Characterization of Endolysin LyJH307 with Antimicrobial Activity against Streptococcus bovis

Abstract

Streptococcus bovis (S. bovis) is one of the critical initiators of acute acidosis in ruminants. Therefore, we aimed to develop and characterize the endolysin LyJH307, which can lyse ruminal S. bovis. We tested the bactericidal activity of recombinant LyJH307 against S. bovis JB1 under a range of pH, temperature, NaCl, and metal ion concentrations. In silico analyses showed that LyJH307 has a modular design with a distinct, enzymatically active domain of the NLPC/P60 superfamily at the N-terminal and a cell wall binding domain of the Zoocin A target recognition domain (Zoocin A_TRD) superfamily at the C-terminal. The lytic activity of LyJH307 against S. bovis JB1 was the highest at pH 5.5, and relatively higher under acidic, than under alkaline conditions. LyJH307 activity was also the highest at 39 °C, but was maintained between 25°C and 55°C. LyJH307 bactericidal action was retained under 0-500 mM NaCl. While the activity of LyJH307 significantly decreased on treatment with ethylenediaminetetraacetic acid (EDTA), it was only restored with supplementation of 10 mM Ca2+. Analyses of antimicrobial spectra showed that LyJH307 lysed Lancefield groups D (S. bovis group and Enterococcus faecalis) and H (S. sanguinis) bacteria. Thus, LyJH307 might help to prevent acute ruminal acidosis.

Specific inhibition of Streptococcus bovis by endolysin LyJH307 supplementation shifts the rumen microbiota and metabolic pathways related to carbohydrate metabolism

“…Therefore, we hypothesized that the supplementation time is important to induce adequate lytic activity in an in vitro system. In the present study, endolysin LyJH307 supplementation at 6 h incubation time markedly decreased the absolute abundance of S. bovis (approximately 70%), and the reduction rate of S. bovis in the present study was similar to previous results presenting reduction rates of the S. bovis group (S. equinus, S. gallolyticus subspecies pasteurianus, S. alactolyticus, and S. infantarius subspecies infantarius) in single cultures [11]. Therefore, the present dosage of endolysin LyJH307 was appropriate for the control of S. bovis under ruminal mixed culture conditions.…”

“…Recombinant LyJH307 was cloned in Escherichia coli BL21 (DE3), and the effect of lytic activity on the S. bovis group was tested following the methodology described by Kim, Lee, Kwon (11). To produce the recombinant endolysin LyJH307, the transformant was grown in Luria-Bertani medium (Difco Laboratories Inc.) until the optical density at 600 nm (OD 600nm ) reached 0.4.…”

“…The number of studies related to endolysin development has been improved by the emergence of multi-drug resistant bacteria, which render many types of antibiotics ineffective. Many researchers have shown that endolysin can lyse gram-positive bacteria when endolysin is administered externally because of the absence of an outer membrane such as a lipopolysaccharide layer [11][12][13]. Therefore, endolysin is a promising replacement for antibiotics.…”

“…In a previous study, we developed endolysin LyJH307 with high lytic activity against S. bovis [11]. Endolysin LyJH307 has a distinct combination of conserved domain consisting of N-terminal NlpC/P60 superfamily (EAD) and C-terminal Zoocin A target recognition domain superfamily (CBD), and the endolysin LyJH307 had potent lytic activity in a variety of condition (pH, 5.0−6.0; temperature, 25−55℃;…”

Diversity and Antimicrobial Resistance in the Streptococcus bovis/Streptococcus equinus Complex (SBSEC) Isolated from Korean Domestic Ruminants

“…Although S. bovis was synonymized with S. equinus and is currently recognized as the SBSEC, the name is still commonly used in ruminant livestock research [ 49 , 50 , 51 ]. However, researchers in the area have already recognized the physiological and genetic diversities of bovine-origin S. bovis isolates [ 20 , 21 ] and genetic heterogeneity in SBSEC has been described recently, especially in S. equinus isolates [ 22 ].…”