Antimicrobial resistance (AMR) is now recognised as a major public health crisis as essential antimicrobial drugs including antibiotics, antifungals, antivirals, antimalarials and anthelmintics [1] become less effective therapeutic options. Continued antimicrobial misuse and overuse in human and animal medicine, and poor prevention and control strategies have proliferated AMR and hurdled the planet into a post antibiotic era. The unwarranted prescription of antibiotics by general practitioners and veterinarians in the absence of diagnostic indicators, as metaphylactics, prophylactics and growth promotors, greatly proliferates AMR. Indeed, poor diagnostics, particularly when disease aetiology for bacterial, fungal, or viral infectious diseases is similar, encourages the misuse and overprescription of antibiotic agents [2]. The immense application of antibiotic agents in food production (agriculture and aquaculture) is also recognised as a major contributor to the emergence and proliferation of AMR. Globally, 100–200 thousand tonnes or 80% of antibiotics are used in food production annually [3], with an increase of 67% predicted by 2030 across all major livestock industries and aquaculture [4]. Europe has implemented bans on the use of growth-promoting antibiotics in food-producing animals, the United States and China, however, are more lenient, with 52% of antibiotics administered in China for growth-promoting activity alone [5]. Globally, AMR results in prolonged morbidity, increased mortality, economic burden, socioeconomic impacts and greatly hampers the success of Sustainable Development Goals, including the provision of maternal and child health, food security, poverty reduction and economic growth [6]. Methicillin-resistant Staphylococcus aureus (MRSA), for example, is the most common Gram-positive multidrug-resistant (MDR) pathogen causing morbidity and mortality globally [7]. Candida auris is an emerging multidrug-resistant nosocomial fungus and is a major threat in healthcare settings [8]. Moreover, global disease outbreaks are becoming a constant threat, as is evident by the emergence of the highly pathogenic human coronaviruses, including SARS-CoV-2 (COVID-19), SARS-CoV-1 (SARS) and the Middle East respiratory syndrome (MERS-CoV). Studies report that COVID-19 can survive and remain infective for approximately 9 days on surfaces [9], making it highly transmissible.

As global initiatives push for research and development into novel antimicrobial agents for use as stand-alone or combination therapy options, there is also a need to establish strategies and preventative measures to reduce AMR. Effective disinfection and sanitation strategies are key in preventing communicable disease transmission in both human and animal environments. Biocides, which are chemicals used as sanitizers and disinfectants, consist of specific formulations containing one or more active ingredients that nonspecifically and fatally target microbial species. Typical commercial biocides used in clinical, industrial and domestic settings consist of quaternary ammonium compounds (QACs), benzalkonium chloride (BAC), chlorine and chlorine-based derivatives, acid anionic agents, hydrogen peroxide (H₂O₂), biguanides (chlorhexidine and alexidine), amphoteric surfactants, bisphenols (triclosan), alcohol, isopropyl alcohol (IPA), aldehydes (e.g., glutaraldehyde), iodine-releasing agents (iodophors), isothiazolones and peracetic acid [10]. As antimicrobial therapeutics become progressively less reliable, there is increasing pressure on effective disinfection protocols to prevent disease transmission in all areas where infectious diseases are a risk. A failure in these protocols will significantly impact on morbidity and mortality globally. The impact of biocidal use on AMR in species is under question however, as evidence suggests biocidal resistance, AMR and MDR mechanisms are interlinked.

