1. PLoS One. 2016 May 26;11(5):e0154816. doi: 10.1371/journal.pone.0154816.
eCollection 2016.

Molecular Analysis of Rising Fluoroquinolone Resistance in Belgian Non-Invasive
Streptococcus pneumoniae Isolates (1995-2014).

Ceyssens PJ(1), Van Bambeke F(2), Mattheus W(1), Bertrand S(1), Fux F(1), Van
Bossuyt E(1), Damée S(1), Nyssen HJ(3), De Craeye S(3), Verhaegen J(4); Belgian
Streptococcus pneumoniae Study Group, Tulkens PM(2), Vanhoof R(1).

Author information: 
(1)Unit of Bacterial Diseases, Scientific Institute of Public Health (WIV-ISP),
1050 Brussels, Belgium. (2)Pharmacologie cellulaire et moléculaire, Louvain Drug 
Research Institute, Université Catholique de Louvain, 1200 Brussels, Belgium.
(3)Unit of Foodborne Pathogens, Scientific Institute of Public Health (WIV-ISP), 
1050 Brussels, Belgium. (4)Laboratory of Clinical Bacteriology and Mycology,
KULeuven, 3000 Leuven, Belgium.

We present the results of a longitudinal surveillance study (1995-2014) on
fluoroquinolone resistance (FQ-R) among Belgian non-invasive Streptococcus
pneumoniae isolates (n = 5,602). For many years, the switch to respiratory
fluoroquinolones for the treatment of (a)typical pneumonia had no impact on FQ-R 
levels. However, since 2011 we observed a significant decrease in susceptibility 
towards ciprofloxacin, ofloxacin and levofloxacin with peaks of 9.0%, 6.6% and
3.1% resistant isolates, respectively. Resistance to moxifloxacin arised
sporadically, and remained <1% throughout the entire study period. We observed
classical topoisomerase mutations in gyrA (n = 25), parC (n = 46) and parE (n =
3) in varying combinations, arguing against clonal expansion of FQ-R. The impact 
of recombination with co-habiting commensal streptococci on FQ-R remains marginal
(10.4%). Notably, we observed that a rare combination of DNA Gyrase mutations
(GyrA_S81L/GyrB_P454S) suffices for high-level moxifloxacin resistance,
contrasting current model. Interestingly, 85/422 pneumococcal strains display
MICCIP values which were lowered by at least four dilutions by reserpine,
pointing at involvement of efflux pumps in FQ-R. In contrast to susceptible
strains, isolates resistant to ciprofloxacin significantly overexpressed the ABC 
pump PatAB in comparison to reference strain S. pneumoniae ATCC 49619, but this
could only be linked to disruptive terminator mutations in a fraction of these.
Conversely, no difference in expression of the Major Facilitator PmrA, unaffected
by reserpine, was noted between susceptible and resistant S. pneumoniae strains. 
Finally, we observed that four isolates displayed intermediate to high-level
ciprofloxacin resistance without any known molecular resistance mechanism.
Focusing future molecular studies on these isolates, which are also commonly
found in other studies, might greatly assist in the battle against rising
pneumococcal drug resistance.

PMID: 27227336  [PubMed - in process]