More Insights about the Efficacy of Copper Ion Treatment on Mycobacterium avium subsp. paratuberculosis (MAP): A Clue for the Observed Tolerance

Abstract

Background: Scientific evidence is scarce for the antimicrobial effect of copper on bacteria characterized as more resistant. Using Mycobacterium avium subsp. paratuberculosis (MAP), a highly resistant microorganism, as a pathogen model, copper ion treatment has shown a significant bactericidal effect; however, the sustainability of MAP against copper toxicity was also reported in several studies. Accordingly, the present study aimed to evaluate the impacts of copper on MAP. Methodology: This study considered physicochemical properties and copper concentration in a buffer since it could modulate MAP response during the application of copper treatment. Results: Despite the efficacy of copper ions in significantly reducing the MAP load in Phosphate Buffered Saline, some MAP cells were able to survive. The copper concentration generated by the copper ion treatment device increased significantly with increasing exposure times. MAP bacterial load decreased significantly when treated with copper ions as the exposure times increased. An increase in pH decreased oxygen consumption, and an increase in conductivity was reported after treatment application. Conclusions: Even with higher concentrations of copper, the efficacy of MAP control was not complete. The concentration of copper must be a key element in achieving control of highly resistant microorganisms.

Are reactive oxygen species (ROS) the main mechanism by which copper ion treatment degrades the DNA of Mycobacterium avium subsp. paratuberculosis suspended in milk?

“…MAP strain ATCC 19698 was cultured in enriched broth 7H9, as reported by Tejeda et al (2022a). In short, MAP growth was monitored weekly, using a Helios Gamma1 spectrophotometer (Thermo Scienti c).…”

Evidence of Homeostatic Regulation in Mycobacterium avium Subspecies paratuberculosis as an Adaptive Response to Copper Stress

“…The sequences of the primers used to amplify a 63-nucleotide fragment of the IS900 gene were 5 -gacgcgatgatcgaggag-3 (L) and 5 -gggcatgctcaggatgat-3 (R). The amplification conditions were as follows: One cycle at 95 • C for 10 min; 45 cycles with three steps of 95 • C for 10 s, 60 • C for 30 s, and 72 • C for 1 sec; and a final cooling step at 40 • C for 30 s. A blank sample (only PCR water) and positive (MAP ATCC 19698) control were used for both the DNA extraction protocol and the PCR reaction [15]. MAP cell counts were estimated according to the genome equivalent principle [13] based on the concentration of MAP DNA that was measured in the same spectrophotometer mentioned above.…”

“…To determine the total copper concentration in the PBS buffer after being challenged, each of the aforementioned treatments was digested with concentrated HCl (37% w/w) and HNO 3 (0.1 N) according to a modified protocol [15]. The measure was then determined by an atomic absorption spectrophotometer (AAS), and the result was reported in ppm.…”

“…A novel treatment based on copper ions was shown to be highly effective in reducing MAP load in liquid matrices, such as PBS and milk [13,14]. Despite this great efficacy, a small proportion of MAP cells managed to tolerate the treatment [14,15].…”