Production of Slime Layer by Staphylococcus

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Production of Slime Layer by Staphylococcus
epidermidis Isolated From Corneal Infection
ARTICLE · JANUARY 2011
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Sinai Waleed
University of Baghdad
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Retrieved on: 28 January 2016

Baghdad Science Journal Vol.8(3)2011
147
Production of Slime Layer by Staphylococcus epidermidis
Isolated From Corneal Infection
Munira CH. Ismail * Fais I. Ali** Sinai W. Mohammed*
Received 31, May, 2009
Accepted 1, July, 2010
Abstract:
A total of 37 Staphylococcus epidermidis isolates, isolated from corneal scraping of
patients with bacterial keratitis and 20 isolates from healthy eyes (as control) (all
isolates, isolated from, Ibn Al- Haietham eye hospital / Baghdad), were tested for
slime production, 52.63% of all isolates were positive-slime production (23 isolates
from patients and 7 isolates from controls). It was found that positive-slime producing
S. epidermidis were exhibited a high resistance to antibiotics as compared to negative-
slime producing isolates.
Key words: Slime Layer, Staphylococcus epidermidis, Keratitis.
Introduction:
The coagulase-negative staphylococci
(CONS) are widely distributed over the
surface of human body, where they
constitute the majority of the common
nasal bacterial micro flora. Among the
CONS, Staphylococcus epidermidis is
the most frequently isolated species
and the most common species
responsible for infection [1]. One
important property of S. epidermidis
which is responsible for its persistence
and / or opportunistic invasion in the
tissues is its ability to produce slime
[2, 3]. Slime not only helps the
organism in adhesion to host cells, but
also protects it from phagocytosis and
from the action of antibiotics [4].
Despite being important ocular
pathogens, S. epidermidis have so far
received little attention in
ophthalmology. The purpose of this
study was to identify, determine
antibiotic susceptibility and slime
production of S. epidermidis isolated
from patients with bacterial Keratitis.
Materials and Methods:
Bacteria: isolates of S. epidermidis
(CONS) from 57 patients, who
attended the Ibn Al-Haietham Eye
Hospital, Baghdad, during October
2001 to October 2002.
Subjects: of the 57 patients, 37 had
come for treatment and investigation of
keratitis and 20 (from healthy eyes)
served as controls.
Methods:
1. Corneal scrapings: were
performed in each case under the slit–
lamp biomicroscope. The scrapings
were taken from the base and the
margins of the ulcer and were then
smeared on glass slides for Gram -
staining. The specimens also
inoculated at 37 ºC on to blood and
chocolate agar.
2. Conjunctival swabs: were
obtained from 20 control subjects. In
order to obtain an ideal swab for
culture.
3. Isolation and identification of
bacteria: culture material from corneal
scrapings and swab were routinely
plated on the following media: Blood
agar and Chocolate agar with 10% CO2
(at 37ºC for 24 hr.). In positive-culture
cases: all bacteria (Staphylococcus
spp.) were identified by API-system
(API-staph) (Bio mereiux).
* Tropical Biological Research Unit / Science College
** Ibn Al- Haietham Eye Hospital / Ministry of Health

147
4. Slime-production test: isolates
were tested for slime production with
the use of a technique described by
Christensen et al., 1982 [5]. In brief, a
loop of organisms from a pure growth
on blood agar plate was inoculated
onto 5ml of trypticase soy broth
(oxoid), and incubated at 37ºC for 48
hr. the contents of the tubes were
aspirated and the tubes were stained
with 1% safranine ( BDH) for 7min. A
visible safranine stained film lining the
wall of the tubes indicated a positive
test.
5. Antibiotic susceptibility: The
susceptibility of S. epidermidis isolates
were performed by Kirby-Bauer disc
diffusion assay [6]. We choose five
effective antibiotics against most
strains of corneal pathogens. The
antibiotics and their
concentrations/disc (µg) were:
[Ciprofloxacin (5 µg), Gentamicin (10
µg), Cephalothin (30 µg), Rifampicin
(5 µg) and Chloramphenicol (30 µg)]
(oxoid).
6. Statistical analysis: Chi-Square
test was used in the analysis of results
[7].
Results and Discussion:
Slime test: A total of 57 CONS
isolates were studied: 23 (62.16%)
isolates from patients and 7(35%) from
controls were positive-slime
production. thus 30 (52.63%) of 57
isolates were positive-slime producers.
Antibiotics susceptibility: The results
of antibiotic susceptibility testing of S.
epidermidis isolates were isolated from
patients with bacterial keratitis are
given in Table (1)& Figure (1). The
results showed a high resistance to
rifampicin and chloramphenicol [16
(69.5%) and 14 (60.8%) respectively].
While cephalothin, ciprofloxacin and
gentamicin had a low resistance [ 7
(30.43%), 8 (34.78%) and 10
(43.47%)] respectively in positive-
slime producing isolates. While
negative-slime producing isolates
exhibited a low resistant against
antibiotics, there was significant
difference (X2
= 25.8, P < 0.05)
between them. Table (2) &Figure (2)
shows antibiotic resistance of control
isolates. Results showed a low resistant
against all antibiotics which are used
particularly in negative-slime
production isolates there was
significant difference (X2
= 42.7, P <
0.05) between them.
Slime layer has been documented to be
one of the virulence markers of S.
epidermidis because of the close
association of slime producing strains
with infections related to indwelling
medical devices including intraocular
lenses [8]. Positive-slime producing
isolates were isolated in high numbers
from patients as compared to control
[9].
We found a positive association
between positive-slime production and
resistance to antibiotics. This is
supported by observation made
presently in some reports [2, 9, 10],
that slime not only helped the
organism to colonize the host tissues,
but it also protected it from the action
of antibiotics.
Cephalothen and Ciprofloxacin a new
broad spectrum antibiotics were found
an effective agent in this study, they
shows a low resistance for most of S.
epidermidis isolates and bacteria had
no chance to develop resistance to
these antibiotics was expected to be
slow because they requires
chromosomal mutation, and resistance
can not to be transferred by plasmid
mediated mechanisms [11]. The results
are similar to those reported by other
authors [12], who found that slime
layer and multi drug resistance were
the important virulence factors of S.
epidermidis in bacterial keratitis.
Studies on biofilms have shown that S.
epidermidis is the most frequently
isolated slime-producing CNS and is

