Efficacy of a quaternary ammonium compound against planktonic and sessile populations of differentLegionella pneumophila strains

Efficacy of Gemacide PN-50^TM (a quaternary ammonium compound) as a commercial formulation recommended for disinfecting heat exchangers was determined for both planktonic and sessile populations of variousLegionella pneumophila strains. The quaternary ammonium compound (QAC) was preferred as an alternative due to the emerging resistance of potentially pathogenic bacteria against different biocides. PlanktonicL. pneumophila strains were suspended in tap water while sessile ones were grown on stainless steel that is used in construction of the cooling towers, then both group of strains were exposed to the biocide. The sensitivity of both planktonic and sessile populations ofL. pneumophila strains to the biocide was different. The biocide was found effective below recommended dosages (1000–2000 mg/L) against planktonic populations ofL. pneumophila, whereas it was determined that higher than the recommended dosages were required for sessile populations. The environmental isolates were more resistant to the biocide than the ATCC isolate was. The results indicated that studying only the planktonic populations ofL. pneumophila for biocide tests might not be sufficient to provide the optimum dosage and contact time information for field trials. Therefore, biocidal activity of a water treatment chemical must be evaluated in terms of dosage and contact times on both planktonic and sessile bacteria.     

The use of biocides to control sulphate‐reducing bacteria in biofilms on mild steel surfaces

Three different types of biocides, viz. formaldehyde (FM), glutaraldehyde (GA) and isothiozolone (ITZ) were used to control planktonic and sessile populations of two marine isolates of sulphate‐reducing bacteria (SRB). The influence of these biocides on the initial attachment of cells to mild steel surfaces, on subsequent biofilm formation and on the activity of hydrogenase enzymes within developed biofilms was evaluated. In the presence of biocides the rate and degree of colonization of mild steel by SRB depended on incubation time, bacterial isolate and the type of biocide used. Although SRB differed in their susceptibility to biocides, for all isolates the biofilm population was more resistant to the treatment than the planktonic population. GA showed highest efficiency in controlling planktonic and sessile SRB compared with the other two biocides. The activity of the enzyme hydrogenase measured in SRB biofilms varied between isolates and with the biocide treatment. No correlation was found between the number of sessile cells and hydrogenase activity.     

A laboratory apparatus for the generation and biocide efficacy testing of Legionella biofilms

P.N. GREEN AND R.S. PIRRIE. 1993. The construction and operation of a laboratory biofilm generator designed to grow sessile populations of legionellas for biocide efficacy testing is described. Some examples of static biocide testing on both sessile and planktonic populations of Legionella bozemanii are discussed along with other potential applications of the apparatus.     

Elsinochrome A production by the bindweed biocontrol fungus Stagonospora convolvuli LA39 does not pose a risk to the environment or the consumer of treated crops

Biological control as an alternative to chemical pesticides is of increasing public interest. However, to ensure safe use of biocontrol methods, strategies to assess the possible risks need to be developed. The production of toxic metabolites is an aspect which has so far largely been neglected in the risk assessment and the registration process for biocontrol products. We have evaluated the risks of elsinochrome A (ELA) and leptosphaerodione production by the fungus Stagonospora convolvuli LA39, an effective biocontrol agent used against bindweeds. The toxicity of the two metabolites to bacteria, protozoa, fungi and plants was evaluated in in vitro assays. The most sensitive bacteria and fungi were already affected at 0.01-0.07 microM ELA, whereas plants were far less sensitive. Leptosphaerodione was less toxic than ELA. Subsequently, it was investigated whether ELA is present in the applied biocontrol product or LA39-treated bindweed and crop plants. In plants ELA was never detected and in the biocontrol product the ELA concentration was far too low to have toxic effects even on the most sensitive organisms. We conclude that the production of ELA by biocontrol strain LA39 does not pose a risk to the environment or to the consumer.     

