How does the misuse of disinfectants contribute to the selection of resistant strains?

Inadequate use of biocides — subinhibitory concentrations, insufficient contact time, irrational product selection, or application on contaminated organic surfaces — exerts documented microbiological selective pressure, favoring the emergence of strains with acquired resistance and cross-resistance to antibiotics. This article synthesizes the molecular mechanisms involved, evidence from the international specialized literature, and recommendations from international organizations, providing clinicians with a practical framework for action.

Table of Contents:

1. The global epidemiological context — why it matters
2. Misuse of disinfectants — frequent clinical scenarios
3. Molecular mechanisms of biocide-induced resistance
4. Biocide–antibiotic cross-resistance: relevant clinical evidence
5. Selective pressure and the ecological dynamics of resistance
6. Best practice principles in disinfectant use — framework for action for specialists
7. Disinfectants are not automatically safe or effective — what the specialist must do

1. The global epidemiological context — why it matters

Antimicrobial resistance (AMR) represents one of the most serious threats to global public health. According to ECDC data, more than 35,000 people die annually in the European Union, Iceland, and Norway as a result of infections caused by antimicrobial-resistant bacteria — a figure comparable to the combined mortality from seasonal influenza, tuberculosis, and HIV/AIDS in the same region.

The debate regarding AMR has traditionally focused on the irrational prescribing of antibiotics and their excessive use in veterinary medicine and animal husbandry — recognized sources of transmissible resistance in humans (ECDC/EFSA/EMA, JIACRA IV, 2024). However, a rapidly expanding body of scientific evidence demonstrates that biocides — including disinfectants used in hospital settings, antiseptics, and sanitizing products in ambulatory medicine — can themselves be an active factor in the selection and amplification of bacterial resistance when used incorrectly.

The European Commission's Scientific Committee on Emerging and Newly Identified Health Risks (SCENIHR) (2009) explicitly concluded that "the selective pressure exerted by biocides may favor bacteria expressing resistance mechanisms... which could create a potential risk of developing co-resistance between antibiotics and biocides" and that prudent and appropriate use of disinfectants is essential to maintain their effectiveness.

•       Insufficient contact time — the disinfectant is wiped off mechanically before the expiry of the action time indicated by the manufacturer

•       Application on surfaces with residual organic load, which chemically inactivates the active substance (blood, exudates, protein material) before it can act

•       Use of the same biocide product on multiple surfaces or in areas with high microbiological pressure without adhering to rotation schedules according to infection control guidelines

•       Incorrect storage of ready-to-use solutions, with gradual degradation of the active substance and unperceived decrease in effective concentration

•       Excessive, unjustified use of antiseptic and disinfectant products in non-critical contexts (surfaces without documented infectious risk), with the consequent contamination of the surrounding environment through sub-lethal biocide residues

3. Molecular mechanisms of biocide-induced resistance

3.1 Efflux pumps — the first line of bacterial defense

The fastest and most efficient mechanism by which bacteria respond to chemical stress is the activation of efflux pumps. The qac (quaternary ammonium compound resistance genes), frequently located on plasmids transmissible via conjugative transfer, encode transport proteins that actively expel the biocide from the cell before it reaches a lethal intracellular concentration.

What makes this mechanism particularly concerning from a clinical perspective is that efflux pumps — particularly the AcrAB-TolC family in Gram-negative bacteria — are polyvalent: they simultaneously expel both biocides and antibiotics (fluoroquinolones, beta-lactams, aminoglycosides, tetracyclines). Thus, a bacterium "educated" by an incorrectly used disinfectant can simultaneously become resistant to multiple classes of antibiotics — a phenomenon termed co-resistance.

A study published in the Journal of Hospital Infection (2024) documents that exposure to subinhibitory concentrations of chlorhexidine increased MICs for ampicillin, tetracycline, vancomycin, gentamicin, cefotaxime, cefuroxime, and ciprofloxacin in a susceptible S. aureus isolate, demonstrating the clinical breadth of this phenomenon.

3.2 Changes in cell membrane permeability

Gram-negative bacteria with increased resistance to chlorhexidine exhibit adaptive changes in the composition of the outer membrane — modification of the lipopolysaccharide profile and porins — which reduce permeability not only to the biocide but also to antibiotics such as ciprofloxacin, imipenem, cefotaxime, ceftazidime, gentamicin, and aztreonam. Studies on clinical isolates of E. coli, K. pneumoniae, P. aeruginosa, and A. baumannii have confirmed this association (Clinical and Experimental Medicine, Springer, 2025).

