ISOLATION AND IDENTIFICATION OF BACTERIAL MICROBES

AMONG DOOR HANDLES, TOILET FLUSH KNOBS AND

AIR CONDITIONING SYSTEMS IN KDCI MAIN
CAMPUS KIDAPAWAN, NORTH
COTABATO, PHILIPPINES

_______________________

Thesis Proposal to the Faculty

of the Medical Laboratory Science Department of
KIDAPAWAN DOCTOR’S COLLEGE, INC.
Ninoy Aquino Road, Kidapawan City

_____________________

In Partial Fulfillment of the Requirements for

Degree of Bachelor in Medical Laboratory Science

By

Catuhao, Seth Wenefred E.

Gallego, Harold James L.
Crodua, Mel Grace O
Ortega, Ivy Grace R.

March 2018
 KIDAPAWAN DOCTORS COLLEGE INC.
Quezon Blvd., Kidapawan City, Province of Cotabato
Tel. No (064) 278-3830 Fax No. (064) 27– 4477
www.kdci.edu.ph

APPROVAL SHEET
In partial fulfillment of the requirements for the degree of Bachelor in Medical
Laboratory Science, this THESIS entitled:
ISOLATION AND IDENTIFICATION OF BACTERIAL MICROBES AMONG
DOOR HANDLES, TOIET FLUSH KNOBS, AND AIR CONDITIONING
SYSTEMS IN KDCI MAIN CAMPUS, KIDAPAWAN, NORTH COTABATO,
PHILIPPINES.
Has been prepared and submitted by Ortega, Ivy Grace, Catuhao, Seth Wenefred,
Crodua, Mel Grace, Gallego, Harold James, who are recommended for the
corresponding ORAL DEFENSE.
Ms. Mary Claire C. Dingal, RMT, MT (AMT), MLS (ASCPi)
Adviser

APPROVED in partial fulfillment of the requirements for the degree of Bachelor in

Medical Laboratory Science,
By the Oral Examination Committee:
PAULINO R. AGRAVE, Ma Ed
Chairman

________________________ ____________________
Member Member

ACCEPTED in partial fulfilment of the requirements for the degree of

Bachelor in Medical Laboratory Science

January 2018 BRYANT C. MARTINEZ, RMT, MD, FPSO-HNS

Date Dean

PAULINO R. AGRAVE, Ma Ed,

Head Research & Publication

SAMUEL H. AQUINO JR., MS, RP, CSIOP

Vice President for Academic Affair

 CHAPTER I

INTRODUCTION

Background of the Study

As stated by Costello et al., (2009) humans can spend the majority of their

time indoors, but little is known about the interactions between the human and

built-environment microbiomes or the forces that drive microbial community

assembly in the built environment. While indoors, we encounter bacterial microbes

on virtually every surface we touch, and this frequent exposure to indoor microbes

carries with it the potential for disease transmission, as well as interactions with

our own commensal microbiome.

As stated by Dr. Stephen Berger, (2017), in the Philippines Shigellosis is

also one of the leading cause of diseases that can be acquired via fecal-oral, water,

fly or fomites. In year 2007, there are 400 cases of infected and 4 deaths are being

reported in Bohol and also Camarines Sur there are 102 citizens infected.

Yet we have very little knowledge and understanding regarding the

ecological processes that drive microbial community assembly indoors, nor do we

understand the degree to which humans share microbes with indoor surfaces and

objects.

Door handles, Toilet flush knobs and air-conditioning systems are objects

which become contaminated with pathogenic bacteria and one of the so called
fomites. Fomites are Inanimate objects and surfaces that are known to play a role

as a reservoir for the transmission of pathogens in the environment either directly,

by surface-to-mouth contact, or indirectly, by contamination of fingers and

subsequent hand-to-mouth, hand-to-eye, hand-to-nose contact and even through

cuts in the skin (Nicas and Best, 2008).

Most outbreaks of infection associated with inanimate objects are caused

by items that should be sterile but have been contaminated. The hypothesis that

environmental microorganism cause human diseases arises from two facts, firstly,

our interaction with the inanimate environment is constant and close, secondly

environmental objects are usually contaminated often with important human

pathogens. Fomites have been implicated in the transmission of acquired

infections because they are in close proximity to the humans and are frequently

touched by hands (Kramer et al., 2006, Murphy et al., 2012, Sexton et al., Siani et

al., 2011, Weber et al., 2010, White et al., 2008).

