Phage vB_Tsc2631 infects the extremophilic bacterium Thermus scotoductus MAT2631 and uses the Ts2631 endolysin for the release of its progeny. The Ts2631 endolysin is the first endolysin from thermophilic bacteriophage with an experimentally validated catalytic site. In silico analysis and computational modelling of the Ts2631 endolysin structure revealed a conserved Zn2+ binding site (His30, Tyr58, His131 and Cys139) similar to Zn2+ binding site of eukaryotic peptidoglycan recognition proteins (PGRPs). We have shown that the Ts2631 endolysin lytic activity is dependent on divalent metal ions (Zn2+ and Ca2+). The Ts2631 endolysin substitution variants H30N, Y58F, H131N and C139S dramatically lost their antimicrobial activity, providing evidence for the role of the aforementioned residues in the lytic activity of the enzyme. The enzyme has proven to be not only thermoresistant, retaining 64.8% of its initial activity after 2 h at 95°C, but also highly thermodynamically stable (Tm = 99.82°C, ΔHcal = 4.58 × 104 cal mol-1). Substitutions of histidine residues (H30N and H131N) and a cysteine residue (C139S) resulted in variants aggregating at temperatures ≥75°C, indicating a significant role of these residues in enzyme thermostability. The substrate spectrum of the Ts2631 endolysin included extremophiles of the genus Thermus but also Gram-negative mesophiles, such as Escherichia coli, Salmonella panama, Pseudomonas fluorescens and Serratia marcescens. The broad substrate spectrum and high thermostability of this endolysin makes it a good candidate for use as an antimicrobial agent to combat Gram-negative pathogens.
Amoebic gill disease (AGD) is a pathogenic disease in salmonids caused by Neoparamoeba perurans. Treatment of AGD infection has been through freshwater bathing of the fish. However, as the availability of fresh water is often limited, hydrogen peroxide has been introduced as an alternative treatment. This study investigated the effect of hydrogen peroxide as treatment for AGD-infected salmon (Salmo salar L.,) at different seawater temperatures and hydrogen peroxide dosages. In total, 600 fish were challenged with N. perurans and the severity of the AGD infection was measured using a gill score scale. After challenge and disease development, the fish were distributed into 12 tanks. The treatment was performed at different seawater temperatures (8°C, 12°C, 17°C) using different hydrogen peroxide doses. Each temperature included an untreated control group. Linear models were used to analyse gill score. A significant effect of treatment was found (À0.68 AE 0.05) regardless of dose and temperature, suggesting that hydrogen peroxide was effective in treating AGD. When the model included dose, a negative linear relationship between dose and gill score was found. The study proved that treatment of AGD with hydrogen peroxide was successful, as gills partially recovered following treatment and further disease development was delayed.
Dietary fish oil is believed to have a beneficial effect in various infections and in autoimmune disorders. This effect may correspond to an altered immune response. In order to discover whether the effect of dietary fish oil is different in various infections, we studied the survival of mice fed fish oil or corn oil supplemented diets and infected in the lungs with either Klebsiella pneumoniae or Streptococcus pneumoniae. 120 NMRI mice were divided into 4 groups, of which 2 groups were fed a fish oil supplemented diet and 2 a corn oil supplemented diet. After 6 weeks the mice were infected in the lungs with Klebsiella pneumoniae (fish oil groups and corn oil groups) or with Streptococcus pneumoniae serotype 3 (both groups). The survival rate was monitored. The experiment was performed twice. The survival of the mice fed fish oil enriched diet and infected with Klebsiella pneumoniae was significantly better compared with the mice fed corn oil enriched diet (p = 0.0001 and p = 0.0013). No difference was found between the mice fed corn oil enriched diet or fish oil enriched diet and infected with Streptococcus pneumoniae serotype 3 (p = 0.74 and p = 0.15). Our results indicate that dietary fish oil has a beneficial effect on survival of mice after experimental pneumoniae when infected with Klebsiella pneumoniae, but not after infection with Streptococcus pneumoniae serotype 3.
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