Optimum Culture Conditions for Production of Exfoliative Toxin by Staphylococcus hyicus

Optimum culture conditions for the production of exfoliative toxin by Staphylococcus hyicus (shET) were examined. High shET activity was obtained from the culture filtrate of HI and TY broth inoculated with S. hyicus. The pH in these two media ranged from 7 to 8.5 during bacterial culture, while the lowest pH in TS and BHI broth was less than 6. shET activity in the culture filtrate from TY broth inoculated with 10⁷ CFU of S. hyicus per ml was higher than that in TY broth inoculated with 10⁶ and 10⁸ CFU of bacteria per ml. When shET activity in the culture filtrate was measured under various shaking conditions, the culture filtrate shaken at 75 oscillations per min had the highest shET activity of the five shaking conditions. shET activity of the culture filtrate of TY broth to which protease inhibitor had been added was the same as that of TY broth without inhibitor. shET activity in a shaking culture in an Erlenmeyer flask was also the same as that in sac culture and that in shaking culture using a shaking (Sakaguchi) flask. shET activity in TY broth supplemented with 100 mM glucose was significantly lower than that in TY broth without glucose. Based on the above results, the optimum culture conditions for the production of shET were as follows: inoculation of 3 × 10⁹ CFU of S. hyicus strain P-1 into 300 ml of TY broth in a 2,000-ml Erlenmeyer flask, and incubation at 37 C with shaking at 75 oscillations per min. Then shET activity of the culture filtrate under appropriate culture conditions was measured after various incubation periods. shET activity was detected 6 hr after inoculation, reached the maximum (2⁵³ exfoliative unit/0.1 ml) at 16 hr and decreased between 20 and 48 hr. Thus, the optimum incubation period was determined to be 16 hr. Then the optimum concentration of ammonium sulfate for isolation of shET from the culture filtrate under appropriate culture conditions was examined. The greatest shET activity was obtained from the fraction salted out with 90% saturated ammonium sulfate. Thus, the optimum concentration of ammonium sulfate for the isolation of shET was determined to be 90% saturation.     

The diet and distribution of Pentacoelum kazukolinda (Kawakatsu & Mitchell, 1984), a maricolan planarian with a freshwater ecology

Among the marine triclads or Maricola, the genus Pentacoelum represents one of the few exceptions in that species occur in brackish water or in freshwater. We examined specimens of a maricolan triclad that externally looked remarkably similar to the continental Spanish representatives of freshwater Pentacoelum hispaniense, albeit that these new samples came from habitats in Germany, the Netherlands, Mallorca and Japan. Comprehensive comparative morphological studies revealed that the animals from these populations are identical to a species described earlier from Hawaii, viz. Oahuhawaiiana kazukolinda Kawakatsu & Mitchell, 1984. It is argued that the species now should be called Pentacoelum kazukolinda (Kawakatsu & Mitchell, 1984) comb. nov. Remarkably, Dutch and German specimens of P. kazukolinda almost completely lack any signs of the presence of the lateral gonopore system, being the defining feature of their taxonomic family, the Bdellouridae. The reason for this reduction or even complete absence of this structure remains obscure. Nonetheless, morphological similarities and 18S rDNA sequences strongly suggest that all animals belong to the same species. Analyses of the mitochondrial gene sequences Cox1 and 16SrDNA of the gut content of P. kazukolinda revealed that the flatworm has a preference for different species of gastropods, while in one population annelids were detected. Our study suggests that gastropods may form the food refuge of P. kazukolinda. Circumstantial evidence suggests that the worldwide distribution of P. kazukolinda may result from human‐mediated transport of the aroid Colocasia esculenta and/or the introduced snail Potamopyrgus antipodarum. The use of P. antipodarum as a food resource may have facilitated the spread and establishment of new populations of P. kazukolinda.