Contamination of an inanimate object is only the first step in transmission

via fomites. As stated by Nicas and Sun (2006) another requirement is that the

etiological agent must remain viable on the fomite for a period sufficient for the

fomite to come into contact with a susceptible individual. If it is able to persist long

enough, the agent must then be able to transfer from the fomite to a point of

entrance on a susceptible individual. For respiratory and gastrointestinal diseases,

the point of entrance is most often a mucous membrane, such as through the

mouth, nose, ears, or eyes. Once transferred, the agent must be able to initiate

infection. In summary, the characteristics of the etiological agent, the fomite, the
infected individual, and the susceptible host, as well as the interactions between

the individuals and the fomite, influence efficacy of fomite- transmission which

leads to various illness.

The researchers are interested in this study because we notice that some

of the students are not responsible enough for their hand sanitation and proper

hygiene which can cause diseases. Fomites, like door knobs, toilet flush knobs, air

condition which are prone and one of the reasons why bacteria can be transmitted

by other people due to poor sanitation.

Statement of the problem

The aim of the present investigation is to isolate and identify the bacterial

microbes present among door handles, toilet flush knobs and air-conditioning

systems in KDCI main campus, Kidapawan, North Cotabato, Philippines.

Specifically, this study aimed to answer the following questions:

1. What are the bacteria present among untreated door handles, toilet

flush knobs and air-conditioning systems?

2. What is the bacterial load of bacterial microbes?

3. Is there a bacterium present among door handles, toilet flush knobs

and air-conditioning system in KDCI main campus, Kidapawan,

North Cotabato, Philippines?

4. What are the possible diseases caused by the isolated bacteria?

5. Which of the fomites contains most microbial load?

Hypothesis of the study

In this study, hypotheses were formulated to determine the difference

between the variable of the study.

1. Alternative Hypothesis: There is a significant difference between

untreated door handles, toilet flush knobs and air-conditioning systems.

2. Alternative Hypothesis: Fomites cannot be the cause of acquired

infections from transmitted diseases.

3. There is no bacterium present among door handles, toilet flush knobs

and air-conditioning systems in KDCI main campus, Kidapawan, North

Cotabato, Philippines

Theoretical Framework

As studied by Chen and Rusin et al. (2002), many elements have appeared

the impact of bacterial transfer between surfaces, including the bacterial species,

humidity levels, pressure and friction between the contact surfaces and inoculums

size on surfaces. Studies have demonstrated that fomites can be sullied with

microscopic organisms can be transmitted by hands and from hands to mouth.

According to Morey et al. (1986), as individuals interacted with surfaces as

work areas, consoles and office furniture, latrine bolt handles, there is probability

of getting microorganisms deposited on them. The point of air-conditioning system

is to give inhabitants a more agreeable condition. By such counterfeit conditions

might be favorable to parasites, microscopic organisms, protozoan and vermin,

which may develop health risk to clients, either by hypersensitivity or infection.

As indicated by the American Society of Heating, Refrigerating and Air

Conditioning Engineers (ASHARAE, 2000), the health effects caused by

microorganisms that are in indoor situations with aerating and cooling system can

be infective or immunological.

Barker and Jones. (2005), stated that contaminated hands of latrine users

can transmit bacterial microbes from their hands to the flushing knobs, entryway

handles and fixtures of the toilets and also household handles and hardware or

equipment. Latrine seats and covers, the encompassing floors, and the nearby

surfaces can be polluted by toilet flush aerosols which are created in generous

amounts amid flushing.

Conceptual framework

The Framework illustrated the determination of the difference between the

independent and dependent variable. The dependent variable pertained to the

bacterial microbes used in the study which was to be isolated and identified. While

the independent variable are the toilet flush knobs, door handles and air-

conditions.
 Independent Variable Dependent Variable

Bacterial Microbes
(Staphylococcus
species,
Streptococcus
species,
Enteroccocus
Toilet flush knobs, door species, and
handles and air-conditions. Enterobacteriaceae.
(Fomites)

Figure 1: Schematic Diagram of the relationships between the variables

Review of Related Literature

The KDCI is composed of 800 students, including faculty and staff. The

sanitation and cleaning maintenance procedure are being done by KDCI utilities

by collecting garbage, mopping the floor, cleaning comfort rooms but not usually

the entire toilet, also the air conditions are not being cleaned daily or weekly.

