Effects of Biocides on antibiotic resistance

6. What are the potential threats of biocide use in terms of bacterial resistance?

• 6.1 How might the use of biocides constitute a direct or indirect threat?
• 6.2 What are potential threats of using biocides in veterinary settings?
• 6.3 What are potential threats of using disinfectants in health care settings?
• 6.4 What are potential environmental threats of using biocides?

6.1 How might the use of biocides constitute a direct or indirect threat?

The SCENIHR opinion states:

3.8. Specific hazards

3.8.1. Direct and indirect hazards

The issue of antibiotic resistance induced by biocidal products is addressed as either a direct hazard or as an indirect hazard through transfer of resistance mechanism(s).

The direct hazard is the selection and dissemination of a resistant bacterium expressing resistance mechanisms active against biocides, antibiotics, or both (e.g. selection of adapted bacteria under selective pressure and change of microflora in some ecological niches, dissemination of this emerging strain and transmission to humans).

The indirect hazard concerns the transfer of mobile genetic elements (plasmid, transposon etc.) carrying genes conferring resistance to biocide, antibiotic or both, to a naturally susceptible strain via genetic exchange (e.g. during contact with commensal flora).

In some cases, both hazards may act together: a resistant bacterium may transfer an additional genetic element to another resistant bacterium enhancing the resistance level. The transfer of genetic element involved in resistance can occur anywhere: in the environment (e.g. water, ground), in the animal, in the food or in the human body (with resident/commensal flora)

Source & ©: SCENHIR,  Assessment of the Antibiotic Resistance Effects of Biocides (2009),
Section 3.8.1.Direct and indirect hazards, p. 48

6.2 What are potential threats of using biocides in veterinary settings?

The SCENIHR opinion states:

3.8.2. Veterinary use and hazard

The use of biocides in veterinary settings could induce resistance against the disinfectants used. This might explain why important zoonotic pathogens like Salmonella spp. disseminate between batches of animals. This may be particularly important where biocides are used at “industrial scale”, for example when animal houses are cleaned and disinfected. Under such conditions areas in the house may not receive optimum levels of active agent. Under conditions like this, the chances of selecting bacteria with increased resistance to the active ingredient are greater.

The same concerns could also apply to foot dips outside animal houses. The levels of the active agent could be diluted by rainfall and it is also quite common for the dips to contain a range of biological and other materials, which could serve to inactivate the active component. As with incorrect dilutions being applied, the chances of selecting resistant bacteria are increased.

If such bacteria are zoonotic like Campylobacter and Salmonella spp. it is possible that antibiotic therapy of infected humans could be compromised (EFSA 2008b). A recent study investigating the effect of cleaning and disinfection procedures in poultry slaughterhouses on the development of or selection for biocide and antibiotic resistance in Campylobacter jejuni and C. coli showed that a very low number (1-2) of genotypes were recovered after cleaning and disinfection under specified conditions and that there was no increase in antibiotic resistance before and after exposure to the disinfection procedures (Peyrat et al. 2008).

Studies, mainly laboratory-based, have shown that some disinfectants can select for bacteria with low level multiple drug resistance (MDR). In pathogens like E. coli and Salmonella spp., MDR can be due to up-regulation of the AcrABTolC efflux pump, although down-regulation of porins may also be involved. This low level resistance could be a possible stepping stone to higher-level antibiotic resistance due to the acquisition of additional resistance mechanisms (Davin-Regli et al. 2008, Piddock 2006). In two recent studies, Salmonella exposed to a range of common farm disinfectants were found to develop a low, but statistically significant, increased risk of selection of mutants with reduced susceptibility to ampicillin, ciprofloxacin and tetracycline. Some of the mutants selected were of the MDR phenotype (Karatzas et al. 2007, Karatzas et al. 2008).

These few data indicate that there is a need for futher studies addressing the potential interaction between the intensive and in some cases long-term use of biocides in animal facilities and the emergence of antimicrobial resistance.

The latter is also important in the light of trends towards an increasing use of antibiotics in modern (intensified) animal husbandy and the demonstrated transfer of resistance pathogens such as MRSA between animals and humans by direct contact and via the food chain (EFSA 2008b).

