Molecular characterization and in vitro- Phytochemical Evaluation of two plants extracts against Multidrug-Resistant and Extended Spectrum Beta Lactamase Producing Salmonella Isolated from birds

This study aims to know the molecular characterization of multidrug-resistant and extended spectrum β-Lactamase Producing Salmonella isolated from birds (chicken feaces) as well as test their susceptibility to two plants extracts namely Azadirachta indica and Ocimum gratissimum. A total of 55 Salmonella enterica isolates were recovered from chicken feaces. Salmonella enterica was identified and characterized using standard methods in the microbiology laboratory. Morphological and biochemical analysis were carried out to identify Salmonella enterica. 16S sRNA sequencing was carried out to also confirm the identification of Salmonella enterica. The Phenotypic and molecular determinants for the production of extended-spectrum β-lactamase was done using the double-disk synergy test and multiplex polymerase chain reaction respectively. Kirby–Bauer disc diffusion method was used to test the antibacterial susceptibility pattern against eight known antibiotics. Phytochemicals were determined by soxhlex extraction method. Aqueous, ethanol and methanol were the solvents used for extraction. Antibacterial activities of Azadirachta indica and Ocimum gratissimum were determined by agar well diffusion assay. The minimum inhibitory concentration of the plant extracts on Salmonella enterica was determined by the agar dilution method. The study showed the presence of Salmonella enterica as well as the presence of extended-spectrum β-lactamase genes responsible for varying resistance of Salmonella enterica to Azadirachta indica and Ocimum gratissimum extracts. Antibiotics were sensitive to Salmonella enterica while some were resistant to Salmonella enterica. The Antibiotic susceptibility test of Salmonella enterica isolated from chicken feaces observed that Aztreonam showed highest (54%) resistance while Cefixime (20%) showed the least resistance. The Phtyochemicals identified were alkaloids, flavonoids, phenols, saponin, steroids and tannin, showing that the plant extracts contains active secondary metabolites that constitutes to the antimicrobial efficacy of plant extracts.  Ocimum gratissimum gave more antibacterial activity of Leave extracts (24mm), followed by roots (22mm) then the least antibacterial activity was the stem (20mm) at  a concentration of 200mg/ml compared to the antibacterial activity of  Azadirachta indica Leave extracts (15mm), followed by roots (13mm) then the least antibacterial activity was the stem (10mm) at also a  concentration of 200mg/ml. The zone of inhibition for Salmonella enterica on the different extracts of Azadirachta indica was between 1mm to 15mm while the zone of inhibition for Salmonella enterica on the different extracts of Ocimum gratissimum was between 2mm to 24mm. The observed antibacterial effects on Salmonella enterica can be as a result of the presence of phytochemicals. This confirms that Azadirachta indica and Ocimum gratissimum have some degree of antibacterial effect and should be used to treat Salmonella enterica infections. There is need to study plants extracts inorder to develop a good bio-therapeutic agents originating from plants.

Keywords: Antibiotics, Antibacterial susceptibility Azadirachta indica, Chicken feaces. Extended-spectrum β-lactamase, Phytochemicals, Salmonella enterica and Ocimum gratissimum.