As mentioned by the Center’s Disease control and Prevention Shigella is

9
estimated to cause 80– 165 million cases of disease and 600,000 deaths annually
because only humans and higher primates carry Shigella, transmission occurs via

the fecal-oral route, including through direct person-to-person or indirectly through

contaminated food, water, or fomites. Recent studies in Australia and Canada

found that 40%– 50% of locally diagnosed shigellosis cases were associated with

international travel. In a study of travel-associated enteric infections diagnosed

after return to the United States, Shigella was the third most common bacterial

pathogen isolated by clinical laboratories. In this study, the risk of infection caused

by Shigella spp. was highest for people traveling to Africa, followed by Central

America, South America, and Asia. In 2014–2015, a large outbreak of

ciprofloxacin-resistant S. sonnei infections occurred in the United States after

travelers to India, Haiti, the Dominican Republic, and other countries returned with

shigellosis. Outbreaks of infections caused by multidrug resistant Shigella,

including isolates resistant to azithromycin or ciprofloxacin, have been reported in

Australia, Europe, Taiwan, Canada, and the United States among men who have

sex with men. Infections caused by Shiga toxin– producing S. flexneri and S.

dysenteriae have been reported repeatedly among travelers to Haiti and the

Dominican Republic.

According to Nwankwo E, 2012, The most common bacteria that will thrive

on fomites are Escherichia coli, Proteus mirabilis, Streptococcus spp., E. faecalis,

Staphylococcus aureus, Coagulase negative staphylococcal and Salmonella

choleraesius.

Fomite described as any non-living object or substance that is capable of

carrying infectious organism hence transmitting them from one individual to

another. There are several factors that can affect the contamination rate of fomites,

such as presence of moisture, frequency of use and general cleanliness.

(Reynolds KA, Watts P, Boone SA, Gerba CP. 2005). One common way by which

organisms that are not resident in the hand are picked up is by contact with

surfaces such as table tops, hospital door handles, toilet handles and taps in the

restrooms as stated by K. R. Bright, S. A. Boone and C. P. Gerba. 2010.

Human skin is an essential wellspring of microscopic organisms in indoor

conditions, and people can transmit bacterial pathogens by touching indoor

spaces. Looking at different parts of the human micro biome with microbial groups

in indoor conditions can recognize bacterial species generally found in the two

situations and along these lines propose the wellspring of pollution or transmission

according to Flores GE, Bates ST, Knights D., et al.2011.

Toilet flush Knob

Bacteria are microscopic organism found everywhere in the Universe as

pathogenic or non-pathogenic. They are found in the environment all around us

and within each one of us, there are trillions and trillions of them. Majority of them

are harmless to human and animals but those few which are harmful can lead to

death of affected individuals (Hooper 2001; Short et al. 2007). Public restrooms

may contain a variety of dangerous bacteria, including from genus Escherichia,

Salmonella, rotavirus, cold virus and Staphylococcus including Methicillin-resistant

Staphylococcus aureus (MRSA) and Streptococcus (Hooper 2001; Peleg and

Hooper 2010; Hooper et al. 2010; Flores et al. 2011; Adewoyin et al. 2013).
 According to Bright Kr, 2005 they get in the restrooms via human excreta.

Improper use of the toilets, inadequate cleanliness of the toilets facilitates can

transmit bacteria from the toilets to the household living rooms. Contaminated

hands of toilet users can transmit the bacteria from their hands.

During Hajj, millions of Muslim pilgrims gather in the holy sites (Arafat,

Muzdalifah and Mina) in Makkah. Although there are plenty public toilet facilities in

the holy sites, these toilets are extremely crowded. If not regularly decontaminated,

however, residual organisms from an infected person can survive in significant

populations. Toilets can provide an ideal environment for spread of pathogens from

gut, respiratory tract and skin via hands and surfaces from one person toss another

(Gerhardts et al., 2012). Toilet handles contamination is one of the common ways

by which organisms that are not resident in the hand are picked up by contact with

surfaces. Due to the unhygienic use of the toilet facilities, fecal matter remains a

major reservoir source of human pathogens, which in adverse situation may bring

about outbreaks of infection (Maori et al., 2013). The pressence of the pathogenic

bacteria on door knobs poses a potential risk to the pilgrims. It has been shown

that hard, non-porous surfaces, such as door handles, have the highest bacterial

transfer rates to hands (Rusin et al., 2002).

