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Discipline: mastitis; Key words: antibiotic resistance, biofilm, Staphylococcus aureus, bacteriocin, Caenorhabditis elegans, in vivo screening platform.
Mastitis-causing pathogens appear to increasingly develop resistance to antibiotics. This implies that regular screening is required to firstly quantify their resistance and resilience, and secondly to establish which antibiotics are still comparatively effective. However, in the long term research needs to move away from antibiotics and consider alternative methods to control these pathogens. Methods which could be considered include usage of bacteriophages and bacteriocins which are “natural” enemies of pathogens. Research is currently been conducted in South Africa to investigate this possibility and therefore it is useful to take note of similar work elsewhere in the world. In this report I discuss results on antibiotic resistance and the effect of bacteriocins which were reported in the Journal of Dairy Science, Volume 99 of 2016, at respectively pages 8607 to 8613 (antibiotics – first author: Dr Aslantis) and 8614 to 8621 (bacteriocins – first author: Dr Son). The titles of the papers are: Investigation of the antibiotic resistance and biofilm-forming ability of Staphylococcus aureus from subclinical bovine mastitis cases and In vivo screening platform for bacteriocins using Caenorhabditis elegans to control mastitis-causing pathogens.
In the antibiotic study, a total of 112 Staphylococcus aureus isolates obtained from subclinical bovine mastitis cases were examined for antibiotic susceptibility and biofilm-forming ability as well as genes responsible for antibiotic resistance, biofilm-forming ability, and adhesin. Antimicrobial susceptibility of the isolates was determined by the disk diffusion method and the biofilm forming ability by the Congo red agar method, the standard tube method and the microplate method. The genes responsible for antibiotic resistance, biofilm-forming ability and adhesion were examined by PCR.
Five of the 112 isolates were identified as methicillin-resistant Staph. aureus. The resistance rates to the antibiotics penicillin, ampicillin, tetracycline, erythromycin, trimethoprim-sulfamethoxazole, enrofloxacin and amoxicillin-clavulanic acid were 45.5, 39.3, 33, 26.8, 5.4, 0.9 and 0.9%, respectively. All isolates were susceptible against vancomycin and gentamicin. The blaZ (100%), tetK (67.6%) and ermA (70%) genes were the most common antibiotic-resistance genes. The three methods used to estimate biofilms respectively suggest that about 71, 67 and 63% of the isolates were biofilm producers. The percentage of icaA, icaD and bap genes in Staph. aureus isolates was 86.6, 86.6, and 13.4%, respectively.
The results indicated that Staph. aureus from sublinical bovine mastitis cases were mainly resistant to β-lactams and, to a lesser extent, to tetracycline and erythromycin. Also, biofilm- and adhesion-related genes, which are increasingly accepted as an important virulence factor in the pathogenesis of Staph. aureus infections, were detected at a high rate.
The bacteriocin study aimed to develop an in vivo (in animal) screening method using Caenorhabditis elegans to identify a novel bacteriocin for controlling the mastitis-causing pathogen Staph. aureus strain RF122 in dairy cows. Using Bacillus spp. the authors developed a direct in vivo screening method that uses 96-well plates and fluorescence image analysis. They then identified a novel bacteriocin produced by Bacillus licheniformis strain 146 (lichenicin 146) with a high in vivo antimicrobial activity using a liquid Clostridium elegans–Staph. aureus assay. They also determined the characteristics of lichenicin 146 using liquid chromatography-mass spectrometry and confirmed that it shared homologous sequences with the bacteriocin family proteins. In addition, RNA-sequencing analysis revealed genes encoding cell surface or membrane proteins that are involved in the bactericidal activity of lichenicin strain RF122 infection.
Bottom line: The direct in vivo screening method which the authors developed, facilitates simple, convenient, cost-effective and reliable screening of potential antimicrobial (bacteriocidal) compounds against mastitis pathogens.