EVALUATION OF ANTIMICROBIAL ACTIVITY AND RESISTANCE PATTERN OF COMMERCIAL HOUSEHOLD BIOCIDES

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ABSTRACT

To access the role of biocide in the environment and the impact of biocide resistance, a panel of biocides was evaluated for antimicrobial activity and biocide resistance patterns.  Equally, plants with traditionally acclaimed biocidal activities were accessed. The samples were subjected to Disc susceptibility testing, minimum inhibitory concentration (MIC) and time-kill determination. Majority of the population use household commercial biocides including benzalkonium chloride, sodium hypochlorite, sodium carbonate, chlorophenol, choroxylenol and ammonium as well as isoprppyl alcohol and other respective combinations.  Preliminary data on zone of inhibition at 1mg/L(v/v) against standard   E. coli and S. aureus strains for benzalkonium (Harpic®), sodium hypochlorite( Jik®), chloroxylenol (Dettol®) and chlorophenol (Ivy fectant®) were all resistant at < 19mm Saureus except dettol and Izal, including Izal towards E. coli >17mm respectively. Xylopia was the only botanical that showed biocidial activity against E.coli >18mm. Xylopia, Azadiracha and Vernonia only showed moderate antistaphylococcal activity at high MIC value (1,024mg/L). Comparatively, Izal B had the highest  to moderate inhibition effect against E. coli  in the following manner Izal> Harpic > Dettol > Jik at 0 hr, while Ivy detergent didnt show biocidal activity at 0 hr contact both at MIC or double MIC value. The double MIC biocide-bacterial time-kill effects were very pronounced and consistent among all the biocides suggesting that higher effect of biocides could only be enjoyed at higher concentrations and longer time contact with the oragnism. This suggests that when applying household biocide, high concentration and more time factor is needed in other to act on the applied area and to achieve inhibition of bacterial growth. The fact that biocides are used extensively in many different products and concentration contribute largely to its efficacy, while at lower dosage usage and less time contact, it remain inactive allowing the organisms to overcome the effect and thereby  contribute to bacterial resistance.







TABLE OF CONTENTS                

Title Page                                                                                                                              i

Certification                                                                                                                         ii

Dedication                                                                                                                            iii

Acknowledgement                                                                                                                iv

Table of Contents                                                                                                                 v

List of Tables                                                                                                                        vi

List of Figures                                                                                                                       vii

Abstract                                                                                                                                viii


CHAPTER ONE                                             1                                                                                            

1.0   INTRODUCTION                                                                                                         1

1.1 Examples showing links between Antibiotic and Biocide Resistance                                    2

1.2What is Biocide?                                                                                                               6

1.3 What is Resistance?                                                                                                         6

1.4 What is biocide resistance?                                                                                              6

1.5 Mechanisms of biocide resistance                                                                                    7

1.6 Mechanisms of intrinsic bacterial resistance to biocide                                                   8

1.7 Mechanism of acquired bacterial resistance to biocides                                                  10

1.8 Common resistance mechanism of both biocide and Antibiotic                                     11

1.1.1 Research Justification                                                                                                    13

1.1.2   Aim and Objectives of the study                                                                                 14


CHAPTER TWO                                                                                                                  15

2.0 Literature Review                                                                                                             15

2.1 chemicals used as ingredient in biocide                                                                           16                   

2.2 Jik                                                                                                                                      17

2.3 Dettol                                                                                                                                18

2.4 Nittol                                                                                                                                 18

2.5 Harpic Active fresh                                                                                                           19

2.2.1 Ivy’s Antiseptic                                                                                                              20

2.2.2 Dettol hand sanitizer and Mum and me hand gel                                                          21

2.2.3 Izal (Z Germicie)                                                                                                            21

2.2.4 Botany of plants investidated for biocidal activity                                                        22

2.2.5 Ricinus communis                                                                                                          22

2.2.6 Jatrophy curcas L.                                                                                                          23

2.2.7Azadirachta indica (Neem)                                                                                             24

2.2.8 Vernonia amydalina                                                                                                       26

2.2.9 Piper guineense                                                                                                              27