Sabra (2013) carried out a study on public female restrooms at Taif,

Kingdom of Saudi Arabia, in order to determine the locality of contamination and

bacterial loads. She found that out of 260 specimens investigated 187(71.9%)

were positive growth. The predominant positive was toilet handles (91.3%),

followed by room door handles 59(73.8%), and room sink 38(63.3%). Isolated
bacteria arranged according their percentage, Staphylococcus. aureus 40.6%,

Escherichia. Coli 22.5%, Bacillus spp. and Klebsiella. pneumoniae 21.4%,

Enterococcus. faecalis 13.4%, Citrobacter spp. 9.6%, Pseudomonas. aeruginosa

8.6% and Proteus. mirabilis 7%.

Door Handles

Fomites include among others door handles or knobs of conveniences,

showers, toilet seats and faucets, sinks, lockers, chairs, and tables, and these are

especially found in public places, hospitals, hotels, restaurants and restrooms.

However, the most common source of fomites infections in most of the hospitals

are door handles or knobs of toilets and bathroom. It is usually believed that

fomites are deposited on door handles/knobs while large traffic of users who throng

in with their own microbial flora and other organisms have picked them elsewhere

while going into the convenience on their way out toilets and bathrooms. In

general, the risk of disease transmission through fomites is determined by the

frequency of site contamination and exposure; level of pathogen excreted by the

host; likelihood of transfer of the infectious agent to a susceptible individual;

virulence of the organism; immuno-competence of the persons in contact; the

practice of control measure such as disinfectant use and personal hygiene.

Consequently, people who can hardly wash their hands after toilets and bathrooms

usage, could result in acquiring community-associated methicillin

resistant Staphylococcus aureus (CA-MRSA) and this can result in outbreak

especially in high prevalence areas. Furthermore, Bright KR, Boone SA, Gerba

CP, et al.,2010 stated that fomites can also serve as bacterial and viral reservoir
through aerosolization and direct transfer from hand to fomite surface. Doors have

large traffic users, who throng in with their own microbial flora and other organisms

they have picked elsewhere and deposit them on door handles while going in and

out. (M. A. Gold hammer, D. P. Dooley, E. W. Ayala, and B. L. Hill. (2006). Those

surfaces in the vicinity of patients termed “high touch surfaces” could serve as high

risk factors for microbial reservoir and route for pathogen transfer. As mentioned

by G. Pellizzer, P. Mantoan and L. Timillero, (2008) Considering the potential risk

of door handles might pose to users when contaminated with pathogens The

spread of infectious diseases through hand contact has been an area of major

concern.

Rusin (2002) stated that the presence of these pathogenic bacteria on

environmental surfaces such as door handles poses a potential risk to vulnerable,

immune-compromised individuals. It has been shown that hard, non-porous

surfaces, such as door handles, have the highest bacterial transfer rates to hands.

Specifically, sought to generate data relating to microbial contamination on

door handles and how this might be related to factors relating to their design and

use. Selected three high acuity environments for study as these are known to act

as hotspots for HCAI transmission. (NHS Scotland National HAI Prevalence

Survey NHS Scotland ,2007).

As studied by Bashir,S. and F, Muhammad, H et.,al.(2016) Staphylococcus

aureus, Bacillus species, Micrococcus species, Escherichia coli, Salmonella

species and Klebsiella species were the primary microscopic organism isolated in

their research work so far. The results of their present study exhibit that public
contact surfaces such as door handles are frequently colonized by several bacteria

and serve as a potential source of infections. Contaminated and improperly

washed hands contaminate door handles, that is to say there is a high level of

bacterial contamination which may lead to high level prevalence of the bacterial

infectious disease due to contaminants. And also isolation of pathogenic bacteria

from the toilets door handles in their study indicates that they could be vehicle of

disease transmission as microbial contamination of toilet door handles surfaces

may be a common means of transfer of potentially pathogenic bacteria among

users.

