Effect of Calcium in Assay Medium on D Value of Bacillus stearothermophilus ATCC 7953 Spores

Abstract

The D value of commercial biological indicator spore strips using Bacillus stearothermophilus ATCC 7953 was increased by higher calcium concentrations in assay media. The calcium concentration in assay media varied among the manufacturers. The calcium concentration in assay media is an important factor to consider to minimize the variation of D value.

Importance of Considering Injured Microorganisms in Sterilization Validation

“…The role of medium constituents in the recovery of injured spores has been discussed (Blocher and Busta, 1982;Shintani, 2006;Sasaki et al, 2000). They described the effects of inhibitors on recovery and the modified metabolic requirements of injured spores.…”

“…Injured spore formers have significant sensitivity to sodium chloride (Feehery et al, 1987), nitrate and nitrite (Chumney and Adams, 1980), surface-active agents including antibiotics (Chumney and Adams, 1980), pH (Feehery et al, 1987), water activity and osmolarity (Mattick et al, 2001), gas atmosphere (Feehery et al, 1987), culture media, media brand and their lots (Pflug et al, 1979;Pflug et al, 1981;Sasaki et al, 2000;Shintani and Akers, 2000;, and incubation temperature and period (Feehery et al, 1987;. The type and extent of these sensitivities vary with the species and the type of injury.…”

“…Recovery is influenced not only by the kind and constituents of the culture medium, but also by the condition of the medium (Pflug et al, 1981;Pflug et al, 1979), the quality of the constituents (Johnson and Busta, 1984;Sasaki et al, 2000), and the brand and the lot of the medium (Pflug et al, 1981;Pflug et al, 1979;Sasaki et al, 2000). Different lots of soybean casein digest, even from the same manufacturer, resulted in different rates of recovery of heated G. stearothermophilus spores (Pflug et al, 1981;Sasaki et al, 2000;Shintani and Akers, 2000;Shintani, 2006). This lot to lot variation TABLE 4.…”

“…The culture medium used to detect injured spore formers will influence their recovery (Johnson and Busta, 1984;Pflug et al, 1979;Wallen and Walker, 1979;Sasaki et al, 2000;Shintani and Akers, 2000;Shintani, 2006). Several media have been used for the recovery of injured spores (Table 4).…”

“…Damage to germination systems by heat has often been characterized by a need for specific germinants or enzymes in the medium (Foegeding and Busta, 1981;Gould, 1984). For example, Ca, glucose or alanine are considered as germinants and thus inclusion of them into the culture medium is indispensable to attain reproducible and successful sterilization validation (Shintani, 2006;Sasaki et al, 2000). Damage to the germination system has been demonstrated with the use of calcium dipicolinate (Edwards et al, 1965a,b), lactate (Foegeding and Busta, 1983a), lysozyme Adams and Busta, 1972;Barach et al, 1974;Adams, 1974;Duncan et al, 1972;Alderton et al, 1974), and mixtures of amino acids (Uchida and Kadota, 1979;Gurney and Quesnel, 1981;Gurney and Quesnel, 1980) and so on.…”