2.3 Xylopia aethiopica                                                                                                           28

2.4 Cassia alata                                                                                                                       28

2.5Thinlayer chromatography                                                                                                29 


CHAPTER THREE                                                                                                                31

3.0   MATERIALS AND METHODS                                                                                     31     

3.1 Study Area                                                                                                                         31

3.2 Biocide/ Plant collection and identification                                                                     31

3.3 Preparation of extracts                                                                                                      31

3.4 Preparation of Biocides                                                                                                    32

3.5 Diameter zone of inhibition determination of the biocides                                              33

3.6 Preparation of test organism                                                                                             33                                                                                     

3.7 Minimum inhibitory concentration (MIC) determination                                                33

3.8 Determination of Time – kill curve method                                                         34

3.9 Thin layer chromatography                                                                                  35

CHAPTER FOUR                                                                                                                36

4.0   RESULT                                                                                                                         36

4.1 Antimicrobial activity of the household biocide                                                              36

4.2 Result of MIC determination of biocides and plant extracts                                            40

4.3 Effect of time- kill determination of the biocides                                                             42

4.4 Thin layer chromatography (TLC)                                                                                   44

CHAPTER FIVE                                                                                                                  47

5.0 DICUSSION AND CONCLUSION                                                                              47       

5.1 Discussions                                                                                                                       47

5.2 Conclusion                                                                                                                        49

5.3 Recommendation                                                                                                             51

 References

 Appendix: Abstract and certificate of scientific presentation








LIST OF TABLE

Table 1: Antimicrobial activity of the selected Household biocide against Escherichia coli and Staphylococcus aureus

Table 2: Antimicrobial activity of the selected plant extract against Escherichia coli and Staphylococcus aureus

Table 3:   Minimum inhibitory concentration of commercial household biocide.

Table 4:   Minimum inhibitory concentration of extract against S. aureus

Table 5:  The effect of time – kill on activities of the biocides

Table 6:  Various retention faction as observed on the TLC plates of different phytochemicals present in the each extract.

 

 

 

                                                                             

 

 

 

 

 

                                                      

LIST OF FIGURES

Fig.1: Bargraph showing the time-kill effect of the E. coli biocides combination at 0 hr at A (single MIC of each biocide) and B (double MIC)

Fig. 2: Bargraph showing the time-kill effect of the E. coli biocides combination at 24 hr at A (single MIC of each biocide) and B (double MIC)

 

 

 

 

 

 

 

LIST OF PLATES

Plate 1: Diagram showing Jik detergent used for household cleaning                                  

Plate2: Diagram showing Dettol detergent used for household cleaning                                 

Plate 3: Diagram showing Nittol powder used as household detergent                                            

Plate 4: Diagram showing Harpic detergent

Plate5: Diagram showing Ivy detergent

Plate 6a &b: Diagram of Dettol and Mum & Me hand gel                

Plate 7: Diagram of Izal germicide                                                                

Plate 8: Diagram Ricinus communis

Plate 9: Diagram Jatropha curcas

Plate 10: Diagram of Azadiracha indica

Plate 11: Diagram of Vernonia amygdalina

Plate 12: Diagram of piper guineense

Plate 13a - 13d: Xylopia aethiopica

Plate 14 A: Diagram of Senna alata

Plate 15: Diagram of different diameter of zones of inhibition 

Plate 16: Experimental set up showing a deep blue coloration. Clear tubes indicates no bacterial        growth and deep blue coloration indicate growth. MIC is the least inhibition concentration .