In the European Union, disinfectants are classified as biocidal products regulated by the Biocidal Products Regulation (BPR) (EU) No 528/2012, ensuring efficacy and safety prior to marketing. Disinfectants can be classified into four overlapping categories including sanitizer, general disinfectant, sporicide and sterilant. Disinfectants, sanitizing agents and cleaning chemical agents have been used to inhibit and prevent microbial growth in pharmaceutical and medical device industries, healthcare, food, drinking water and domestic settings for decades. Effective cleaning and disinfection strategies are enforced to prevent disease transmission and control infectious disease by sanitising surfaces, fomites and personnel. In terms of disinfection, there are differences between disinfectants, sanitizers, antiseptics and sterilizing agents based on the desired objectives, the composition and concentration of the biocide, the contact time, residual levels and the area being disinfected [11]. In healthcare settings, the requirement for disinfection is determined by the nature of the item in terms of patient care. Medical devices are categorised as critical, semicritical and noncritical in terms of the risk of transmission of infectious diseases to patients. Critical items, including implants, must be purchased sterile or steam-sterilised, whereas high-level chemical disinfectants glutaraldehyde, hydrogen peroxide, ortho-phthalaldehyde (OPA), peracetic acid with hydrogen peroxide, and chlorine are suitable for semicritical items such as endoscopies [12]. Noncritical items that only come in contact with skin require disinfection with low-level disinfectants such as QACs. In food production, disinfectants used in animal settings are strong, and often toxic biocidal chemicals are applied to contaminated surfaces, whereas biocides used in food processing and domestic environments are usually less toxic and more diluted. To achieve microbial death using biocidal solutions, cleaning must precede treatment to eliminate organic and inorganic material. Additionally, specific guidelines for chemical concentration, contact time, temperature and pH must be adhered to. Disinfectant efficacy is impaired by interfering substances, typically organic matter, temperature, pH, contact time and the concentration. For instance, the pH affects the reaction kinetics of the disinfectant and thus influences the antimicrobial activity by altering the disinfectant molecule or cell surface. Hence, while an increase in pH will improve the antimicrobial activity of certain disinfectants, including, QACs and glutaraldehyde, it will decrease the activity of others, such as iodine, hypochlorite and phenols. In addition, many disinfectants work optimally at higher temperatures (typically 20 °C), where a lower temperature can lead to loss of disinfectant efficacy, particularly for QAC and aldehyde-containing disinfectants [13]. On the other hand, oxidising agents such as chlorine- or iodine-based disinfectants are not as affected by low temperatures [14]; however, they are more prone to inactivation by organic matter. Importantly, alcohol-based disinfectants are not significantly hindered by the presence of organic matter contamination [15], unlike many other disinfectant types. Unlike antimicrobial therapeutics that specifically target microbial cell components, such as cell walls, specific enzymes and genetic material, biocides interact nonspecifically with microbes, having multiple targets [10] and varying efficacies dependant on the target microorganism. For example, QACs disrupt the lipid bilayer structure of cell membranes, leading to membrane destabilisation, loss of function/structure and cytoplasmic leakage. Consequently, vegetative bacterial and fungal cells, and enveloped viruses are most affected, where QACs are ineffective against nonenveloped viruses and spores. Moreover, Gram-negative bacteria are less affected by these agents, due to the presence of their outer membrane and glycolipid endotoxin component, when compared to that of Gram-positive species. In addition, higher concentrations of QACs are generally required to be effective against yeasts and mould species. On the other hand, oxidising agents such as iodine and chlorine exert a broader spectrum of activity, being active against bacteria (including recalcitrant Gram-negative pathogens), fungi and viruses. Indeed, biocides often differ in their relative efficacies against the myriad of microorganisms, mainly due the biocidal formulation, the efficacy of the active component, the use and contact time, and the adsorption and uptake by cells (where chemical composition and architectural structure vary among different microbes). Intracellularly, biocides cause cell damage by disrupting metabolic processes, coagulating cellular components, and disrupting proteins and/or genetic material [16]. The antimicrobial activity of biocides is either through growth inhibition (bacteriostatic and fungistatic) or as a killing agent (sporicidal, bactericidal, fungicidal and virucidal). As mentioned, susceptibility to biocidal activity varies amongst microorganisms and typically follows the order from least to most susceptible: prions, coccidia, endospores, mycobacteria, Gram-negative bacteria, fungal species and Gram-positive bacteria [17]. Biocidal activity against viruses depends on their structure, specifically on the presence of an envelope, where enveloped viruses are more sensitive than nonenveloped viruses [18]. To ensure efficacy, testing of disinfectants to determine antimicrobial activity via suspension tests such as the European standards EN 1276, 1650 and 1656 (amongst others) are conducted. These tests generally require a 5-log reduction of viable cell numbers within a set number of minutes [19]. Nonetheless, suspension tests do not mimic the growth conditions of microbial species present in environmental samples, do not assess microbial growth phases such as log or stationary phases and do not account for resistant species. The EN 13,697 is a surface test to determine efficacy on varying surface materials but does not account for biofilm formation. The use of biocidal solutions at subtoxic concentrations, times or other treatment parameters leads to the survival of subpopulations of microbial species. This selective pressure promotes biocidal resistance, which is becoming increasingly recognised as a risk to public health safety, particularly when observed in species displaying multidrug resistance to antimicrobial therapeutics. Of greatest concern is the promotion of therapeutic resistance following exposure to biocidal solutions, termed cross-resistance [20].

The emergence of disinfectant-resistant microbes raises many issues, from disease transmission in healthcare settings and food production, to the manufacture of sterile pharmaceutical drugs and medical devices. The definition of biocidal resistance remains somewhat uncertain, some suggest resistance is a decrease in susceptibility as determined by an increase in the minimum inhibitory concentration (MIC) while others suggest bacteria surviving biocidal exposure at any usable concentration are deemed resistant [20].

