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Egypt. Acad. J. biolog. Sci., 3(1): 75- 81(2011) G. Microbiology
Email: egyptianacademic@yahoo.com ISSN: 2090-0872
Received: 20/11/2011 www.eajbs.eg.net
Antibacterial Effect of some Mineral Clays In Vitro
Shehab A. Lafi1 and Mohammed R. Al-Dulaimy2
1- Microbiology Department ,College of Medicine / Anbar University West of IRAQ
2- Microbiology Department , College of Dentistry, Anbar University.
Email Shehab_58@yahoo.com
ABSTRACT
Background:- The use of geological mineral clay to heal bacterial infections has
been evident, since the earliest recorded history, and specific clay minerals may prove
valuable in the treatment of bacterial diseases, including infections for which there are
no effective antibiotics, such as multi-drug resistant infections.
Objectives:- The aim of this study is to identify new inhibitory agents in an era when
bacterial antibiotic resistance continues to challenge human health and the availability
of new antimicrobial compounds is limited.
Materials and methods:- Two bacterial isolates were taken from patient with
different infections (UTI, skin infection). The two isolates were used to study the
antibacterial effect of four types of mineral clays (gray clay, yellow clay, white clay,
and pink clay).
Result:- Three types of clays (white clay, gray clay, and yellow clay,) have an
antibacterial effect against Staphylococcus aureus that was isolated from skin
infection. And have no effect against Pseudomonas aeruginosa that was isolated from
patient with UTI. On the other hand, pink mineral clay explained an antibacterial
effect against Pseudomonas aeruginosa, and has low effect against Staphylococcus
aureus.
Conclusions:- Our results indicated that mineral clay could provide an alternative
treatment against numerous human bacterial infections.
Key words: Mineral clay, Infections, Antibacterial agents.
INTRODUCTION
The intentional consumption of
earth materials, such as clays, by humans
and animals is known as geophagy, a
complex behavior, largely attributed to
religious beliefs, cultural practices,
psychological disorders, cosmetics,
dietary/nutritional needs, and medicinal
benefits Wiley, Andrea S. (2006). The
oldest evidence of geophagy practiced by
humans comes from the prehistoric site
Kalambo Fallo on the border between
Zambia and Tanzania, where a calcium-
rich white clay, believed to have been
used for healing geophagical purposes
Root-Bernstein, R. S. et al. (2000).
Throughout history a large number of
writers and physicians have commented
on geophagy. In the first century AD,
physicians mention a famous
medicament Known as terra sigillata
(earth that has been stamped with a seal)
otherwise known as Lemnian Earth.
Lemnian Earth was used for many
maladies (but most notably for
poisoning), and so great was the demand
from the 13th to the 14th centuries, that
almost every country in Europe strove to
find within its boundaries a source of
supply, Thompson , C.J.S. et al. (1913).
More recently, the well known
German naturopaths, Kneipp, Kuhn, Just,
Felke, and others of the last century have
contributed to clay's revival in natural
treatments. Although geophagy can be
found relatively easily in many
Shehab A. Lafi and Mohammed R. Al-Dulaimy
76
developed countries and among the more
tribally oriented people, Abrahams, P.W.
and Parasons, J. A.(1996).
The term "clay" refers to a
naturally occurring material composed
primarily of fine-grained minerals, which
is generally plastic at appropriate water
contents and will harden with dried or
fired. Although clay usually contains
phyllosilicates, it may contain other
materials that impart plasticity and
harden when dried or fired. The
associated phases in clay may include
materials that do not impart plasticity
and organic matter Guggenheim S.
(1995).
Mineral Clays come in many
colors and hues depending on their
sources and mineral content, and their
healing properties may vary accordingly.
While the trace element and mineral salts
in clays partly account for their healing
properties, clay is thought to act as a
catalyst rather than the actual healing
agent Wilson M.J. (2003).
Research in modern uses of high
quality healing clays has indicated that it
can be effective used to treat skin
affections, stomach ulcers, arthritis.
These clays can help in the treatment of
various conditions such as hemorrhoids,
viral infection, mucus colitis, open
wounds, anemia, and acne among other
ills. Because clays contains both types of
dietary iron–ferrous and ferric- in an
easily assimilated form, they can help
treat anemia. It alleviates allergic
discomfort by neutralizing allergen
Carretero, M. I. (2002); Plumlee, G.S.
and Ziegler, T. L. (2003).
