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Germ Kill (Skin Drying) Testing

MEASURING SKIN ANTIBACTERIAL ACTIVITY BY THE GLOVE JUICE METHOD
By Nichole Juarez

Dr. Angela Hudson California State University, Fresno

Background
According to the Center for Disease Control (CDC), the number of resistant organisms within the community is increasing. The presence of resistant microbes, in the hospital, is not a new or unexpected occurrence. However, these resistant microbes, once seen only in nosocomial infections, are now being seen, in the community, at a rate that continues to increase on a daily basis. It is believed that resistant organisms such as methicillin resistant staph aureus (MRSA), vancomycin resistant enterococcus (VRE), and necrotizing fascitis are becoming more prevalent in the community due to factors such as: non-compliance with hand hygiene practices, cross-transmission from patients to family members, and incorrect or inadequate prescription and administration of antibiotic therapy.

Hand washing“remains the single most effective and cost-efficient method for preventing and reducing the transmission of nosocomial infections” (Antoniak, 2004, p.1) However, non-compliance with hand washing has been a factor in continuing the spread of resistant organisms. This issue was, initially, addressed with the invention of rinse-free, alcohol-based hand sanitizers. The alcohol-based hand sanitizers have proven to be more effective than hand washing in regards to disease prevention (Moralejo, 2003, p. 1). However, studies have shown that the alcohol-based sanitizers are not only flammable, posing a risk to any healthcare facility that utilizes this product; the alcohol-based sanitizer can also cause drying and cracking of the hands, becoming less effective with increased use. This study hopes to test the alcohol-based sanitizer against a newer, novel surfactant, allantoin, benzalkonium chloride (SAB) hand sanitizer. The SAB hand sanitizer is water-based, thus, eliminating many of the adverse effects associated with hand sanitizer use.

Purpose
The purpose of this study is to determine the sustained effectiveness of two different types of hand sanitizers, alcohol-based and alcohol-free, on bacterial growth over a forty eight hour period of time by utilizing the federally approved glove juice protocol amongst student volunteers at California State University, Fresno.

Aims
The three specific aims of the study are to: a) identify the relationship between the chemical basis of two different types of hand sanitizers, alcohol vs. alcohol-free, and kill rate of bacterial growth b) identify the relationship between the frequency and duration of sanitizer use and subsequent bacterial growth and c) to determine the relationship between the amount of friction used to apply rinse-free hand sanitizer and kill rate of bacterial growth.

Review of Literature

A. History

In 1929, Alexander Fleming discovered the first antibiotic, penicillin. Since that time, the discovery of medications used to fight various forms of bacteria has skyrocketed. “In the 1940s, the widespread availability of penicillin and the subsequent discovery of streptomycin led to a dramatic reduction in illness and death from infectious diseases” (Center for Disease Control and Prevention, 2005, p. 9) With the increased availability of antibiotic medications, especially penicillin, providers and consumers began to use penicillin unabashedly; at times, prescribing or taking the medication for unintended uses. Not long after, the bacteria began to show signs of resistance, either by the building of a stronger bacterial cell wall or through mutation. Between the years of 1946 and 1956, reported incidence of penicillin-resistant staph aureus in one hospital rose from 14% to 59% (History, n.d., p.5). In an attempt to control the resistant organisms being produced, scientists began to search for different forms of antibiotics. One way that this was done was to chemically modify the antibiotics already in use until an effective form was discovered. This type of research is responsible for producing many of the second, third, and fourth generation antibiotics that we utilize on a daily basis. Although, the resistant forms of bacteria have remained under control in the past, statistics have shown that the number of resistant organisms is continuing to rise at a frightening pace, placing healthcare workers in an even more precarious position.

B. Disease Prevention

The healthcare worker has a great responsibility in prevention of disease. Every day, healthcare workers come into contact with numerous resistant microorganisms. Microorganisms can easily be transmitted from patient to patient, if careful hand hygiene practices are not implemented. Hand washing has long been accepted as the primary means for prevention of infection. However, healthcare workers have either become too busy or lackadaisical in their adherence to their institution’s hand hygiene practices, thus, proliferating the growth and transmission of resistant microorganisms. Another factor that has increased the growth of microorganisms, is the continued misuse of antibiotic medications on the part of the provider, as well as, the consumer.

