Syndemics of fluoroquinolone resistance in N. gonorrhoeae, E. coli and K. pneumoniae: a global ecological analysis

It is unclear how important it is to reduce fluoroquinolone consumption in the general population to prevent the spread of fluoroquinolone resistance in Neisseria gonorrhoeae (bystander selection). Methods We assessed bystander selection by using Spearman’s correlation to assess if the country-level prevalence of fluoroquinolone resistance in N. gonorrhoeae was correlated with the prevalence of fluoroquinolone resistance in four other gram-negative species - Acinetobacter baumannii, Escherichia coli, Klebsiella pneumoniae and Pseudomonas aeruginosa. Results Fluoroquinolone resistance in N. gonorrhoeae was positively associated with homologous resistance in all 4 species - A. baumanii. (ρ=0.61, P=0.0003, E. coli (ρ=0.67, P<0.0001), K. pneumoniae (ρ=0.52, P=0.0004) and P. aeruginosa (ρ=0.40, P=0.0206). Positive associations were also found between the national prevalence of fluoroquinolone resistance and fluoroquinolone consumption in the general population in the preceding year for 4 of the 5 species. Conclusions Gonococcal fluoroquinolone resistance can be productively viewed as being part of a syndemic of fluoroquinolone resistance. Strengthening antimicrobial stewardship programs may help retard the spread of fluoroquinolone resistance in N. gonorrhoeae.


Abstract Background
It is unclear how important it is to reduce fluoroquinolone consumption in the general population to prevent the spread of fluoroquinolone resistance in Neisseria gonorrhoeae (bystander selection).

Methods
We assessed bystander selection by using Spearman's correlation to assess if the country-level prevalence of fluoroquinolone resistance in N. gonorrhoeae was correlated with the prevalence of fluoroquinolone resistance in four other gramnegative species -Acinetobacter baumannii, Escherichia coli, Klebsiella pneumoniae and Pseudomonas aeruginosa.

Background
Understanding the drivers of antimicrobial resistance (AMR) in N. gonorrhoeae is vital to prevent the emergence of untreatable gonorrhoea [1,2]. Until recently, it was thought that the major determinant of gonococcal AMR was exposure to antibiotics used to treat gonorrhoea [3][4][5]. Subsequently it was appreciated that N. gonorrhoeae is asymptomatic for much or most of the time it circulates in a population. This means that antibiotics used for other indications may cause AMR (bystander selection) [6].
Since gonorrhoea infections tend to cluster with other STIs, this bystander selection may be predominantly due to antibiotics used to treat other STIs (termed the STI bystander theory) [1,7,8]. Others have argued that antibiotic consumption for all indications plays a role (community bystander theory; Figure 1) [6,[9][10][11].
It is vital to establish which of these is the predominant driver of gonococcal AMR. If it is antimicrobials used to treat STIs, then preventing the further emergence of gonococcal AMR could be accomplished by interventions such as antimicrobial stewardship limited to within STI services. If total antibiotic consumption was an important determinant, then stewardship efforts to reduce antibiotic consumption in the whole community would be important [9].
Some studies have found evidence that antimicrobial consumption in the general population is associated with gonococcal AMR [9][10][11], whereas other have found no association [12]. We recently approached this problem from a novel angle. We tested for a country-level association between the prevalence of fluoroquinolone AMR between N. gonorrhoeae and 3 other pathobionts in European countries. We found that the prevalence of fluoroquinolone resistance in N. gonorrhoeae was positively associated with that in Pseudomonas aeruginosa, A. baumanii. and Klebsiella pneumoniae as well as being associated with fluoroquinolone consumption in the general population [13]. It was unclear how robust these findings were and if they applied outside the continent of Europe, where the variation in prevalence of fluoroquinolone resistance is much greater [2]. We therefore undertook to repeat the analysis using a global database of countries.

Antimicrobial Resistance Data
The AMR data for A. baumanii, Escherichia coli, Klebsiella pneumoniae and Pseudomonas aeruginosa was taken from the Center for Disease Dynamics, Economics & Policy's (CDDEP) ResistanceMap database. CDDEP aggregates data on antibiotic resistance from several sources. The data are then harmonized to present similar definitions of resistance across countries and regions to enable comparisons between countries. Further details pertaining to the methodology and definitions used to define antimicrobial resistance can be found at [14]. The list of sources used to obtain the data is provided in STable 1. CDDEP provides data on fluroquinolone resistance for 10 bacterial species but only 4 of these have data for more than 15 countries. We limited our analyses to these 4 species. For each of the species a resistance prevalence estimate from a single year for each country was provided in the dataset. This typically applied to the year 2017 (Table 1).