There is a clinical need for new
treatment options for serious Gram-
positive and Gram-negative infections.
Recently, introduced agents such as the
newer fluoroquinolones and the ketolide
telithromycin have limited use as they do
not cover methicillin-resistant
Staphylococcus aureus (MRSA), or
glycopeptide-resistant enterococci
(GRE). Clay minerals may prove success
in the treatment of these problematic
resistance bacterial pathogens Lynda B.
W. et al. (2008); Shah PM. (2005) &
Williams, L. B. et al. (2004).
MATERIALS AND METHODS
This research was fulfilled in the
bacteriology laboratory in the College of
Dentistry of Al-Anbar University. Four
types of mineral clays, {gray clay
(Kaolinite) yellow clay (Fuller’s earth),
white clay (Bentonite), and pink clay
(Common clay)}, were examined to
detect their antibacterial activity against
two bacterial pathogens that were
reported as significant pathogens of skin
infections: the first pathogen is
Staphylococcus aureus that was isolated
from patient (adult male) with skin
infection (furunculosis). And the second
pathogen is Pseudomonas aeruginosa
that was isolated from patient (adult
female) with urinary tract infection.
Before use in any susceptibility testing,
all mineral samples were sterilized by
autoclaving at 121°C for 15 min.
Bacterial strains were grown overnight
on suitable medium and diluted with
fresh medium to achieve an approximate
density of 1 × 107 cfu/400 L. Total
viable count (TVC) was done to confirm
the initial bacterial counts, serially-
diluted bacterial cultures were plated on
the Brain heart infusion agar plates and
enumerated. After dilution, sterilized
mineral clays (200 mg) were introduced
into 400 L of media containing the
initial pathogen inoculums. The
bacteria–mineral mixtures were
incubated for 24 h at 37°C. Positive
controls for growth of bacteria in the
absence of clay minerals were included
in each series of independent
experiments. Negative control growth
experiments with clay minerals in Brain
heart infusion broth were performed
several times throughout the course of
the study. The same tests were
performed using sterile Brain heart
infusion broth as negative control
Antibacterial Effect of some Mineral Clays In Vitro 77
0
10
20
30
40
50
60
70
80
90
100
positive control 0h
positive control 24h
white clay mixture 24h
gray clay mixture 24h
pink clay mixture 24h
yallow clay mixture 24h
type of clays
bacterial viability CFU
without clays. The bacteria – clay
mixtures were incubated for 24 h at
37°C., Shelley E.H. (2008).
After incubation, the mixtures
were subjected to successive 10-fold
serial dilutions in the normal saline,
mixed with a vortex shaker to ensure
dispersion and quantitatively cultured in
duplicate onto agar plates to determine
the number of viable bacteria Collee J.G.
et al. (1998).
RESULTS
Incubation with the (white clay,
gray clay, and yellow clay) revealed
incomplete killing of Staphylococcus
aureus. That means these clays
demonstrated a bactericidal effect against
this bacterium. In contrast, incubation
with the pink mineral clay resulted in
low growth of Staphylococcus aureus.
That means these clays demonstrated a
bacteriostatic effect against this bacterial
isolate (Fig. 1).
Fig. 1: Antibacterial activity of mineral clays that used in this study against Staphylococcus aureus
isolate.
In the presence of the white and gray mineral clay, growth of P. aeruginosa
isolate was significantly enhanced while growth of this bacterium in presence of the
yellow clay was not significantly different compared to bacterial growth in media
alone. On the other hand, Pink mineral clay demonstrated a bactericidal effect against
P. aeruginosa isolate (Fig. 2).
Fig. 2: Antibacterial activity of mineral clays that used in this study against P. aeruginosa isolate.
0
10
20
30
40
50
60
70
80
90
positive control 0h
positive control 24h
white clay mixture 24h
gray clay mixture 24h
pink clay mixture 24h
yallow clay mixture 24h
type of clays
bacterial viability CFU
Shehab A. Lafi and Mohammed R. Al-Dulaimy
78
DISCUSSION
Worldwide, antibiotic resistant
bacterial pathogens have become a
serious problem in both the developed
and underdeveloped nations. In certain
settings, such as hospitals and some
child-care locations, the rate of antibiotic
resistance is so higher than the usual,
low-cost antibiotics are virtually useless
for treatment of frequently seen
infections. This leads to more frequent
use of newer and more expensive
compounds, which in turn leads to the
rise of resistance to those drugs. A
struggle to develop new antibiotics
ensues, to prevent losing future battles
against infection, Andersson DI. and
Levin BR.(1999); Newman D.J. and
Cragg GM.(2007), mineral clays could
prove an inexpensive bactericidal
compounds against resistant bacterial
pathogens.
