Assessing the risk of resistance to cationic biocides incorporating realism-based and biophysical approaches

The control of microorganisms is a key objective in disease prevention and in medical, industrial, domestic, and food-production environments. Whilst the effectiveness of biocides in these contexts is well-evidenced, debate continues about the resistance risks associated with their use. This has driven an increased regulatory burden, which in turn could result in a reduction of both the deployment of current biocides and the development of new compounds and formulas. Efforts to balance risk and benefit are therefore of critical importance and should be underpinned by realistic methods and a multi-disciplinary approach, and through objective and critical analyses of the literature. The current literature on this topic can be difficult to navigate. Much of the evidence for potential issues of resistance generation by biocides is based on either correlation analysis of isolated bacteria, where reports of treatment failure are generally uncommon, or laboratory studies that do not necessarily represent real biocide applications. This is complicated by inconsistencies in the definition of the term resistance. Similar uncertainties also apply to cross-resistance between biocides and antibiotics. Risk assessment studies that can better inform practice are required. The resulting knowledge can be utilised by multiple stakeholders including those tasked with new product development, regulatory authorities, clinical practitioners, and the public. This review considers current evidence for resistance and cross-resistance and outlines efforts to increase realism in risk assessment. This is done in the background of the discussion of the mode of application of biocides and the demonstrable benefits as well as the potential risks.     

Volatiles from Marina neglecta: Biocide effect on insect vectors of tropical diseases in Southern Mexico

Preventive measures based in the control of insect vectors are considered as the best choice to decrease the incidence of insect-borne diseases. Herein we report on the volatile content of the leaf essential oils from Marina neglecta, a medicinal plant distributed in the tropical regions of southern Mexico. In order to investigate the chemical variation of the essential oils, a volatile screening was performed during the four seasons of the years 2016–2019. Simultaneously, their biological activity was tested on distinct life stages of Meccus pallidipennis, M. bassolsae, Aedes aegypti and A. albopictus. Essential oils were mainly constituted of β-pinene (>30%) β-caryophyllene (>25%) and germacrene D (>13%). Dorsal-abdomen application of essential oils on triatomines, revealed an efficient LC₅₀ for nymphs of the stages I to III (4 µg/insect), nymphs of the stages IV to V (5–6 µg/insect), and adults (7–8 µg/insect). The LT₅₀ for the stages I to III was between 6 and 8 h, whereas that for the stages IV to V and adults oscillated between 12 and 16 h and 22 to 26 h, respectively. Fumigation experiments performed on nymph V, demonstrated that 300 µg L^?1 air produced 100% mortality after 72 h post-treatment. Among tested volatiles, β-pinene and β-caryophyllene produced a comparable mortality rate (p < 0.01) than that of essential oils in the stages assayed. Essential oils showed strong larvicidal (LC₅₀, 24–36 µg mL^?1) and adulticidal (35–48 µg mL^?1) activities in mosquito species with an LT₅₀ of 4.5 h and 25–35 min, respectively. The evaluation of β-pinene produced a significant mortality rate (p < 0.01) in larvae whereas germacrene D was the most effective volatile (p < 0.01) against adults of both mosquito species. According to our results, β-pinene was the most effective volatile against the four insect species evaluated and its effect was comparable to that of the essential oil.     

Biodeterioration of crude oil and oil derived products: a review

Biodeterioration of crude oil and oil fuels is a serious economic and an environmental problem all over the world. It is impossible to prevent penetration of microorganisms in oil and fuels both stored in tanks or in oilfields after drilling. Both aerobic and anaerobic microorganisms tend to colonise oil pipelines and oil and fuel storage installations. Complex microbial communities consisting of both hydrocarbon oxidizing microorganisms and bacteria using the metabolites of the former form an ecological niche where they thrive. The accumulation of water at the bottom of storage tanks and in oil pipelines is a primary prerequisite for development of microorganisms in fuels and oil and their subsequent biological fouling. Ability of microorganisms to grow both in a water phase and on inter-phase of water/hydrocarbon as well as the generation of products of their metabolism worsen the physical and chemical properties of oils and fuels. This activity also increases the amount of suspended solids, leads to the formation of slimes and creates a variety of operational problems. Nowadays various test-systems are utilized for microbial monitoring in crude oils and fuels; thus allowing an express determination of both the species and the quantities of microorganisms present. To suppress microbial growth in oils and fuels, both physico-mechanical and chemical methods are applied. Among chemical methods, the preference is given to substances such as biocides, additives, the anti-freezing agents etc that do not deteriorate the quality of oil and fuels and are environmentally friendly. This review is devoted to the analysis of the present knowledge in the field of microbial fouling of crude oils and oil products. The methods utilized for monitoring of microbial contamination and prevention of their undesirable activities are also evaluated. The special focus is given to Russian scientific literature devoted to crude oil and oil products biodeterioration.     