As stated by Hedieh et al. 2012 led an investigation if there is presence of

microbial contamination of door handles in two busy intensive care units and one

high dependency unit was related to their design, location, and usage. They found

a noteworthy relationship between the frequency of movement through a door and

the degree to which it was contaminated. Furthermore, the doors location, design

and method of utilization all influenced contamination. When compared to push

plate designs, pull handles uncovered by average of a fivefold higher level of

contamination; lever handles, however, showed the highest levels of bacterial

contamination when adjusted for frequency of use.

As studied by Nwori et al. (2012) investigated 180 door handles/knobs of

public conveniences of selected public offices, motor parks and markets in Abuja

Metropolis, Nigeria. They found that 156 (86.7%) were positive for bacterial growth.

The bacteria were Staphylococcus. aureus (30.1%), Klebsiella . pneumonia

(25.7%), Escherichia coli (15.6%), Enterobacter species (11.2%), Citrobacter

species (7.1%), Pseudomonas aeruginosa (5.9%) while Proteus species had the

least prevalence,(4.5%).

Air conditioning system

As stated by Nester et al., 1996 air – conditioning system are appliances

that controls temperature humidity and purity at all times regardless of the

climate conditions. It has a blower equipped with refrigerating unit that cools the

air and filtered. Han et al., 1998 Brown and wake, 2000 mentioned that air -

conditions modifies indoor air quality and are installed in homes, offices, vehicles

and industries for various reasons including good operational output of

equipment and comfort.

Abdel HAA, Farag SA (1999) stated that indoor air quality has turned into a

vital public health concern as many people spend over 90% of their time in inside

like houses, workplaces and schools. Air in the indoor condition can be polluted by

various pollutant among which airborne microorganism (bacteria and fungi) are

most important.

Air Conditioning South East (2013) mentioned the developmental process

of utilizing ventilation systems by humans in houses, classrooms, and different

places however brought about the assistance of facilitating life for people amid

times when climate is harsh, but, their utilization tragically acquired unfortunate

impacts that they turned into a decent source and additionally media for bacterial

or fungal/mold contamination.
 As indicated by Aydogdu, H., A. Asan, Otkun, M.T. (2008) improperly

maintained air conditions may introduce these microbial pieces and metabolites

can be a wellspring of wide variety of diseases going from hypersensitivity side

effects to risky conditions, for example, respiratory, dermatological infections,

epidemics and food pollution.

Adnan A. (2013) stated that individuals with poor health or immune

compromised patients are more serious hazard to these microorganisms that could

be present in this environment.

M.N. Modebelu and A. F. Modebelu (2013) in their study revealed that four

bacterial species were predominantly and frequently isolated from various

ventilation devices, which they were examined in their study. Those ventilation

devices include air-conditioners, electric fans and open doors/windows are used

in various establishments as a means of control and enhancement of air quality.,

Bacterial microbes that they have been isolates which include Micrococcus luteus,

Arthrobacter globiformis both of which belong to the family of actinomycetes,

Staphylococcus aureus and Staphylococcus epidemidis were discovered as

serious contaminants of the various ventilation devices. The isolates co-exist as

habitants of the ventilation devices utilizing available nutrients circulating in

environmental aerosols for survival.

As indicated by Gide Anas, David Sunday Aligbe et., al. (2016). Lint and

liquid drops are the important medium for microorganisms to spread. It is important

to clean the parts that are easy to be infected in air-conditioning systems (e.g.,

filter, heat exchanger and muffler) and to replace them in time in order to avoid the
aggradations of pollutants. In addition, the condensing water ought to be

eliminated in time in air-conditioning systems to prevent bacteria from propagating

and spreading. Filtration is a very practical and effective strategy of improving air-

conditioning system, the air filtration systems represent a good solution for the

improvement of Indoor Air Quality (IAQ), and the Antimicrobial treatments of filters

might be an answer for these issues. It is possible to avoid the aggregation and

distribution of microorganisms by adding anti-microbial agents on the surfaces of

filter, which add to the improvement of air quality.

Meadow et al. (2014) found that microbial communities in indoor air were

significantly influenced by ventilation and occupancy. Although community

structure in indoor air was closely associated with that of outdoor air, human-

associated bacteria were over two times more abundant in an occupied indoor

environment. Bouillard et al. found that Micrococcus species., Staphylococcus

spp., and Streptococcaceae species. were the most common species found in the

air of a healthy office building. These bacteria are representative of the normal

human flora, providing further evidence that human occupancy shapes the

bacterial communities in indoor air to some degree. Kloos and Musselwhite

showed that Staphylococcus spp., Micrococcus spp., Acinetobacter spp., Bacillus

spp., and Streptomyces spp. are part of the normal human skin flora. Charlson et

al. found high relative abundances of Staphylococcaceae species.,

Propionibacteriaceae species., Corynebacteriaceae species., Streptococcaceae

species., Veillonellaceae species., Prevotellaceae species., Fusobacteriaceae

species., and Neisseriaceae species.

