Letters in Applied Microbiology 1999, 28, 461–465 Effect of higher pasteurization temperatures, and longer holding times at 72°C, on the inactivation of Mycobacterium paratuberculosis in milk I.R. Grant, 1 H.J. Ball 2 and M.T. Rowe 1 1 Department of Food Science (Food Microbiology), The Queen’s University of Belfast, and 2 Veterinary Sciences Division, Department of Agriculture for Northern Ireland, Belfast, UK 2031/98: received and accepted 27 January 1999 I.R. GRANT, H.J. BALL AND M.T. ROWE. 1999. Raw cow’s milk spiked with 10 6 cfu ml -1 of Mycobacterium paratuberculosis was subjected to heat treatments of 72, 75, 78, 80, 85 or 90 °C for 15 s, and 72 °C for 20 and 25 s, using laboratory pasteurizing units. Three bovine strains of Myco. paratuberculosis were studied (NCTC 8578, B2 and DVL 943). Each strain was subjected to all the heat treatments indicated on three separate occasions. Although each of the heat treatments achieved a substantial (5–6 log 10 ) reduction in numbers of viable Myco. paratuberculosis, small numbers of the organism (4–16 cfu 10 ml -1 ) survived in a proportion of the milk samples at each of the higher temperatures investigated, right up to 90 °C for 15 s. A longer holding time of 25 s at 72 °C was found to be more effective at inactivating Myco. paratuberculosis. Only one of the three strains studied, B2, yielded small numbers of survivors after heating at 72 °C for 20 s, but it was completely inactivated by extending the holding time at 72 °C by a further 5 s to 25 s. It was concluded that a longer holding time is more likely to achieve the complete inactivation of Myco. paratuberculosis in milk than a higher pasteurization temperature. INTRODUCTION Mycobacterium paratuberculosis causes paratuberculosis, com- monly known as Johne’s disease (JD), in cattle, sheep, goats and other ruminants (Cocito et al. 1994). Although not cur- rently classified as a zoonotic agent, it has been suggested that Myco. paratuberculosis may also be implicated in an intestinal disease in humans known as Crohn’s disease (Chiodini 1989). Milk has been suggested as a possible vehicle of transmission of the organism from cattle to humans (Hermon-Taylor 1993; Thompson 1994). Little is known about the levels of Myco. paratuberculosis that may be naturally present in infected milk from cattle with Johne’s disease. Three studies reported the isolation of Myco. paratuberculosis from milk aseptically obtained from cows with asymptomatic (Sweeney et al. 1992; Streeter et al. 1995) and clinical (Taylor et al. 1981) Johne’s disease. One of these studies (Sweeney et al. 1992) reported a Myco. paratuberculosis titre of 2–8 cfu 50 ml -1 of milk from an asymptomatic animal. However, as the udders were Correspondence to: Dr I. Grant, Department of Food Science (Food Microbiology), The Queen’s University of Belfast, Newforge Lane, Belfast BT9 5PX, UK (e-mail: I.Grant@qub.ac.uk). © 1999 The Society for Applied Microbiology thoroughly decontaminated prior to milk collection, none of these studies took any account of the contribution made by faecal contamination of the milk during normal milking procedures. Previous research carried out in this laboratory showed that Myco. paratuberculosis in milk may not be completely inactivated by current high-temperature, short-time (HTST) pas- teurization conditions (72 °C/15 s) unless present at levels of ³ 100 cfu ml -1 (Grant et al. 1996a, 1998). These findings were recently corroborated by D value data for Myco. paratuberculosis in milk published by Sung and Collins (1998). These studies suggest that pasteurization at 72 °C for 15 s may only achieve the inactivation of around 10 2 cfu Myco. paratuberculosis ml -1 of milk. The question of what heat treatment would completely inactivate Myco. paratuberculosis in milk was addressed in the present study. The effect of higher heating temperatures (75, 78, 80, 85 and 90 °C) for 15 s, and longer holding times (20 and 25 s) at the current pasteurization temperature (72 °C), on the thermal inactivation of Myco. paratuberculosis in milk was inves- tigated. The aim was to inactivate 10 6 cfu ml -1 of three of the most heat-resistant strains of Myco. paratuberculosis identified in the course of earlier studies (Grant et al. 1996a, 1998).