The cytoplasmic membrane as the site of the antimicrobial action of N-octylethanolamine

On agar spread plates, N-octylethanolamine was biocidal at comparable minimum inhibitory concentration (MIC) values (3–4mm) against Pseudomonas aeruginosa (two strains), Escherichia coli, Staphylococcus aureus, Bacillus subtilis, Candida tropicalis, and Acremonium sp. which had been grown on a number of different media. The inhibition was greater at higher pH values. In liquid culture, growth inhibition by 3mm N-octylethanolamine was accompanied by cell lysis. Both effects could be prevented by the presence of 1mm spermine or spermidine, but only in bacteria, and not at high pH values. These effects of the polyamines were shown to be non-specific, being shared by other polycations and Mg²⁺ ions. N-Octylethanolamine at concentrations above its MIC caused total inhibition of the oxidation of 1mm glucose by Ps. aeruginosa (CAS1 and PAO1), E. coli, or C. tropic an effect that was partially reversible by Mg²⁺ ions. At concentrations below the MIC, there was little inhibit ion of glucose oxidation but a potent inhibition of the extrusion of ethidium bromide from intact cells of E. coli, suggesting that at such concentrations N-octylethanolamine is uncoupling oxidative phosphorylation. The data presented confirm the view that the biocidal effects are due to action on the cytoplasmic membrane.     

Comparative antibacterial potential of selected aldehyde-based biocides and surfactants against planktonic Pseudomonas fluorescens

The antimicrobial efficacy of two aldehyde-based biocides (glutaraldehyde, GTA, and ortho-phthalaldehyde, OPA) and two surfactants (cetyltrimethyl ammonium bromide, CTAB, and sodium dodecyl sulphate, SDS) was tested against planktonic Pseudomonas fluorescens. The antimicrobial effects were evaluated by respiratory activity as a measure of the oxygen uptake rate, adenosine triphosphate (ATP) release, outer membrane proteins (OMP) expression and cellular colour changes. The results were compared with the bacterial characteristics without chemical treatment. Tests in the presence of bovine serum albumin (BSA), in order to mimic a disinfection process in the real situation under dirty conditions, were performed according to the European Standard EN-1276. P. fluorescens was completely inactivated with OPA (minimum bactericidal concentration, MBC = 0.5 mM) and CTAB (MBC = 5 mM) and was resistant to GTA and SDS. Only CTAB promoted cellular disruption and consequent ATP release. The antimicrobial action of the chemicals tested was significantly reduced when BSA was introduced into the bacterial cultures, increasing markedly the MBC values. Additionally, the presence of BSA acted as a disruption protective agent when CTAB was applied and stimulated the bacterial respiratory activity when lower concentrations of SDS were tested. The OMP of the bacterial cells was affected by the application of both surfactants. OMP expression remained unaltered after biocide treatment. Bacterial colour change was noticed after treatment with biocides and surfactants. In summary, P. fluorescens was extremely resistant to GTA and SDS, with antimicrobial action being quenched markedly by the reaction with BSA.     

Biodegradability of end-groups of the biocide polyhexamethylene biguanide (PHMB) assessed using model compounds

Polyhexamethylene biguanide (PHMB), a biocide used in a wide variety of disinfection and preservation applications, is a polydisperse mixture in which the end-groups may be any combination of amine, guanidine and cyanoguanidine. Using PHMB model compounds (1,6-diaminohexane; 1,6-diguanidinohexane; 1,6-di(cyanoguanidino)hexane; 4-guanidinobutyric acid), we have determined the biodegradation characteristics of each end-group in several strains of bacteria isolated for their ability to utilise PHMB as a sole source of nitrogen. Bacteria were screened for growth at the expense of each model compound (at non-inhibitory concentrations) as sole nitrogen source. None of the isolated bacteria was capable of utilising a cyanoguanidine end-group as growth substrate, whereas several bacteria were shown to utilise amine or guanidine end-groups. In particular, a strain of Pseudomonas putida was capable of extensive growth with 1,6-diguanidinohexane as a sole nitrogen source, with complete removal of guanidine groups from culture medium within 2 days, and with concomitant formation of unsubstituted urea, which in turn was also utilised by the organism. We conclude that whilst amine and guanidine end-groups in PHMB are likely to be susceptible to biodegradation, cyanoguanidine end-groups are likely to be recalcitrant.     