 Recent epidemiological studies have reported fomites in the transmission

of human pathogens within high-exposure environments such as hospitals, child-

care facilities, long-term care facilities, and sports facilities. Staphylococcus

epidermidis (34%), followed by Enterococcus faecalis (26%), Staphylococcus

aureus (14%), Escherichia coli (10%). Streptococcus pyogenes and

Pseudomonas alkaligenes (4%). Streptococcus agalactiae, Klebsiella oxytoca,

Klebsiella pneumonia and Pseudomonas aeruginosa (2%). Eighty percent of all

swab samples cultured were gram positive ad 20% were gram negative as stated

by Eickhoff TC (1994).

Scope and Limitation

This study will be focusing on the isolation and identification of bacterial

microbes (gram negative), if it is present among door handles, toilet flush knobs

and air-conditions specifically at the adjustment louver. The method that the

researchers will use in identifying and isolating bacterial microbes through bacterial

culture, gram- staining, and biochemical test. Bacterial microbes like Proteus

species doesn’t need an additional test to determine the kind of species. The

experiment will be conducted at Kidapawan Doctors College Inc. Thesis laboratory

and at Kidapawan Doctors Hospital laboratory.

Significance of the Study

As stated by Bustillo (personal communication ,2017), our hands act as a

bridge for the transmission of certain bacterium that grows, because we use our

hands in performing things like urinating and excreting, wiping our anus with the

use of tissue paper or water, and using it for flushing at the same time. Comfort

room flush handles and door knobs will be contaminated through the hands of the

user then it can be transmitted by other students/faculty and staff.

Classrooms and Faculty offices have installed air-conditioning system

which they use to cool themselves from the hot weather. When air conditioners are

not cleaned thoroughly and filters changed, a breeding ground for all kinds of

bacteria is created. These systems especially can be home to black mold, as

moisture can build up in the adjustment louvers and ducts from condensation that

forms when the cool air passes through. When these microorganisms go air-borne,

they can lead to a multitude of breathing problems, including a potentially fatal

infectious pneumonia.

School administration of Kidapawan Doctors College Incorporated must be

informed of the results from the research that the researchers will be conducting

about the Isolation and Identification of bacterial microbes from comfort rooms

specially in toilet flush knobs and air-conditioning systems in KDCI Main Campus

that carries infectious organisms which can occur especially when there is no

proper hygiene.
 The importance of our study is to inform the administration of the school to

make an action to the specified concern. This is not only for the students but also

for everyone.

Therefore, the researchers are increasingly turning their attention for

sterility check of the school because prevention is better than cure.

Operational Definition of terms

Air-condition system – An apparatus that controls the temperature and

humidity of an enclose system.

Bacteria – Referred as some eubacteria that is a single – celled organisms lack

true nucleus and multiply through binary fission.

Biochemical test - Test such as oxidase test, which uses an artificial electron

acceptor, and can be used to distinguish organisms on the bases of presence or

absence of respiratory enzymes.

Door handle- A door handle is an attached object or mechanism used to manually

open or close a door. These door handles are being one of the fomites that will be

our independent variable. Door handles on main campus.

Fomites - Fomite described as any non-living object or substance that is capable

of carrying infectious organism hence transmitting them from one individual to

another. There are several factors that can affect the contamination rate of fomites,
such as presence of moisture, frequency of use and general cleanliness. Those

are door handles, toilet flush knobs and air-condition systems.

Gram- stain - It is a method of staining of microorganism by the use of violet stain

then iodide solution, decolorizing alcohol or acetone solution, and counter staining

with safranin. The changing of the violet color stain or pink color of the counter

stain serves as a primary means of identifying and classifying bacteria. This will be

use as one of our steps in actual data gathering.

