Tuberculocidal activity of a peroxymonosulphate-based disinfectant-detergent (Virkon)

Recently, even in Europe and North America, there has been a marked increase in the cases of tuberculosis due to various factors including immigration from third-world areas, poverty, promiscuity and, last but not least, lowered immune system defenses in HIV patients. Besides Mycobacterium tubercolosis, other non tubercolosis mycobacteria (NTM) such as Mycobacterium avium, chelonae and abscessum are held responsible for human pathologies. In compliance with EN 14348, a test suspension (1.5 x 109 –5 x 109 UFC/ml) of Mycobacterium terrae CIP104321 (N) was initially prepared. The validation suspension (NV) was obtained by diluting the test suspension (N) until a concentration of 3 x 102 –1.6 x 103 cfu/ml was obtained. Further dilution of the NV yielded a concentration of countable cfu after seeding (NV0). An additional validation of the experimental conditions was also required, e.g. verification that the test conditions presented no lethal effects (A), verification that the neutralizing agent showed no signs of toxicity (B) and validation of the neutralization dilution method (C). To evaluate the tuberculocide capacity of the product in question, a diluted solution of the interfering substance (0.3 g/l of fraction V of cow albumina to simulate “clean” conditions; 3 g/l of fraction V of cow albumina and 3 ml/l of defibrinate erythrocytes to simulate “dirty” conditions) was added to the initial suspension of Mycobacterium terrae and this was then added to a sample of the Virkon diluted with hard water at a concentration that varied from 0.25 to 4% at a temperature of 20 or 40 ˚C for 60 minutes. It was then neutralized with sodium thosulphate (Na). Aliquots of N, NV, N0, NV0, Na and A, B, C ― suitably diluted to contain a countable number of cfu ― were sown on plates containing a Middlebrooke and Cohn medium agarized with 10% OADC enrichment and incubated at 36 ˚C  1 ˚C for 21 days. After this period the cfu developed were counted. For each experimental series the basic limit was always checked and corresponded to the following values: a) N between 1.5x109 cfu/ml and 5.0x109 cfu/ml (9.17≤lgN0≤9.70) and N0 between 1.5x108 cfu/ml and 5.0x108 cfu/ml (8.17≤lgN0≤8.70) b) NV0 between 30 and 160 cfu/ml (3.0 x101 and 1.6 x102) and NV between 3.0x102 and 1.6x103 cfu/ml c) A. B. C were equal to or greater than 0.5x NV0 The tuberculocide activity was deemed proven when there was a reduction in the living mycobacteria contained in the test suspension at the end of the contact time established and after neutralization (Na) of at least 4 log. Our results show that, at a temperature of 40°C and for a contact time of 60 minutes, at 0.25%, 0.5%, 1% and 2% concentrations in hard water, the product tested showed tuberculocide activity under clean conditions since it already yielded a 4.71 log reduction at a concentration of 0.5% and 4.70 log at 1% while, under dirty conditions, it gave a >5.79 log reduction for the 2% concentration. Moreover, it must be pointed out that, under clean conditions, at a the concentration of 0.25% the reduction was 3.75 log while under dirty conditions, concentrations of 0.25%, 0.5% and 1% all showed reductions of less than 4 log, thus meeting the standards which require that at least one of the concentrations tested present a concentration lower than 4 log and at least one concentration present a reduction greater than 4 log. Therefore, if the tuberculocide activity of Virkon is clearly proven, in agreement with EN14348, a limit to its use is given by the temperature of 40°C, the only temperature that ensures safe biocide activity vs. mycobacteria. Hence it is not possible to envisage its being used to disinfect surfaces contaminated by Micobacterium tuberculosis or NTM that are normally disinfected at room temperature. This product could instead be profitably used to disinfect surgical and endoscopic instruments that come into contact with Mycobacterium tuberculosis or with non tubercular mycobacteria.