UNIVERSITY OF COPENHAGEN Faculty of and Medical Faculty

UNIVERSITY OF COPENHAGEN Faculty of and Medical Faculty of Health Helth and Medical Sciences A NOVEL METHOD TO MEASURE ANTIBIOTIC USE IN HUMANS ANNE METTE LERBECH M. SC. (PHARMACY) Dias 1 Department of Pharmacy

Department of Pharmacy UNIVERSITY OF COPENHAGEN COAGULASE-NEGATIVE STAPHYLOCOCCI (CNS) Part of the normal flora - human skin and mucus membranes Predominant species: S. epidermidis, S. hominis, S. haemolyticus Source: www. science photo. com Contaminants of urine samples Potential pathogens that have become resistant to many known antibiotics Resistance in CNS are suggested to develop due to positive selective pressure following antibiotic treatment → Antimicrobial resistance of CNS → indicator for antibiotic drug consumption Precise picture Dias 2

Department of Pharmacy UNIVERSITY OF COPENHAGEN INCORPORATING CHEMISTRY INTO THE STUDY Antibiotics are excreted through the kidneys → present in urine Urine sample Analysis by SPE-HPLC-MS/MS Person exposed to antibiotics? Pharmacy at Shai Osudoku District Hospital Dias 3

UNIVERSITY OF COPENHAGEN Department of Pharmacy AIM Occurrence of antimicrobial resistance in CNS and the presence of 9 commonly used antimicrobial agents in human urine. Relation between the incidence of resistant CNS and the presence of antibiotics in urine. Dias 4

UNIVERSITY OF COPENHAGEN SITES IN GHANA Kumasi Faculty of Pharmacy, KNUST Dodowa Shai Osudoku District Hospital Accra (capital) Korle-Bu Teaching Hospital, Central Lab, UGMS Soruce: Google Maps Dias 5 Department of Pharmacy

Department of Pharmacy UNIVERSITY OF COPENHAGEN METHOD Sampling Adult patients with a request for urine, blod or stool analysis. Korle-Bu Dodowa Qualitative interviews Demographics, medical history, antibiotic consumption Mary-Anne Ahiabu in interview-room, Dodowa Patients provide ~150 m. L urine 2 m. L for culture and susceptibility testing ~150 m. L for analysis of antibiotics in urine 246 urine samples were collected between February and April 2012 Dias 6

UNIVERSITY OF COPENHAGEN CULTURE AND SUSCEPTIBILITY TESTING Growth Inoculated on Discovery agar Isolated on MH agar Identification of CNS Gram staining, coagulase tube test MALDI-To. F/MS Antimicrobial susceptibility Disk diffusion method/ 12 antimicrobials 96 CNS were isolated from 84 patients Dias 7 Department of Pharmacy

UNIVERSITY OF COPENHAGEN PREVALENCE OF STAPHYLOCOCCI ISOLATED FROM URINE SAMPLES 8 different species of CNS among the 96 isolates Lerbech et al. PLo. S One, 2014 Dias 8 Department of Pharmacy

UNIVERSITY OF COPENHAGEN Department of Pharmacy PREVALENCE OF RESISTANT CNS ISOLATED FROM URINE SAMPLES Penicillin: 98% Trimethoprim: 67% Tetracycline: 63% Resistant CNS is a indicator for antibiotic cumsumption Lerbech et al. PLo. S One, 2014 Dias 9

Department of Pharmacy UNIVERSITY OF COPENHAGEN % PATIENTS VS. NUMBER OF ANTIBIOTIC MARKERS 70 % 27 % Data from Ghana: Lerbech et al. PLo. S One, 2014 Dias 10 100% of patients exhibited resistance to at least 1 antibiotic Multiple drug resistant CNS: resistant to more than 3 antibiotics

UNIVERSITY OF COPENHAGEN ANALYSIS OF ANTIBIOTIC IN URINE Sample preparation - Internal standard added / Frozen (Korle. Bu/Dodowa) - transported to KNUST - Loaded on SPE columns / Frozen - Transported to Denmark - Elution of samples Analysis by HPLC-MS/MS 121 urine samples were analysed Dias 11 Department of Pharmacy

