Cefotax-magnetic nanoparticles as an alternative approach to control Methicillin-Resistant Staphylococcus aureus (MRSA) from different sources

The prevalence of S. aureus isolation from the different examined samples

Staphylococcal infections represent a public health issue in hospitals and health care settings as well as a major economical and welfare problem in dairy animal farming²⁵. The prevalence of S. aureus isolation from the farm under the study (Table 2) showed that 63 (33.1%) out of 190 different samples were bacteriologically positive. Moreover, the isolation was mainly obtained from manager swabs followed by milk machine swabs, nasal swabs and hand swabs (60.0, 53.3, 40.0 and 28.0%, respectively), and to a lesser extent in milk samples (24.0%). Meanwhile it was not isolated at any percent from water trough swabs, at X² = 48.8, P < 0.001. These findings spotlight the great role played by the environment as a reservoir of the bacterial infection to the farm workers and later on consumers everywhere also suggested the great role they play in the dissemination of the infection and the running of the pathogen in the nature between animals and humans, where Fluit²⁵ proved that the presence of human MRSA isolates in both mastitic milk or infected dairy cattle suggested the role of the environment in transmission of the pathogen between animal and human populations through contaminated utensil, Fessler et al.²⁶ also reported that 20% of personnel hanging the poultry in the slaughter were MRSA positive compared to 1.9% from other personnel, while 8% of poultry arriving to the slaughter were MRSA positive that increased to 35% during the day through contamination which suggested the role played by the environment in spreading of the infection, Spohr et al.²⁷ also proved environmental pollution with MRSA through detection of MRSA in different farm animals, Antoci et al.²⁸ mentioned that prevalence of MRSA was 36 and 61% from humans and bulk milk tank, respectively that might be attributed to low milk hygienic measures. Furthermore, according to Joubert et al.²⁹ MRSA in humans, can be maintained and colonized in the mucosa of healthy individuals, especially nasal mucosa and their skin as a potential carrier of the organism that coincide with our results where we were able to isolate the pathogen by (40.0%) from nasal swabs of farm workers; our finding were in harmony with Juhász-Kaszanyitzky et al.³⁰, Suelam et al.³¹ and Suranindyah et al.³².

Sensitivity pattern (%) of S. aureus traits to the tested antibiotics

The pattern of antibiotic sensitivity of examined isolates (Table 3) showed variable results where all isolates were resistant to the majority of tested antibiotics where all isolates were significantly resistant to amoxicillin clavulanic acid (P < 0.001) with variable degree (feed staff, nasal, hand, milk and milk machine, respectively) (61.1, 60.0, 57.1, 41.7 and 37.5% respectively), beside all isolates showed resistance to cefotax including feed staff (77.8%), milk samples (75.5%), milk machine (75.0%), hand swabs (71.4%) and finally nasal swabs (70.0%). Also, most of the obtained isolates from milk machine, nasal swab, feed staff and milk showed resistance to oxytetracycline (68.7, 50.0, 44.4 and 41.7%, respectively). Meanwhile all isolates were significantly sensitive to cefotax magnetic nanocomposite (P < 0.05) where the sensitivity by milk machine reached up to (87.7%), milk and feed staff (83.3% each), hand swabs (71.4%), and nasal swabs (60.0%). Moreover, all isolates obtained (milk, feed staff, hand swabs, milk machine and nasal swabs, respectively) were also significantly sensitive to vancomycin at P < 0.05 (83.3, 66.6, 57.1, 56.3 and 50.0%, respectively). On the other hand, the recovered isolates showed variable degree of resistance and sensitivity to cefepime and kanamycin but mainly resistance especially among environmental traits. The illustrated results suggested that antibiotic resistant traits of S. aureus are no longer exclusive issue but still in the same time pose a high risk due to the zoonotic importance of this pathogen also the possibility of transmission of resistance genes of both disinfectants as well other antibiotics between man and animal populations³³. The obtained results are to some extent in agreement with Tong et al.³⁴ who indicated that drug-resistant strains are growing to represent serious health threats specially MRSA, and Kayvan et al.³⁵ who proved that S. aureus isolates were resistant to oxacillin (71.15%), cefoxitin (67.31%) and tetracycline (69.81%) and Tenhagen et al.³⁶ who mentioned that 100.0% of S. aureus were resistant to cefoxitin and penicillin. In contrast Cihalova et al.³⁷ found that MRSA were resistant to second-line treatment such as vancomycin and doxycycline. And Jamali et al.³⁸ reported lower rates of resistance to oxacillin, penicillin and cefoxitin (13.0, 44.4 and 4.9%, respectively).

