| In consideration of the particular requirements of the end users of infant formula,the quality and safety monitoring of these products have raised much concern.With the gradual refinement of the safety review process of infant formula products,physical and chemical pollution factors have been eliminated completely.However,due to the rich nutrient content of the raw materials and the special requirements for the manufacturing technique,microbial contamination may still be an existing factor that can jeopardize the product standard and safety.The presence of inactivated bacteria and their metabolites in the milk powder will also significantly affect the flavor,solubility,shelf life and safety of the products.It is hard to detect such spectrum of microbes using traditional detection methods.Therefore,it is imperative to establish a preventive platform specifically targeting to microbial contamination.The full-length 16S rRNA gene sequencing by the PacBio SMRT sequencing technology,culture-dependent technology,quantitative polymerase chain reaction(qPCR)technology,enzyme linked immunosorbent assay(ELISA),salting-out method and immunoaffinity high performance liquid chromatography(IHPLC)were applied to assess the bacterial contamination of 30 infant formula samples in a more comprehensive and systematic way.ELISA results showed that the higher content of immunologic active material in the domestic infant formula,which was correlated with some bacteria.By applying IHPLC,the mycotoxin content was not detected in any of the investigated samples.The results released by PacBio SMRT sequencing technology showed that the existence of structural differences in the bacterial composition between domestic and imported infant fonnula.The result might indicate the severity of contamination in the imported subgroup 1.Psychrotrophic bacteria and thermophilic bacteria,e.g.Bacillus cereus and Anoxybacillus flavithermus,were at some point present in the infant formula samples.These bacteria might affect the product shelf life.With applying both qPCR and PacBio SMRT sequencing technology,a very low number of 16S rRNA genes corresponding to Escherichia coli,Staphylococcus aureus and Salmonella were detected in most infant formula samples,which did not detected by culture-dependent technology.We further analyzed the relationship between heat treatment and bacterial composition in infant formula.It was found that the whey protein nitrogen index(WPNI)was negatively correlated with the abundances of some of the Bacillus.For example,the high amounts of Bacillus cereus and Geobacillus stearothermophilus might suggest repetitive heat treatment.The above methods can be used to assess the microbial contamination of infant formula samples in a more comprehensive way.Additionally,PacBio SMRT sequencing platform,based on the full-length 16S rRNA gene,is able to accurately analyze the composition of microbial contaminants of infant formula at species level;thus,it is particularly suitable for use in studying the microbial content in dairy products.To explore the sources of microbial contamination and accurately describe the microbial content in the samples,this study established methods in microbial identification and tracking.By applying a total of 6 sampling points along the process of’enclosed’ production mode(with good containment from raw milk collection,to the final milk powder products,completing the production in 8 hours)that zero distance from pasture to manufactory,including raw milk,milk sample before and after pasteurization,after the first and second milk concentration steps,and until milk powder production.The results released that the direct powder spraying production mode resulted in greater variations in the microbial structure.However,the ’enclosed’production mode led to relatively little changes in the microbiota composition with the stable microbial structure of the final products.In addition,the ’enclosed’ production model has maintained high content of the immunologic active material through the processing.The results of the microbial source tracking study revealed that the process of pasteurization influenced the bacterial diversity of milk powder apparently and the bacterial diversity of pasteurized milk was lowered than that of other manufactory stages.The main source of the dominant species,Bacillus cereus and Bacillus flexus,in the milk powder samples was tracked to the raw milk.Their relative abundances increased after pasteurization,possibly due to intrinsic high heat-enduring properties of Bacillus.These data indicate the significance of implementing microbiological control for raw milk and its processing.The’enclosed’ production mode is particularly suitable for producing MP and infant formula milk powder.This work has accurately described the sample microbial contamination at species level,especially in detecting the low abundant,uncultivable,and inactivated taxa.This study has established integrative methods in identifying and tracking the sources of microbial contamination.of infant formula.It enables the identification of source of microbial contaminants meanwhile enhances the control of bacterial contamination.The present study has developed a powerful tool for effective monitoring and control of microbial quality and safety of milk powder and infant formula production.It serves also as theoretical and technical references for the quality and safety control of products for large dairy enterprises. |
PDF Full Text Request