3.3 Biofilm formation

Exposure to sublethal concentrations of disinfectants can paradoxically stimulate biofilm formation. Bacteria inside a mature biofilm are 150 to 3,000 times more resistant to chlorine-based disinfectants than planktonic cells (data reproduced from LeChevallier et al.). The biofilm acts as a physicochemical barrier: disinfectant penetration is mechanically inhibited by the extracellular polysaccharide matrix, and local consumption of the active substance (through reactions with organic matter) prevents it from reaching an effective concentration at the level of deep cells.

From a clinical perspective, infectious biofilm — on vascular catheters, prostheses, implants, ventilation lines — generates chronic infections that are extremely difficult to eradicate with systemic antibiotic therapy, and bacterial resistance within the biofilm is frequently not detected by standardized susceptibility tests on planktonic cultures.

3.4 Horizontal gene transfer (HGT) — disseminating resistance beyond a single strain

Perhaps the most relevant mechanism from the perspective of hospital epidemiology is the ability of bacteria to horizontally transfer resistance genes — via plasmid conjugation, phage transduction, and transformation. Biocide exposure actively stimulates HGT, allowing resistance genes acquired by one strain to be rapidly disseminated to other bacterial species within the same microbiological ecosystem.

Studies published in Communications Medicine (Nature, 2022) demonstrate that exposure to sub-MIC concentrations of disinfectants triggers bacterial stress responses that induce an increased mutation frequency and stimulate HGT, allowing for the rapid accumulation of new resistance mechanisms. Resistance genes tend to co-reside on plasmids or integrative conjugative elements, transferring together — creating the premises for the simultaneous co-selection of resistance to multiple antimicrobials.

3.5 Bacterial dormancy states — persister cells and VBNC

An often-overlooked aspect is the ability of bacteria to survive biocide exposure by entering metabolically dormant states: persister cells (superficial dormancy) and VBNC — Viable But Non-Culturable cells (deep dormancy). These cells survive biocide exposure, are not detected by standard cultures, but can return to an active phenotype — a process called resuscitation — with the frequent acquisition of additional mutations that confer stable resistance.

4. Biocide–antibiotic cross-resistance: relevant clinical evidence

The term cross-resistance refers to the situation where a resistance mechanism acquired against a biocide simultaneously confers reduced susceptibility to one or more antibiotic agents, even in the absence of direct antibiotic exposure. This is by far the most important clinical consequence of the incorrect use of disinfectants.

Quaternary ammonium compounds (QAC / BKC — benzalkonium chloride)

QACs are the most used surface disinfectants in healthcare settings. Studies published in Clinical and Experimental Medicine (Springer, 2025) document that MRSA strains with increased resistance to BKC exhibit increased resistance to various beta-lactams and ofloxacin. At the mechanism level, the overexpression of efflux pumps in response to QACs simultaneously expels antibiotics from completely different families.

Chlorhexidine (CHX)

Chlorhexidine is the reference antiseptic in surgical hand hygiene and surgical field preparation. However, accumulated data highlights that exposing Gram-negative bacteria to subinhibitory concentrations of CHX increases MICs for colistin, ceftazidime, amikacin, meropenem, gentamicin, and piperacillin/tazobactam — last-line antibiotics for multidrug-resistant bacteria. A study on Klebsiella pneumoniae (Wand et al., 2017) demonstrated that adaptation to CHX increases resistance to colistin — the last therapeutic resort for some MDR Gram-negative infections.

Sodium hypochlorite

Widely used for surface disinfection, sodium hypochlorite has been associated, in experimental studies, with increased MICs in P. aeruginosa isolates against colistin, ceftazidime, amikacin, meropenem, gentamicin, and piperacillin, after incubation with subinhibitory concentrations of the biocide.

5. Selective pressure and the ecological dynamics of resistance

The concept of selective pressure is essential for understanding how the incorrect use of disinfectants amplifies resistance at the scale of the hospital microbiological ecosystem. Any antimicrobial agent — be it an antibiotic or a biocide — applied at a concentration not high enough to kill all members of a microbial population does not produce sterilization, but rather Darwinian selection: strains with reduced susceptibility survive preferentially, multiply, and become dominant.

Data published in van Dijk et al. (Communications Medicine, 2022) and Murray et al. (npj Antimicrobials and Resistance, 2024) add a critical ecological perspective: disinfectants reach surface waters and soil through sewage systems and sewage sludge fertilization. Progressive dilution generates precisely the sub-MIC concentrations that constitute the most efficient determinant of adaptive selection. Thus, the healthcare facility is not an isolated system — hospital disinfection practices contribute to a global environmental reservoir of resistance genes, with documented circulation between the environment, animals, and humans (the One Health perspective).

