Determination Of The Microbial Contamination Of Disinfectant And Antiseptic Produced In Luth

Project and Seminar Material for Pharmaceutical Science

Determination Of The Microbial Contamination Of Disinfectant And Antiseptic Produced In Luth


This study was conducted to determine the prevalence of bacteria in disinfectants/antiseptics produced and used in three major wards in Lagos University Teaching Hospital (LUTH) and to determine their microbial load in these ward. A questionnaire component was used within LUTH, and a laboratory study was carried out on samples of disinfectant and antiseptic samples collected in three (3) departments/wards. Of the total of 60 tested samples, 7 samples (11.7%) were contaminated by aerobic bacteria. Of these, chlorhexidine gluconate and cetrimonium bromide (1% concentration) were the most contaminated, with 15% of the samples positive for aerobic bacteria (P0.05). Of the 24 isolates in our study, 58.3% of them were resistant to ciprofloxacin, again emphasizing the possible therapeutic consequences of infections that result from exposure to contaminated disinfectants. In order to minimize the risk posed to patients by disinfectant contamination in health facilities in Trinidad and Tobago, we recommend that: (1) reduce the use of disinfectants in the hospital; (2) reduce the amount of bacterial contamination in the ward; and (3) reduce the bacteriological quality of the disinfectants; and (4) increase the sensitivity of the bacteria to antimicrobial agents.

Chapter One


1.1 Background of the Study

Antiseptic and disinfectants are chemical agents that inhibit or destroy microorganisms on living tissue (antiseptics) and inanimate surfaces and objects (disinfectants). These chemical agents are used extensively in hospitals and other health care settings for a variety of topical and hard surface applications. In particular, they are an essential part of infection control practices and aid in the prevention of nosocomial infections. This wide spread use of antiseptic and disinfectant products has prompted some speculation on the development of microbial resistance, in particular, whether antibiotic resistance is induced by antiseptics or disinfectants.

A wide variety of active chemical agents (biocides) are found in these products, many of which have been used for hundreds of years for antisepsis and disinfection including; alcohols, phenols, iodine, and chlorine. Most of these active agents demonstrate a broad spectrum of antimicrobial activity and they are used to reduce microbes on the living tissue or surface of medical equipments and other inanimate objects. The mechanism of resistance to antiseptic and disinfectant solutions includes cellular impermeability, biofilm formation, efflux and mutations at the target site or over expression of a target site.

As a therapeutic significance a number of bacterial contaminants isolated from antiseptics and disinfectants have exhibited resistance to commonly used antimicrobial agents. It has also been reported that contaminated antiseptics and disinfectants exhibit decreased efficacy and effectiveness. The response of different types of microorganisms to antiseptics and disinfectants vary and result in micro-biostatic or microbiocidal effects. Multiple nosocomial out breaks have resulted from a lack of intrinsic antimicrobial activity of antiseptics, resistant pathogen, over dilution of the antiseptics, or the use of contaminated antiseptics.

Bacterial contamination of disinfectant solutions is common with preparation by unskilled personnel, use of unsterilized containers and prolonged use with other contributing factors for the high levels of contamination (dilution of disinfectants with tape water, inadequate care of stock solution bottles and long storage of the diluted disinfectants in the wards).

In adequate disinfection of medical devices or environmental surfaces may result from lack of intrinsic antimicrobial activity of the disinfectant, an incorrect choice of chemical disinfectant, a resistant pathogen, over dilution of the disinfectant, inadequate duration of disinfection, lack of contact between the disinfectant and the microbes, or the use of contaminated disinfectants. With the emergence of pathogens such as Methicillin Resistant Staphylococcus Aureus (MRSA), Vancomycin Resistant Entericocci(VRE) and gram negative bacteria, for example, Pseudomonas aeruginosa and acinetobacterspecies which are resistant to multiple antibiotics, there is an increased need for effective antisepsis and disinfection.

