Department of Molecular Biology and Biophysics, UConn Health, Farmington, CT, USA.
Department of Microbiology and Immunology, University of Iowa, Iowa City, IA, USA.
Lett Appl Microbiol. 2020 Jun;70(6):407-412. doi: 10.1111/lam.13290. Epub 2020 Mar 26.
Inactivation of Bacillales and Clostridiales spores is of interest, since some cause food spoilage and human diseases. A recent publication (mSphere 3: e00597-1, 2018) reported that glycerol monolaurate (GML) in a non-aqueous gel (GMLg) effectively killed spores of Bacillus subtilis, Bacillus cereus and Clostridioides difficile, and Bacillus anthracis spores to a lesser extent. We now show that (i) the B. subtilis spores prepared as in the prior work were impure; (ii) if spore viability was measured by diluting spores 1/10 in GMLg, serially diluting incubations 10-fold and spotting aliquots on recovery plates, there was no colony formation from the 1/10 to 1/1000 dilutions due to GMLg carryover, although thorough ethanol washes of incubated spores eliminated this problem and (iii) GMLg did not kill highly purified spores of B. subtilis, B. cereus, Bacillus megaterium and C. difficile in 3-20 h in the conditions used in the recent publication. GMLg also gave no killing of crude B. subtilis spores prepared as in the recent publication in 5 h but gave ~1·5 log killing at 24 h. Thus, GMLg does not appear to be an effective sporicide, although the gel likely inhibits spore germination and could kill spores somewhat upon long incubations. SIGNIFICANCE AND IMPACT OF THE STUDY: Given potential deleterious effects of spores of Bacillales and Clostridiales, there is an ongoing interest in new ways of spore killing. A recent paper (mSphere 3: e00597-1, 2018) reported that glycerol monolaurate (GML) in a non-aqueous gel (GMLg) effectively killed spores of many species. We now find that (i) the Bacillus subtilis spores prepared as in the previous report were impure and (ii) GMLg gave no killing of purified spores of Bacillales and Clostridiales species in ≤5 h under the published conditions. Thus, GMLg is not an effective sporicide, though may prevent spore germination or kill germinated spores.
芽孢杆菌目和梭菌目的孢子失活很有研究意义，因为其中一些会导致食物变质和人类疾病。最近的一篇出版物（mSphere 3: e00597-1, 2018）报道称，甘油单月桂酸酯（GML）在非水凝胶（GMLg）中可以有效地杀死枯草芽孢杆菌、蜡状芽孢杆菌和艰难梭菌的孢子，而炭疽芽孢杆菌的孢子则较少受到影响。我们现在发现：（i）如前一篇报告所述制备的枯草芽孢杆菌孢子不纯；（ii）如果通过将孢子在 GMLg 中稀释 1/10 来测量孢子活力，连续将孵育物稀释 10 倍，并将等分试样点在回收平板上，则由于 GMLg 的携带污染，从 1/10 到 1/1000 的稀释液中都没有菌落形成，尽管对孵育孢子进行彻底的乙醇清洗可以消除这个问题；（iii）在最近的出版物中使用的条件下，GMLg 在 3-20 小时内没有杀死高度纯化的枯草芽孢杆菌、蜡状芽孢杆菌、巨大芽孢杆菌和艰难梭菌的孢子。GMLg 在 5 小时内也没有杀死如最近的出版物中制备的粗糙的枯草芽孢杆菌孢子，但在 24 小时时给出了约 1.5 个对数的杀灭。因此，GMLg 似乎不是一种有效的杀孢子剂，尽管凝胶可能抑制孢子萌发，并在长时间孵育时可能会杀死一些孢子。
鉴于芽孢杆菌目和梭菌目的孢子可能产生有害影响，人们一直对新的孢子杀灭方法感兴趣。最近的一篇论文（mSphere 3: e00597-1, 2018）报道称，甘油单月桂酸酯（GML）在非水凝胶（GMLg）中可以有效地杀死许多种孢子。我们现在发现：（i）如前一篇报告所述制备的枯草芽孢杆菌孢子不纯；（ii）在已发表的条件下，GMLg 在 ≤5 小时内没有杀死芽孢杆菌目和梭菌目的纯孢子。因此，GMLg 不是一种有效的杀孢子剂，尽管它可能阻止孢子萌发或杀死已萌发的孢子。