Plate 17: Spotted plate showing various separation in TLC tank

Plate 18: TLC plates showing separation of extracts

 

 

 

 

 

 

 

CHAPTER ONE

 

INTRODUCTION

The term biocide includes disinfectants, antiseptics, and preservatives. Biocides are chemicals designed to kill all sizes and life stages of organisms, especially micro- organisms, effectiveness of biocide varies with the concentration of a biocide and duration of exposure (Akimitsu et al., 1999). Species that are exposed to sub- lethal concentrations, or for too short of time may be injured but may survive. Biocides are used for drinking water treatment, waste water treatment, ship ballast water treatment, disinfectants and as antifouling agents that prevent mollusks from accumulating in industrial pipes. Biocides are produced in liquid and powder forms, in ready- to- use formulations or as concentrates, and are applied using a variety of techniques. It does not include antibiotics, which in spite of being in the strictest sense, tend to be categorized separately. In recent years there has been a trend towards use of biocides in the home environment, schools (especially in day care centers), and the number of chemicals in antibacterial products are enormous, probably at least 10,000 with 1,000 commonly used in the hospitals and homes (Levy, 2001). These products have been marketed for decontamination of food preparation surfaces (e.g. Dettox), area perceived to be microbial contaminated (e.g. toilets) and general improvement cleanliness in the home. A product called microban is a biocide (triclosan) that is incorporated into chopping boards, knife handles and wellington boots. Several workers have suggested that widespread use of biocides may impact on the prevalence of antibiotics resistance micro- organisms.

Biocide resistance was first recognized nearly 70 years ago by ( Heathman et al., 1995a), which identified chlorine resistance in Salmonella typhi while antibiotic resistance was identified shortly after the availability of penicillin, (Heathman et al.,  1995b) but links between the two have only been recognized more recently. It is remarkable that there is a large amount of data on antibiotics resistance, yet there is a comparatively small number of workers worldwide who are investigating the mechanism of biocides resistance. Because biocides tend to act concurrently on multiple sites within the micro – organism, resistance is often mediated by non- specific means.


1.1 EXAMPLES SHOWING LINKS BETWEEN ANTIBIOTIC AND BIOCIDE RESISTANCE

(Manzoor et al., 2009) demonstrated ethambutol resistance in strains of Mycobacterium chelonae that has selected in vitro to be resistance glutaraldehyde. This resistance was associated with changes in the composition of the cell well, indicating that reduced permeability may be the mechanism for this cross resistance. These links between biocide and antibiotic resistance are not confined toa typical Mycobacteria, and it has been shown that resistance to the benzalkonium chloride is closely linked to oxacilline resistance in Staphylococcus aureus. (Akimitsu et al.,, 1999) reported that benzalkonium chloride resistant mutants of methicillin resistant Staphylococcus aureus (MRSA) had oxacillin MICs as high as 512mg/l, composed with 16mg/l for the parent strain and 0.3mg/l for methicillin susceptible Staphylococcus aureus (MSSA). Furthermore, resistance to plasmid can be mediated. A strain of gentamicin resistant (MRSA) was shown to contain a multidrug resistant plasmid (PSAJI) that conferred resistance to aminoglycosides, ethidium bromide, benzalkonium chloride and chlorhexidine. This plasmid, when transferred to E. coli, continued to express resistance to the same biocides as when in the original host. Plasmid mediated resistance to biocides is a well-recognized phenomenon. Such as to quaternary ammonium compounds and other biocides has been identified in Staphylococcus aureus ,Pseudomonas spp and members of the Enterobacteriaceae, and is mediated by specific genes (qacA,B,C,D and E) qacA,BandC (described in S.auerus) mediated resistance by an active efflux mechanism and have sequence homology with tetracycline efflux genes. qacE is a plasmid mediated resistance gene found in gram negative organisms that also codes for an energy dependent multidrug efflux mechanism. These resistance determinants are associated with resistance to a variety of antibiotics including trimethoprim, sulphonamides, oxacillin and aminoglycosides.

The public is bombarded with advertisements advocating the use of biocides in the home. The implication is that homes are dangerous places, heavily contaminated with virulent microorganisms, and the only way to ensure the safety of one’s children is to use disinfectant liberally. This engenders a false sense of security in the public mind by suggesting that the widespread use of biocide reduces the chance of acquiring infectious diseases. There are no data to support this stance; rather, as biocide use in the home environment continues to increase. There is a real risk that widespread biocide use could exacerbate the already worrying trend towards increased antimicrobial resistance in clinically relevant organisms. The problem is that we do not yet know how great the problem is, or even if a problem exists. There are no good epidemiological data on the impact of biocide use on antimicrobial resistance, and our knowledge of the prevalence of antimicrobial resistance is still poor.

