The application of flow cytometry in determining the bacteriological quality of raw sheep’s milk in Slovakia

Abstract

-In this study enumeration of bacterial cells in raw sheep's bulk milk by an automated flow cytometry method (Bactoscan FC) was evaluated. The results were discussed with respect to the results obtained by the analysis of cow's milk. Within-laboratory reproducibility, which depended on the counting level, was slightly worse than that of raw cow's milk (valued in the interval > 50 Individual Bacterial Cell (IBC)·µL -1 ). In routine procedure, if preceding samples had counts > 20 000 IBC·µL -1 , then this was observed to increase the count of the following sample significantly in many cases ("carry-over"). Bactoscan counts were moderately enhanced with increasing numbers of somatic cells in sheep's milk. However, this trend was indicated only in samples with lower plate counts (< 10 5 colony-forming units (CFU)·mL -1 ). The regression equation between the results of the anchor plate method and the Bactoscan method was computed from the sample set which represented sheep's milk production in Slovakia during 2004 (data between percentiles 1 and 99, n = 877): log 10 CFU·mL -1 = 1.088 log 10 IBC·µL -1 + 2.292; with an accuracy of the estimate, expressed as standard deviation of regression s y,x = 0.305 log 10 CFU·mL -1 . The actual regression for raw cow's milk was verified, introducing both raw cow's milk samples and raw sheep's milk samples. The transformation of IBC·µL -1 to CFU·mL -1 realised by various published conversions was evaluated by testing of the parallelism of the regression lines and also by comparison of obtained differences on a model data set. The conclusions support the requirement to set up a special conversion for the intended field of application at any time. Résumé -L'application de la cytométrie de flux pour déterminer la qualité bactériologique du lait cru de brebis en Slovaquie. Dans cette étude, la numération des cellules bactériennes a été évaluée dans le lait cru de brebis de grand mélange par la cytométrie de flux automatisée (Bactoscan FC). Les résultats ont été comparés à ceux obtenus sur le lait de vache. La reproductibilité interne au laboratoire augmentait avec le niveau de la numération et était légèrement inférieure à celle obtenue sur lait cru de vache (pour les valeurs > 50 cellules bactériennes individuelles (CBI)·µL -1 ). La méthode de routine a montré que si les échantillons précédents avaient un niveau de contamination > 20 000 CBI·µL -1 , cela pouvait fréquemment augmenter significativement les valeurs dénombrées dans l'échantillon suivant ("carry over"). L'augmentation des cellules somatiques dans le lait de brebis entraînait une légère augmentation des valeurs de dénombrements par Bactoscan. Cependant cette tendance n'a été observée que dans les échantillons ayant les plus faibles niveaux de dénom-brements sur boîte (< 10 5 unités formant des colonies (UFC)·mL -1 ). L'équation de régression entre les résultats de dénombrements sur boîte et de dénombrements par Bactoscan a été calculée à partir de l'ensemble des échantillons représentant la production du lait de b...

The relationship of flow cytometry results with classical measures of bacterial counts in raw refrigerated milk

“…Other authors observed similar correlations between the two methods (Suhren and Walte 1998; Ninane et al. 2000; Tomáska et al. 2006; Cassoli et al.…”

“…Other studies that evaluated the same equipment showed standard deviation varying from 0.25 to 0.30 log 10 CFU/mL (Ninane et al. 2000; Suhren and Reichmuth 2000; Tomáska et al. 2006; Cassoli et al.…”

Seasonal dynamics and possible development of total count of microorganisms in sheep’s milk

“…The results show a high variability of TCM which clarify that sheep's milk is microbiologically more complex than cow's milk (Tomáška et al 2006). The above expressed high variability is a good proof of normal frequency distribution absence in the used data files.…”

“…The bacterial count was determined by a method based on laser flow cytometry (Tomáška et al 2006) using BactoScan FC (Foss, Hillerød, Denmark). The method was accredited according to criteria of ISO 17025, regularly controlled via internal (pilot samples) and external (Huefner standards; MIH Milchwirtschaftliches Institut Dr. J. Huefner, Hergarz, Germany) and by participation in interlaboratory comparisons (MIH Milchwirtschaftliches Institut Dr. J. Huefner).…”

Application of flow cytometry to industrial microbial bioprocesses

“…On the other hand, manufacture of dairy products requires the constant and rigorous analysis of the microbial quality of raw materials, end-products as well as the production process itself [220] to guarantee the absence of risks. FC is presently used by many dairies to ensure that milk or its derivatives (heat-treated milk, powder milk and whey) fulfill quality requirements [18,[221][222][223]. Limitations around the identification of potential pathogens or spoilage microorganisms that might be present in milk are removed by combining FC with fluorescent in situ hybridization (FISH) to rRNA [104].…”