Multidrug-resistant bacteria are present throughout the world [1, 3, 4]. The global movement of these pathogens through leisure travel, medical tourism, and military casualty movement is concerning [2–4]. For military personnel, one proposed source of MDR bacteria has been pathogen introduction into deployed military treatment facilities through host nation patients, with subsequent nosocomial transmission to US personnel. However, most of the data supporting this conclusion were from studies completed over five years ago and focused on MDR Acinetobacter[8, 9]. During 2009 and 2010, increasing numbers of casualties were noted to be colonized with ESBL-producing E. coli, although no source has been elucidated for this possible epidemiological shift in MDR organisms . In this study, 2% of US-based personnel (without recent antibiotic exposure, overseas travel, or active infection) were colonized with MDR E. coli in the perirectal region, whereas Afghanistan-based personnel showed an 11% colonization prevalence of MDR E. coli. This finding demonstrates significantly higher (5.5-fold, p<0.01) colonization in individuals based primarily on their deployed status, as their prior healthcare system exposure was also minimal. This finding is consistent with a prior study demonstrating prevalent and incident GNR colonization in recently deployed, hospitalized individuals compared to non-deployed hospitalized patients . In contrast to this previous study, which demonstrated a 60% clonality of incident MDR A. baumannii isolates and 25% clonality of incident ESBL-producing isolates , our study did not show significant strain-relatedness or isolate clonality in non-hospitalized, uninjured, healthy Soldiers, regardless of deployed status. The difference in these findings may be attributed to the increased risk of specific MDRO exposure and antimicrobial pressure in healthcare facilities during casualty evacuation in the previous study. No other bacterial species showed broad antimicrobial resistance in our study. While increased colonization rates could explain the increase in ESBL-producing isolates recovered from combat casualties, a comparison of 465 E. coli isolates originating from combat-injured patients in Iraq, Afghanistan and the US to isolates in this study found no matching PFGE patterns (data not shown).
The finding of asymptomatic, healthy people colonized with MDR E. coli, despite minimal contact with the healthcare system, is concerning. Although numerous studies have reported community associated ESBL-producing E. coli urinary tract infections, most have some risk factor for a MDR-GNB infection: genitourinary pathology, previous bacterial infections, prior intravenous antibiotic treatments, and hospitalization in the previous 12 months [5, 18]. A study from Madagascar reported a 10% stool colonization rate among healthy people, but the study reported numerous MDR bacterial species (e.g. E. coli, Klebsiella pneumoniae, Enterobacter cloacae, and Citrobacter freundii) and an association of colonization with socioeconomic status . Other studies have reported MDR stool colonization rates between 1-7%; however, these studies included patients with substantial exposure to the healthcare system and not young, healthy patients as observed in our study [20, 21].
We noted that non-MDR E. coli recovered from Afghanistan-based study participants was associated with higher antimicrobial resistance, with potential clinical implications associated specifically with ampicillin-sulbactam and tetracycline resistance. As has been previously reported in patients with traumatic abdominal wounds, Enterobacteriaceae may demonstrate significant resistance to ampicillin-sulbactam, risking inadequate therapy in up to 50% of patients . This is consistent with our study finding of 54% ampicillin-sulbactam resistance in non-MDR E.coli isolates from the Afghanistan-based participants. This increased antimicrobial resistance might be associated with exposure to doxycycline for malaria chemoprophylaxis. A previous study assessing the impact of tetracycline and doxycycline exposure on stool bacteria showed no increase in pathogen quantities, to include Klebsiella, Enterobacter, Pseudomonas, Proteus, Serratia, or E. coli. Although the bacteria had increased antimicrobial resistance after tetracycline exposure, this was not observed after doxycycline exposure . Other studies have reported increased resistance after doxycycline exposure and some change in gut flora, but not significantly different than prior to therapy [24, 25]. We did not identify genes consistently responsible for tetracycline resistance across the isolate spectrum, but those primarily involved were tet(A) and tet(B), which is similar to what is described in the literature [26–28].
There is also concern that antimicrobial use in animal husbandry practices might be leading to greater resistance. This has been shown in tetracycline-focused studies that linked tet(M), tet(A), and tet(B) genes to introns and dairy feed . In the current study, it is worth noting that there was greater tetracycline resistance of the non-MDR E. coli isolates associated with the tet(A) gene, but no clonality was detected by PFGE. There are reports of TEM-1, CXT-M-15 and tet(A) genes carried in the same plasmid, raising concern that doxycycline use might not only be selecting its own resistance, but also ESBL-mediated resistance [29, 30]. Increased antibiotic use in food production processes may play a role in this microbial epidemiological finding and warrant further investigation.
It is unclear what role the environment played on the MDR colonization rate of personnel deployed to Afghanistan. Although the participants resided in a remote area of Afghanistan, they had more than adequate potable water for all activities, including bathing, cooking and drinking. The study participants likely had minimal to no consumption of local food, as off-base exposure did not correlate with colonization status. It is also unclear if the use of different swab type for MDR pathogen collection, or the delay in laboratory processing of swabs from Afghanistan, impacted the recovery of pathogens. The overall similarity in total number of pathogens recovered between the two study sites, the similar body areas colonized between the two study sites, and data supporting stability of bacteria on swabs for extended periods of time, suggest that the observed differences are valid. It may be of interest to further assess seasonal variation of MDR pathogen colonization, as our study evaluated individuals primarily in spring and summer months, and may not be representative of colonization prevalence during other seasons. Previous studies have shown seasonal variation in GNB infection rates, with increased incidence of K. pneumoniae, A. baumannii, and E. coli infections in summer months [31, 32]. Further study limitations include the relatively low percentage of female participants (31% in the US-based study population and no females in the Afghanistan-based population), along with the lack of other potentially relevant demographic data, such as race. These limitations may preclude generalization of our findings to other populations.
Finally, the current study identified a point prevalence of MDR colonization in geographically distinct military populations. Prospective studies on military service members prior to deployments, and prior to malaria chemoprophylaxis/antibiotic exposure, are needed to determine the true incidence of colonization with potential pathogens over time. This study identified antimicrobial resistance patterns that may have implications on clinical treatment decisions, particularly in light of recent literature reports on β-lactamase inhibitor use for ESBL-producing bacteria. For example, we identified MDR E. coli colonization isolates that show relative susceptibility to piperacillin-tazobactam, but resistance to ampicillin-sulbactam. These findings would seem consistent with a recent study suggesting clinical efficacy against ESBL-producing E. coli bloodstream infections with these antimicrobial agents . The findings in the current study may also impact the choice of early empiric antimicrobial therapy for traumatic injuries during war and possibly humanitarian/disaster relief missions, which may have similar post-injury infection risks. Historically, early antimicrobial therapy in these environments has included agents without enhanced coverage against ESBL-producing bacteria, due to concern for further selection of antimicrobial resistance, and the fact that we have not typically encountered those resistant pathogens early after injury . The recommendation to avoid broader early antimicrobial therapy might need to be reconsidered, especially for injuries involving the perineal and perirectal areas, as these are associated with higher ESBL-producing E. coli colonization rates than other body regions.