In order to understand and control biocide resistance it is essential that more effort is put into surveillance so that we can understand the impact of the use of antimicrobial agents on the epidemiology of resistance organisms. Resistance to biocide is less common and likely reflects the multiplicity of targets within the cell as well as the general lack of known detoxifying enzymes.

Resistance typically results from cellular changes that impact on biocide accumulation, including cell envelope changes that limit uptake, or expression of efflux mechanisms. Still, target site mutations leading to biocide resistance, though rare, are accommodating biocide (e.g. triclosan) such that strains expressing these are both antibiotic and biocide resistant (Russell, et al., 2002).  Efflux pumps have the potential to act on a range of chemically dissimilar compounds and have been implicated in both biocide and antibiotic- resistance bacteria. Cell wall changes may also play a role in the observed cross- resistance between biocides and antibiotics, probably by reducing permeability. Microbial changes that result in resistance to biocides should therefore cause concern. However, of equal significance, is the possibility of genetic linkage between genes of biocides resistance and those for antibiotic resistance.

Much later, iodine found use as wound disinfectant, chlorine water in obstetrics, alcohol as a hand disinfectant and phenol as a wound dressing and in antiseptic surgery. In the early part of the twentieth century, other chlorine releasing agents (CRAs), and acridine and other dyes where introduced, as were some quaternary ammonium compounds (QACs, although these were only used as biocides from the 1930s). Later still, various phenolic and alcohols, formaldehyde and hydrogen peroxide were introduced and subsequently (although some had actually been produced at an earlier date)biguanides, iodophors, bisphenols, aldehydes, diamidines, isocyanurates, isothiazolones and peracetic acid. It has been claimed that the chronological emergence of qacA and qacB determinants in clinical isolates of Staphylococcus aureus mirrors the introduction and usage of cationic biocides, (Russell et al.,, 1997).If everything else is constant, then the more concentrated the biocide the greater its efficacy and the shorter the time necessary to kill all the micro-organisms. Not all biocide are similarly affected by concentration adjustments. In order words, halving the concentration may double the exposure time for some biocides but quadruple it for others. It is particularly important to ensure that disinfectants do not become diluted with excess water remaining on endoscopes after rinsing, as the activity of the disinfection process will be significantly compromised. Thus, it is important that water is purged from all channels after cleaning and before disinfecting and to regularly monitor the concentration of biocide when it is being reused. The potency of a biocide affects the exposure time required to achieve the same level of microbial kill. Several physical and chemical factors influence the efficacy of biocides. These include; temperature, ph, relative humidity and water hardness.

Generally, the activities of most biocides increase as the temperature increases as the temperature increases. There is a point at which the chemical degrades if heated too much. An increase in ph improves the antimicrobial activity of some agents as with glutaraldehyde, but decreases the activity of others such as hypochlorite. This effect is caused either by alteration of the germicidal molecule or the cell surface of the micro organism. Relative humidity influences the activity of gaseous agents such as ethylene oxide. Water hardness reduces the rate of kill of biocide because divalent cation such as magnesium and calcium interact with soap to form insoluble precipitates. Organic matter such as serum, blood, pus or faecal material may interfere with the activity of biocides in at least two ways; first, a chemical reaction between biocide and organic matter may result in a complex that is les germicidal or non germicidal leaving less of the active agent to attack the micro organisms. Secondly, organic material many protect microorganisms from attack by acting as a physical barrier. This is another reason for the meticulous cleaning of object before any sterilization or disinfection procedure.

To adequately sterilize or disinfect an item, it must be exposed to the appropriate concentration of biocide for a certain minimum contact time. All surfaces of the item must come in contact with the biocide for that period of time. This means that for endoscopic equipment the biocide must be introduced into all lumens and channels. Presence of air pockets and incomplete immersion in the biocide means item will not be reliably or completely processed.


1.3 WHAT IS BIOCIDE?

 According to the direction 98/8/EC of the European parliament and council of the 16 February 1998. Biocides are defined as active substance and preparations containing one or more active substance, put up in the form in which they are supplied to the user, intended to destroy, render harmless, prevent the action of or otherwise exert a controlling effect on any harmful organism by chemical or biological means.

