TY  - JOUR
AU  - Burghout, P.J.
AU  - Cron, L.E.
AU  - Gradstedt, H.
AU  - Quintero, B.
AU  - Simonetti, E.R.
AU  - Bijlsma, J.J.
AU  - Bootsma, H.J.
AU  - Hermans, P.W.M.
PY  - 2010
UR  - https://hdl.handle.net/2066/88184
AB  - The respiratory tract pathogen Streptococcus pneumoniae needs to adapt to the different levels of carbon dioxide (CO(2)) it encounters during transmission, colonization, and infection. Since CO(2) is important for various cellular processes, factors that allow optimal CO(2) sequestering are likely to be important for pneumococcal growth and survival. In this study, we showed that the putative pneumococcal carbonic anhydrase (PCA) is essential for in vitro growth of S. pneumoniae under the CO(2)-poor conditions found in environmental ambient air. Enzymatic analysis showed that PCA catalyzes the reversible hydration of CO(2) to bicarbonate (HCO(3)(-)), an essential step to prevent the cellular release of CO(2). The addition of unsaturated fatty acids (UFAs) reversed the CO(2)-dependent in vitro growth inhibition of S. pneumoniae strains lacking the pca gene (Deltapca), indicating that PCA-mediated CO(2) fixation is at least associated with HCO(3)(-)-dependent de novo biosynthesis of UFAs. Besides being necessary for growth in environmental ambient conditions, PCA-mediated CO(2) fixation pathways appear to be required for intracellular survival in host cells. This effect was especially pronounced during invasion of human brain microvascular endothelial cells (HBMEC) and uptake by murine J774 macrophage cells but not during interaction of S. pneumoniae with Detroit 562 pharyngeal epithelial cells. Finally, the highly conserved pca gene was found to be invariably present in both CO(2)-independent and naturally circulating CO(2)-dependent strains, suggesting a conserved essential role for PCA and PCA-mediated CO(2) fixation pathways for pneumococcal growth and survival.
TI  - Carbonic anhydrase is essential for Streptococcus pneumoniae growth in environmental ambient air.
EP  - 4062
SN  - 0021-9193
IS  - iss. 15
SP  - 4054
JF  - Journal of Bacteriology
VL  - vol. 192
N1  - 1 augustus 2010
DO  - https://doi.org/10.1128/JB.00151-10
L1  - https://repository.ubn.ru.nl/bitstream/handle/2066/88184/88184.pdf?sequence=1
ER  - 
TY  - JOUR
AU  - Simoes, A.S.
AU  - Sa-Leao, R.
AU  - Eleveld, M.J.
AU  - Tavares, D.A.
AU  - Carrico, J.A.
AU  - Bootsma, H.J.
AU  - Hermans, P.W.M.
PY  - 2010
UR  - https://hdl.handle.net/2066/88330
AB  - While performing surveillance studies in Oeiras, Portugal, designed to describe the impact of pneumococcal conjugate vaccine on colonization, we observed an increase from 0.7% in 2003 to 5% in 2006 in the prevalence of penicillin resistance (MIC of 2 to 6 mg/liter) among presumptively identified pneumococcal isolates. Although 15 of the 22 penicillin-resistant isolates obtained in 2006 were optochin resistant, they were bile soluble and thus considered to be bona fide pneumococci. This study aimed to clarify the nature of these isolates by using a combination of phenotypic and genotypic approaches that included routine strategies for pneumococcal identification, multilocus sequence analysis (MLSA), and comparative genomic hybridization (CGH). By MLSA, all isolates were classified as "streptococci of the mitis group" that, however, were distinct from typical Streptococcus pneumoniae or Streptococcus mitis. A single isolate was identified as Streptococcus pseudopneumoniae. CGH confirmed these findings and further indicated that a considerable part of the proposed pneumococcal core genome is conserved in these isolates, including several pneumococcal virulence genes (e.g., pavA, spxB, cbpE, and cbpD). These results suggest that among pneumococci and closely related streptococci, universal unique phenotypic and genetic properties that could aid species identification are virtually impossible to define. In pneumococcal colonization studies, when atypical strains are found, MLSA and CGH are informative tools that can be used to complement routine tests. In our study, after correct identification of the penicillin-resistant true pneumococci, we found that penicillin resistance levels among pneumococci remained stable from 2003 to 2006.
TI  - Highly penicillin-resistant multidrug-resistant pneumococcus-like strains colonizing children in Oeiras, Portugal: genomic characteristics and implications for surveillance.
EP  - 246
SN  - 0095-1137
IS  - iss. 1
SP  - 238
JF  - Journal of Clinical Microbiology
VL  - vol. 48
DO  - https://doi.org/10.1128/JCM.01313-09
L1  - https://repository.ubn.ru.nl/bitstream/handle/2066/88330/88330.pdf?sequence=1
ER  - 
TY  - JOUR
AU  - Vries, S.P.W. de
AU  - Hijum, S.A.F.T. van
AU  - Schueler, W.
