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The Impact of Nanosilver Particles On Foot -Odor Producing Microbes

Article in Current World Environment · December 2014

DOI: 10.12944/CWE.9.3.09

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Current World Environment Vol. 9(3), 615-622 (2014)

The Impact of Nanosilver Particles

On Foot - Odor Producing Microbes
Bassam Mashat and Abdel Hameed A. Awad*

Department of Environmental and Health Research, the Custodian of the

Two Holy Mosques Institute for Hajj and Umrah Research,
Umm Al Qura University, Makkah, P.O. Box 6287, Makkah 21955, Saudi Arabia.

http://dx.doi.org/10.12944/CWE.9.3.09

(Received: July 09, 2014; Accepted: August 29, 2014)

Abstract

Nanosilver applications have been implicated in paint, plastic, textile and medical industries.
Nanosilver particles absorb sweat and in turn avoid pungent smell of feet. The present study aims to
determine the antibacterial activity of Ag-coated different sock fabrics and comparing their efficacy
in reducing foot-borne resident bacteria. Synthesize of nanosilver particles was performed in various
concentrations by wet reduction method using silver nitrate and tri-sodium citrate. Coating of different
sock fabrics, cotton, nylon, and mixed cotton and nylon, was carried out by exposing these fabrics
to various silver concentrations for 24 hrs. The antibacterial efficacy of the nanosilver-finished
fabrics was checked by zone inhibition test and antibacterial test. The coated nylon fabric showed
better antimicrobial activity than other fabrics. There was no significant difference found between
antibacterial activity and type of fabric. Klebsiella pneumoniae exhibited the smallest inhibition zone,
and the inhibition zone increased with increasing silver concentration. The inhibition zones varied
according to bacterial species, each species had its own minimum inhibition concentration. The
percentages of reduction ranged between 18-80%. Sarcina lutea was highly sensitive to nanosilver
particles. The effect of temperature, relative humidity, dirt and oil on the antibacterial activity of silver
coated fabrics should be studied in the future.

Key words: Nanosilver particle, Antimicrobial activity, Sock fabric,

Foot-borne bacteria, Efficiency of reduction.

introduction surface area, thus increasing their contact time

with bacteria and fungi, and vastly improving its
The antimicrobial activity of the silver ions was first bactericidal and fungicidal effectiveness7. The use
identified in the 19th century, and colloidal silver was of metal nanoparticles for water disinfection is
accepted by the US Food and Drug Administration relatively new8,9. Nanosilver coating surface of textile
(FDA) as being an effective for wound management or footwear is one approach to produce highly active
in the 1920s1,2. The antibacterial effects of Ag- salts surfaces to have UV blocking, antimicrobial, and self
have been noticed since antiquity 3. Silver is an cleaning properties.
antifungal and antibacterial agent for antibiotic
resistant microbes4, it has beneficial healing and Suppression of microbial growth is
anti-disease properties5, and it prevents infection, accomplished by exposing the feet to air to enhance
and anti-inflammatory6. evaporation and reduce moistures’ growth-stimulating
effect. The resolution of a severe infection requires
Nanotechnology is a rapidly growing prescription drugs, salves, or foot soakings. Baking
science of producing and utilizing nano-sized soda, basil oil, tea tree oil, sage oil, and clove oil have
particles. Nanosilver particles have large specific been reported to inhibit the growth of both aerobic
616 Mashate & Awad et al., Curr. World Environ., Vol. 9(3), 615-622 (2014)

