Cereus Spores Research Articles

Influence of water activity and acidity on Bacillus cereus spore inactivation during combined high pressure-thermal treatment

Understanding the inactivation kinetics of bacterial spores during combined pressure-thermal processing is essential for process design. The knowledge on the influence of pH and water activity (aw) during combined pressure-thermal treatment is limited. Experiments were conducted using Bacillus cereus spores suspended in citrate-phosphate buffers, adjusted to different levels of pH (3.8–7.3) and aw (0.80–0.99). Spores were then treated with a pressure-heat combination (600 MPa, 70 °C) using a laboratory-scale high-pressure kinetic tester. Linear regression and Weibull models were used to fit the experimental data. The kinetic parameters of the spores inactivation estimated from the models were plotted as a function of pH and aw. Results showed that the inactivation kinetic curves were characterized by tailing, particularly at low aw levels. The log reduction of B. cereus spores decreased with the decrease of aw. During pressure-thermal treatment, the changes in acidity values for a given water activity have limited influence on decimal reduction values of the spores. Study findings will be helpful for designing pressure-thermal processes to produce microbially safe products with varying water activity and acidity levels.

Inactivation of Bacillus cereus spores using a combined treatment of UV-TiO2 photocatalysis and high hydrostatic pressure

Bacillus cereus spores are resistant to high hydrostatic pressure (HHP) processing treatment. A combination of UV-TiO2 photocatalysis (UVTP for 10 min) and two cycles of 600 MPa HHP treatment for 10 min for the first cycle and 1 min for the second cycle (UVTP-2HHP) at ambient temperature was applied to inactivate B. cereus spores inoculated on a solidified agar matrix (SAM) used as a model matrix. Two cycles of HHP treatment were used as a strategy for induction of spore germination, followed by inactivation. UVTP and 2 cycles of HHP resulted in a 5.0-log CFU/cm2 spore reduction (initial spore count was 6.6 log CFU/cm2), including an approximate 0.8-log CFU/cm2 reduction due to a synergistic effect. The inactivation mechanism of UVTP pretreatment was related to lipid peroxidation of the spore membrane based on the level of malondialdehyde (MDA) making spores susceptible to the HHP treatment. Flow cytometry and transmission electron microscopic (TEM) analyses showed severe physiological alteration and structural damage to spores after the combined treatment. UVTP and 2 cycles of HHP showed potential for effective inactivation of B. cereus to ensure food safety from B. cereus spores on food products. Practical applicationsInactivation of bacterial spores remains a technical challenge for HHP and other interventions because spores are highly resistant to high pressure. Pretreatment with UVTP followed by two cycles of HHP resulted in reduction in B. cereus spores due to a synergistic effect. This hurdle technology of UVTP and HHP can help food industry in ensuring food safety against the Bacillus spores.

Evaluation of the contamination by bacteria of the Bacillus cereus group and the determination of some physico-chemical parameters of Attiéké garba sold in Daloa (Central West, Côte d'Ivoire).

In Côte d'Ivoire as elsewhere in Africa, attiéké plays an important role in feeding the population. The objective of the work is to conduct a study on the sanitary quality of the attiéké garba sold in Daloa. Fifty (50) samples of attiéké garba were collected by purchase from various high-traffic outlets in different districts of Daloa town. Samples are collected in the morning around 10 a.m. every 2 days at a rate of 10 per site. Microbiological analyzes and the determination of some physico-chemical parameters of attiéké garba were carried out. The bacterial load of Bacillus cereus present in the samples tested in the majority of districts is greater than or equal to 105 CFU/g of food. About 60% of the samples tested was contaminated with B. cereus spores against 72% contaminated with the vegetative form of B. cereus. However, physicochemical analyzes showed that attiéké garba was acidic and the humidity was between 37.36 ± 15.9 and 55.26 ± 20.5%. Attiéké garba could pose a danger to consumers.

Inactivation kinetics of Bacillus cereus vegetative cells and spores from different sources by antimicrobial photodynamic treatment (aPDT)

Antimicrobial photodynamic treatment (aPDT) is a promising alternative to conventional thermal inactivation methods. This study aimed to evaluate Bacillus cereus vegetative cells' inactivation and spores by aPDT with new methylene blue (NMB) as photosensitizer (PS) using red light. The efficacy of aPDT was determined initially by minimal inhibitory concentration (MIC) of NMB at different concentrations and fluences. Cluster analysis from the results of MIC grouped B. cereus strains according to their aPDT susceptibility. Then, four strains (B63, B3, 436, and ATCC 14579) were selected for a survival study of aPDT with NMB in three concentrations (5, 50, and 100 μM). The viability of B. cereus vegetative cells and spores was reduced in all tested fluences. The strain ATCC 14579 (vegetative cells and spores) was the most susceptible to aPDT at relatively low fluence (25 J/cm2). The Weibull model presented a good fit for the inactivation data estimating the kinetic parameters δ (first decimal reduction) and p (shape parameter). This study contributes to the knowledge of the behavior of different strains of B. cereus regarding an emerging method of microbial inactivation. The variability of inactivation among strains will allow the development of more reliable processes.

