European Journal of Biomedical and Pharmaceutical Sciences

SYNERGISTIC EFFECT OF COLOSTRUMS LACTOFERRIN AND PURIFIED CHARACTERIZED BREVICIN ON ESBL PRODUCING PANTOEA AGGLOMERANS ISOLATED FROM BLOOD CULTURES OF INFANTS IN BAGHDAD.

Likaa Hamied Mahdi Al Musawii*, Nadheema Hammood Hussein and Buthainah Mohammed Taha

ABSTRACT

In the present study, seven Lactobacillus brevis (L. brevis) strains isolated from human breast colostrums identified by API 50WERE USED. To evaluate the potential use of these strains as probiotics, the effect of gastric juice, pH and bile salts tolerance were conducted. Cell tolerancey to acidity and bile salt are important factor that affect the probiotics to remain and exert their potential functionalities in a host. Brevicin produced from L. brevis strain no.5 purified by ammonium sulfate precipitation, Sep-pack C18 cartridge and reverse-phase HPLC chromatography on C18 Nucleosil column and characterized. Lactoferrin had been isolated from human colostrums purified by gel filtration and ion-exchange chromatography. Antibacterial activity of crude and purified brevicin against sixteen Pantoea agglomerans strains was determined moreover synergestic effect of lactoferrin and brevicin against these strains was conducted. All L. brevis strains showed acid resistance which survive at pH 3 and 2.5, bile salt concentration until 0.5% and all the strains had the ability of aggregation. Brevicin was considered to be heat stable in different temperature degrees and also it was stable at pH 6 and 7 which gave 100% activity. Brevicin was insensitive to lipase and α-amylase but sensitive to pronase E, pepsine and proteinase K, the chromatogram by HPLC showed a single symmetric peak by absorbance at 280 nm and the specific activity of the purified brevicin was increased 1.68*104 fold with a final yield of 0.4%. Also the molecular weight of purified brevicin by gel filteration was estimated at approximately (3.8) K Daltons. Our results showed that Pantoea agglomerans strains were had high resistant rates for most antibiotics under study and the most effective antibiotics against these bacteria were imipenem and netilmicin. Also 12(75%) of these bacteria were positive ESBL producing strains by screening test. Out of 12 positive ESBL producing strains, 8(66.67%) harboring blaCTX-M gene by genotypic detection while all strains don’t harboring blaSHV gene. Detection of ESBL bacteria is importance for infection control and prevention of dissemination of ESBL producers by cross-transmission and for epidemiological purposes. Also increasing in number of ESBL resistant bacteria have been reported worldwide and alternative compounds should be used against these bacteria. Our results showed that the purified brevicin was more effective than crud brevicin in all concentrations (p≤ 0.05) and there was a synergic antimicrobial effect of brevicin and lactoferrin when they were tested together, even in the concentration of 30 μg/mL brevicin and 30 μg/mL of lactoferrin (the highest values evaluated). Pantoea agglomerans strains were affected by lactoferrin and significantly more affected by the synergic use of both proteins (brevicin and lactoferrin) (P<0.05).

Keywords: Lactobacillus brevis, brevicin, lactoferrin, Pantoea agglomerans, ESBL producing bacteria and antimicrobial activity.