147
also the most common cause of
nosocomial infections in patients with
catheters, medical implants or other
invasive devices [13].
In conclusion, our findings showed
that the slime layer, was responsible
for resistance to antibiotics
Table (1): Antibiotic resistance of
keratitis isolates (No. of isolates=37).
Antibiotics
Positive Slime
production
(NO. of
isolates=23)
%
Negative
Slime
production
(NO. of
isolates=14)
%
Chloramphenicol 14 60.8 2 15
Ciprofloxacin 8 34.78 2 15
Cephalothen 7 30.43 5 35
Rifampicin 16 69.5 4 29
Gentamycin 10 43.47 4 29
Fig. (1): Antibiotic resistance pattern
of Keratitis isolates.
Table (2): Antibiotic resistance of
control isolates ( No. of isolates=20).
Antibiotics
Positive
Slime
production
(NO. of
isolates=7)
%
Negative Slime production
(NO. of isolates=13)
%
Chloramphenicol 4 59 5 39
Ciprofloxacin 3 39 3 25
Cephalothen 3 44 5 39.5
Rifampicin 2 29.5 4 31
Gentamycin 2 29 3 23
Fig. (2): Antibiotic resistance pattern
of Control isolates.
References:
1. O'Gara, J.P., Humphreys, H. 2001.
Staphylococcus epidermidis
biofilms: importance and
implications (Review article). J.
Med. Microbiol. 50: 582-587.
2. Nayak, N., Satpathy, G. 2000.
Slime production as a virulence
factor in Staphylococcus
epidermidis isolated from bacterial
keratitis. Indian J. med. Res. 111:
6-10.
3. Beachey, E.H. 1981. Bacterial
adherence: adhesion-receptor
interactions mediating the
attachment of bacteria to mucosal
surface. J. Infect. Dis. 143: 325-
345.
4. Quie, P.G., Belani, k. k. 1987.
Coagulase-negative staphylococcal
adherence. J. Infect. Dis. 156: 543-
547.
5. Christensen, G. D., Simpson, W.
A., Bisno, A. L. and Beachey, E.
H. 1982. Adherence of slime-
producing strains of
Staphylococcus epidermidis to
smooth surfaces. Infect. immune.
37: 318-326.
6. Baur, A. W., Sheris, J. G. and
Truck, M. 1966. Antibiotic
susceptibility testing by
0
10
20
30
40
50
60
70
Percentageofantibioticsresistance
Chloram. Cipro. Cepha. Rifam. Gentam.
Antibiotics
Keratitis isolates
Slime+(NO. of isolates=23)
Slime-(NO. of isolates=14)
0
10
20
30
40
50
60
Percentageofantibiotics
resistance
Chloram. Cipro. Cepha. Rifam. Gentam.
Antibiotics
Control isolates (Commensals)
Slime+(NO. of isolates=7)
Slime-(NO. of isolates=13)

