This study aimed to characterize Staphylococcus aureus and detect the tst1 gene from clinical and food samples in Silchar, India. A total of 96 samples were collected, of which 42 were clinical and 54 were food. S. aureus was isolated from 34 clinical samples and 18 food samples using biochemical tests. The isolates showed high resistance to erythromycin but sensitivity to vancomycin and teicoplanin. Polymerase chain reaction detected the tst1 gene in 34 clinical and 18 food isolates. This study demonstrates the presence of virulent S. aureus strains in foods in Silchar that can potentially cause toxic shock syndrome.

1. Characterization of Staphylococcus aureus and assessment of tst1 gene
from different clinical and ready-to-eat food samples
UNDER THE SUPERVISON OF
DR. INDU SHARMA
ASSISTANT PROFESSOR
DEPARTMENT OF MICROBIOLOGY
ASSAM UNIVERSITY, SILCHAR
PRESENTED BY:
RUHELY NATH
ROLL: 041614 NO: 22180118
REGISTRATION NO: 18-140061953 OF
2015-2016
DEPARTMENT OF MICROBIOLOGY
ASSAM UNIVERSITY, SILCHAR

2. INTRODUCTION:
• What is Staphylococcus aureus?
 Discovered in Aberdeen, Scotland in 1880 by the surgeon Sir Alexander Ogston in pus from surgical
abscesses.
How does S.aureus affect?
S.aureus is one of most common causes of infection which cause variety of manifestations, including
life-threating blood infections such as, (TSS) Toxic Shock Syndrome.
TOXIC SHOCK SYNDROME:
 Toxic shock syndrome toxin 1 (TSST-1) is an exotoxin produced by Staphylococcus aureus . It is thought
to play a central role in the pathogenesis of toxic shock syndrome (TSS). TSS is a multi-system illness
characterized by fever, hypotension or shock, skin rash, desquamation of both hands and feet skin, and
multiorgan involvement
 Staphylococcal TSS may manifest in two general forms, menstrual or non-menstrual.
 The toxins multiply and spread into blood stream.
 Infections usually occurs when bacteria enter your body through an opening in your skin such as a cut,
sore or other wound.

3. METHODS
1. COLLECTION OF SAMPLES
Coagulase slide test and coagulase tube test both are positive
for S.aureus ,because S.aureus known to produce coagulase
enzyme.
3. Microscopic observation:
Gram staining is done for all isolates
5. PHENOTYPICALLY CONFIRMATORY TEST FOR
S.aureus
6. ANTIBIOTIC SUSCEPTIBILITY TEST:
PCR screening were done for all the coagulase positive samples for
the detection of tst1 gene.
4. BIOCHEMICAL REACTIONS
2. ISOLATION & IDENTIFICATION OF CLINICALAND FOOD SPECIMENS
Disc diffusion method is used for isolates
7. PCR ASSAY:
TOTAL 96 SAMPLES
CLINICAL SAMPLE:42 Clinical samples were collected in sterile containers from community-acquired and private diagnostic laboratory(SRL Silchar)
FOOD SAMPLES: 54 samples were collected aseptically from different areas of Silchar city.
IMViC tests and rapid biochemical tests i.e., catalase &
oxidase were done for the isolates

4. OBJECTIVES OF THE PRESENT STUDY:
• Identification and biochemical characterization of Staphylococcus aureus
from clinical and ready to eat food samples.
• Antibiotic susceptibility test for coagulase positive Staphylococcus aureus.
• Detection of tst1 gene from isolates identified as Staphylococcus aureus.

5. ISOLATION OF CLINICAL AND FOOD SAMPLES
0
2
4
6
8
10
12
14
16
18
20
pus(19) sputum(9) urine(6) throat swabs
clinical samples
no.ofsamples
0
2
4
6
8
10
12
14
16
pasteurized
milk
powder milk amul lassi gulab jamun cheese meat fried rice fermented
pickles
FOOD samples
no.ofsamples

6. Morphological characterization of the isolates:
MORPHOLOGICAL TEST RESULTS
GRAM SATINING
GRAM CHARACTERSTICS
GRAM POSITIVE
GRAM POSITIVE, PURPLE
COLOR, COCCI IN SHAPE
CULTURAL
CHARACTERSTICS
RESULTS
MANITOL SALT AGAR PLATES
BLOOD AGAR PLATES
Shape: circular
Elevation: convex
Size: large
Appearance: shinny
Pigmentation: golden yellow
Shape: circular
Elevation: pin head
Size: large
Appearance: shiny
Pigmentation: white creamy β
hemolysis

7. Biochemical characterization:
Biochemical test Results
Oxidase test negative
Catalase test positive
Indole production test negative
Methyl Red test positive
Voges Proskauer test positive
Citrate utilization test positive
Urease test positive
Triple Sugar (TSI) agar test A/A ; G(-), H2S(-ve)
Phonotypical confirmatory test for S.aureus:
Coagulase test results
Slide coagulase positive
Tube coagulase positive