The purpose of disinfection in clinical, veterinary, domestic and medical sectors (medical and pharmaceutical) is to reduce the viable microbial load on surfaces and fomites that are directly responsible for pathogen transmission. Biocidal efficacy, however, is impacted by the presence of interfering substances, typically, organic matter, temperature fluctuations, pH, contact time and the concentration applied. The spread of infectious diseases where AMR pathogens often result in patient mortality represents a serious public health risk. The presence of biocidal resistance in AMR species represents an increased risk where disease transmission may not be preventable. The presence and mechanisms of biocidal resistance have not been elucidated for many disinfectants and clinically relevant species. There is also a lack of detailed information on which biocidal agents are more prone to inducing AMR in species than others. Currently, there are numerous zoonotic pathogens transmissible to humans via direct animal contact or food contamination, including AMR species of Cryptococcus, Candida, Aspergillus, Campylobacter, Listeria, Salmonella, E. coli O157, Vibrio, Clostridium and Streptococcus [79], which, like the nosocomial ESKAPE pathogens, display antibiotic and biocidal resistance [80]. For example, studies have described antibiotic-resistant clinical E. coli strains that require higher concentrations of BAC for disinfection, and foodborne Pseudomonas strains demonstrating resistance to BAC and ampicillin, amoxicillin, erythromycin and trimethoprim [81]. These Gram-negative aerobic bacilli are the main pathogens associated with nosocomial (hospital-acquired) infections, including pneumonia, bacteraemia and UTIs, and are particularly associated with infectious disease in intensive care units [82]. Morbidity rates of 61% for Pseudomonas [83] and 11.5% for E. coli [84] apply. Moreover, sublethal exposure of the zoonotic Salmonella typhimurium to QACs promoted resistance to chloramphenicol, tetracycline, ampicillin and acriflavine [85]. Salmonella species showing resistance to sodium hypochlorite have displayed resistance to ceftazidime (S. enteritidis) and amikacin, tobramycin, cefazolin and cefotaxime in S. typhimurium [81]. The CDC estimates that Salmonella results in 1 million cases of infectious diseases yearly in the US and is the second most common foodborne pathogen in Europe (after Campylobacter). The incidence of nosocomial fungal infections associated with treatment failure is increasing, globally. Invasive fungal pathogens, including Cryptococcus, Candida and Aspergillus, result in 90% of life-threatening fungal disease in immunocompromised persons [51]. Candida auris, an emerging nosocomial MDR fungus, was responsible for 50 and 33 cases of disease in the UK and Spain, respectively, in 2016 [86], where C. auris has a 30-day mortality rate of 35%. There is a lack of information specifically detailing the susceptibility of clinically relevant fungi to common disinfectants or detailing mechanisms of resistance present. Zoonotic fungal infections, including dermatophytosis, sporotrichosis and histoplasmosis, are an important public health issue globally, however there is a lack of information on adequate preventative measures to control transmission [87]. Similar to bacterial species, the presence of fungal biofilms allows microbial species to persist in the environment and resist disinfection solutions. Currently, there is a lack of information on the susceptibility of fungal biofilms and multispecies biofilms to disinfection regimes. Many viruses, including hepatitis B and C, rotavirus, enteroviruses and cytomegalovirus, are associated with nosocomial transmission. Respiratory viruses, including respiratory syncytial virus, adenovirus, rhinoviruses, SARS-CoV-2 and influenza, are the main nosocomial viruses where direct contact between patients, healthcare staff, fomites and air and water droplets promotes transmission where they can cause or contribute to patient mortality [88]. Studies indicate that children are more susceptible to nosocomial viruses, with 49% of viral infections occurring in premature infants, while 24% of nonventilated pneumonia was viral in nature [89]. Of influenza cases in hospitals, 5.65% are related to nosocomial transmission and result in chronic illness and mortality. Preventative measures, including suitable disinfection regimes and parameters ensuring viral inactivation or evidence of resistance, are also essential.

To prevent nosocomial transmission, effective infection control systems that are heavily reliant on disinfection control measures must be in place. To be effective in a clinical setting, disinfectants must demonstrate efficacy against a broad range of microbial pathogens from bacterial, fungal and viral species. A “one fits all” disinfection solution is not realistic however, as variations in environmental factors and microbial species will impact efficacy. Antiseptics used clinically for skin disinfection often contain alcohol or IPA, with newer solutions containing additional agents such as chlorhexidine, povidone iodine or benzalkonium chloride. The added benefit of these additional biocides is uncertain however, and no added efficacy has been demonstrated for BAC or povidone [81], and BAC runs the risk of inducing AMR in species. While the emergence of antimicrobial resistance in microbes may become evident due to a lack of response to drug therapy, the emergence of biocide resistance can go unrecognised indefinitely. In 2015, the WHO announced its Global Action Plan aiming to combat AMR, which included limiting the application of numerous critically important antibiotics in veterinary applications. Perhaps a focus on the correct use and optimal application of key biocidal solutions must also be considered, particularly in clinical and veterinary settings where disease transmission is high. The safety implications of the misuse and overuse of disinfectants must also be considered, as certain disinfectants (sodium hypochlorite, sodium chloride, chlorine and QACs) are irritants and corrosive to the respiratory and intestinal mucous membranes of humans and animals [90], where chlorine is carcinogenic. Currently, there are no comparable guidelines in place for monitoring the use of disinfectants on a large scale [91] in terms of environmental safety.