Toilet flush knob - A flush toilet (also known as a flushing toilet, flush

lavatory or water closet (WC)) is a toilet that disposes of human excreta (urine and

feces) by using water to flush it through a drainpipe to another location for disposal,

thus maintaining a separation between humans and their excreta. Flush toilets can

be designed for sitting (in which case they are also called "Western" toilets) or for

squatting, in the case of squat toilets. The opposite of a flush toilet is a dry toilet,

which uses no water for flushing. The toilet flush knobs on KDCI Main Campus,

being used by students, faculty and staffs.

Bacterial culture – Growth of microorganisms on a culture medium, such as

nutrient agar and use it for identification.

 CHAPTER II

METHODOLOGY

This chapter contains the research design, locale of the study focused on the
ideas, methods, and procedures of the research.

Research Design

Complex factorial research design will be used in this study. The research

aims to isolate and identify the kinds of bacterial microbes present among the toilet

flush knobs, door handles and air-conditioning systems. The microbes will be

identified through their characteristics and morphological features.

Locale of the Study

The researchers will be conducting their study at the Thesis Laboratory in

Kidapawan Doctors College Incorporated, Quezon Boulevard, Kidapawan City.

The isolation and identification of the bacterial microbes will be conducted at

Kidapawan Doctors College Incorporated, Quezon Boulevard, Kidapawan City by

the researchers. The swab samples will be collected by the researchers

themselves. The researchers will conduct their Bacterial Culture, gram-staining,

microscopy and biochemical test in Kidapawan Doctors College Inc.

Data Gathering Procedure

The following will be the step to be done on the actual gathering of data.

Cotton Swab

Collect swabs from door handles, toilet flush knobs and air-conditions and

place in trypticase soy broth in a test tube sealed with a cotton buds, then incubate

for 24 hours.

Check the turbidity

After incubation check if it turns turbid or clear, if it is turbid inoculate

the sample in the MacConkey agar and if it is clear don’t proceed for

inoculation process.5

Identifying Lactose, non-lactose and late lactose fermenter

After 24 hours of incubation in the MacConkey agar, identify the color

of growth. If the color is pink it’s a lactose fermenter, if the color is light pink it’s

a late lactose fermenter then if colorless it’s non-lactose fermenter.

Gram Staining and Biochemical test

In biochemical test, the use of nutrient broth like Triple Sugar Iron, Lysine

Iron Agar, Citrate, Spot Indole Motility, Urease and reagents that can identify the

microorganisms, that produce enzymes or fermentation occurs which lead to

changes of the nutrient broth. First dry the material on the slide so that it does not

wash off during the staining procedure. Adherence can be improved by the fixation

in 70 percent to 95 percent alcohol or by gently warming the slide to remove all

water from the material. Place the smear on a staining rack, and overlay the

surface of the material to be stained. Place the smear in an upright position in a

staining rack, allowing the excess water to drain off and the smear to dry. Never

blot a critical smear. Never put immersion oil on a smear until it is completely dry.

Examine the stained smear using the low power objective, and select an area to

examine more closely using a 40 to 60 oil objective. Suspicious areas are

evaluated using the 100 oil objective of the microscope.

References

1. Anas, David Sunday Aligbe et., al. Studies on Microorganisms Associated

with Air-Conditioned Environments. (2016)

2. Adnan A, Bacteria is the most useful microorganism in the environment and

beneficial for human beings. But it also has some

harmfuleffectsonhumanbody,biotecharticles.com.http://www.biotecharticle

s.com /Biology-Article/Beneficial-and-Harmful-Bacteria-312.html. (2013)

3. Abdel HAA, Farag SA, An indoor bio-contaminants air quality. Int J

Environ Health Res 9: 313. (1999)

4. Bashir,S. and F, Muhammad, H et.,al Isolation and Identification of

Bacterial Contaminants from Door Handles of Public Toilets in Federal

University Dutse, Jigawa State Nigeria (2016)

5. Barker and Jones,” The potential spread of infection caused by aerosol

contamination of surfaces after flushing a domestic toilet,” 2005.

6. Biochemical markers on public surfaces. (2005)

7. Bright KR, Boone SA, Gerba CP. “Occurrence of bacteria on elementary

classroom surfaces and the potential role of Elementary Classroom

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 Fomites

Swab Samples

Inoculate on TSB

Check for Turbidity

Turbid Clear

Inoculation Stop

MacConKey agar

Growth Colony Count

Gram Stain Microscopy

Biochemical Test