N. gonorrhoeae:
The prevalence of ciprofloxacin resistance per country was obtained from the WHO Global Gonococcal Antimicrobial Surveillance Programme (GASP).
GASP is a collaborative global network of regional and subregional reference laboratories that monitors gonococcal AMR in participating countries. The full GASP methodology, including suggested sampling strategy, laboratory techniques, external quality assurance, and internal quality control mechanisms has been published elsewhere [2]. A minimum inhibitory concentration (MIC) breakpoint of 1μg/ml was used to define resistance to ciprofloxacin [2]. GASP typically provides data for a number of years for each country. We selected the data applying to the same year as that used to provide the resistance estimate for the comparator species from the CDDEP data. estimates are reported as the number of standard doses (a dose is classified as a pill, capsule, or ampoule) per 1000 population per year [9].

Analyses
For each comparison, Spearman's correlation was used to assess the country-level association between the prevalence of AMR in N. gonorrhoeae and each of the other bacterial species in the same year. To test the association between fluoroquinolone resistance in each species and fluoroquinolone consumption Spearman's correlation was performed using the fluoroquinolone consumption preceding the year the resistance prevalence estimates were obtained from. The statistical analyses were performed in Stata 16.0. A P-value of <0.05 was regarded as significant.

Results
Data was available for between 42 and 60 countries for each of the resistance prevalence estimates per bacterial species (

Syndemics of fluoroquinolone resistance
The prevalence of ciprofloxacin resistance in N. gonorrhoeae was positively  Table 2; Fig. 3]. In the case of N. gonorrhoeae this association was not statistically significant (ρ=0.35, P=0.0620, N=28); Table 2].

Discussion
We found that the prevalence of gonococcal fluoroquinolone resistance was associated with fluoroquinolone resistance in all four of the gram-negative bacteria assessed. Fluoroquinolone AMR was significantly associated with consumption for four of the five bacteria assessed. These findings strengthen the conclusion from our previous study limited to Europe, that found that the fluoroquinolone resistance in N.
gonorrhoeae can be productively viewed as a syndemic of resistance in other gram negatives [13].
The most parsimonious way to explain these findings is that high fluoroquinolone consumption in the general population is partly responsible for driving fluoroquinolone resistance in all these species. This finding has important consequences. It suggests that reducing fluoroquinolone consumption to below certain thresholds could reduce the probability of resistance sprreading in all of these bacterial species [15,16]. This may be particularlry important for N. gonorrhoeae. As combined high-level resistance to ceftriaxone and azithromycin emerges in N. gonorrhoeae, interest is mounting in treating gonorrhoea with single dose oral ciprofloxacin [17,18]. Studies have confirmed that this strategy results in close to 100% eradication, if preceded by rapid screening for molecular markers of fluoroquinolone resistance [17]. This strategy has also been found to result in reduced use of ceftriaxone thereby reducing selection pressure for cephalosporin resistance [17]. This strategy is, however, not possible in numerous countries in East Asia, where the prevalence of ciprofloxacin resistance is close to 100% [2]. Studies from Europe have noted declines in ciprofloxacin resistance in the last decade related to changes in treatment protocols and reductions in fluoroquinolone use [13,19]. These findings provide additional motivation for high fluoroquinolone consumption countries/populations to reduce their fluoroquinolone use in order to preserve these agents as a treatment option for gonorrhoea.
Fluoroquinolone use is also rapidly increasing in a number of relatively low consumption countries, such as countries in sub Saharan Africa where the prevalence of gonococcal fluoroquinolone resistance is low [2,9,20]. Our findings suggest that these countries may be advised to limit this increase to safer limits.
As in our previous study fluoroquinolone resistance in N. gonorrhoeae was most strongly associated with that in E. coli. Likewize the association between fluoroquinolone consumption and resistance was strongest in E. coli. These findings are congruent with the fact that E. coli is the most prevalent of all the bacteria considered in the general human population [6]. This suggests that monitoring fluoroquinolone susceptibility in E. coli may function as a useful early warning system to monitor excess fluoroquinolone consumption in populations at risk for AMR [21,22].
Our analysis is subject to a number of limitations. Chief amongst these are that the AMR prevalence estimates from both GASP and CDDEP are based on various methodologies making cross country comparisons problematic. We did not adjust our analyses for either differences in susceptiblity testing strategies or breakpoints between countries or over time as this information is not provided by GASP or CDDEP.
These limitations should however result in a misclassification bias which would typically result in a bias towards the null hypothesis [23]. This would be expected to reduce the strength of any association found.
The epidemiology of resistance is complex and factors other than the amount of fluroquinolones consumed by humans may influence the level of fluoroquinolone resistance. These include consumption of other classes of antimicrobials, clonal spread of resistance independent of fluoroquinolone consumption, travel by humans and consumption of antimicrobials for acquaculture and livestock production [1,2,20,24,25]. Future research should strive to develop models that incorporate all these factors so as to be able to better predict and ultimately prevent the further emergence of fluoroquinolone resistance in N. gonorrhoeae and other bacteria. In the absence of these studies, the evidence provided by this analysis provides additional evidence to motivate policy makers, prescribers and consumers in high fluoroquinolone consumption populations to reduce consumption.     and Neisseria gonorrhoeae (E).