bacteria and yeast-mold-fungi10,11. Several studies Evaluating the antimicrobial activity of nanosilver
have been carried out on bactericidal activity of nano particles against bacterial species
particles and their applications in plastic, health, The antimicrobial activity of nanosilver
textile and paint industries12-14. Silver is a medically particles coated sock fabrics was studied against
proven natural antibacterial agent, which acts to kill Sarcina lutea, Pseudomonas aer uginosa ,
most bacteria that cause foot odor and other sweaty Staphylococcus epidermidis, and Klebsiella
foot problems15. Silver ions are passed from the silver pneumoniae. These bacteria are frequently isolated
fibers to infiltrate odor emitting bacterial cells and from human skin. The tested bacterial species were
bond with their DNA chains7. Silver nanoparticles inoculated in test tubes containing trypticase soya
destabilize plasma membrane potential and depletion broth (Hi-Media Laboratories, India) medium, and
of levels of intercellular adenosine triphosphate by incubated at 37oC for 18 h. The culture suspensions
targeting bacterial membrane resulting in bacterial were serially diluted in sterilized distilled water
cell death16. reaching a final concentration of 104 colony forming
unit per milliliter (CFU/mL).
The common skin microbes rapidly
proliferate in a moisture-rich and enclosed The antimicrobial efficacy of nanosilver particles
environment resulting in limited or no athletic was evaluated by the following tests:
participation. Hot weather, sweating, exercise, and Zone inhibition test
shoes generate a moisture-rich environment that Antimicrobial activity of the prepared
stimulates overgrowth of both aerobic bacteria and nanosilver particles was confirmed by zone inhibition
fungi. The present study aims to determine the test19. A sterile cotton swab was dipped into the
antibacterial activity of Ag-coated different sock culture suspensions, and streaked onto the surface
fabrics and comparing their efficacy in reducing of Muller Hinton agar medium. Square pieces of
foot-borne resident bacteria. both the coated (test) and uncoated (control) sock
fabrics were gently placed on the Muller Hinton
Materials and methods agar. This procedure was repeated for all the tested
bacterial species with different sock fabrics and
Synthesis of nanosilver particles Ag- concentrations. The plates were incubated
Chemical wet reduction method was used at 37°C for 24 h, and the zone of inhibition was
to synthesize nanosilver particles. The synthesis measured20.
of Ag-citrate was done according to the literature
procedure17. Breifly: 25 mL of 0.005 molar (M) Antimicrobial test
stock solution of silver nitrate in water was diluted The coated (test) and uncoated (control)
to 125 mL, and heated until it begins to boil. Then fabrics with 400 mm2 ± 0.1 cm2 diameters were
5 mL of 1% tri-sodium citrate solution was added; placed in a 250-ml sterile flask, and allowed to absorb
heating continued until the color was pale yellow. 0.5 ml of different bacterial culture suspensions.
The yellowish brown color indicates formation of The flasks were incubated at 37oC for 24 h. After
nanosilver particles. The solution was cooled at room incubation, 20 ml of sterile distilled water was added
temperature, and serial dilutions were prepared, into the flasks and shaken vigorously for 3 min.
(0.005 M, 0.01 M, and 0.05 M). Serial dilutions up to 10"3 were prepared. Aliquots,
0.2 ml, of the original sample, and its serial dilutions
Coating of sock fabrics with nanosilver were spread-plated, in triplicate, onto the surface
particles of nutrient agar for counting of the tested bacteria.
Pieces of 20 x 20 mm size (400 mm2 The percentage of reduction of viable organism was
±0.1mm ) of each sterilized sock fabrics, nylon,
2
calculated according to Shastri and his coauthors7.
cotton and mixed cotton and nylon, were soaked into
various concentrations of Ag-citrate overnight. The Isolation and identification of foot-borne
fabric pieces were washed with sterilized distilled bacteria
water to remove any adsorbed ion particles, and The samples of sock and foot skin were
dried at 120°C in a hot air oven for 30-60 min18. collected from 10 volunteers by swab sampling
 Mashate & Awad et al., Curr. World Environ., Vol. 9(3), 615-622 (2014) 617

technique. The volunteers were healthy people, Results and discussion

without wounds, diabetic, and athlete’s foot
infections; but suffer from pungent foot odors. The Identification of bacterial isolates
swabs were suspended in phosphate buffer solution, A total of 315 bacterial isolates belonging
and shaking vigorously for 15 min. Serial dilutions to 14 genera were identified in Table 1. Kocuria and
were prepared, and aliquots (0.5 ml) were spread- Sarcina were the dominant Gram positive cocci, and
plated onto the surface of nutrient agar medium. The Gram-negative bacteria represented by Klebsiella,
bacterial plates were incubated at 37°C for 48 h. Escherichia coli, Pseudomonas and Acinetobacter.
Pseuddomonas species were the dominant Gram
Three to five isolates of different colony negative bacterial species isolated. Gram positive
morphology which appear in more than 5% of bacteria were the dominant isolates, because they
the nutrient agar medium plates were picked up, are associated with skin scales and every day human
purified and subcultured for further identification. activities. Most of the isolated bacterial genera
Bacterial isolates were identified using Gram stain, are non-medical important for the healthy people,
oxidation fermentation, oxidase and catalase tests however some genera produce pungent foot odor
described in the Bergey’s Manual of Systematic and play an important role in wound infection. Gram
Bacteriology21. negative bacteria may be dangerous if they are
detected in high concentration22.
Statistical analysis
ANOVA (one-way and post hoc double Nanosilver particles
comparison tests) were used for determining Nanosilver par ticles were produced
antibacterial activity on bacterial growth with various by chemical reduction of silver nitrate (AgNO3)
Ag coated fabric concentrations. Statistical analysis and trisodium citrate according to the following
was performed using SPSS 18 (PASWStatistics 18). reaction:
P<0.05 was considered as significant.
4Ag + + C 6 H 5 O 7 Na 3 + 2H 2 O→4Ag 0 +
C6H5O7H3 + 3Na+ +H+ +O2