Department of Pharmacy UNIVERSITY OF COPENHAGEN OCCURRENCE OF ANTIBIOTICS IN URINE 64% of the samples tested showed the presence of at least one antibiotic Lerbech et al. PLo. S One, 2014 Dias 12

UNIVERSITY OF COPENHAGEN Department of Pharmacy % PATIENTS VS. NUMBER OF ANTIBIOTIC FOUND/TAKEN Antibiotics were detected in 77 out of 121 patients but only 16 patients responded to have taken antibiotics More antibiotics measured than reported (p = 0, 003) Lerbech et al. PLo. S One, 2014 Dias 13

Department of Pharmacy UNIVERSITY OF COPENHAGEN INCIDENCE OF RESISTANT CNS AND THE PRESENCE OF ANTIBIOTICS S. haemolyticus (R) non-S. haemolyticus (R) Total Antibiotic detected No antibiotic detected Total 267 106 373 47 37 314 143 84 457 p = 0. 0063 The prevalence of resistant S. haemolyticus was more frequent than the prevalence of resistant non-S. haemolyticus when antibiotic were detected in the urine. Lerbech et al. PLo. S One, 2014 Dias 14

UNIVERSITY OF COPENHAGEN Department of Pharmacy PERSPECTIVES Lerbech et al. : - participants who were either seeing a physician for a minor health ailment, such as suspected UTI or pregnant women coming in for routine check-ups. - Samples collected at referral hospitals and patients may have sought care elsewhere and given antibiotics before reporting. Sofie Trads and Sophie Hane-Weijman: - Healthy Ghanaians from Korle Gonno and Dodowa. - Inclusion criteria: NOT taken antibiotics within the past 14 days Dias 15

UNIVERSITY OF COPENHAGEN Department of Pharmacy PREVALENCE OF STAPHYLOCOCCI ISOLATED FROM URINE SAMPLES Dias 16

UNIVERSITY OF COPENHAGEN Department of Pharmacy PREVALENCE OF RESISTANT CNS ISOLATED FROM URINE SAMPLES Dias 17

UNIVERSITY OF COPENHAGEN OCCURRENCE OF ANTIBIOTICS IN URINE Lerbech et al. also analysed for cefurocime and doxycycline Dias 18 Department of Pharmacy

UNIVERSITY OF COPENHAGEN Department of Pharmacy CONCLUSION Lerbech et al. : - The prevalence of resistant S. haemolyticus was more frequent than other resistant CNS species when antibiotic were detected in the urine Sophie Hane-Weijman and Sofie Trads: - Samples with the presence of antibiotic were significantly more likely to contain resistant CNS Overall: - The prevalence of detected antibiotics in human urine was more frequent than the use reported by the participants - Selective pressure favoring resistant CNS An urgent need to: - identify potentially diffuse sources of antibiotic exposure - more appropriate selection and use of antibiotics in Ghana Dias 19

UNIVERSITY OF COPENHAGEN ACKNOWLEDGMENTS AND THANK YOU Bjarne Styrishave, Niels Frimod-Møller, Japheth A. Opintan, Martin Hansen, Britt Pinkowski Tersbøl Samuel O. Bekoe, Mary-Anne Ahiabu, Noah O. Nkrymah, Gifty Sunkwa-Mills Nana Adade, Michael Olu-Taiwo, Emmanuel Osisiogu, Dela Agboado, Amos Akumulena Kennedy T. C. Brightson, Samuel Ametepeh Staff at the laboratories at Korle-Bu Teaching Hospital and Shai Osudoku District Hospital All the sweet Ghainans whom participated in the study Søren Bak, Jonas Abdel-Khalik, Kenneth Munk Pedersen, Erland Björklund, Susanne Hermansen, Sophie Hane-Weijman, Sofie Trads Dias 20 Department of Pharmacy

UNIVERSITY OF COPENHAGEN THANK YOU FOR LISTENING Dias 21 Department of Pharmacy