Use of dissolved ozone for controlling planktonic and sessile bacteria in industrial cooling systems

Cooling water treatment requires effective, environmentally-safe biocides compatible with system operation. The unique combination of high biocidal activity during use with no toxic discharge, could render dissolved ozone a safe biocide for cooling water treatment. Planktonic and sessile cells of Pseudomonas fluorescens (a frequent microbial contaminant of industrial systems) were used in this work to assess the biocidal effectiveness of ozone. Dissolved ozone showed to be effective at concentrations between 0.1 and 0.3 ppm, to eliminate completely the levels of planktonic cells used in this paper (10⁷–10⁸ cell/ml) within a range of contact times between 10 and 30 min. However, ozone at 0.15 ppm was only able to diminish sessile cell population by two or three orders of magnitude. This minor biocidal effectiveness of ozone against bacterial biofilms is discussed in this paper, taking into account recent concepts on structure and dynamics of biofilms. Different metallic substrata were assayed to verify if there was any effect of metal nature on the biocidal action. Open circuit potentials vs. time experiments and potentiodynamic polarization curves were made for assessing the effect of dissolved ozone on the corrosion behavior of the metals tested.     

Biofilm formation on copper and its control by inhibitor/biocide in cooling water environment

The present study has successfully identified the nitrate reducing bacteria present in the cooling water system and also investigated the performance of industrially applied biocide and inhibitor on the bacterial inhibition. In order to carry out the objective of this study, facilities and methods such as 16S rRNA gene sequencing, Lowry assay, SEM, EIS, ICP-MS and weight loss analysis were being utilized. In this study, two out of the five morphologically dis- similar colonies identified through 16S rRNA gene sequencing, namely the Massilia timonae and the Pseudomonas, were being utilized in the biocorrosion study on copper metal. From the surface analysis using SEM demonstrated the phenomenon of biofilm formation on the copper surface. 2-methylbenzimidazole has the addition of methyl group in the diazole ring position of benzimidazole it has create basicity environment and inhibit the metal deterioration. Meanwhile, it is also deducible from the EIS and protein analysis that com- bination of biocide with either of the inhibitors gives rise to better biocorrosion suppression (0.00178 mpy and 0.00171mpy) as compared to the sole effect of either biocide or inhibitor (0.00219 mpy, 0.00162 and 0.00143). Biocorrosion system biocide with MBM was found to exhibit 65% corrosion inhibition efficiency. Moreover, adoption of 2-Methylbenzimidazole seems to display better performance as compared to Multionic 8151, which is adopted in cooling water system.     

Microbial degradation of formaldehyde in white mineral dispersions preserved with formaldehyde-releasing biocides

Two different formaldehyde-degrading microorganisms, Pseudomonas putida and Methylobacterium extorquens, were isolated from calcium carbonate slurry containing the formaldehyde-releasing biocide (ethylenedioxy) dimethanol. Their relative formaldehyde biodegradation and formic acid production kinetics were studied in broth and in calcium carbonate slurry for each microorganism individually, as well as in mixed cultures. Furthermore, the minimal inhibition concentration (MIC) was determined. The results indicated that in slurry, M. extorquens is more tolerant of formaldehyde than P. putida. In slurry, microbial-induced oxidation of formaldehyde caused a temporary accumulation of formic acid, which is presumed to be responsible for pH drop and destabilisation of the calcium carbonate slurry suspension systems. In addition, the residual formaldehyde concentration was observed to drive dominance and recovery of individual formaldehyde-resistant microorganisms in the slurry. Overall, this investigation indicated that biodegradation of formaldehyde in calcium carbonate slurry is brought about by alternating dominance of bacterial genera of mixed formaldehyde-resistant microbial populations.     

Biodeterioration of stone: a review

The alteration and weathering of stone is basically determined by natural and anthropogenic impacts influencing various physical, chemical and biological damage factors at the object site. Whether as direct or catalytically enhancing factor, the biodeterioration of stone is coupled with nearly all environmentally induced degradation processes: the presence of the one makes deterioration by the other all the more effective. The bioreceptivity of stone is described by its structure and chemical composition, while the intensity of the microbial contamination is determined by the referring climatic conditions and the anthropogenic euthrophication of the atmosphere. The microflora improves the nutrient and moisture-restricted growth conditions on building stones by the formation of surface-covering biofilms. Besides the aesthetical impairment caused by the coloured biopatina, the biofouling effect promotes even “abiotic” deterioration processes due to the alteration of the material structure as well as their thermo-hygric properties; in addition, mechanical pressure due to the shrinking and swelling of the colloidal biofilms might cause a further weakening of the mineral lattice. Acidolytic and oxido-reductive biocorrosion processes complete the biodeteriorating attack of stone acting as a preliminary precursor for the latter formation of detrimental crusts. Suitable and reliable methods for the detection of biodeterioration processes are available, but only the interdisciplinary diagnosis and evaluation of the entire decay process of stone allows the formulation of adaequate countermeasure strategies. In case the significance of biodeterioration impacts is proven, the possible effects of the microbial contamination on cleaning procedures, protective treatments as well as biocidal applications has to be considered. This paper will give a comprehensive overview to the biodeterioration of stone and stresses the practical relevance for the conservation.