The active substance are without concern {Annex IA of the directive} or with concern about their inherent capacity to cause an adverse effect on humans, animals or the environment.


1.4 WHAT IS RESISTANCE?

Resistance is the ability of micro-organisms of certain species to survive or even to grow in the presence of a given concentration of an antimicrobial agent that is usually sufficient to inhibit or kill micro-organisms of the same species.


1.5 WHAT IS BIOCIDE RESISTANCE?

Biocide resistance occurs when a biocide lost its effectiveness to kill or inhibit microbial growth. Thus, the bacteria are resistant and continue to multiply.

 

1.6 MECHANISMS OF BIOCIDE RESISTANCE

Biocides have multiple target sites against microbial cells. Thus, the emergence of general bacterial resistance is unlikely to be caused either; (i) by a specific modification of a target site or (ii) by a by- pass of a metabolic process.  It emerges from a mechanism / process causing the decrease of the intercellular concentration of biocide under the threshold that is harmful     to the bacterium. Several mechanisms based on this principle (mode of action) have been well described including change in cell envelope change in permeability, efflux and degradation. It is likely that these mechanisms operate synergistically although very few studies investigating multiple bacterial mechanisms of resistance following exposure to a biocide have been performed. The efficacy of biocides depends on a range of intrinsic and extrinsic factors, (EFSA, 2008a) Intrinsic factors are characteristics of the biocidal agent and its application, concentration and contact time are crucial. Furthermore, the combination of contact time and concentration determine the result in term of microbial reduction. This is called the CT concept, and within certain limits of time and concentration, there is a relationship with a defined constant characterizing efficacy. Thus, the same result could be obtained with a high concentration of disinfectant during a short contact time, or a lower concentration during a longer contact time. The stability of the active compounds of the biocide in the environment also influences the efficacy. Extrinsic factors derive from the environment during application. The temperature of the environment is important, as mort substance have a lower efficacy at low temperature.  The presence of protein reduces efficacy as they interact with the substance. The mode of contact also influences the efficacy, as does the contact time (mechanical effects). The ph is another important factor.  The concentration of the micro organism, the age of the bacterial community and protection by attachment on particular matter, and the presence of biofilms play an increasing important role.


1.7 MECHANISMS OF INTRINSIC BACTERIAL RESISTANCE TO BIOCIDES

Several mechanisms conferring bacterial resistance to biocides have been described; some are inherent to the bacterium, others to the bacterial population. In addition, some of the resistance mechanisms are intrinsic (or innate) to the micro organism while others have been acquired through forced mutations or through the acquisition of mobile genetic elements (Poole 2002a). Innate mechanisms can confer high level bacterial resistance to biocides. In this case, the term insusceptibility is used. The most described intrinsic resistance mechanism is changes in the permeability of the cell envelope, also referred to as “permeability barrier”. This is not only found in spores (Cloete 2003,and Russell et al.,, 1997), but also in vegetative bacterial such as mycobacteria and to some extent in Gram negative bacteria.  The permeability barrier limits the amount of a biocide that enters the cell, thus decreasing the effective biocide concentration (Denyer and Lambert 2002). In mycobacteria the presence of a mycolarabinogalactan layer account for the impermeability to many antimicrobials (Lambert 2002, Russell 1996, Russell et al.,, 1997). In addition, the presence and composition of the arabinogalaitan ?arabinomannan cell also plays a role in reducing the effective concentration of biocide that can penetrate within mycobacteria (Manzoor et al.,, 1999).

