., 2019; van Prehn et al., 2016; Varghese et al., 2017). Using combination of different methods, the bile solubility test (BST) has been recommended for accurate species identification within SMGS (Satzke et al., 2013; Slotved et al., 2017).
Direct identification of pneumococcal serotypes in blood cultures by a PCR-reverse-hybridisation technique
2020, Enfermedades Infecciosas y Microbiologia ClinicaCitation Excerpt :The Quellung reaction is the gold standard for serotyping pneumococci. In recent years, some alternative serotyping procedures that include PCR, real-time PCR, sequencing analysis of specific genes, and whole genome sequencing, have been developed.1 Some of these techniques are reduced to a limited number of detectable serotypes2 or require expensive equipment (sequencing analysis).1