Skin infections, such as Buruli
ulcer, is recognized as a global health
threat, and recent observations showing
that the clay minerals were effective at
healing necrotic Buruli ulcer disease
prompted our investigations of these
geological nanomaterials, Brunet de
Courrsou L. (2002). The use of clay
minerals in the treatment and healing of
skin lesions represents great promise for
the development of an inexpensive cure
for many skin diseases and topical
infections, Williams L. B. et al. (2004).
To assess the effect of the clays
minerals on the growth of clinically-
relevant Gram-negative and Gram-
positive bacteria, susceptibility testing of
Staphylococcus aureus and
Pseudomonas aeruginosa was performed
in liquid cultures. To achieve poultice
consistencies similar to those used to
treat skin infection patients, initial
bacterial cultures (∼107 bacteria/400 L)
were mixed with 200 mg of clay
minerals and incubated at 37°C for 24
hours, Shelley E.H. (2008). In the
present study three out of four mineral
clays (white clay, gray clay, and yellow
clay) were seen to have a good
bactericidal effect against Gram-positive
bacteria, other hand, pink mineral clay
demonstrated a bacteriostatic effect
against the same bacteria, and has a
bactericidal effect against Gram-negative
bacteria . Lynda et al. and Necip) stated
that Bentonite may provide a natural
pharmacy of antibacterial agents, Lynda
B. W. et al. (2009). and Necip G.
Bentonites (2009).
Shelley et al. reported that a
specific mineral clays have a clear
bacteriostatic effect against antibiotic-
resistant S. aureus, a single antibiotic-
resistant strain, PRSA, and a multidrug
resistant strain, MRSA, and he stated
that the same mineral clays have a good
bactericidal effect against several
antibiotic-sensitive Gram-negative
bacteria: E. coli, S. enterica serovar
Typhimurium and P. aeruginosa, Shelley
E.H., (2008). Zeev et al. reported that the
test microorganisms (i.e. Escherichia
coli, Staphylococcus aureus,
Propionibacterium acnes, Candida
albicans) rapidly lost their viability when
added to the mineral mud20.
Jackson et al (1974) stated that,
because of the special quality of specific
mineral clays for incorporating various
ions, the surface can be either
hydrophilic or hydrophobic depending
on the charge and available solutes. A
hydrophobic surface is inherently
organophyllic and could harbors an
organic substance that is lethal to
bacteria, Rong X. et al. (2008) and
Tertre, E. et al. (2006).
Stotzky, G. (1996), reported that
clay particles carry a negative electrical
charge while toxins in the body carry a
positive charge. With the usage of clay,
whether internally or externally, the
positively charged ions of various toxin
are attracted to the negatively charged
surface of the clay particle and an
exchanged of ions takes place. In this
Antibacterial Effect of some Mineral Clays In Vitro 79
process, the toxins are bound and held
onto until they can be eliminated
(Detoxification).
Enhanced growth of Gram-
negative bacteria upon incubation with
clay minerals, as demonstrated with
white and gray mineral clays, is not an
uncommon occurrence, as
microorganisms inherently require
numerous trace elements to facilitate
growth, Jiang, D. (2007).
Gabriel, B. et al. (1972), reported that
clay minerals have a positive effect on
the survival of Gram-negative Klebsiella
aerogenes., clay minerals may serve to
protect environmental bacteria from UV
irradiation or toxic substances.
Additionally, by providing structural
support and organic or inorganic nutrient
acquisition via its high cationic exchange
capacity, clay minerals may protect
bacteria and serve as a minimal
nutritional sphere for bacterial
proliferation, Bitton G, et al. (1972);
Lu¨nsdorf, H. et al. (2000) and Yanbo,
W. et al. (2010).
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Antibacterial Effect of some Mineral Clays In Vitro 81
ARABIC SUMMARY
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)
.(
:
) (
Staphylococcus aureus
Pseudomonas aeruginosa .
Pseudomonas aeruginosa
Staphylococcus aureus.
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