U ovom radu istražili smo prisutnost mikroorganizama u usnoj šupljini bolesnika prije i nakon transplantacije bubrega. Naime, mikroorganizmi u usnoj šupljini su važan dio normalne oralne flore, ali isto tako neki od njih su važan uzrok različitih bolesti kako same usne šupljine, tako i organizma u cjelini. Za identifikaciju mikroorganizama koristili smo matricom potpomognutu ionizaciju laserskom desorpcijom s vremenskim proletom i masenim spektrometrom. Radi se o vrlo osjetljivoj metodi kojom se može identificirati veliki broj različitih mikroorganizama.
Ciljevi istraživanja su bili analizirati prisutnost mikroorganizama u usnoj šupljini prije i nakon transplantacije bubrega te korelirati prisutnost mikroorganizama s oralnim zdravljem i oralnim statusom, imunosupresijom, antibioticima i šećernom bolesti.
Istraživanje je provedeno na 50 bolesnika koji su imali transplantaciju bubrega u Klinici za urologiju Kliničkog bolničkog centra Zagreb. Ispunili smo upitnike o oralnom zdravlju i oralnom statusu, analizirali broj vrsta mikroorganizama te vrstu imunosupresije, upotrebu antibiotika kao i prisutnost šećerne bolesti.
Sukladno ciljevima pokazali smo da u ustima bolesnika na dijalizi, odnosno nakon transplantacije postoji velik broj različitih mikroorganizama, ali i da se prisutnost nekih vrsta mikroorganizama mijenja nakon transplantacije. Također smo ustanovili da oralna higijena, odnosno oralni status nisu značajno utjecali na promjene mikroorganizama u usnoj šupljini kod naših bolesnika vjerojatno zbog toga što oni nisu promijenili svoje navike vezane uz oralnu higijenu niti im se promijenio oralni status nakon transplantacije bubrega. Nismo našli značajne promjene mikroorganizama vezane uz upotrebu antibiotika, odnosno prisutnost šećerne bolesti, vjerojatno zbog toga što se radilo o relativno malom broju bolesnika koji su uzimali antibiotike, odnosno koji su imali šećernu bolest.
Pokazali smo povećanje broja i vrsta Gram pozitivnih koka (aerobnih i anaerobnih) i smanjenje Gram negativnih štapića. Moguće je da su ove promjene povezane s imunosupresivnom terapijom, ali i s profilaktičkom primjenom antimikrobnih lijekova. Potrebna su daljnja istraživanja oralne mikrobiote kod bolesnika s transplantiranim bubrezima.
|Abstract (english)|| |
Background of research: The oral cavity is a complex system with very important functions, but it is also a home to a huge number of microorganisms, constantly changing during the lifetime of a person, and which depends on a numerous factors. More than 200 microbial species have been cultured from the oral cavity and 400 to 500 additional taxa (or phylotypes) have been detected by the 16S rRNA genetic analysis. Many of these microorganisms are responsible for oral and general health, but also involve different oral as well as different diseases in the entire organism.
The number of patients with an end-stage renal disease (ESRD) is increasing and the disease is becoming an important health care problem. Patients with ESRD (i.e., dialysis) have worse oral health and oral status and an increased risk of different oral and related systemic diseases. Kidney transplantation is the best treatment option for patients with ESRD, however, immunosuppression has serious side effects. Immunosuppressive drugs may increase the risk of different infections caused by various microorganisms including those present in the oral cavity. It can also change the oral status and predispose kidney transplant recipients to different oral lesions, as well as to different diseases of the entire organism such as diabetes mellitus or cancer.
Aim of the study: In this study we investigated the microorganism’s diversity in oral cavity before and after kidney transplantation using matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF MS).
Methodology: In all, 50 patients who had underwent a kidney transplant at the Department of Urology, Clinical Hospital Centre Zagreb (Croatia) from October 2013 to December 2014 were included in this prospective study. The oral swabs (using sterile cotton sticks) and washouts (using 5 ml of sterile saline) were taken from patients a few hours before the kidney transplantation (Sample A), 7-14 days after (early post-transplant period, Sample B) and then 3-6 months after the kidney transplantation (late post-transplant period, Sample C). Before the kidney transplantation, patients filled out a questionnaire about their oral hygiene and were examined by a dentist, their oral status was recorded and as was oral pathology, which can be a contraindication for kidney transplantation but which was excluded.
Results: Overall, 187 different microbial species were identified, and a total of 1812 different microorganisms. The average number of microbial species in Sample A was 13.0, in Sample B 12.6, and 13.8. (p=0.2) in Sample C. Microorganisms were grouped (based on micromorphology and biochemical characteristics) into 10 different groups. Our study shows that although there was no statistically significant change in the total number of microorganism species, nonetheless, a statistical significant change in different groups of microorganisms does exist. These changes relate to oral health (i.e., oral diseases) and oral hygiene, immunosuppressive therapy, antibiotics and other drugs that kidney transplant recipients receive, as well as other diseases. We did not show significant changes relating to the oral health and oral hygiene, nor the number of bacterial species in our patients probably because of the fact that their oral status and oral habits did not change after the kidney transplantation. The same also applies to the majority of their diseases (i.e., they were present before and after kidney transplantation) and the drugs used to treat them.
Furthermore, we did not found any correlation between the number of microbial species, and the type of immunosuppression, which is probably due to the small number of subjects when the group of 46 patients was divided into two and then to four additional subgroups.
What we did observe was a steady increase in the number of species in the group of gram-positive cocci and related bacteria, and in the group of anaerobic gram-positive cocci. What is also apparent is the over-3-period decrease in the number of species in the group of gram-negative rods. This is pretty much opposite of what happens during aging: the number of gram-positive bacteria is highest in childhood and decreases with the age as the number of gram-negative rods increases.
Conclusion: There is a significant change (increase/decrease) in certain groups of microbial species after kidney transplantation probably due to immunosuppressive therapy and/or use of antimicrobial drugs. Further studies are needed to clarify these changes in kidney transplant recipients.