Role of penA polymorphisms for penicillin susceptibility in Neisseria lactamica and Neisseria meningitidis
Introduction
Neisseria meningitidis (N. meningitidis, meningococci) is a facultative pathogenic bacterium that asymptomatically colonizes the nasopharynx of approximately 10% of the human population (Gold et al., 1978, Cartwright et al., 1987). Occasionally, N. meningitidis causes severe diseases like meningitis and septicemia (Rosenstein et al., 2001). The apathogenic species N. lactamica is closely related to meningococci and shares the same habitat (Guibourdenche et al., 1986). However, N. lactamica colonizes predominately infants and toddlers, whereas carriage of meningococci is much more frequent in adolescents and young adults (Gold et al., 1978, Cartwright et al., 1987, Bennett et al., 2005, Kristiansen et al., 2012).
Penicillin G remains a first-line agent against invasive meningococcal infections (Nadel and Kroll, 2007), but in the last years the number of meningococcal isolates that displayed reduced penicillin susceptibility increased in Europe and other parts of the world (Richter et al., 2001, Tapsall et al., 2001, Vazquez et al., 2007, Bertrand et al., 2012, Bijlsma et al., 2014). Between 2002 and 2007, 14.1% of the invasive meningococci in Germany were intermediate susceptible or resistant towards penicillin (Vogel and Frosch, 2008). Five specific amino acid polymorphisms in the transpeptidase region of penicillin binding protein 2 (PBP2), encoded by the penA gene, showed high correlation with reduced penicillin susceptibility (Thulin et al., 2006, Taha et al., 2007). Commensal Neisseria spp. have been suggested to be a source for altered penA alleles (Spratt et al., 1989, Spratt et al., 1992, Saez-Nieto et al., 1990, Lujan et al., 1991). In a Spanish study, most N. lactamica isolates showed decreased susceptibility to penicillin (Arreaza et al., 2002). There are, however, no data on penA sequences of N. lactamica.
In this study, penicillin susceptibility and penA alleles were analyzed for 123 N. lactamica and 129 meningococcal strains isolated in Germany. Furthermore, transformation experiments were conducted to investigate the effect of penA transfer from N. lactamica to N. meningitidis.
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Bacterial isolates and antimicrobial resistance testing
N. lactamica strains (n = 123) were collected during the Bavarian meningococcal carriage study in winter 1999/2000 (Alber et al., 2001). N. meningitidis strains (n = 129) were randomly selected from invasive isolates received by the German reference laboratory for meningococci in 2006. For transformation experiments the capsule null locus (cnl) meningococcal strain α14 (Claus et al., 2002, Schoen et al., 2008) was used.
MICs of penicillin G were determined with Etest® strips (bioMérieux, Nürtingen,
Penicillin susceptibility of N. lactamica and N. meningitidis
In total, 123 N. lactamica strains and 129 invasive meningococci were analyzed by Etest® to determine their penicillin G susceptibility. MIC values in N. lactamica (range: 0.064–2.0 mg/L, MIC50 = 0.38) were significantly higher than those in N. meningitidis (range: 0.016–0.25 mg/L, MIC50 = 0.064) (p ≤ 0.001) (Fig. 1A, Table 2). Forty-five of the N. lactamica strains, for which β-lactamase production was ruled out, but none of the meningococcal strains were resistant to penicillin. Only three N.
Discussion
For a collection of Spanish N. lactamica isolates it was shown that penicillin MIC values were higher than those found in meningococci (Arreaza et al., 2002). These results could be confirmed in the present study for a German strain collection, suggesting that at least in Europe penicillin MICs are higher in N. lactamica than in N. meningitidis. Genetic analysis of the 3′ region of the penA gene revealed that it was much more variable in N. lactamica than in meningococci and both species had
Acknowledgements
The German reference laboratory for meningococci and Haemophilus influenzae is supported by the Robert Koch-Institute with funds of the German Federal Ministry of Health (funding code 1369-237). Moreover, we are grateful to Eva Hong, the curator of the penA database. This publication made use of the Neisseria Multi Locus Sequence Typing website (http://pubmlst.org/ neisseria/) developed by Keith Jolley and sited at the University of Oxford (Jolley and Maiden, 2010). The development of this site
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