Table. 1: The percentages of bacterial genera The surface of sock fabric is highly porous
isolated from sock fabric and foot skin which may results in homogeneity of nanoparticles
inside fabric. Silver coated fabric is anti-bactericidal
Genus No. % to a wide range of pathogens, absorbs sweat and
helps eliminate unpleasant foot odor by killing
G +ve bacteria bacteria23.
Kocuria 45 14.28
Bacillus 38 12.06 Antibacterial efficacy of Ag-coated fabric against
Staphylococcus 39 12.38 foot borne bacteria
Sarcina 44 13.96 The antibacterial activity varied depending
Tetrads 35 11.11 on the type of sock fabric, Ag-concentration, and
Diplococci 40 12.69 bacterial species (Table 2). The inhibition zone of
Sporeforming bacilli 6 1.9 Ag-coated sock fabrics on bacterial growth are
Streptomycetes 3 0.95 shown in figures 1-3. The one-way descriptive
Corynebacteria 8 2.54 analysis of variance (ANOVA) showed no significant
Streptococcus 4 1.27 differences between inhibition zones with Ag-coated
G -ve bacteria cotton, nylon, and mixed cotton fabrics (p > 0.5).
Klebsiella pneumoniae 6 1.9 However, nylon fabric exhibited better antimicrobial
Escherichia coli  7 2.22 activity (Fig. 3). Ag-coated sock fabrics exhibited the
Pseudomonas 34 10.8 highest antibacterial activity against Pseudomonas
Acinetobacter 6 1.9 aeruginosa and the lowest against Klebsiella
Total 315 100 pneumoniae.
618 Mashate & Awad et al., Curr. World Environ., Vol. 9(3), 615-622 (2014)

The effect of Ag-concentration on bacterial to 0.05M increased inhibition zone (Figs. 4-6).
growth Pseudomonas aeruginosa inhibited with Ag-
The antibacterial activity varied with coated cotton and mixed fabrics at concentration
Ag-concentration. Increasing concentration of of 0.005M. Pseudomonas aeruginosa was found
Ag-coated sock fabrics over a range of 0.005 M to be the most sensitive to nanosilver particles.

Table. 2: Zones of inhibition (mm) of foot borne bacteria obtained

with sock coated nanosilver-particles

Ag- Sarcina Klebsiella Pseudomonas Staphylococcus

Concentration lutea pneumoniae aeruginosa epidermidis
/molar
C C+N N C C+N N C C+N N C C+N N

0.005 0 21 22 0 0 0 22 21 22 0 0 0
0.01 27 24 27 25 23 24 25 22 26 25 24 26
0.5 28 28 29 26 24 26 27 27 30 26 26 29

C: cotton, C+N: mixed cotton and nylon, N: nylon

M =0.005 M =0.01 M =0.05

30

25

20
zone size/mm

15

10

0
Sarcina lutea Klebsiella pneumoniae Pseudomonas aeruginosa Staphylococcus
epidermidis

organism

Fig. 1: Inhibition zones obtained by Ag-cotton sock against different bacterial species

M=0.005 M=0.01 M=0.05

30
25
20
zone size/mm

15
10
5
0
Sarcina lutea Klebsiella pneumoniae Pseudomonas aeruginosa Staphylococcus
epidermidis

organism

Fig. 2: Inhibition zones obtained by Ag-coated mixed cotton and nylon sock against bacterial
species
 Mashate & Awad et al., Curr. World Environ., Vol. 9(3), 615-622 (2014) 619

M=0.005 M=0.01 M=0.05

35
30
25
zone size/mm

20
15
10
5
0
Sarcina lutea Klebsiella pneumoniae Pseudomonas aeruginosa Staphylococcus
epidermidis

organism

Fig. 3: Inhibition zones obtained by Ag-coated nylon sock against bacterial species

Sarcina lutea Klebsiella pneumoniae Pseudomonas aeruginosa Staphylococcus epidermidis

30

25

20
zone size

15

10

0
M=0.005 M=0.01 M=0.05

conccentration

Fig. 4: Inhibition zone of bacterial species obtained by various Ag-concentrations coated cotton
sock

Sarcina lutea E.coli Pseudomonas aeruginosa Staphylococcus epidermidis

30

25
zone size/mm

20

15

10

0
M=0.005 M=0.01 M=0.05

concentration

Fig. 5: Inhibition zone of bacterial species obtained by various Ag-concentrations coated mixed
cotton and nylon sock
620 Mashate & Awad et al., Curr. World Environ., Vol. 9(3), 615-622 (2014)

Sarcina lutea Klebsiella p neumoniae Pseudomonas aeruginosa Stap hy lococcus ep idermidis

35

30

25
zone size/mm

20

15

10

0
M=0.005 M=0.01 M=0.05

concentration

Fig. 6: Inhibition zone of bacterial species obtained by various Ag-concentrations coated nylon
sock.