C. Rinse-Free Hand Sanitation

In order to decrease the transmission of resistant organisms between patients, many hospitals have adopted the use of alcohol free hand sanitizers. These hand santizers have proven to be even more effective than hand washing when used to clean hands that are not visibly soiled. However, according to a study published in the
AORN journal in 1998, continued use of an alcohol-based sanitizer produces a “decrease in antimicrobial effectiveness over time and is accompanied by swelling, erythema, a discomfort of the palmar surface of subjects’ hands” (Dyer, 1998, p. 249). This study was performed utilizing the “glove juice” method or hand sanitizer effectiveness protocol. In this protocol, each hand sanitizer is applied to the subject’s hands a total of ten times within a two-hour time frame. Although, this research study has provided useful information regarding the effects of rinse-free hand sanitizers on bacterial growth, it is hoped that even more information will be gained by examining the effectiveness of each rinse-free hand sanitizer over a longer period of time, specifically, a forty-eight hour period.

Conceptual Framework
The model chosen to guide this research study is the conceptual framework of Epidemiology. Epidemiology “is the study of how various states of health are distributed in the population and what environmental conditions, life-styles, or other circumstances are associated with the presence or absence of disease” (Epidemiology, 1986, p. 3). This particular model deals with seven specific categories.

1. Investigation of Disease Etiology and Determination of the Natural History of Disease
As stated in the literature review, the number of resistant organisms has grown due to factors such as: inadequate or improper use of antibiotic therapy and non-compliance with hand hygiene practices, resulting in patient to patient transmission within the hospital setting.

2. Identification of Risks
In this particular study the factors responsible for the increased risk of disease amongst people in the community are: poor hygiene practices, lack of knowledge regarding antibiotic therapy, and decreased health-promoting activities. More specifically, the risks associated with the hospitalized patient are magnified. The hospitalized patient usually presents with an illness that may predispose them to some type of infection (i.e.. cancer, cardiovascular disease, or diabetes). The hospital is a reservoir for bacteria, therefore, any patient entering this type of setting is at risk for infection. If the health care worker providing direct patient care to the already at-risk patient is non-compliant with the facility’s hand hygiene practices, then, the patient’s risk for infection is dramatically increased.

3. Identification of Syndromes and Classification of Disease
One of the primary goals of this research study is to evaluate the effectiveness of rinse-free, alcohol-free hand sanitizers so that they might be used to prevent the transmission of resistant bacteria, in particular. It is these resistant microbes or “super bugs” that have been known to result in increased severity of illness, as well as, patient mortality.

4. Differential Diagnosis and Planning Clinical Treatment
Research has shown that the increased transmission of resistant forms of bacteria is primarily due to poor hand hygiene practices, as a means of disease prevention. Therefore, in order for the transmission of these dangerous microbes to be halted, good hand hygiene practices must be implemented and reinforced. A goal of this study is to identify the benefits of an alcohol-free, rinse-free hand sanitizer in hopes that it will be used as an alternative to alcohol-based sanitizer.

5. Surveillance of the Health Status of Populations
The first signs of resistance to antibiotic medications were noted in 1945. Since then, the number of people infected with a resistant form of bacteria has continued to rise, primarily among hospitalized patients and people with compromised immune systems. Although resistant microbes are prevalent within these two specific populations, these microbes have shown that they are no respecter of persons. The incidence of bacterial infections continues to rise, regardless of socioeconomic status, health insurance, or age. Therefore, it is important that the issue of good hand hygiene practices is widely addressed and implemented.

6. Community Diagnosis and Planning of Health Services
Due to the fact that the majority of patient populations deal with the issue of bacterial resistance, the implementation of good hand hygiene practices should be addressed in the same magnitude. The Center for Disease Control (CDC), has recognized the need for such an initiative and is responsible for launching a “Public Health Action Plan to Combat Antimicrobial Resistance” (Center for Disease Control and Prevention, 2005, p.2). This document outlines the needs and goals required to halt the incidence of resistant antimicrobials. One of the action items listed states that a public health campaign should be conducted to “promote hand hygiene practices, as well as other behaviors that prevent the transmission of infectious organisms” (Center for Disease Control and Prevention, 2005, p. 26). It is hoped that the results of this research study provides the community with useful information regarding two different forms of rinse-free hand sanitation.

