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EXPERIMENTAL ANALYSIS OF POLYHERBAL FORMULATED FUMAGATION

PRACTICES OVER THE GROWTH OF STAPHYLOCOCCUS AUREUS

Article · May 2020

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Mukt Shabd Journal ISSN NO : 2347-3150

EXPERIMENTAL ANALYSIS OF POLYHERBAL FORMULATED

FUMAGATION PRACTICES OVER THE GROWTH OF
STAPHYLOCOCCUS AUREUS
M. Babu*, Manikandan. M1, Yogashree. G1, Rekha. D1 Ashok.K*
**1
Department of Microbiology and Biotechnology,
Faculty of Arts and Science,
Bharath Institute of Higher Education and Research, Chennai,Tamil Nadu, India.

ABSTRACT

In the present study, environmental disinfection efficiency of polyherbal fumigation practice

has been evaluated by using natural plant products such as neem leaves, basil leaves,
peppermint leaves, eucalyptus leaves, dried garlic and ginger powder, cloves powder, chinese
chaste tree leaves and a few drops of lavender oil. The efficiency of poly herbal fumigation
using these natural products to disinfect air and surface was evaluated. In addition, the
ability of the fumigation using polyherbal to disinfect inert surface was evaluated using
Staphylococcus aureus. Polyherbal fumigation with 60 minutes exposure reduced more
bacterial colonies than the control and 20, 30 and 45 minutes exposures by cfu. The result of
the study strongly suggests that 60 minutes exposure not only reduced the bacterial colonies
but also showed more DNA damage. If future these kind of polyherbal fumigation is wanted
to control many dread full diseases than the usage of chemical fumigating substances.

Keyword: polyherbal fumigation, Staphylococcus aureus and COMET assay.

INTRODUCTION

Health-related infections are a global problem that affects 15% of hospitalized patients
worldwide [1-5]. Hospital-acquired infections (HAI) results in increased antibiotic resistance,
finical burden, morbidity and mortality [2]. Transmission usually occurs through health
workers, patients, hospital equipment, or interventional procedures. The most common areas of
infection are the blood stream, urinary tract and surgical trauma. Although any bacterium causes
HAI, multidrug-resistant (MRD) pathogens cause HAI. This increase can be partly explained by
the indiscriminate use of antibiotics and lack of hygiene measures, especially among medical
personnel [6-11]. The most commonly found MRD pathogens are those shown in Table 1.

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Table 1: Common causative microorganisms of HAI

Type of infections Microorganisms

Blood borne infections Hepatitis B virus (HBV) and hepatitis C virus (HCV) and HIV
Urinary tract infection (UTI) Staphylococcus saprophyticus, Klebsiella species, Enterococcus
faecalis, Enterobacteriaceae and yeast
Surgical site infections Staphylococcus aureus, Escherichia coli, Pseudomonas
aeruginosa, Citrobacter freundii, Citrobacter koseri,
Acinetobacter baumannii, Acinetobacter lowfii, Klebsiella
pneumonia, Klebsiella oxytoca, Proteus mirabilis, Proteus
vulgaris, Morganella species and Bacteroides fragilis,
Propionibacterium acnes.
Ventilator associated infections Pseudomonas aeruginosa, Staphylococcus aureus, Acinetobacter
species, Serratia marcescens, Stenotrophomonas maltophilia,
Burkholderia cepacia and Klebsiella pneumonia.
Air borne infections Mycobacterium tuberculosis, Varicella zoster virus, A(H1N1),
A(H3N2), SARS and carnivorous.
Water borne infections Legionella pneumophila, Acanthamoeba species, Hartmannela
species, Burkholderia cepacia, Cyclospora cayatenensis,
Cryptosporidium parvum and Naegleria species.

Modern hospitals and operation theatres must have a proper design of the ventilation system
with special functions such as air handling and airflow direction, including> 2.5 Pa and a
HEPA-filter if room air is recirculated [12]. The operating and maintenance costs of these
ventilation systems are very high. Thus, underdeveloped and developing countries do not have
as expensive technologies and common conventional chemical disinfection as those used in the
healthcare environment, which is associated with toxic side effects. Therefore, there is a need to
investigate new and unconventional methods of disinfecting the health care environment.
Disadvantages of chemical disinfectants are the development of drug resistance, toxicity and
causes hypersensitivity. Therefore, natural polyherbal incense today plays a major role as a
disinfectant because it is environmentally friendly and inexpensive.

In India, traditional fumigation practices using neem leaves, basil leaves, peppermint
leaves, eucalyptus leaves, dried garlic and ginger powder, cloves powder, chinese chaste tree

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leaves and a few drops of lavender oil provides many health benefits such as antirepellent,
antimicrobial, anticancer and anti-inflammatory activity [13-17]. In view of the above, we
decided to verify the efficacy of the poly herbal fumigation method using these plant products
to improve the microbiological quality of air and disinfect the surface to reveal its potential
benefits in chemical smokers.

MATERIALS AND METHODS

To evaluate the disinfection efficiency of poly herbal fumigation, we exposed

Staphylococcus aureus streaked pert plates to poly herbal fumigation (0, 20, 30, 45 and 60
minutes) in a closed box of 5 "X 3’ sizes and the non-fumigated perti plate are considered as
control. The fumigation exposed petriplates were serially diluted and then the culture was
spread on the nutrient agar plates. Further the plates were incubated at 37⁰C for 24 hrs and after
the incubation the bacterial colonies were counted to identify the survival of the cells by cfu.
Moreover, the fumigated different time intervals of bacterial cells were quantified using
COMET assay to identify the qualitative DNA damage to the cells. The qualitative DNA
damage to the cells was done according to the method of Singh et al. (1988) [18].

