Occurrence of Emphasis Type="Italic"Clostridium

Authors: B Turchi S Pero B Torracca F Fratini S Mancini A Galiero F Pedonese R Nuvoloni D Cerri

Publish Date: 2016/07/20

Volume: 96, Issue: 5, Pages: 693-701

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Abstract

The presence of gasproducing clostridia in ewe’s milk can lead to the occurrence of lateblowing defects in cheeses However data on this aspect are limited In the present study using the most probable number MPN method clostridial spores were enumerated in 527 ewe’s milk samples collected in the Grosseto Province Tuscany Italy from autumn 2014 to summer 2015 In addition using polymerase chain reaction PCR we identified the species most frequently involved in lateblowing defects in cheese Clostridium tyrobutyricum Clostridium butyricum Clostridium beijerinckii and Clostridium sporogenes and of Clostridium perfringens Gasproducing clostridial spores were detected in 99 of samples Spore concentrations ranged from 360 to more than 110000 spores∙L−1 We observed that 86 of samples had a spore load higher than 1000 spores∙L−1 During autumn and summer spore concentrations were significantly higher than in winter and spring P  0001 A total of 222 isolates obtained from 77 MPN positive tubes from different milk samples were subjected to PCR Colonies from 63/77 82 MPN positive tubes were taxonomically identified Among the 63 PCRpositive samples C perfringens was the most frequently detected species 56 followed by C sporogenes 44 C tyrobutyricum 79 C butyricum 16 and C beijerinkii 16 In addition in 11 of the MPN positive tubes at least two clostridial species were found to be present simultaneously This work highlights the presence of clostridial spores in ovine milk from central Italy Tuscany and led to the identification of some of the clostridia species involved in such high spore loadsThe presence of clostridial spores in ewe’s milk could represent an emerging problem due to several factors related to climate and herd management changes In particular the increasing number of wolves Canis lupus in central Italy Bassi et al 2015b Vitali 2014 has led to an increase in semiextensive farming with an increased use of fenced pastures and housing facilities which promote the spread and concentration of environmental spores especially when the bedding is not managed appropriately In this regard in some areas of Spain and France a ewe’s milk quality payment system has been adopted in order to promote farm management practices that are effective in reducing the clostridial spores load of the milk A maximum limit of 1000–1300 spores∙L−1 has been established Pirisi et al 2007Despite the fact that not only bovine milk cheeses but also ewe’s milk cheeses can be affected by lateblowing defects GómezTorres et al 2015 little is known regarding the occurrence of clostridial spores in ewe’s milk Indeed the current literature regarding the presence and relevance of clostridia spores in Italian ewe’s milk is limited Scintu et al 2004 However a few surveys carried out in Spain are available Garde et al 2011 Arias et al 2013 GómezTorres et al 2015 Concerning data from reports analyzing milk samples for the production of Manchego cheese in Spain Garde et al 2011 detected lactatefermenting clostridial spores in 97 of samples Despite these data these authors also reported a low annual incidence 028 of lateblowing defects in Manchego cheese and ascribed this result to the use of lysozyme in cheese manufacturing In fact lysozyme can reduce lateblowing especially when the spore concentration is lower than 500 L−1 However some drawbacks have been reported such as its allergenicity and the variable degree of susceptibility of the spores Brändle et al 2016 In addition Garde et al 2011 observed the highest mean milk spore count in the summer 26800 spores∙L−1 with C sporogenes as the most abundant species in cheeses with lateblowing defects This is in accordance with Arias et al 2013 who also found that C sporogenes was the most commonly isolated species followed by C beijerinkii and C tyrobutyricum Arias et al 2013 also highlighted that the use of farmmade forage and wet brewer grain as animal feed as well as the presence of dust in the milking parlor favors the presence of Clostridium spp spores in milkRecent studies have also highlighted the presence of Clostridium perfringens in raw milk and curds Chaturvedi and Shukla 2015 Feligini et al 2014 Since this species is able to produce gas from the metabolism of carbohydrates Pigato et al 2012 this could have an impact on the enumeration of gasproducing clostridia using the MPN methodOur aim was to enumerate gasproducing clostridial spores in ewe’s milk samples collected in the province of Grosseto Tuscany Italy by the MPN method An additional aim was to evaluate by PCR the presence of some of the clostridial species most frequently involved in the occurrence of lateblowing defects in cheeses C beijerinckii C butyricum C sporogenes and C tyrobutyricum and of C perfringens as well

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