7. Evaluation of Health Services
A research study performed in 1998, testing the effects of alcohol-based and alcohol-free hand sanitizers on bacterial growth, showed that the repeated use of alcohol-based sanitizers resulted in minor skin irritation, as well as, an increase in bacterial growth. This information was collected after ten repeated uses of the alcohol-based sanitizer. One of the goals of this research study is to determine the effects of the same two types of hand sanitizers over an extended period of time, specifically forty eight or more hours, in order to further evaluate their effectiveness in prevention of infection.

Methodology Research Design
Each test subject will be examined for any pre-existing dermal irritation or skin problems prior to being enrolled in the study. At enrollment the subject will be assigned an alcohol gel treatment group or an alcohol-free treatment group. A Bernoulli distribution randomization chart will be created for the random assignments. Once assigned, the disinfectant (alcohol or alcohol-free) will remain the treatment group for the duration of the study.

Sample/Minimum Data Set
A subject pool sufficient to gain statistically significant data will be selected and used. It is expected pool will contain between 12 and 36 subjects. Subjects will be gathered from student volunteers at California State University of Fresno via convenience sampling. Eligibility criteria for this non-probability sample include factors such as

• intact skin integrity of the hands (no abrasions, wounds, etc.)
• transportation to and from study site
• at least 18 years of age and abov
• willingness to sign informed consent to participate in study

Exclusion criteria will mirror the eligibility criteria in most respects:

• no artificial nail
• no non-removable jewelry
• visible signs of hand trauma
• immunocompromised subjects or subjects in close contact with an immunocompromised individual (ie. persons with cystic fibrosis or cancer, newborn children, or the elderly
• inability to participate in complete study

This research study will take place in a laboratory at California State University, Fresno. The sample for this study will be obtained by the distribution of a flyer through the CSUF campus. General information regarding the purpose of the research study, eligibility and exclusion criteria, as well as, researcher contact information will be listed on the flyers. Respondents to the distributed information will be carefully screened for eligibility, with special consideration taken to exclude any vulnerable subjects.

Instruments
The instruments used in this research study include the bacteria Serratia marcescens, polyethylene gloves, tritpicase soy agar (TSA) mediums, and two hand sanitizers, alcohol-based and the benzalkonium chloride (alcohol free) hand sanitizer. The glove juice protocol will be used as a guideline for this research. Bacteria collected from the research subjects will be cultured overnight and read the next day.

Procedure

The procedure for this research study was adapted from the federally approved glove juice sampling method. However, a few alterations have been made to enhance the findings in this research study. The procedure is as follows (Dyer, 1998, p. 244):

Day One:

1. Cultures of Serratia marcescens are prepared to a concentration of approximately 108 bacteria per milliliter of inoculum, and effectiveness is established through a series of contamination and washing cycles.
2. Subjects will be asked to remove any adornments from hands. Subjects will, at the proper times, apply the indicated hand sanitizer, providing friction for either ten or twenty seconds.
3. A baseline is determined by inoculation of the subjects’ hands with five mL of Serratia marcescens for 45 seconds.
4. Allow hands to dry for two minutes.
5. Polyethylene glove containing 50 mL of collection fluid will be placed on the subjects’ hands and secured above the wrists with rubber bands.
6. Collection fluid is then spread over the subjects’ hand sand massaged for one minute in a standardized manner to ensure uniform recovery of the collection sample.
7. Collection samples from the hands are then pooled and immediately plated onto tripticase soy agar (TSA) media with both neat samples and serial dilution cultures to guarantee accurate colony counts.
8. After a baseline is obtained, the test subjects then proceed through a series of 10 wash cycles with the test solution with ten minutes passing between each wash cycle.
9. Glove juice samples will be taken after the first, third, seventh, and tenth contamination and wash cycles, as required by the US Food and Drug Administration
10. It is estimated that the entire procedure, from start to finish, will take approximately four to six hours on day one of the study

Data Analysis
Total counts after exposure to the hand disinfectant will be collected for comparison of the two treatment groups. Data will be analyzed in two different ways. Each of the two treatment groups will be separately analyzed to determine a confidence interval. This will determine if they each reach at least a three log reduction. At least a 3 log reduction must be achieved to meet the standard for an effective hand sanitizer.