The role of the lipopolysaccharides (LPs) as a permeability barrier in Gram negative bacteria has been well documented (Denyer and Maillard 2002, McDonnell et al.,) There have also been a number of reports of reduced biocide efficacy following changes in other components of the outer membrane ultra structure including proteins (Brozel and Cloete 1994). Fatty acid composition (Guerin-Mechinet al.,, 1999) and phospholipids. It must be noted that in the above mentioned examples, an exposure to biocides was followed by changes in ultra structure related to a decrease in biocidal susceptibility, usually at a low concentration (under the MIC value).  The charge property of the cell surface also plays in bacterial resistance mechanisms to positively charged biocides such as QACs. It is likely that bacterial resistance emerges from a combination of mechanisms (Tattawasart et al., 2000), even though single specific mechanisms are often investigated. The presence of efflux pumps is another mechanism that has been well described in the literature. It has gained increased recognition as a resistance mechanism over the past decade. Efflux pumps decrease the intercellular concentration of toxic compounds, including biocides (Borges - Walmslay 2001, Levy2002and Putman et al.,, 2000). They are widespread among bacteria and five main classes have been identified: the small multidrug resistance (SMR) family (now part of the drug / metabolite transporter (DMT) super family), the major facilitator super family (MFS),the ATP – binding cassette ( ABC) family, the resistance nodulation division ( RND) family and the multidrug and toxic compound extrusion (MATE) family (Borges – Walmsley, 2001 and,Poole 2004,). The importance of efflux pumps in terms of bacterial resistance to biocides might be considered as modest since the increase in bacterial susceptibility to select biocides as the results of the expression of efflux pumps is usually measured as an increase in MICs rather than as resistance to a high concentration of an active substance. Efflux pumps have been shown to reduce the efficacy of a number of biocides including QACs, phenolics parabens and intercalating agents notably in staphylococcus aureus with identified pumps such as QacA-D, QacG and QacH s and in Gram negative bacteria such as pseudomonas aeruginosa, with MexAB – OprM, MexCD – OprJ, MexEF – OprN and MexJK(Poole 2004,) and Escherichia coli with AcrAB – ToIC, AcrEF – ToIC and EmrE  (McMurry et al., 1998a ).  The enzymatic transformation of biocide has also been described as a resistance mechanism in bacteria, notably to heavy metal (e.g. silver and copper, enzymatic reduction of the cation to the metal, (Cloete2003); parabens, aldehydes formaldehyde dehydrogenase, (Kummerle et al., 1996), peroxygens catalase, super oxide dismutase and alkyl, hydroperoxidases mopping up free radicals, (Demple 1996). Environmental bio-degradation of various compounds has been well described notably among pseudomonas and complex microbial communities.  However, the importance of degradation as a bacterial resistance mechanism to “in use” concentrations (high concentrations) of biocides remain under.  As for efflux, increased resistance following degradation of biocides has been measured as s decreased in MICs but not necessarily as a decreased in lethal activity.  The modification of target sites has been described on rare occasions and does not seem to be widespread among bacterial, although there is a paucity of information on this subject. The bisphenoltriclosan has been shown to interact specifically with an enoylacyl reductase carrier protein at a low concentration (Healthet al., 1999).  The modification of this enzyme has been associated with low level bacterial resistance (Health et al., 2000, McMurry et al., 1999). It has been noted that at a high concentration triclosan must interact with other targets within the cell, the alteration of which justified the lethal effect of the bisphenol (Gomez Escalada et al., 2005a).


1.8 MECHANISMS OF ACQUIRED BACTERIAL RESISTANCE TO BIOCIDES

The development of bacterial resistance through acquired mechanisms such as mutation and the acquisition of resistance determinants are of concern since a bacterium that was previously susceptible can become insusceptible to a compound or a group of compounds (Russell 2002a).   The acquisition of resistant genes has been well described in the literature (and it is particularly important to consider, this as it might confer cross or co – resistance on occasion (Poole 2004). However, there is little information on the effect of biocides on the transfer of genetic determinants. One study in particular highlighted that while some biocides at a sub-inhibitory (residual) concentration could inhibit genetic transfer, others increased genetic transfer efficiency. There have been investigations on co- transfer of resistant markers in epidemic methicillin resistant Staphylococcus aureus following antibiotic treatment to decolonize patients (Cookson et al., 1991a). The authors reported that there was no evidence of increased in chlorhexidine MICs six years after the first isolation of the epidemic strains, although the strain carried a qac gene (Cookson 2005).