Characterization of barium ferrite nanoparticles and barium ferrite cefotax nanocomposite

Figure 3 clarifies X-ray diffraction spectroscopy that was used to investigate the crystalline structure of the samples. The X-ray diffraction pattern of barium ferrite showed the specific peak at 2θ at 17.79, 18.91, 23.09, 30.39, 32.24, 35.57, 37.17, 40.51, 42.73, 55.19, 56.68 and 63.34 which are corresponding to (101), (102), (006), (110), (107), (200), (203), (205), (206), (217), (2011) and (220) planes of BFO³⁹. This result indicates that the barium hexaferrite powder is well crystallized. Regarding barium ferrite/cefotax, the X-ray diffraction pattern (Fig. 3) clearly evidences a slight shifting to a high diffraction angle, a small change in the relative intensity, and a broadening of the diffraction peaks of pure BFO owing to BaF/cefotax formation. This phenomenon is due to the cefotax loading on BFO. And the specific peak of cefotax appeared at 2θ = 33.2, reflections indicating that cefotax is successfully composited with BFO nanoparticles. All the samples exhibit minor peak for hematite phase (marked with asterisks) appeared at 2θ = 31.2 probably because of the preparation method^40,41. It can be also seen in Fig. 3 that the intensity of the hematite peak shrinks with loading cefotax⁴². Average crystallite sizes of the samples were calculated using Scherer equation and are found to be in range of 23 nm–29 nm.

Fourier-transform infrared spectrum was used to identify the functional groups present in the synthesis of samples. The spectrum of pure BFO (Fig. 4a) mainly illustrates characteristic peaks, including 429 cm⁻¹ and 583 cm⁻¹ corresponded to the lattice vibrations of octahedral and tetrahedral metal ions respectively^43,44. The intense peak that appeared at 896 cm⁻¹ belonged to vibrations from Fe–O or Ba–O⁴⁵. On the other hand, BFO/cefotax, Fig. 4b, spectrum b showed the appearance of important bands of cefotax in the spectra of the BFO /cefotax composite as following: stretching of O–H groups appeared at 3456 cm⁻¹, C=O stretching vibration of (COO)^2– centered at 1852.76 cm⁻¹, N–H stretching is shown around 1569 cm⁻¹, the peaks at 1623, 1494.10, 1367.96, 1032, 772, and 610 cm⁻¹ are assigned to the presence of O of an amide group, N–H, C=O, C–N, C–O, CH₂, and C–S groups, respectively⁴⁶.

Zeta potential analysis is a measure of the charge attraction/repulsion between particles in solution, suspension stability, and gives us information about the reason of aggregation and dispersion. Figure 5 illustrated the zeta potentials of aqueous dispersions of the BFO and BFO/Cefotax were -31.40 and -38.80 mV, respectively. The results show that the lower zeta potential value of the BFO/Cefotax can be due to the interactions between the inorganic material and organic molecules (cefotax). The decrease is expected to be based on electrostatic considerations. The size distribution intensities of the BFO and BFO/cefotax were 642.10 and 839.9 nm, respectively.

The surface morphologies of the prepared BFO and BFO loaded with cefotax were studied by SEM, and ideal images of the samples were presented in Fig. 6. As expected, the loading with cefotax resulted in clearly visible microstructure differences of the obtained materials. For BFO formation of ultrafine, irregular-shaped grains (Fig. 6a). The particles are plate like hexagonal /hexagonal in structure. Even though the particles are not uniform hexagonal but mixed of plate like hexagonal/hexagonal. However, for BFO loaded with cefotax, the surface morphology changes to regular grains formed regular hexagonal platelets (Fig. 6b). Therefore, BFO/cefotax composite is expected to be promising catalyst.

Figure 7 shows the high-resolution transmission electron microscopy (HRTEM) micrographs of the BFO and BFO loaded with cefotax. It can be clearly seen that there was an obvious difference between the HRTEM images. Pure BFO consists of layered structure with an irregular shape Fig. 7a. BFO loaded with cefotax image Fig. 7b shows cefotax is distributed on the surface of the barium ferrite nano sheet. The face-to-face assembly of BFO and cefotax optimizes their contact area which is advantageous for controlling MRSA.

At this study a trial for increasing the antibiotic cefotax, one of the methicillin resistant antibiotics, efficiency against MRSA by loading it on magnetic NPs as barium ferrite. The results in the current study showed a good bactericidal activity for the antibiotic loaded on NPs against the resistant bacteria through the MIC, MBC and disc diffusion assay while the magnetic material itself (barium ferrite) showed no antimicrobial activity. The main reason for improvement the antimicrobial activity against the resistant bacteria depends upon mainly their high surface area to volume ratio which gives a chance for the antibiotic to come in contact well with more surface area of the bacterial cell membrane⁴². Magnetic nanoparticles in current nanotechnology approaches is one of the major topics of interest^14,15. Loading antibiotics on magnetic nanoparticles had an effective strategy in the treatment of infectious diseases caused by resistant microorganisms specially if targeted with external magnetic field as this fasten form the treatment and minimizes the side effects with enhances efficiency^16,17. Ferrites in particular are non-toxic of low price, biocompatible and their magnetism is very important in different biomedical applications. Also loading of antibiotics on large surface area of magnetite NPs increases the interaction and the penetration of the loaded antibiotics with the bacterial cell surface besides ferrites causes oxidative stress and free radical formation which help in the bactericidal activity of the loaded antibiotics^14,46,47. In addition, the efficacy of this loading on NPs materials depends mainly upon various factors such as surface modification, natural characteristics of particles, particle composition and the targeted bacterial species. The safety of barium ferrite had been previously studies and indicated the safety of these particles to be used for treatment of infections¹⁵.

Source: Ecology - nature.com