As with antibiotic resistance, resistance to these germicides may be an intrinsic property or may arise either by chromosomal gene mutation or by the acquisition of genetic materials. As a result of this health care associated infections are important causes of morbidity and mortality all over the world. The center for disease control and prevention (CDC) has estimated that health care associated infection accounts for an estimated 1.7 million infections, 99,000 deaths and $ 4.5 billion in excess health care costs annually. Despite best efforts to eliminate these infectious microorganisms, they continue to emerge and re-emerge which contribute to human illness and death especially as a result of hospital acquired infections. The successful eradication of these pathogens with antiseptic/disinfectant solutions has been complicated by the development of highly resistant strains. As a result of extensive use of antiseptic/disinfectant solutions, a significant proportion of the pathogens have not only developed resistance, but they also grow in the solution of these biocides. The activity of biocides against microorganisms is not always consistent due to several basic methodological problems as well as high intrinsic resistance due to differences in membrane structure. Persistent reports have shown that disinfectants designed for the control of infectious microorganisms are themselves subjected to microbial contamination. It has become increasingly obvious that infections acquired in hospitals lead to increase morbidity and mortality which has also added noticeable to economic burden. As different researchers show that contaminated antiseptics or disinfectants pose a health risk to patients particularly in the pediatric and surgical wards. Bacteria isolated from contaminated antiseptic/ disinfectant solutions exhibit increased resistance to commonly used antibiotics which contributes a serious public health problem, giving the fact that bacteria have the ability to share resistance markers, and once a resistance develops for one agent, a cross-resistance to other agents can occur. Nosocomial infections associated with contaminated antiseptic products are difficult to assess. Several factors may limit the identification of infections related to antiseptics or disinfectant products, however; it is a recognized public health problem worldwide with the prevalence rate of 5-10%. Generally these infections pose a problem of enormous magnitude globally by prolong hospitalization, increase cost of health care, and decrease the effectiveness of the treatment.

1.2 Statement of the Problem

One major cause of hospital infection is the use of contaminated disinfectants and antiseptics. Reports of epidemics of nosocomial bacterial infections have implicated contaminated disinfectants and antiseptics both applied directly to the skin of animals or humans and used to decontaminate instruments and appliances used for diagnosis and treatment.

Hospitals are been using various disinfectants and antiseptics extensively, but there is no report on the microbial contamination of these biocides from any referral hospital from this part of the country.

1.3 Objective of the Study

The aim of this study is to determine the microbial contamination of disinfectants and antiseptics produced and used in three major wards in Lagos University Teaching Hospital (LUTH) and to determine the microbial load in these wards.

1.3 Research Questions

  1. What is the effect of microbial contamination of disinfectants and antiseptics produced and used in LUTH?
  2. How can the level of microbial load of the contamination of disinfectants and antiseptics in three wards in LUTH be determined?

1.5 Research Hypotheses

H0: There is no significant effect of microbial contamination of disinfectants and antiseptics produced in LUTH.

H1: There is significant effect of microbial contamination of disinfectants and antiseptics produced in LUTH.

1.6 Significance of the Study

This study is an attempt to improve the delivery of public health care in Lagos University Teaching Hospital, including reducing or eliminating contamination of disinfectants and antiseptics in pharmacy departments, pediatric/neonatal wards as well as of surgical equipment in surgical wards in LUTH.

This study will be of immense benefit to other researchers who intend to know more on this topic and can also be used by non-researchers to build more on their work.

1.7 Scope / Limitations of the Study

This study was conducted to determine the prevalence of bacteria in disinfectants/antiseptics at three departments/wards in Lagos University Teaching Hospital (LUTH):

  1. Pharmacy departments (pharmacy stock and diluted samples before delivery to wards).
  2. Pediatric/neonatal wards (after delivery from the pharmacy department, diluted pre-use and in-use samples), and;
  3. Surgical wards (after delivery from the pharmacy department, diluted pre-use and in-use samples).

The investigation also determined the sensitivity of the bacteria to antimicrobial agents.

1.8 Limitations Of Study

This study will have some limitations most especially in the area of data collection. Financial constraints as well as time available for the completion of the study among other factors will limit the scope of the study.

1.9 Background Study Of Lagos University Teaching Hospital (Luth)

Lagos University Teaching Hospital (LUTH) is a tertiary referral hospital in Idi-Araba, Surulere, Lagos State, Nigeria.

The hospital was established in 1962 and is affiliated with the University of Lagos, College of Medicine.

1.10 Definition Of Terms


An agent that inhibits or destroys microorganisms on living tissue including skin, oral cavities, and open wounds.

Chemical Disinfectant:

A chemical agent used on inanimate surfaces and objects to destroy infectious fungi, viruses, and bacteria, but not necessarily their spores. Sporicidal and antiviral agents may be considered a special class of disinfectants.

Disinfectants are often categorized as high-level, intermediate-level, and low-level by medically oriented groups based upon their efficacy against various microorganisms.

Cleaning Agent:

An agent for the removal from facility and equipment surfaces of product residues that may inactivate sanitizing agents or harbor microorganisms.


The removal of microorganisms by disinfection or sterilization.