Decreased biocide susceptibility of adherent Legionella pneumophila.

In a study of the in vitro effectiveness of biocides against Legionella pneumophila, some aspects of the cooling tower environment were replicated in the laboratory, paying particular attention to water hardness and pH. Pieces of Douglas fir and polyvinyl chloride were colonized in a recirculating system and the comparative efficacy of two biocides (Bronopol and Kathon) against the sessile and planktonic populations was examined. While the biocides were relatively effective against the planktonic L. pneumophila population over a short period of time (minimum 9-12 h), substantially longer periods of time (maximum greater than 48 h) were required to reduce the number of cultivable bacteria to below detectable levels in the adherent population. The results indicate that failure to monitor the sessile population of L. pneumophila in laboratory studies of biocides may result in the use of incorrect dosages and/or contact times in field trials with apparently reduced in situ efficacy.     

Reviewing microbial electrical systems and bacteriophage biocontrol as targeted novel treatments for reducing hydrogen sulfide emissions in urban sewer systems

Microbially induced concrete corrosion (MICC) is a costly, and ongoing problem affecting the infrastructure of water utilities worldwide. Traditionally MICC has been treated with chemicals and physical techniques that inhibit the release of hydrogen sulfide (H₂S), preventing sulfuric acid formation and the consequent corrosion. However, these methods require continual dosing and monitoring to ensure process efficiency and prevent undue costs. This review focuses on recent research into two potential novel treatments for MICC: re-engineering the sewer sulfur cycle by removing sulfide using electrodes in microbial electrical systems as an alternative electron acceptor and; altering the microbial community using targeted bacteriophage biocontrol to reduce specific sulfide-producing bacteria. These novel treatments hold the potential to reduce water utilities reliance on continual chemical dosing providing a long-lasting treatment I believe necessitates further laboratory and field-trial investigation.     

Decreased biocide susceptibility of adherent Legionella pneumophila

J.B. WRIGHT. I. RUSESKA AND J.W. COSTERTON. 1991. In a study of the in vitro effectiveness of biocides against Legionella pneumophila , some aspects of the cooling tower environment were replicated in the laboratory, paying particular attention to water hardness and pH. Pieces of Douglas fir and polyvinyl chloride were colonized in a recirculating system and the comparative efficacy of two biocides (Bronopol and Kathon) against the sessile and planktonic populations was examined. While the biocides were relatively effective against the planktonic L. pneumophila population over a short period of time (minimum 9-12 h), substantially longer periods of time (maximum > 48 h) were required to reducc the number of cultivable bacteria to below detectable levels in the adherent population. The results indicate that failure to monitor the sessile population of L. pneumophila in laboratory studies of biocides may result in the use of incorrect dosages and/or contact times in field trials with apparently reduced in situ efficacy.     

Studies on the behaviour of Pseudomonas fluorescens biofilms after Ortho-phthalaldehyde treatment

A relatively novel biocide, ortho-phthalaldehyde (OPA), was tested to control biofilms formed by Pseudomonas fluorescens on stainless steel surfaces. The toxic action of OPA was assessed in terms of inactivation and removal of the biofilm by means of, respectively, the determination of the respiratory activity and the variation in the dry weight of the biofilms. For comparison, the activity of OPA against suspended bacteria was also evaluated. The results showed that higher concentrations of OPA and longer exposure times are needed to inactivate P. fluorescens biofilms than planktonic populations, thus denoting that sessile bacteria have a reduced susceptibility to OPA. This appears to be associated with the reaction with the proteins of the matrix, as demonstrated by the reduction of the antimicrobial action of OPA in the presence of a protein (bovine serum albumin). The application of OPA appeared to cause little effect in the removal of biofilms from the metal slides since the mass of biofilm that remained on the surfaces, after biocide treatment, was within the same range as those observed in the control tests. These results suggest that, with OPA application, biofilms can be inactivated but stay attached to the surfaces, decreasing thereby the success of the chemical treatment.     