However, this was not the case with triclosan, where clinical isolates of Staphylococcus aureus showed high level mupirocin resistance and low level triclosan resistance (MlC 2- 4 mg/l) (Cookson et al., 1991b). The authors described that resistance to both chemically unrelated compounds was transferred and cured together (Cookson 2005).


1.9 COMMON RESISTANCE MECHANISMS OF BOTH BIOCIDE AND ANTIBIOTICS

Considerable controversy surrounds the use of biocides in an ever increasing range of consumer products and the possibility that their indiscriminate use might reduce biocide effectiveness ant alter susceptibilities towards antibiotics (Gilbert and McBain 2003). These concerns have been based largely on the isolation of resistance mutants from in-vitro monoculture experiments. Some of the evidence suggests that exposure to biocides may be leading to increased antibiotic resistance, but the number of studies in the clinical or environmental setting is low.

However, a recent study performed in the community highlighted a significant relationship between high QAC MICs, high MICs to triclosen and resistance to one or more antibiotics. Further research is needed to establish a correlation between biocide exposure(s) and development of antibiotic resistance. Biocides that tend to act concurrently on multiple sites within the micro organism and thus resistance is often mediated by non-specific means. Efflux pumps have been shown to cat on a range of chemically dissimilar compounds and have been implicated in both biocide and antibiotic resistance bacteria (Maillard 2007 and Poole 2007). Cell wall changes by reducing permeability may also play a role in the observed resistance to biocide. The possibility of genetic linkage between genes for biocide resistance and for antibiotics resistance has also been described.

Several publications and reviews have presented the cell target of biocides and the various mechanism used by the bacterial cell to evade the toxic activity of biocides (Denyer and Maillard 2002, Lambert 2004, Maillard 2007, Poole 2007).it is important to note that antibiotics and biocide antibacterial actions show many similarities despite some differences in terms of target, killing, behavior and clinical aspects (Poole 2007).among the similarities, we can mention;

      i.         The penetration/uptake through bacterial envelope by passive diffusion;

     ii.         The effect on the membrane integrity and morphology;

   iii.         The effect on diverse key steps of bacterial metabolism (replication, transcription, translation, transport, various enzymes).

Faced with this toxic effect and stress, the responses, adaptation of bacterial cells presents some similar defense mechanisms that can overlap the original functions to confer resistance against structurally non-related molecules. Among the biocide resistance strains intrinsic and acquired mechanisms are described. Intrinsic resistance is an innate property conferred by the bacterial genome (species-dependent) and includes impermeability, efflux, biofilms and transformation of toxic compounds. To decrease the intercellular concentration of noxious molecules, Gram-negative bacteria can regulate the permeability of their membranes by decreasing the synthesis of porins (membrane pore- forming proteins involved in antibiotic uptake) and modifying the lipopolysaccharide structure (Poole et al., 2002) or over expressing the efflux pumps (membrane portentous complexes involved in antibiotic expulsion) (Poole 2007). These strategies are involved in the resistance against antibiotics and biocides. In parallel, the acquired resistance occurs via mutation and acquisition of mobile DNA (transposon, plasmids) coding for resistant element (enzyme transporter).

Similarly, the acquired processes may protect against antibiotics and biocides (Millard 2004). In addition, some of the mechanisms that play a major role in resistance are controlled by diverse genetic cascade regulations that share common gene regulations (Sox S, Mar A) (Poole 2007).


1.9 RESEARCH JUSTIFICATION

Several research works have been carried out on assessment of antimicrobial activities but very few have been documented on the efficacy of commercially available household biocides and their impact on resistance drive in the environment and homes. Hopefully, some botanicals may contain biocidial properties as they are used traditionally for such purpose with little or no scientific background or documentation for their activity.  Thus, this study seeks to access the antimicrobial efficacy of household biocides and their resistance patterns as well some botanicals used as local biocides. 

 

1.10  AIM AND OBJECTIVES OF THE STUDY

1.To evaluate antimicrobial activities of commercial biocidal products and their resistance pattern.

2.To provide information to the public of the resistance pattern of microorganism to biocide, through scientific presentation or publication

3.To compare the effect of botanicals and biocides on selected pathogenic organisms.

 

 

 

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