RESULTS AND DISCUSSION

Polyherbal fumigated plates showed decrease in colonies number when compared to

non-fumigated control plate (Fig. 1A). Fig. 1E showed more decrease in colonies than the fig.
1B, C and D. The result shows that maximum decrease in bacterial colony was observed only in
the 60 minutes exposure of Polyherbal fumigation as shown in the table 2. The present cfu study
has shown that regular polyherbal fumigation in the hospital environment or in the houses may
help to reduce the virulence of aerobic bacteria such as Staphylococcus aureus.

Table2: Effect of polyherbal fumigation on microbes present in air evaluated by cfu.

Treatment Bacterial colony cfu/m3

Non-fumigated pert plate (Control) 80±4.949
20 minutes 35±4.242
30 minutes 23.7±14..990
45 minutes 4.8±0.141
60 minutes 1.08 ±0.0282

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Values are mean + SD of five individual observations

Fig. 1: CFU of polyherbal fumigated plates A- Control non-fumigated plate, B- plates

exposed to 20 minutes fumigation, C- plates exposed to 30 minutes fumigation, D- plates
exposed to 45 minutes fumigation and E-plates exposed to 60 minutes fumigation

The application of polyherbal fumigation involves large samples of medicinal plants of

traditional Indian flora that may find real application in controlling dread full microbes present
in air, as they work as effective weapons against the air borne infections such as
Mycobacterium tuberculosis, Varicella zoster virus, A(H1N1), A(H3N2), SARS and
carnivorous in future. Further, long-term studies in virto and in vivo involving more subjects are
needed to clarify if this approach could represent a cost effective strategy in controlling the
microbes present in air. The advantage of using polyherbal fumigation not only kills the
microbial load present in but it also cures health disorders of human being by inhaling or
exposing to it [19]. Neem (Azadirachta indica) is used as fumigating substances against house
hold, storage and crop pests and exposing the neem leaves fumes in the house helps to get rid of

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microbes due to its high antimicrobial, antimalarial, anti-inflammatory, febrifuge, vermifuge,

antiseptic, anticancer and antihelminthic property [20-28]. Benefits of basil (Ocimum sanctum)
can be attributed to its internal home care and its ability to assist in protecting the body from
toxic-inducing damage. These functions are often attributed to its high polyphenolic compounds
and it possess various biomedical properties such as antimicrobial, anticancer, antioxidant, anti-
larvicidal, anti-inflammatory, anti-allergic, anti-stress and anti-tussive [29-32].

Inhalation of peppermint (Mentha piperita) leaves fumigation gives relief from cold,
sinusitis and bronchitis [33]; Fumigation of Eucalyptus (Eucalyptus obliqua) leaves give an
aroma odour which solves the problems of flu and respiratory disorders [34-38]. Ginger
(Zingiber officinale), garlic (Allium sativum) and clove (Syzygium aromaticum) powder
fumigation gives relief from antiflu drug, Tamiflu and bird flu H5N1 [39] and it also possess
biomedical property such as antimicrobial, antioxidant and anticancer [40-42]. Vitex negungo
leaves are effective used as a larvicidal and also used as an antimicrobial, anticancer and
antioxidant [43]. Lavender oil is highly used for pest control and antimicrobial against
Streptococcus sobrinus, Porphyromonas gingivalis, Streptococcus mutans, Staphylococcus
epidermides, Candida albicans, Lactobacillus casei and Bursera aloexylon [44].

The cost of isopropyl alchol, formaldehyde, glutaraldehyde, chlorohexxidine,

digluconate, hydrogen peroxide, ethanol, methyl bromide, phosphine and benzalkonium
chloride fumigation, which are expensive, toxic and process various side effects such as
irritating to skin, eyes and respiratory tract. Whereas by using poly herbal fumigation method
does not produce any toxic effect to human being and the preparation is very easier and cheaper.

Poly herbal fumigation produces lethal effects in Staphylococcus aureus bacterial cells when
exposed to 60 minutes than the 20, 30 and 45 minutes by the qualitative DNA damage assay. 60
minutes exposure bacterial cells showed more DNA damage in terms of formation of tail and
head in the cells, while no such changes were observed in non-fumigated bacterial cells
(control). Moreover, it was evidently observed that DNA damage was incubation time-
dependent. DNA break was observed in the formation of tail formation in 20, 30, 45 and 60
minutes exposure to poly herbal fumigation as shown in the figure 2. Similar observation was
observed by Bhatwalkar et al. (2019) [45] and they reported that SEM analysis showed reduced
number of bacteria in garlic peel fumigated surface samples. This is the first report to discuss
about the DNA damage induced by the polyherbal fumigation and in future more research is
needed to identify the mechanism involved in controlling the growth of bacterial colonies.

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Fig. 2: Qualitative DNA damage of polyherbal fumigated plates A- Control non-fumigated

plate, B- plates exposed to 20 minutes fumigation, C- plates exposed to 30 minutes
fumigation, D- plates exposed to 45 minutes fumigation and E-plates exposed to 60
minutes fumigation

CONCLUSION

The study found that neem leaves, basil leaves, peppermint leaves, eucalyptus leaves,
dried garlic and ginger powder, cloves powder, chinese chaste tree leaves and a few drops of
lavender oil reduce aerobic bacteria. In addition, polyherbals are effective in disinfecting
bacterial resistance which is one of the main factors responsible for hospital-acquired infections.
In addition, they have the potential to be used as an alternative to conventional fumes harmful
and toxic in the disinfection in the epidemic situations.

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