0.005 0.01 0.05

90
80 77
80
70 72 71
67 66
60 62
56 53 53
50 49 48 46
41
%

40 37 39 36
40 40 37
33 32
30
20 22
18
10
0
c n c n c n c n

Sarcina Staphylococcus Pseudomonas Klebsiella pneumoniae

epidermidis
bacteria/sock type

Fig. 7: Percentages of reduction of bacteria obtained against both Ag-coated sock fabrics and
various concentrations

Ag-coated nylon fabric effectively inhibited Sarcina Pseudomonas and Sarcina were inhibited at the
and Pseudomonas (Fig. 6). Klebsiella pneumoniae lowest Ag-concentration.
exhibited more resistant to nanosilver; however
S. aureus and Pseudomonas aeruginosa were Antibacterial test
more sensitive. This can be attributed to the cell The degree of antibacterial activity was
wall structure of Klebsiella. Gram-positive and carried out against bacterial species with Ag-coated
Gram-negative bacteria differ in their membrane and uncoated cotton and nylon sock fabrics (Fig.
structure, the thickness of the peptidoglycan layer. 7). Ag-coated nylon fabric showed higher rate of
Klebsiella tends to be rounder and thicker than killing than Ag-coated cotton fabric. However non
other members of the Enterobacteriaceae family. significant difference was found between Ag-coated
Klebsiella forms a capsule or slime layer24. These nylon and cotton sock fabrics (P> 0.05). The rate of
results suggested that the antimicrobial activity of killing increased gradually with increasing Ag-ion
nanosilver particles depended on characteristics concentration. A significant difference (P<0.05)
of bacterial species, and every species had its was found between Ag-coated and uncoated sock
own minimum inhibition concentration. In this study fabrics. Ag-coated cotton and nylon sock fabrics
 Mashate & Awad et al., Curr. World Environ., Vol. 9(3), 615-622 (2014) 621

effectively inhibited Sarcina lutea, with increasing Ag- ion to irreversibly damage key enzyme systems in
concentration, the percentage of reduction reached the cell membranes of pathogens29, and depletion
77% and 80%, respectively. Klebsiella pneumoniae of levels of intracellular adenosine triphosphate by
exhibited low killing rate, with percentages of targeting bacterial membrane16.
reduction ranged between 18-46%
Nanosilver particles effectively inhibit growth
Gupta and Cooper et al.25 observed that of various microorganisms, making them applicable
the growth of bacterial colonies around silver-loaded to diverse medical devices and antimicrobial control
cotton fabric was inhibited. One potential advantage systems. Coating of socks fabric with nanosilver
of silver nylon is its ability to release silver ions particles can be used as an effective way to
continuously over the entire time period 26. This combat foot-borne pathogens and thereby reducing
confirms the greater inhibition zones of nylon sock discomfort like foot odor, and athlete’s foot, without
fabric coated with nanosilver particles. Hong et al.23 toxic or allergenic effects.
evaluated the antibacterial activities of the heat-
treated PVA/AgNO3 nano-fibres against S. aureus Conclusion
and K. pneumonia. They found a reduction in the
number of colonies of S. aureus and K. pneumoniae Nanosilver proved to be effective in inhibiting
(> 99.9%) after 18 h of incubation. These results bacterial growth. Ag-coated sock fabrics sufficiently
are not agreement with the results in the present inhibited bacterial growth. Ag- coated fabric acts
study, this difference is attributed to the materials as antimicrobial agent killing foot-odor producing
and techniques were used. bacteria. The percentage of reduction increased
gradually with increasing silver concentration,
The mechanism of the inhibitory effect of reached ~80% for Sarcina lutea. Ag-coated nylon
Ag ions on microorganisms is partially known. Some sock fabric exhibited greater antimicrobial activity
studies have reported that the positive charge on the than cotton fabric. Klebsiella pneumoniae was highly
Ag- ion is crucial for its antimicrobial activity through resistant to nanosilver particles. Coating of socks
the electrostatic attraction between negative charged fabric with nanosilver particles can be used as anti-
cell membrane of microorganism and positive odor, and to combat foot-borne bacteria.
charged nanoparticles27. The antimicrobial activity
of nanosilver particles on Gram-negative bacteria Acknowledgment
depended on Ag nanoparticle concentration, and
formation of pits in the cell wall of bacteria28. The This work was funded by Scientific
effectiveness of silver compound as an antiseptic is Research and Revival of Islamic Heritage Institute,
based on the ability of the biologically active silver Umm Al Qura University, Makkah, Saudi Arabia

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