In addition to the confidence level analysis, the alcohol gel will be compared to the alcohol-free product by matched pair test of significance. In this comparison, the T-test for significance will be used to determine the difference between the two treatment groups.

Human Subjects Protection
Prior to any action taken to begin this study, this research proposal will be submitted to the California State University, Fresno Institutional Review Board. Upon approval, a flyer, listing the purpose of the research study as well as eligibility criteria and researcher contact information will be distributed to students at California State University, Fresno. Students considering participation in the study will be carefully screened for any risk factors associated with contact with Serratia marcescens, as well as, any of the exclusion criteria previously mentioned. Information received concerning immune status or other risk factors will be kept confidential even if the potential subject decides not to participate in the study.

Data obtained from this study will be kept either in a locked file in the researcher’s home or on a computer requiring a password to access information. It is expected that the majority of information obtained from this study will be statistical in nature, however, protection of this information is a priority and will be adhered to.

RESULTS
Data gathered from this study are the plate count numbers of colony forming units of the test bacteria, Serratia marcescens. Reduced numbers if organisms after soap washed or by application of the test sanitizers are measured against the initial inoculums gathered from the unwashed or untreated hand in the baseline count. The differences are measured in log (base 10) reductions. Counts are made on each subject after baseline inoculation after a soap wash and after the 1st, 3rd,7th and 10th application of the sanitizer. Results of the counts, the calculations in log 10, and the log reductions are shown in the following tables and charts.

Results are tabulated in sixteen subjects with SafeHands applied to the contaminated skin. Average log reductions from the six separate subjects are compared to six separate subjects with Purell Alcohol Gel (62%) applied to contaminated hands in like manner.

To determine log reduction, colony forming units (line bacteria capable of forming colonies when plated on agar plates and counted by measured dilutions spread evenly over an agar plate made of nutrient broth which, like the dilution fluid, also contains neutralizing solution to stop any previous antimicrobial action. After 24 hours of incubation, plates are counted. Plate counts are multiplied by the appropriate dilution to get the actual number represented in the collection and that number is converted to log 10 for averaging and subtracting from the baseline to get the log reduction.

DISCUSSION
Log averages of colony counts (see table) show log reductions from application on SafeHands alcohol-free hand sanitizer to be approximately 3 log reduction – equal to that of hand washing by the end of the 10-wash cycle. 61% alcohol gel performed much less effectively throughout the wash cycle, but was significantly less than SafeHands (approx. 1.5 log reduction) by the end of the 10 wash cycle. Other similar studies (Dyer, 1998 ) have shown similarly poor responses of alcohol sanitizers by the end of the wash cycle, presumably due to delipidization and damage to the skin.

It is important to keep in mind that the control wash included a rinsing phase before collection (normal hand washing procedure), but the alcohol gel and the non-alcohol hand sanitizer washes did not include a rinse before collection, representing their normal application procedure in field use. The protocol calls for high contamination – much higher than most any field conditions and medical protocols, for example call for thorough hand wash any time the hands are visibly soiled. Even using high challenge numbers and been without rinsing. SafeHands performed as well as hand washing in this study.

SafeHands consistently out-performed the alcohol-gels, which have gained popularity and are commonly considered effective. The results of this study indicates that SafeHands is more effective than alcohol-gels, especially in situations calling for multiple applications.

SKIN DRYING RESULTS
Ninety participants were enrolled in the hand washing study and all 90 finished the study. Sixty of the participants were randomly selected to use the alcohol-free solution only. The other 30 used only the alcohol-based hand sanitizer. All 90 participants were students at California State University, Fresno.