A green biocide enhancer for the treatment of sulfate-reducing bacteria (SRB) biofilms on carbon steel surfaces using glutaraldehyde

Generally speaking, a much higher concentration of biocide is needed to treat biofilms compared to the dosage used to for planktonic bacteria. With increasing restrictions of environmental regulations and safety concerns on large-scale biocide uses such as oil field applications, it is highly desirable to make more effective use of biocides. In this paper a green biocide enhancer ethylenediaminedisuccinate (EDDS) that is a biodegradable chelator, was found to enhance the efficacy of glutaraldehyde in its treatment of sulfate-reducing bacteria (SRB) biofilms. Experiments were carried out in 100 ml anaerobic vials with carbon steel coupons. The ATCC 14563 strain of Desulfovibrio desulfuricans was used. Biofilms on coupon surfaces were visualized using scanning electron microscopy (SEM). Experimental results showed that EDDS reduced the glutaraldehyde dosages considerably in the inhibition of SRB biofilm establishment and the treatment of established biofilms on carbon steel coupon surfaces.     

The corrosion behaviour of galvanized steel in cooling tower water containing a biocide and a corrosion inhibitor

The corrosion behaviour of galvanized steel in cooling tower water containing a biocide and a corrosion inhibitor was investigated over a 10-month period in a hotel. Planktonic and sessile numbers of sulphate reducing bacteria (SRB) and heterotrophic bacteria were monitored. The corrosion rate was determined by the weight loss method. The corrosion products were analyzed by energy dispersive X-ray spectroscopy and X-ray diffraction. A mineralized, heterogeneous biofilm was observed on the coupons. Although a biocide and a corrosion inhibitor were regularly added to the cooling water, the results showed that microorganisms, such as SRB in the mixed species biofilm, caused corrosion of galvanized steel. It was observed that Zn layers on the test coupons were completely depleted after 3?months. The Fe concentrations in the biofilm showed significant correlations with the weight loss and carbohydrate concentration (respectively, p?<?0.01 and p?<?0.01).     

Biocide treatment of biofilms

Biofilms are ubiquitous in nature and microorganisms often exist as members of complex consortia, rather than as pure cultures. Their localised metabolic activity can create diffusion gradients of nutrients, fermentation byproducts and possible associated corrosion products within the biofilms; together with cell lysis, these cause a mosaic of microenvironments which may be totally different to the bathing phase. Such habitats pose a major, and often ignored, constraint on the interpretation of results obtained from laboratory disinfection models which can be physically, environmentally and physiologically inappropriate. For example, the most commonly used model for inactivation of microorganisms by biocides utilises the so-called ‘Chick- Watson law’: this implies that biocide concentration and contact time, the (C × T) factor, are the two key variables determining biocide efficacy. However, applications of the ‘law’ have assumed complete and uniform mixing of microorganisms and biocide, ignoring that diffusion might be rate limiting and that biocide concentration might decrease with time. Recent results suggest that many of the viable bacteria in chlorinated potable water are attached to surfaces and under these circumstances coliforms have withstood at least 12 ppm free residual chlorine. The use and efficacy of alternative biocides such as monochloramine against aquatic biofilms is discussed.     

Expanding the microbial monitoring toolkit: Evaluation of traditional and molecular monitoring methods