Source & ©: SCENHIR,  Assessment of the Antibiotic Resistance Effects of Biocides (2009),
Section 3.8.2. Veterinary use and hazard, p. 48 - 49

6.3 What are potential threats of using disinfectants in health care settings?

The SCENIHR opinion states:

3.8.3. Health care use and hazard

Studying environmental isolates from automated endoscope washer disinfector (AWD) provides a different perspective. Micro-organisms are being isolated with increasing frequency from washer disinfectors and processed endoscopes (Fraser et al. 1992, Gillespie et al. 2000, Griffiths et al. 1997, Kressel and Kidd 2001, Maloney et al. 1994, Nomura et al. 2004, Schelenz and French 2000, Takigawa et al. 1995). There are several reports about the emergence of 2% glutaraldehyde resistant Mycobacterium chelonae (Griffiths et al. 1997, Kingeren and Pullen 1993, Nomura et al. 2004).

Other bacteria, such as vegetative cells of Bacillus subtilis, Microcooccus luteus, Streptococcus sanguinis, Streptococcus mutans, Staphylococcus intermedius, were isolated from AWD following a high level disinfection process using chlorine dioxide. It was noted that most of these isolates remained sensitive to another oxidising agent when their susceptibility was investigated using a standard suspension efficacy test (Martin et al. 2008). The low concentration of the disinfectant (Griffiths et al. 1997, Maillard 2007, van Klingeren and Pullen 1993) or the presence of biofilms (Babb 1993, Pajkos et al. 2004, Smith and Hunter 2008), are considered important factors in determining the reduced susceptibility to biocides.

The presence of bacterial biofilms is one of the main challenges in terms of antimicrobial resistance with relevance for medical pratice, particularly for medical devices (Donlan and Costerton 2002, Dunne 2002). Pajkos et al. (2004) ascribed the failure of high-level disinfection in endoscope reprocessing to the presence of biofilms which can be very common and extensive on surfaces of endoscope tubings. Shackelford et al. (2006) observed that even the effective high-level disinfectant ortho-phthalaldehyde showed reduced activity against mycobacterial biofilms in vitro, but not against Pseudomonas aeruginosa biofilms. Even though most HAI are caused by bacteria associated with biofilms, most laboratories do not use biofilm tests to assess the efficacy of biocides and no European standards for the testing of disinfectants against biofilms in health care applications exist (Cookson 2005).

The linkage between biocides and antibiotic resistance in health care settings is a topic of great concern. However, clinically relevant resistance was only occasionally demonstrated, and when present, involved antibiotics of limited current use (e.g. chloramphenicol resistance in E. coli and tetracycline resistance in P. aeruginosa) (Weber and Rutala 2006). With regard to washer disinfectors, Nomura et al. (2004) studied the susceptibility of Mycobacterium chelonae isolated from bronchoscope washing disinfectors to 2% glutaraldehyde and antibiotics, and found an association of glutaraldehyde with antibiotic resistance.

Several studies have been carried out to evaluate the susceptibility of antibiotic-resistant bacteria to disinfectants. Antibiotic-resistant bacterial isolates were found to be as susceptible to disinfectants as their antibiotic-susceptible counterparts (Anderson et al. 1997, Rutala et al. 1997, Sakagami et al. 2002). Based on these data, antibiotic resistance was not deemed to require changes in disinfection protocols (Byers et al. 1998, Rutala et al. 2000).

The evidence base relating to biocide resistance and its relation with antibiotic resistance needs to be improved. An international consensus on the correct tests for determing biocide resistance and well designed surveillance systems are required. Antibiotic use and resistance should be continuously monitored. Reference and research laboratories should evaluate biocide resistance in any important new or multiple antibiotic resistant organisms (Cookson 2005).

The need for proper use of disinfectant and antiseptics should be stressed and health care workers should be trained to comply with clear and agreed policies and practices, avoiding unnecessary and incorrect use of biocides (e.g. choice of the appropriate product on the basis of the risk assessment; application of the product with regard to proper duration, concentration, pH or temperature; removal of organic debris before disinfection). A more appropriate use of antibiotics for therapy and prophylaxis also needs to be implemented. Gilbert and McBain (2004) believed that the risk associated with overuse of biocides in the health care environment is overstated, but recommended that to improve hygiene, applications that have demonstrable benefits should be emphasised

Source & ©: SCENHIR,  Assessment of the Antibiotic Resistance Effects of Biocides (2009),
Section 3.8.3. Health care use and hazard, p. 49 - 50

6.4 What are potential environmental threats of using biocides?

The SCENIHR opinion states:

3.8.4. Environment and hazard

Prior to determination of multi-resistance in micro-organisms in the environment it is of essence to determine whether or not biologically meaningful, i.e. not simply measurable, concentrations of biocides occur in the immediate environment such as sewage treatment plants and their immediate outflows.