144
standardized single disk method.
Am. J. Clin. Path. 43: 493-496.
7. Negi, K. S. 2008. Biostatistics with
latest MCQs. A.I.T.B.S. publishers,
2nd
. India pp146.
8. Raskin, E. M., Speaker, M. G.,
McCormick, S. A., Wong, D.,
Menikoff, S. A. and Pelton. H. k.
1993. Influence of hepatic
materials on adherence of
staphylococci to intraocular lenses.
Arch. Ophthalmol, 111: 250-253.
9. Peters, G., Locci, R. and Pulverer,
G. 1980. Adherence and growth of
coagulase-negative staphylococci
on surface of intravenous catheters.
J. Infect. Dis. 146: 479-482.
10. Arslan, S., Özkardes, F. 2007.
Slime production and antibiotic
susceptibility in staphylococci
isolated from clinical samples.
Mem. Inst Oswaldo Cruz, Rio de
Janeiro. 102(1): 29-33
11. Tuft, S. J., Metheson, M. 2000. In
vitro antibiotic resistance in
bacterial keratitis in London. Br. J.
Opthalmol. 84: 687-691.
12. Nayak, N., Nag, T.C., Satpathy, G.
& Ray, S.B. 2007. Ultrastructural
analysis of slime positive & slime
negative Staphylococcus
epidermidis isolates in infectious
keratitis. Indian J Med Res 125:
767-771.
13. Oliveira, A., Cunha, M. L. R. S.
2008. Bacterial Biofilms with
Emphasis on Coagulase-Negative
Staphylococci. J. Venom. Anim.
Toxins Incl. Trop. Dis., 14, 4, 588
‫أصابت‬ ‫من‬ ‫المعزولت‬ ‫للجلذ‬ ‫العنقوديه‬ ‫المكوراث‬ ‫بكتريا‬ ‫من‬ ‫اللزجت‬ ‫الطبقت‬ ‫أنتاج‬
‫القرنيت‬
*‫محمذسعيذ‬ ‫وليذ‬ ‫سيناء‬ **‫علي‬ ‫اسماعيل‬ ‫فائز‬ *‫جلوب‬ ‫اسماعيل‬ ‫منيرة‬
‫االبحاد‬ ‫وحدة‬ *‫العلىم‬ ‫كليت‬ / ‫الحارة‬ ‫للوٌاطق‬ ‫البايىلىجيت‬
‫الصحت‬ ‫وسارة‬ / ‫للعيىى‬ ‫الهيثن‬ ‫ابي‬ ‫هسخشفى‬ **
‫الخالصت‬:
‫لـ‬ ‫الكلي‬ ‫الوجوىع‬ ‫هي‬37‫بالخهـا‬ ‫هصـابيي‬ ‫هزنـى‬ ‫مزًيـاث‬ ‫هـي‬ ‫عشلـج‬ ‫للجلـد‬ َ‫يـ‬ ‫العٌيى‬ ‫الوكـىراث‬ ‫لبكخزيـا‬ ‫عشلـت‬
‫و‬ ‫البكخيــز‬ ‫ـت‬‫ـ‬‫اليزًي‬20‫زة‬ ‫الســي‬ ‫ـابت‬‫ـ‬‫هص‬ ‫(يــز‬ ‫ـىى‬‫ـ‬‫عي‬ ‫مزًيــاث‬ ‫ـي‬‫ـ‬‫ه‬ ‫عشلــج‬ ‫ـت‬‫ـ‬‫عشل‬‫هــي‬ ‫ـج‬‫ـ‬‫عشل‬ ‫العــشالثل‬ ‫ـا‬‫ـ‬‫جوي‬
‫ل‬ ‫ـدا‬‫ـ‬ ‫ب‬ / ‫ـىى‬‫ـ‬‫للعي‬ ‫ـثن‬‫ـ‬‫الهي‬ ‫ـي‬‫ـ‬‫اب‬ ‫ـفى‬‫ـ‬‫هسخش‬‫ـز‬‫ـ‬‫ظه‬52.63‫ـا‬‫ـ‬‫هٌه‬ %23‫و‬ ‫ـابيي‬‫ـ‬‫الوص‬ ‫ـي‬‫ـ‬‫ه‬ ‫ـت‬‫ـ‬‫عشل‬7‫ـابيي‬‫ـ‬‫الوص‬ ‫ـز‬‫ـ‬‫(ي‬ ‫ـي‬‫ـ‬‫ه‬
‫ومد‬ ‫اللشجت‬ ‫بيت‬ ‫لل‬ ‫هٌخجت‬‫أظهزث‬‫الٌخائج‬‫إى‬‫اث‬ ‫ـا‬ ‫الو‬ ٍ‫حجـا‬ ‫عاليـت‬ ‫هياوهـت‬ ‫لهـا‬ ‫بيـت‬ ‫ال‬ ٍ‫لهذ‬ ‫الوٌخجت‬ ‫العشالث‬ ‫جويا‬
‫الوٌخج‬ ‫(يز‬ ‫بالعشالث‬ ‫هيارًت‬ ‫الحيىيت‬.‫ت‬

Production of Slime Layer by Staphylococcus

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