Evaluation of microbial populations in oilfield systems is critical to understand the risk of microbiologically influenced corrosion, reservoir and surface souring (hydrogen sulfide production), and biofouling. Although traditional culture based methods have dominated oilfield microbial monitoring for years, use of molecular tools is becoming more common for both field and laboratory evaluations. The implementation of these additional tools is in response to some of the disadvantages of culture-based methods such as long incubation times and underestimation of actual microbial populations. The current work provides a direct comparison of culture-based methods (serial dilution for sulfate reducing, acid producing, and general heterotrophic bacteria) with molecular methods including adenosine triphosphate (ATP) and adenosine monophosphate (AMP) quantification and quantitative polymerase chain reaction (qPCR). The results demonstrate that these technologies provide nearly identical results in untreated samples with known culturable organisms, but show some differences when used in the context of a planktonic kill study. The pros and cons of each technology were addressed with respect to their use in field monitoring, laboratory monitoring, and microbial kill studies. The authors found that none of the technologies described in this work provide an all-inclusive answer, but together they provide significant insight into the microbial population in an oilfield system. In short, the authors demonstrate that it is advantageous for oilfield stakeholders to expand their microbial monitoring toolkit with these new technologies to ensure sustainable, cost-effective operation.     

Biofilm formation in aqueous metal working fluids

Planktonic and biofilm flora in circulating coolant systems were examined in engineering workshops and in a laboratory model. In a CNC lathe, thorough cleaning and disinfection reduced microbial counts, but within five weeks substantail numbers of organism were detected in both the coolant and, especially, the biofilm. Physical disruption of the biofilm in a milling machine, using a wire brush, increased the planktonic bacterial counts up to twenty times. Results from the model system indicate that biocides may increase the time before the development of high microbial numbers, both in the biofilm and the coolant. The use of a metal-working fluid containing a triazine biocide was shown to result in the development of substantial fungal contamination with most being associated with the biofilm.     

A Systematic Approach to Capacity Strengthening of Laboratory Systems for Control of Neglected Tropical Diseases in Ghana,Kenya, Malawi and Sri Lanka

Background

The lack of capacity in laboratory systems is a major barrier to achieving the aims of the London Declaration (2012) on neglected tropical diseases (NTDs). To counter this, capacity strengthening initiatives have been carried out in NTD laboratories worldwide. Many of these initiatives focus on individuals'' skills or institutional processes and structures ignoring the crucial interactions between the laboratory and the wider national and international context. Furthermore, rigorous methods to assess these initiatives once they have been implemented are scarce. To address these gaps we developed a set of assessment and monitoring tools that can be used to determine the capacities required and achieved by laboratory systems at the individual, organizational, and national/international levels to support the control of NTDs.

Methodology and principal findings

We developed a set of qualitative and quantitative assessment and monitoring tools based on published evidence on optimal laboratory capacity. We implemented the tools with laboratory managers in Ghana, Malawi, Kenya, and Sri Lanka. Using the tools enabled us to identify strengths and gaps in the laboratory systems from the following perspectives: laboratory quality benchmarked against ISO 15189 standards, the potential for the laboratories to provide support to national and regional NTD control programmes, and the laboratory''s position within relevant national and international networks and collaborations.

Conclusion

We have developed a set of mixed methods assessment and monitoring tools based on evidence derived from the components needed to strengthen the capacity of laboratory systems to control NTDs. Our tools help to systematically assess and monitor individual, organizational, and wider system level capacity of laboratory systems for NTD control and can be applied in different country contexts.     

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.     

Registration of Phlebiopsis gigantea as a Forest Biocontrol Agent in the UK: Recent Experience

The registration of one of the world's first biological control agents for a plant pathogen, in January 1998, after 30 years of field use, is discussed in relation to its approval under the UK Control of Pesticides Regulations 1986. The control agent, Phlebiopsis gigantea (formerly known as Peniophora gigantea), is a common wood-rotting basidiomycete which is applied to the freshly cut surfaces of pine stumps to prevent their colonization by the pathogenic rootrotting fungus Heterobasidion annosum . Unlike many other biocontrol agents, P. gigantea is not a biocide that kills the target organism. Rather, it competes for resources that the pathogen would otherwise use, providing an extension to a naturally occurring process. Experience in securing approval is reviewed, with special reference to the identity of the organism and its natural distribution, and to assessments of the risk its use might pose in the environment. It is suggested that before the formal process of applying for approval of a biocontrol agent begins, there should be procedures for agreeing precise data requirements. These should be based on the type of crop to be treated and the mode of operation and characteristics of the biocontrol organism. This would streamline the registration process.     

