Prevalence of antibiotic resistant mastitis pathogens in dairy cows in Egypt and potential biological control agents produced from plant endophytic actinobacteria

Abstract

Dairy production is threatened by antibiotic resistant pathogens worldwide, and alternative solutions to treat mastitis are not available. The prevalence of antibiotic resistant strains is not well known in less developed countries. The prevalence of pathogenic bacteria and their resistance to 21 commercial antibiotics were studied in milk samples taken from 122 dairy cows suffering from the symptoms of mastitis in Egypt. The bacterial species were identified with molecular methods, and antibiotic resistance was studied with disc diffusion method. The prevalence of Streptococcus aureus, Escherichia coli and Pseudomonas aeruginosa were 30%, 17% and 3.5%, respectively. Most (90%) of the S. aureus strains showed resistance to penicillin whereas only 10% of the strains were resistant to oxacillin. Nearly half (40%) of E. coli strains showed resistance to streptomycin. Six P. aeruginosa strains showed resistance to several antibiotics, including ceftriaxone, enrofloxacin and levofloxacin. This points out that despite P. aeruginosa was not common, it should be followed up carefully. Potential biocontrol agents against antibiotic resistant mastitis bacteria were searched among 30 endophytic actinobacterial strains derived from wild medicinal plants. Three plants, namely Mentha longifolia, Malva parviflora and Pulicaria undulata were chosen for a more detailed study; their endophytic actinobacteria were used to prepare metabolic extracts. The crude metabolites of the actinobacteria were extracted with ethyl acetate. All metabolic extracts inhibited the growth of S. aureus, methicillin-resistant Staphylococcus aureus (MRSA), E. coli and P. aeruginosa in vitro. The 16S rRNA sequence analysis revealed that the most efficient actinobacterial strains were two Micromonospora sp. and one Actinobacteria bacterium. We conclude that the combination of the metabolites of several endophytic actinobacteria derived from several medicinal plants would be the most efficient against pathogens. Different metabolite cocktails should be studied further in order to develop novel biocontrol agents to treat antibiotic resistant mastitis bacteria in dairy cows.

Keywords: Antibiotics; Biological control; Cattle; Drug resistance; Milk; Pathogenic bacteria; Traditional medicine.

Fig. 1

Antibiogram of pathogenic bacteria S. aureus (A) P. aeruginosa (B) and E. coli (C) isolated from milk samples. Bacteria is divided into resistant, intermediate and sensitive groups according to the sensitivity to antibiotics.

Fig. 2

Agarose gel electrophoresis of the amplified mecA gene PCR product (310 bp) for the five methicillin-resistant S. aureus (MRSA).

Fig. 3

Agarose gel electrophoresis of the amplified phoA gene PCR product (720 bp) for pathogenic E. coli.

Fig. 4

Phylogenetic tree of ITS rDNA sequences of the endophytic actinobacteria used to prepare metabolic extracts against pathogenic bacteria causing mastitis in dairy cows. The tree was constructed by the neighbor-joining algorithm using the maximum composite likelihood model. Bootstrap percentages from 1000 replicates are shown.