The ECDC has repeatedly emphasized that "the use of antimicrobials exerts ecological pressure on microorganisms and contributes to the emergence and selection of antimicrobial-resistant microorganisms in populations" and that AMR management explicitly includes the prudent use of all antimicrobials, including biocides.

6. Best practice principles in the use of disinfectants — action framework for specialists

6.1 Rational selection of the biocidal product

The choice of disinfectant must be based on the documented spectrum of activity against target microorganisms identified in the facility, the risk class of the surface or medical instrument/device (Spaulding classification: critical/semi-critical/non-critical), and efficacy evidence from validated clinical studies, not exclusively on cost or availability.

Biocidal products authorized in the EU are regulated by Regulation (EU) 528/2012 concerning biocidal products. The exclusive use of products with valid authorization and those appearing on the positive list of approved active substances reduces the risk of selection for resistance through exposure to substances with partially validated antimicrobial activity.

6.2 Compliance with usage parameters

Concentration: Preparation of working solutions strictly according to manufacturer instructions, using calibrated dosing systems. Empirical dilution — even with a variation of ±20–30% from the recommended concentration — can transform an effective bactericidal disinfectant into a bacterial selection agent.

Contact time: Compliance with the action time indicated on the product technical data sheet is mandatory. Disinfectant applied and wiped off immediately does not act correctly — it exerts sublethal selective pressure. Medical staff must be explicitly instructed not to wipe surfaces before the contact time has expired.

Surface preparation: Effective disinfection requires, mandatorily, prior mechanical cleaning to remove organic load. Applying disinfectant to surfaces visibly contaminated with organic matter dramatically reduces the effective concentration available through chemical inactivation.

6.3 Rotation and diversification of biocidal products

The chronic, exclusive use of the same biocidal product in the same healthcare facility creates optimal conditions for the gradual adaptation of the resident microbiome. Rotation programs for classes of disinfectants — implemented based on the recommendations of infection control teams — reduce the probability of selecting mutants with stable resistance to a single active substance.

6.4 Avoiding unjustified use in non-critical contexts

CDC and ECDC point out that disinfection of non-critical surfaces (routine flooring, surfaces without direct contact with patients or sterile materials) does not provide demonstrated clinical benefits over routine cleaning with detergents. The use of disinfectants on large areas, without a clear clinical indication, increases environmental selective pressure and contributes to the contamination of the environment with sublethal biocidal residues.

6.5 Susceptibility monitoring and resistance surveillance

Implementing an active microbiological surveillance program — including determining MICs for biocides frequently used in the facility, in parallel with routine antibiograms — allows for early detection of phenotypic changes in clinical isolates. Identification of strains with reduced susceptibility to biocides must trigger an immediate analysis of disinfection practices and a potential protocol restructuring.

7. Disinfectants are not automatically safe or effective — what the specialist must do

Health professionals have the responsibility to treat the disinfection protocol with the same rigor with which they treat the prescribing of an antibiotic. Product selection, working concentration, contact time, surface preparation, and product rotation are not administrative details — they are clinical decisions with direct epidemiological consequences.

Infection control teams must include in periodic audits not only antibiotic consumption (classical antimicrobial stewardship) but also biocide stewardship — assessing the rationality of the use of disinfectants and antiseptics, compliance with protocols, and possible correlations with local patterns of microbiological resistance.

Continuous training of all medical and paramedical staff in the field of correct disinfection practices represents an investment with a direct return in reducing the incidence of healthcare-associated infections (HAIs) and in preserving the effectiveness of the available therapeutic arsenal.

The relationship between the incorrect use of disinfectants and the selection of resistant strains is supported by a solid and growing body of scientific evidence, validated by international reference organizations (ECDC, SCENIHR/European Commission, WHO, CDC). The mechanisms involved — overexpression of efflux pumps, changes in membrane permeability, biofilm formation, horizontal gene transfer, and induction of bacterial dormant states — are documented at the molecular level and have clinical consequences demonstrable through increased MICs for critically important antibiotics.

Preventing these consequences does not require exceptional resources — it requires procedural discipline, continuous training, and active surveillance. The rational use of disinfectants, integrated into the broader strategy of combating antimicrobial resistance, is a collective responsibility of the entire medical community.

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