AU  - Riesbeck, K.
AU  - Hays, J.P.
AU  - Hermans, P.W.M.
AU  - Bootsma, H.J.
PY  - 2010
UR  - https://hdl.handle.net/2066/88539
AB  - Moraxella catarrhalis is an emerging human-restricted respiratory tract pathogen that is a common cause of childhood otitis media and exacerbations of chronic obstructive pulmonary disease in adults. Here, we report the first completely assembled and annotated genome sequence of an isolate of M. catarrhalis, strain RH4, which originally was isolated from blood of an infected patient. The RH4 genome consists of 1,863,286 nucleotides that form 1,886 protein-encoding genes. Comparison of the RH4 genome to the ATCC 43617 contigs demonstrated that the gene content of both strains is highly conserved. In silico phylogenetic analyses based on both 16S rRNA and multilocus sequence typing revealed that RH4 belongs to the seroresistant lineage. We were able to identify almost the entire repertoire of known M. catarrhalis virulence factors and mapped the members of the biosynthetic pathways for lipooligosaccharide, peptidoglycan, and type IV pili. Reconstruction of the central metabolic pathways suggested that RH4 relies on fatty acid and acetate metabolism, as the genes encoding the enzymes required for the glyoxylate pathway, the tricarboxylic acid cycle, the gluconeogenic pathway, the nonoxidative branch of the pentose phosphate pathway, the beta-oxidation pathway of fatty acids, and acetate metabolism were present. Moreover, pathways important for survival under challenging in vivo conditions, such as the iron-acquisition pathways, nitrogen metabolism, and oxidative stress responses, were identified. Finally, we showed by microarray expression profiling that approximately 88% of the predicted coding sequences are transcribed under in vitro conditions. Overall, these results provide a foundation for future research into the mechanisms of M. catarrhalis pathogenesis and vaccine development.
TI  - Genome analysis of Moraxella catarrhalis strain RH4, a human respiratory tract pathogen.
EP  - 3583
SN  - 0021-9193
IS  - iss. 14
SP  - 3574
JF  - Journal of Bacteriology
VL  - vol. 192
N1  - 1 juli 2010
DO  - https://doi.org/10.1128/JB.00121-10
L1  - https://repository.ubn.ru.nl/bitstream/handle/2066/88539/88539.pdf?sequence=1
ER  - 
TY  - JOUR
AU  - Leegaard, T.M.
AU  - Bootsma, H.J.
AU  - Caugant, D.A.
AU  - Eleveld, M.J.
AU  - Mannsaker, T.
AU  - Froholm, L.O.
AU  - Gaustad, P.
AU  - Hoiby, E.A.
AU  - Hermans, P.W.M.
PY  - 2010
UR  - https://hdl.handle.net/2066/89734
AB  - Accurate differentiation between pneumococci and other viridans streptococci is essential given their differences in clinical significance. However, classical phenotypic tests are often inconclusive, and many examples of atypical reactions have been reported. In this study, we applied various phenotypic and genotypic methods to discriminate between a collection of 12 streptococci isolated from the upper respiratory tract of HIV-seropositive individuals in 1998 and 1999. Conventional phenotypic characterization initially classified these streptococci as Streptococcus pneumoniae, as they were all sensitive to optochin and were all bile soluble. However, they did not agglutinate with anti-pneumococcal capsular antibodies and were also far more resistant to antimicrobial agents than typeable pneumococci isolated in the same period. Genotypic characterization of these isolates and control isolates by both multilocus sequence analysis (MLSA) and comparative genomic hybridization (CGH) showed that only a single isolate was genetically considered to be a true S. pneumoniae isolate, and that the remaining 11 non-typable isolates were indeed distinct from true pneumococci. Of these, 10 most closely resembled a subgroup of Streptococcus mitis isolates genetically, while one strain was identified as a Streptococcus pseudopneumoniae isolate. CGH also showed that a considerable part of the proposed pneumococcal core genome, including many of the known pneumococcal virulence factors, was conserved in the non-typable isolates. Sequencing of part of the 16S rRNA gene and investigation for the presence of ply by PCR corroborated these results. In conclusion, our findings confirm the close relationship between streptococci of the Mitis group, and show that both MLSA and CGH enable pneumococci to be distinguished from other Mitis group streptococci.
TI  - Phenotypic and genomic characterization of pneumococcus-like streptococci isolated from HIV-seropositive patients.
EP  - 848
SN  - 0026-2617
IS  - iss. Pt 3
SP  - 838
JF  - Microbiology (New York)
VL  - vol. 156
N1  - 1 maart 2010
DO  - https://doi.org/10.1099/mic.0.035345-0
L1  - https://repository.ubn.ru.nl/bitstream/handle/2066/89734/89734.pdf?sequence=1
ER  -