Avaliação in vitro do efeito bacteriostático do plasma rico em plaquetas (PRP) em Staphylococcus aureus sensível a meticilina (MSSA) e Staphylococcus aureus resistentes a meticilina (MRSA) pelo teste kill curve assay

Abstract

Staphylococcal infections are relevant for their frequency and severity. Staphylococcus aureus (S. aureus) is a relevant Gram-positive bacterium because it commonly causes infections, because of the increase in resistance cases, it develops quickly and represents a serious public health problem in the world. Platelets stand out in their participation in infectious and inflammatory processes due to their interaction with the pathogen, in particular bacteria and defense cells can be obtained in higher concentration in platelet-rich plasma (PRP), a concentrate of blood with different concentrations of factors of growth, platelets and leukocytes, and that has been used in a multidisciplinary way, in tissue regeneration and with antimicrobial properties. Given the hypothesis that platelets are responsible for the bacteriostatic effect of PRP against S. aureus, this study aimed to evaluate the in vitro bacteriostatic effect of canine platelets on methicillin-resistant S. aureus (MRSA) and methicillin-sensitive S. aureus (MSSA) using compounds containing different concentrations of platelets and canine leukocytes. A randomized block experimental study was designed, comparing the bacteriostatic effect of platelets, in 6 experimental groups against MRSA (G1-G6) and MSSA (G7-G12): G1 and G7, saline; G2 and G8, negative control; G3 and G9, culture medium; G4 and G10, PRP, G5 and G11, pure platelet-rich plasma (P-PRP), and G6 and G12, isolated leukocytes (WBC). The kill curve assay model was used to assess bacterial multiplication by counting colony-forming units (CFU/mL) in Mueller-Hinton (MHB) culture medium, after incubation of the bacteria with their respective treatments in MHB broth by 0h, 1h and 2 hours. PRP had a leukocyte contamination rate of 1:12.59 platelets, not observed in P-PRP, allowing the individualized evaluation of platelets on bacteria. For both bacteria, it was observed that leukocytes (WBC) and platelet:leukocyte interaction induced a reduction in bacterial multiplication throughout the experiment, with no significant difference between time points (p>0.05). The effect of treatments at each time point and compared to the basal control group (G1 and G7), a similar behavior was observed for both bacteria in the first hour of incubation, with a reduction in bacterial multiplication in the groups treated only with WBC (G5 = -2 .86x; G11 = -8.79x), followed by treatment with PRP (G4 = -1.69x; G10 = -3.96x), a fact observed in the second hour of incubation, but with reduced bacterial multiplication in G5 (-1.07x) and G11 (-2.36x), and an intense reduction in the PRP group only for MSSA (G10 = -8.03x), a fact not observed for MRSA (G5 = 1.33x). Interestingly, the isolated platelets stimulated bacterial multiplication both in the first hour of incubation (G6 = 1.53x; G12 = 1.24x) and in the second hour (G12 = 1.50x), with the exception of G6 (-1.16x ). This effect may be due to physical platelet:leukocyte interaction, such as the content of platelet granules and leukocytes. Thus, platelets play an important role in the inhibition/reduction of bacterial multiplication in vitro of MRSA and MSSA when in the presence of leukocytes, emphasizing the synergistic interaction, especially in the first moments of exposure to the pathogen.