An automated system for biocide testing on biofilms*

The paper presents and discusses a novel on-line real-time non-destructive continuous-flow system for biocide testing on industrial biofilms. This laboratory system is capable of monitoring changes in growth, accumulation and respiratory activity of biofilms in response to biocidal treatment. The system incorporates a fouling monitor for continuous measuring of the rate of biofilm accumulation (heat transfer resistance), a sensor for monitoring of microbial activity (oxygen meter for monitoring the rate of biofilm respiratory activity), and subsystems necessary for microbial life support and control of operation parameters. Examples of system operation and testing of oxidizing and non-oxidizing biocides are presented. Received 25 May 1997/ Accepted in revised form 25 November 1997     

Selector systems in recording physiological and behavioural activity in sedentary aquatic animals

The design and use of selector systems in recording physiological rate functions and behavioural activity in sessile invertebrates, particularly bivalves, is described for use in the laboratory and the field. The experimental limitations of the technique are discussed and the use of switch systems with other types of aktograph is described. Switch systems permit large groups of animals to be used in activity studies, whilst reducing both time spent in data handling and cost per animal of the recording system.     

Effect of some biocides on non-tuberculous mycobacteria

The incidence of disease and nosocomial infections produced by non tuberculosis mycobacteria (NTM) has increased in immunocompetent patients, but also and more frequently, in immunosuppressed patients. Several studies have disclosed that mycobacteria are more resistant to biocides than non-sporulating bacteria; in addition, some species are particularly resistant. The biocide action of sodium hypochloride and glutaraldehyde on Mycobacterium aviumintracellulare, Mycobacterium gordonae, Mycobacterium fortuitum and Mycobacterium chelonae was studied, using a modified Kelsey Maurer test. For the different species, both the minimal inhibitory concentration (MIC) and the minimal action time were determined. Effectiveness of sodium hypochloride and glutaraldehyde against the different mycobacterial species varied. The same was true for different isolates of the same species. Sodium hypochloride effective MIC and exposure time (killing of 99.99% of all NTM) were 0.2% and 5 minutes, respectively. In order to achieve 100% killing, 0.5% MIC and 15 minute exposure were needed. In the case of glutaraldehyde, 99.99% of the bacteria were killed with 1% MIC and a 15 minute exposure. An effectiveness of 100% was achieved with a 2% MIC of glutaraldehyde and a 15 minute exposure. Sodium hypochloride and glutaraldehyde are effective biocides for mycobacteria. The first biocide is cheap and effective at low concentrations, but its corrosive and oxidant nature makes it impossible for use in hospitals or with laboratory equipment. Glutaraldehyde (neither corrosive nor oxidant) is a safe alternative for disinfection of this type of equipment. However, it is important to bear in mind that these pathogens may develop resistance to biocides.     

Efficacy of three prevention strategies against legionella in cooling water systems

The efficacy of three different prevention strategies on legionella in cooling systems was studied. The strategies were as follows: (1) water temperature was lowered; (2) water quality was improved; or (3) the system was disinfected with polyhexamethylene biguanidechloride (PHMB) biocide or with 2-bromo-2-nitropropane-1,3-diol (BNPD) biocide. Lowering of water temperature was the most effective method to reduce the concentration of legionella in cooling systems. Improving of water quality resulted in a transitory disinfection effect. The additions of PHMB or BNPD decreased the concentrations of both legionella and heterotrophic bacteria in cooling water. The effect of biocides, however, lasted at the most only a few months. If possible, lowering water temperature and improving the water quality should be the primary practices for controlling bacterial growth in cooling systems. Regular biocide treatments should be incorporated into the maintenance procedures if technical improvements cannot be done or if their efficiency is too low.