Bloodstream infection caused by bacterial pathogen

Bloodstream infection caused by bacterial pathogen and their sensitivity and resistance antibiotic pattern in Dhaka area

1. 1 Course Code: MPG-513 Course Title: Thesis Dept. of Microbiology Primeasia University M .Sc Project Presentation on Bloodstream infections caused by bacterial pathogens and their sensitivity and resistant to antibiotics pattern in patients in Dhaka areas hospital Presented By: Md.Masud Rana ID: 231032061

2. 2 Index Bloodstream infection Aims and Objectives Materials Method Sample Site of sample collection Method of sample collection Procedure Isolated bacterial strains from blood Sample Gram Staining Biochemical test Antimicrobial susceptibility Conclusion

3. 3 Study Objective Isolation and identification of bacteria from Blood culture using culture-based method. Determination of gender and age-wise distribution of Bloodstream infection in blood samples collected from Gulshan Clinic LTD Progoti Sarani,Shahjadpur , Dhaka. Analysis of antibiotic sensitivity pattern of isolated bacteria isolated from clinical samples commonly prescribed antibiotics in that region

4. 4 Bloodstream Infection: A bloodstream infection, also known as a bacteraemia , blood poisoning or septicaemia , is an infection caused by bacteria entering the bloodstream. It occurs when a bacterial infection somewhere in the body, such as in the lungs, intestines, urine or skin, enters the bloodstream. 

5. 5 Aims and Objective: Mostly when a patient is suffering from an unknown disease. To confirm bacteremia in patients we isolated Staphylococcus aureus . and Salmonella spp. from blood sample from different areas in areas hospital . Identification of the isolated using culture, microscopic and biochemical tests . Antibiotic sensitivity test by disk difussion method..

6. 6 Materials: Petri dishes Test tubes Inoculating loop Sterile cotton Bunsen Burner Sterile forceps Autoclave Incubator Hand Gloves Microscope

7. Method: Collection of Blood sample ( 100 ) for a total 1 month period from Gulshan Clinic LTD Progoti Sarani,Shahjadpur , Dhaka. ꜜ Blood culture bottle entered into the blood culture machine (BACT/ALART 3D) ꜜ Sub cultured from positive blood culture. ꜜ Isolation of the strains using selective media and enriched media. ꜜ Gram staining. ꜜ Biochemical test. ꜜ Antimicrobial susceptibility test.

8. Sample: Whole Blood Amount of Sample: For Adult: Minimum 5 ml For infant & Children : 1-2 ml: Neonate 2-3 ml: 1 month – 2 years. 3-5 ml: older children

9. 9 Site of sample collection: Arterial or Venous Blood Indwelling arterial or venous lines Central or Peripheral

10. 10 Methods of sample collection: A minimum of 5 ml blood is taken through vain puncture and injected into blood culture bottles. The blood is collected using clean technique .This requires that both the tops of the culture and the vain puncture site of the patient are cleaned prior to collection with alcohol swabs containing 2% chlorohexidine and 70 % isopropyl alcohol.

11. 11 Figure : Blood Culture bottle

12. 12 Procedure: Incubated blood culture bottle at 37˚C for 24 – 72 hours in blood culture machine ( Bactec 9050 ) for positive or negative signal. When the machine showed positive signals, then subculture was done from the culture bottle on the MacConkey agar media and Blood agar media for further testing (For subculture we used one selective medium & one enriched medium). The blood was subcultured by streaking plate technique. Incubated all plates at 37˚C for 24-48 hours. After 24 hours we observed the growth of microorganisms.

13. 13 Figure : Blood Culture Machine ( Bactec 9050)

14. 14 Isolated bacterial strains from blood Sample : Suspected bacterial culture on different enriched media and selective media:- Staphylococcus spp. on Blood agar media Salmonella spp. on MacConkey agar media

15. 15 Out of 100 samples 52 samples were detected positive growth and 128 samples were negative growth detected by culture method. Total patients Samples Positive growth Sample percentage Negative growth Sample percentage 100 26 26% 74 74% Figure: Percentage of Positive and negative samples

16. 16 Result: Analysis of blood Sample Out of 26 positive samples, male patients were 15 & female samples were 11 . blood Samples Male 15 Female 11 Total 26 Figure : Gender-wise distribution of positive case numbers in blood sample

17. 17 Result: Analysis of Blood Sample Blood Sample Age Samples Infected rate 11 to 20 04 14% 21 to 30 05 19% 31 to 40 04 15% 41 to 50 05 17% Above 50 08 35% Figure : Age-wise distribution of positive cases in Blood samples