8. ANTIBIOTIC SUSEPTIBILITY PROFILING
VAN
Product
code
Antimicrobial
agents
Total
samples=70
Resistant /
(%)
Total
samples=70
Intermediat
e / (%)
Total
samples=70
Sensitive /
(%)
SDO13 Erythromycin
(E)
69 (99.99%) ___ ___
SD219 Cefepime
(CPM)
67 (95.71%) 3 (4.28%) ___
SD737 Gatifloxacin
(GAT)
52 (74.28%) 10 (14.2%) ___
SD213 Teicoplanin
(TEI )
62 (88.57%) ___ 5 (7.14%)
SD045 Vancomycin
(VA)
59 (84.2%) ___ 8 (11.4%)
Fig: Disc diffusion plate showing antimicrobial
activity by antibiotics

9. Molecular characterization
• PCR ASSAY
• A genotype detection of tst1 gene was made by polymerase chain reaction for all the 96 test isolates. Among all the
isolates those 52 (34 clinical samples and 18 from food samples) phenotypically confirmed isolates of S.aureus were also
found to be tst1 gene positive 1 2 3 4 5 6 control
819 bp 800 bp
Primer pairs Sequence(5’- 3’) Amplified product size Reference
Tst1-f
Tst1-r
ACGTTTACACATTTGAATGAAGG
CGTTATAAAGATAAAAGGGAGAACG 819BP
Moneckeet et al., 2007

10. DISCUSSION
• The objective of this study was the screening of clinical and food samples for the identification of coagulase positive
s.aureus strains and detection of tst1 gene, obtained from different local areas of silchar city, India. Staphylococcus
aureus is a one of the most important and significant human pathogens,
• Out of 42 clinical samples, 34(80.9%) samples were positive for slide coagulase and tube coagulase. Whereas, 18 food
samples were coagulase positive out of 53 samples(33.9%). Coagulase positive strains produce a variety of toxins and
are therefore potentially pathogenic.
• The key factor for the success of s.aureus as a pathogen is its remarkable capacity to acquire antibiotic resistance
the present study showed that susceptibility against Vancomycin (11.4%) was highest followed by Teicoplanin (7.14%)
then Gatifloxacin (14.2%) and cefepime (4.28%) are intermediate, whereas Erythromycin (99.99%) resistant for
staphylococcus strain. Hence, the prevalence and degree of antimicrobial resistance are increasing worldwide
• Thus, the current study indicates that although the presence of tst1 gene is less positive in food samples but currently
the major problem that physicians have to face when treating S.aureus infections is antibiotic resistance
• Thus, based on the present and limited study data attention should be paid to the occurrence of multi-drug resistant
Staphylococcus aureus, the current results also shows that due to the large phenotypic switching and altering capability
in its gene expression the reservoirs for S.aureus is increasing its field so, that foods which are ready to eat might be
also potential hazardous source of Staphylococcus aureus.
• This study reveals that the presence of the high virulence toxic gene in food samples and the pathogenicity of S.aureus
which exists in local areas of Silchar city. Thus strict control measures must be applied to minimize the contamination in
food

11. Conclusion:
• This variations in prevalence of S. aureus may be due to poor hygienic measures taken by food handlers
(Zakaryet al., 2011). As the reservoirs of S.aureus are humans and contaminated fomins, therefore the
foods which are prepared, it should be under proper hygienic conditions for the better health of the
consumers. This study revealed that the resistance of S.aureus is increasing & dominating its virulence
factors in different environments even though things are maintained in pasteurized conditions.
 FUTURE SCOPE:
The finding of this study gives some important areas of future research which are outlined below.
 The mechanism behind the occurrence of S.aureus in packed food, also the genetics behind TSS caused by
direct consumption of contaminated food remains to be clarified.(Schlievert in 2001)
 TSST-1 is common among invasive S.aureus strains, especially methicillin resistant and need to develop
vaccines (Michie c et al., 2002) Investigation whether vaccination with nontoxic mutant toxic shock
syndrome toxin 1 (Mtsst-1) can protect against S.aureus infection, (Mckenney D et al., 1999) and
therapeutic approaches regarding to S.aureus require further study.

12. References:
• Cruickshank, R., J.P Duguid, B.p. Marmion and R.H.A. Swain, 1975. medical
microbiology. 12th ed., vol. ii chruchill livingstone edinburg london and new york
• Crass, B.A. and bergdoll, MS. 1986. toxin involvement in toxic shock syndrome. J. infect.
DIS 153,918
• Dinges mm, orwin pm, schlievert pm. Exotoxins of staphylocoocs aureus. Clinical
microbiol- rev 2000; 13:16-34
• Martin m 13.1.16-34. 2000. clin microbiol. Rev 2000,13(1):16.doi.
• Sorum h, sunde m (2001) resistance to antibiotic in the normal flora of animals vet.res.
13:227-241

13. THANK YOU
FOR YOUR ATTENTION