A chemical or physical agent that destroys or removes vegetative forms of harmful microorganisms when applied to a surface.

Sanitizing Agent:

An agent for reducing, on inanimate surfaces, the number of all forms of microbial life including fungi, viruses, and bacteria.

Sporicidal Agent:

An agent that destroys bacterial and fungal spores when used in sufficient concentration for a specified contact time. It is expected to kill all vegetative microorganisms.


An agent that destroys all forms of microbial life including fungi, viruses, and all forms of bacteria and their spores. Sterilants are liquid or vapor-phase agents.


Are microscopic organisms that exist as unicellular, multi-cellular, or cell clusters. Micro organisms are widespread in nature and are beneficial to life, but some can cause serious harm. They can be divided into six major types: bacteria, archaea, fungi, protozoa, algae, and viruses.


Is the invasion of an organism’s body tissues by disease-causing agents, their multiplication, and the reaction of host tissues to these organisms and the toxins they produce. Infectious disease, also known as transmissible disease or communicable disease, is illness resulting from an infection.

Infection Control:

Is the discipline concerned with preventing nosocomial or healthcare-associated infection, a practical (rather than academic) sub-discipline of epidemiology. It is an essential, though often under recognized and under supported, part of the infrastructure of health care.

Chapter Five

Discussion Of Findings


The 6.1% prevalence of contaminated disinfectant samples from the four hospitals in Trinidad compares favorably with the 3% prevalence found in 11 Danish hospitals (18) and the 7.9% reported for Malaysian hospitals (19). Considerably higher prevalence levels have been reported for other countries, including 34.4% in Nigeria (7) and 43% in Japan (8). However, when comparing the frequency of contamination, one should consider the types and concentrations of disinfectants since resistance varies among microorganisms (2, 4). The total aerobic plate count that we found, which ranged from under 10 CFU/mL to 3.6 2 10 4 CFU/mL, is similar to reports by others, where the counts ranged from 10 2 to 10 8 CFU/ mL (8, 9). The relatively high count of aerobic bacteria in the samples is indicative of the probability of attaining an infective dose and of establishing an infection (20, 21). The risk is further magnified when contaminated disinfectants are used in sensitive areas of the health facilities such as surgical wards and pediatric/neonatal wards. Clearly, any resulting nosocomial infections could have grave consequences (20, 22, 23). Of the three disinfectants/antiseptics investigated, chlorhexidine gluconate and cetrimonium bromide (which was a 1% concentration) was the most contaminated, with 15% of the samples positive for aerobic bacteria. Since similar practices took place in the preparation of the three disinfectants/antiseptics studied, it was expected that they would all have been exposed to the same degree of contamination. However, the difference in the frequency of contamination may be explained in part by the low concentration (1%) of chlorhexidine gluconate and cetrimonium bromide, which obviously was unable to inhibit or kill the contaminants. Given that this substance was the one that the 11 hospitals used most frequently to clean wounds, the health risks for patients in Trinidad are clear. Contaminated chlorhexidine gluconate and cetrimonium bromide was responsible for an outbreak of Pseudomonas maltophilia amongst Australian patients (20). Of the 60 chlorhexidine gluconate samples in our study, only 2 of them (3.3%) were contaminated by aerobic bacteria. Nevertheless, the fact this disinfectant was applied on skin surfaces did not entirely eliminate the risk that it poses to the patients in Trinidad.