18. 18 Gram Staining: We performed gram staining to identify gram positive or negative cocci /bacilli. 1) Salmonella spp . is gram negative organism 2) Staphylococcus spp. is gram positive organism. Gram Negative Gram Positive

19. 19 Gram staining Sample Type Positive Sample Isolated bacterial strains Gram staining Number Blood 26 Staphylococcus aureus . Gram positive cocci 16 Salmonella spp. Gram negative bacilli 10

20. 20 Biochemical tests: Suspected Strains Biochemical profile of isolated bacterial strains Catalase Tests Oxidase Tests Urease Tests Methyl Red (MR) Citrate test Positive Negative Positive Negative Positive Negative Positive Negative Positive Negative Staphylococcus spp. + - + + + Salmonella spp. + - - + +

21. 21 Catalase Test Oxidase Test Urease Test

22. 22 Citrate Test Methyl Red Test

23. 23 Antibiogram Antimicrobial susceptibility profile of isolated bacterial strains on Mueller Hinton Agar. At first, a single isolated colony was inoculated in the Mueller Hinton Broth Inoculated on the Mueller Hinton agar media Placed antimicrobial disk on the Mueller Hinton agar media plate by Kirby Bauer method. Observed sensitivity or resistance after 24 hours incubation at 37˚ C temperature

24. 24 Antibiotic discs used for antibiogram are: 1. Amoxicillin 12. Vancomycin 2. Doxycycline 13. Tetracycline 3. Levofloxacin 14. Linezolid 4. Ceftraxone 15. Ampicillin 5. Clindamycin 16. Azithromycin 6. Ciprofloxacin 17. Ceftazidime 7. Co- trimoxazole 18. Cefuroxime 8. Erythromycin 19. Colistin 9. Imipenem 20. Meropenem 10. Gentamicin 21. Nalidixic Acid 11. Rifampicin 22. Tigecycline Sensitive Resistant Intermediate Figure : Antibiotic sensibility test

25. 25 Result: Analysis of Blood Sample Antibiotic Susceptibility Pattern of Staphylococcus aureus in Blood Samples The antimicrobial sensitivity pattern in Blood samples showed that  Staphylococcus aureus  isolates were highly sensitive to doxicycline (88.0 %), Levofloxacin (76.0 %) , Clindamycin (85.0 %) and co- trimoxazole (100.0 %) while resistant to Amoxicillin (58.0 %), cifrofloxacin (70.0 %) and erythromycin (91.0 %),

26. 26 Result: Analysis of Blood Sample Antibiotic Susceptibility Pattern of Staphylococcus aureus in Blood Samples The antimicrobial sensitivity pattern in blood samples showed that  Staphylococcus aureus  isolates were highly sensitive to imipenem (100.0 %), Gentamicin (100.0 %) , rifampicin (91.0%), vancomycin (85.0%), tetracycline (100.0%) and linezolid (10.0%).

27. 27 Result: Analysis of Blood Sample Antibiotic Susceptibility Pattern of Salmonella spp in Blood Samples The antimicrobial sensitivity pattern in blood samples showed that  Salmonella spp  isolates were highly sensitive to ampicillin (90.0 %), ceftazidime (95.0 %) , ceftraxone (79.0%), cefuroxime (74.0%), and colistin (100.0%) while resistant to azithromycin (95.0%) ).

28. 28 Result: Analysis of Blood Sample Antibiotic Susceptibility Pattern of Salmonella spp in Blood Samples The antimicrobial sensitivity pattern in blood samples showed that  Salmonella spp  isolates were highly sensitive to imipenem (100.0 %), meropenem (100.0 %) , nalidixic acid (100.0%) and tigecycline (90.0%), and while intermediate to ciprofloxacin (79.0%) ).

29. 29 Discussion : Case study reported that there was a significant number ( 26 out of 10 0 patients) of patients were found where male patients were 15 & female patients were 11 . Among 26 patients found that 2 bacterial strains. Such as Staphylococcus spp. (Positive 16 samples. ) , Salmonella spp. (Positive 10 samples. ).

30. 30 Conclusion The study was conducted on patients visiting the Gulsan clinic LTD area in Dhaka areas hospital where 26 patients were analyzed. Among them, Staphylococcus aureus and Salmonella spp was found to be the most common isolate in blood samples, These findings highlight the importance of regular monitoring of bacterial isolates in clinical settings to prevent and manage infections effectively.

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