The 60 subjects who used the alcohol-free hand sanitizer showed no visual skin breakdown before the start of the study. After a five cycle wash with the alcohol-free hand sanitizer, there were no visual changes to the subjects’ hands. In addition, there were no verbal ratings of the skin irritation to the hands by the participants. After a 10 cycle wash with the alcohol-free hand sanitizer and the final Chlorhexidine Gluconate (CHG) scrub, the hands were inspected again by the investigator. Of the 60 participants, one complained of a very mild itchiness over the knuckles, which was rates 1/3 itchiness. All 60 participants showed no visual signs of skin irritation to the tissue of the hands.

The 30 participants’ hands that used the alcohol-based product were inspected before the start pf the study and showed no visual skin breakdown. After a five cycle wash with the alcohol-based hand sanitizer, 2 of the 30 subjects presented with visible redness to the tops of the hands. Both subjects were rated 1/3 for mild redness to the knuckles. After a 10 cycle wash with the alcohol-based hand sanitizer and the final CHG scrub the hands were inspected again by the investigator. The same 2 participants showed more redness to the tops of the hands, knuckles, and fingers, and were rated 2/3 for moderate redness. The other participant complained of itchiness and dryness to the knuckles of the hands. It was rated 1/3 for mild itchiness to the knuckles, and 1/3 for the mild dryness to the knuckles.

DISCUSSION AND CONCLUSION
Hand sanitizers are becoming a more popular antiseptic treatment than the traditional use of soap and water. Their ability to provide instant protection by reducing bacteria on the hands has an impact on the healthcare industry. The literature suggests alcohol-based agents interfere with the skin’s natural barriers, thus causing dermal irritation. In addition, alcohol-based products possess flammability properties and may be absorbed into the skin. Alcohol-free hand sanitizers contain surfactants and emollients that prevent dryness and irritation to the skin and reduce the number of microorganisms.

This study evaluated two types of hand sanitizers, one containing an alcohol-based agent and the other an alcohol-free product. As suggested in previous research, the alcohol-based products presented more dermal effects on the tissue of the hands compared to the alcohol-free hand sanitizer. The most common visual skin change after repeated use with the alcohol-based hand sanitizer was redness to the top of the hands. There were no visual skin changes after repeated use with alcohol-free hand sanitizer.

In the hospital setting, hand washing with soap and water is recommended if hands are visibly soiled, otherwise instant hand sanitizers are an acceptable alternative. Alcohol-free hand sanitizers provide efficient antiseptic coverage without the irritations to the tissue of the skin. And with the high volume of patient contact, alcohol-free hand solutions allow healthcare workers to provide hygenic care to their patients. Reducing the rate of hospital-acquired infections and preventing the amount of occupational contact dermatitis are two ways alcohol-free hand sanitizers are making a name for themselves in healthcare.

In the healthcare environment, hand hygiene guidelines are established to decrease the spread of disease from person to person. Both alcohol-based hand sanitizers and alcohol-free hand sanitizers provide initial bacteria killing abilities, while the alcohol-free product offers more satisfying effects on the skin. Frequent use of alcohol-based agents for hand antisepsis can cause irritation and dryness to the skin. In the hospital setting, irritant contact dermatitis is an important concern for healthcare workers, which directly impact compliance with hand washing

Education is a key factor in improving patient compliance. Increased compliance and awareness are essential steps toward improving patient care in the health care industry. This research study focused on ways to decrease skin irritation on the hands by using alcohol-free hand products, rather than alcohol-based agents. This study serves to inform professionals about the irritating dermal effects alcohol-based hand sanitizers possess and the advantages of using alcohol-free hand products. The results from this study are offered to assist healthcare workers select an alternative hand wash which provides pleasing effects on the skin and optimal bacteria-killing abilities.

References

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Center for Disease Control and Prevention. (2005). A public health action plan to combat
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Dyer, D.L., Gerenraich, K.B., & Wadhams, P.S. (1998). Testing a new alcohol-free
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Epidemiology: What is it about. (1986). In Epidemiology in Nursing and Health Care
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