Other research (8) has reported a considerably higher prevalence, 60%, for contaminated chlorhexidine gluconate samples. All the samples of methylated spirit that we studied from all the sources were negative for aerobic bacteria. This suggests that the concentration of this antiseptic and its mode of activity completely eliminated all the contaminating aerobic bacteria that we found in the two other disinfectants that we studied. By its inherent nature, methylated spirit would not harbor microorganisms, and it is used in its original concentration from the supplier. It was hardly a surprise that all stock solutions of the three disinfectants/antiseptics tested in all the pharmacies were free of aerobic bacteria. This could be explained in part by the high concentration of the active ingredient in the disinfectants/antiseptics prior to dilution, as well as the fact that they had not yet been exposed to potential environmental contamination. It has been established that unsanitary practices during the preparation and distribution of disinfectants/antiseptics contribute significantly to their contamination in a hospital environment (10, 20, 22–24). For example, using inappropriate sources of water to dilute the disinfectants/antiseptics as well as failing to maintain adequate cleanliness of the disinfectant/antiseptic containers are important sources of bacteria. Deionized water used in diluting chlorhexidine gluconate and cetrimonium bromide was the source of P. maltophilia in a nosocomial epidemic in Australia (20). In Trinidad, pipeborne water has been reported to have a low residual chlorine level and high coliform counts (25, 26), contrary to the recommended zero tolerance for coliforms in potable water (27). Nevertheless, the pipeborne water at these four hospitals did not appear to be the main source of aerobic bacteria contamination. This is because aerobic bacteria were not detected in any of the disinfectant/antiseptic samples from the two of the four hospitals where disinfectant containers are subjected to a high temperature while undergoing two cycles each of wash and rinse, followed by autoclaving at 255 °F at 15 lb per square inch of pressure (124 °C at 1.055 kg/cm2 ) for 7 to 10 min. Contaminated containers can contaminate disinfectants (21, 28). Studies elsewhere have shown that in-use disinfectants are more heavily contaminated than diluted samples at the pharmacy level (7, 19, 20). That is because the containers and the length of time that diluted samples are used in the wards affect contamination and the growth of bacteria. However, in our study, we did not individually trace the disinfectant/antiseptic samples in a longitudinal fashion from the pharmacy departments to the pediatric/neonatal wards or the surgical wards of the hospitals; we took samples at the hospitals only at weekly intervals. This limitation may partially explain our failure to detect a higher prevalence or counts of aerobic bacteria in the in-use disinfectant/antiseptics in the pediatric/neonatal wards and the surgical wards as compared with diluted disinfectant/antiseptic samples in the pharmacy departments awaiting distribution in the hospitals. It was noteworthy that Pseudomonas spp. accounted for all the aerobic bacteria isolated from the disinfectant samples studied.

Pseudomonas spp. have been reported to be the predominant aerobic bacteria recovered from contaminated disinfectants/antiseptics elsewhere (8, 19). It has also been reported that different Pseudomonas spp. vary in their resistance to disinfectants (2, 5). However, bacteria other than Pseudomonas have been isolated from contaminated disinfectants and have been implicated in nosocomial epidemics (19, 29). Of therapeutic relevance is our finding that all 24 isolates of Pseudomonas spp. tested exhibited resistance to one or more of the 14 antimicrobial agents tested. In the health care delivery system in Trinidad and Tobago, in treating Pseudomonas spp. infections, gentamicin is the empirical choice for intravenous treatment of inpatients who have good renal functions, whereas ceftazidime is prescribed for patients with renal impairment (30). It is worrisome that 41.7% of the 24 isolates in our study were resistant to gentamicin and that 100% of them were resistant to ceftriaxone. In Trinidad and Tobago one of the quinolones, ciprofloxacin, is the first line of oral therapy for Pseudomonas infections. Of the 24 isolates in our study, 58.3% of them were resistant to ciprofloxacin, again emphasizing the possible therapeutic consequences of infections that result from exposure to contaminated disinfectants/antiseptics. In the high-risk areas of the hospitals (the pediatric/neonatal wards and the surgical wards) 5 out of the 72 samples of pre-use or in-use disinfectants/antiseptics were contaminated. This 6.9% prevalence is obviously unacceptable because of the health risk posed to patients in these sensitive areas. Combined with the relatively high counts of Pseudomonas spp. isolated, the high prevalence of resistance to antimicrobial agents commonly used in their control is definitely a source of concern. In order to minimize the risk posed to patients by contaminated disinfectants/antiseptics in health facilities in Trinidad and Tobago, we recommend that:

  1. Standardized guidelines for preventing microbial contamination of disinfectants/antiseptics be instituted and enforced in all hospital pharmacies.
  2. functional autoclaves be provided by the Government to all public hospitals or health institutions that prepare solutions such as disinfectants/antiseptics for utilization in critical areas, e.g., pediatric/neonatal wards and surgical wards, in order to sterilize all disinfectant/antiseptic containers; in cases where the desired sterilization temperature is not achievable, all containers must be subjected to at least 100 °C prior to filling with disinfectants/antiseptics.
  3. Health facilities should be encouraged to use freshly prepared disinfectants/antiseptics and to institute measures to monitor (e.g., by expiry dating) diluted disinfectants/antiseptics to minimize prolonged use in the postdilution period; wherever possible, single-use containers, with a maximum volume of 500 mL, should be introduced.
  4. Systems should be instituted to determine the microbial load of disinfectants/antiseptics, particularly of in-use preparations, in order to minimize the potential health risk that they pose. This can be done by routine random microbiological sampling of disinfectant/antiseptic samples before leaving the pharmacy departments and while in use at their destinations in the hospitals.

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