One of the best examined examples remains triclosan, for which 79% of the incoming triclosan in sewage treatment plants was shown to be removed via biodegradation and 15% via sorption to activated sludge, thus resulting in approximately 6% of the incoming triclosan being released into the receiving streams (Singer et al. 2002). Despite this rather high removal rate in sewage treatment plants, effluent concentrations of triclosan ranged between 42-213 ng/L, thus resulting in concentrations of 11-98 ng/L in receiving waters for the particular sewage treatment systems investigated. The latter concentrations represent the lower range of triclosan concentrations reported from previous investigations in wastewaters (0.07 – 14 000 µg/L), possibly reflecting major differences in the technical capabilities of sewage treatment systems as well as in analytical capability (Jungclaus et al. 1978, Lindström et al. 2002, Lopez-Avila and Hites 1980, McAvoy et al. 2002). Correspondingly, between 50-2300 ng/L triclosan are reported for surface waters (streams) (Kolpin et al. 2002, Lindström et al. 2002), in seawater (50-150 ng/L) (Okumura and Nishikawa 1996), and in sediments (1-35 µg/kg) (Steffen and Lach 2000).

A comparable environmental investigation determined the density, heterotrophic activity, and biodegradation capabilities of heterotrophic bacteria in situ in a lake ecosystem following exposure to long-chain (C12 to C18) quaternary ammonium compounds (QACs) (Ventullo and Larson 1986). Monoalkyl and dialkyl substituted QACs were tested over a range of concentrations (0.001 to 10 mg/liter) and demonstrated that none of the QACs tested had significant adverse effects on bacterial densities in either acute (3 h) or chronic (21 day) studies. Moreover, chronic exposure of lake microbial communities to a specific monoalkyl QAC resulted in an adaptive response and recovery of heterotrophic activity. This adaptive capability was investigated further by Nishihara et al. (2000), who demonstrated that Pseudomonas fluorescens TN4 isolated from sewage treatment plants degraded didecyl-dimethyl-ammonium chloride (DDAC) to produce decyl-dimethyl-amine and subsequently, dimethylamine, as the intermediates.

The TN4 strain also assimilated other quaternary ammonium compounds (QACs), alkyl-trimethyl- and alkyl-benzyl-dimethyl-ammonium salts, but not alkylpyridinium salts. TN4 was highly resistant to these QACs and degraded them using an n-dealkylation process (Nishihara et al. 2000). Despite this adaptive response and probably because of the enormous consumption of these compounds, high concentrations of QACs, especially C12 chain benzalkonium chloride (BAC-C12) as well as long C-chain dialkyl-dimethyl-ammonium chloride (DDAC-C18), can be found in sediments of surface waters with a maximum concentration of 3.6 mg/kg and 2.1 mg/kg, respectively (Martínez-Carballo et al. 2007).

The above data demonstrate that significant amounts of biocides readily reach both the immediate environment (kitchen sink) and the more distant environment (sewage treatment plants and surface waters). The question of whether these environmental concentrations will lead to resistance in micro-organisms was addressed for triclosan by McBain et al. (2004a) using a gradient plate technique. They exposed several bacterial strains, including inter alia Streptococcus oralis, Streptococcus sangula, Streptococcus mutans, Neisseria subflava and triclosan resistant Escherichia coli (ATCC 8739) to increasing, sublethal concentrations of triclosan. MIC values towards chlorhexidine, metronidazole and tretracyclin were determined before and after biocide exposure. The experiments failed to demonstrate a biologically significant induction of drug resistance in triclosan-exposed bacteria, beyond that demonstrated for E. coli, thus suggesting that triclosan-induced drug resistance is not generally readily inducible nor is it transferred across bacterial species.

A similar investigation by McBain et al. (2004b) investigated the effects of short-term (12 days) and long-term (3 months) QAC-containing detergent exposure on biofilms from house-hold sink drains. Denaturing gradient gel electrophoresis analysis identified the major microcosm genera as Pseudomonas, Pseudoalteromonas, Erwinia and Enterobacter, and demonstrated that aeromonads increased in abundance under 10-50% QAC-containing detergent exposure. Long-term QAC-containing detergent exposure did not significantly change the pattern of antimicrobial susceptibility, thus suggesting that even though antimicrobial susceptibility changes (multi-resistance) have been reported in isolated bacterial cultures, such changes do not necessarily occur within complex micro-organism communities

Source & ©: SCENHIR,  Assessment of the Antibiotic Resistance Effects of Biocides (2009),
Section 3.8.4 Environment and hazard, p. 50 - 51