Growth Characteristics of Bacillus cereus in Sake and during Its Manufacture

Sake (Japanese rice wine) has been recognized as being low risk in terms of its microbiological safety. However, a confirmation of the food safety aspects of sake based on scientific evidence is important for establishing consumer confidence, in part because consumer concerns regarding food safety have increased. The presence of Bacillus cereus spores in refined rice wine has been reported, and in light of consumers' growing concern over food safety, the establishment of food and beverage safety is important for consumers' reassurance. Herein, to confirm the microbiological safety of sake, we investigated the content and growth of B. cereus. We conducted a spore addition test to determine whether B. cereus spores grow during sake production, and we observed no growth or germination of B. cereus spores during the manufacturing process. We also observed that processes such as solid-liquid separation and filtration help remove the risk posed by B. cereus. We then conducted a survey to assess the density of B. cereus in various commercial sake products. We analyzed 162 samples of commercial sake and observed that 11 of the products had ≥1 CFU of living cells in 1 mL of sake (detection rate, 6.8%). There was no product in which ≥100 CFU of living cells per 1 mL of sake was detected. Our findings confirmed that the density of these bacteria in sake is lower than that in other foods and that the probability of infection is very low. The emetic toxin produced by B. cereus was not detected in any of the sake samples. This is the first study based on experimental data demonstrating that B. cereus is not able to grow in sake or during the sake manufacturing process. We, thus, conclude that the safety risk of B. cereus in sake is negligible. Our findings indicating that B. cereus is not a significant hazard in the sake brewing process will contribute to food hygiene management based on scientific evidence in sake breweries.

PATHOGENICITY OF FEED-BORNE BACILLUS CEREUS AND ITS IMPLICATION ON FOOD SAFETY

Bacillus cereus (B. cereus) is a novel emerging pathogen contaminated extensively in animal feed and food chains, posing a huge economic loss for animal industry and high risk for human health. This pathogen is a robust omnipresent heat resistant spore former, able to form biofilm and isolated from different environments such as food and atmosphere that occur all year round without any particular geographic distribution. The potential of survival for B. cereus spores in unfavorable conditions pose a considerable threat to food safety and also cause economic losses to the food industry. B. cereus aggravates acute diarrhea and malnourishment in poultry by inducing gizzard erosion and ulceration (GEU). It will facilitate persistent other bacterial infection in the lungs via damaging gastric intestine tract. Also, it can cause serious food safety because it seems difficult to fully prevent their presence in food. It may cause gastrointestinal diseases that trigger emetic and diarrheal symptoms as well as general and local infections related to the respiratory tracts of immunologically threatened individual and newborns. B. cereus produces a wide range of potential virulence factors, including heat stable/labile toxins (cerulide, NHE, HBL, CytK, Ent-FM, bc-D-ENT, CLO, HlyII, HlyIII) and tissue-destructive enzymes (PI-PLC, PC-PLC, SMase, ?-lactamase, InhA1, NprA), but their roles and molecular mechanism in specific infections have not been clarified yet. This review provided a historical record of possible risk factors and pathogenesis of animal industry and highlights the implications for animal industry and food safety by ingestion of the feed-borne Bacillus cereus.

Effects of intense pulsed light and gamma irradiation on Bacillus cereus spores in mesquite pod flour

This study was conducted to evaluate the inactivation of Bacillus cereus spore in mesquite flour with intense pulsed light (IPL) and gamma radiation. The physical, chemical, and toxicity of treated mesquite flour were also investigated. The results showed that up to 3.51 log10CFU/g B. cereus spore inactivation was achieved with 8 kGy of gamma radiation, and up to 1.69 log10CFU/g reductions could be achieved after 28s of catalytic IPL exposure. Although chemometric analysis showed 9-hydroxy-10,12-octadecadienoic acid was slightly increased after a 28s-catalytic IPL treatment, the concentration is within the acceptable range. No significant increase in acetic or propionic acids (typical off-flavor volatile compounds) was observed after either treatment. For cytotoxicity, the Caco-2 cell viability analysis revealed that these two technologies did not induce significant cytotoxicity to the treated mesquite flour. Overall, these two technologies exhibit strong potential for the decontamination of B. cereus in mesquite flour.

Optimization of the Antimicrobial Effects of Surfactin against Bacillus cereus Spores

The purpose of this study was to establish a three-variable bactericidal model of temperature, time, and concentration to determine the optimal conditions for Bacillus cereus spore inactivation by surfactin. To obtain the binary regression equation of the inactivated spore model, a total of 17 simulations were performed using response surface methodology. The experimental results showed that the three factors each had a discernible but nonequal impact on the inactivation response value. Multiple regression analysis of experimental results using Design-Expert software yielded the following equation: Y = 1.47 + 0.39ξ1(temperature) + 0.38ξ2(time) + 0.39ξ3(concentration) - 0.20ξ1ξ2 + 0.22ξ1ξ2 - 0.12ξ2ξ3 - 0.23ξ12 - 0.11ξ22 - 0.40ξ32. Optimal inactivation of spores was achieved by treatment with surfactin at a concentration of 4 mg/mL for 40 h at 53°C, with the response value reaching 1.8. The spores were treated with surfactin under these conditions; the microstructural changes of spores were observed by use of scanning electron microscopy. We found that the structures of the outer wall of the spores were damaged, whereas the spores in the control sample showed no visible damage.

Extended shelf life milk processing: Effect of simulated cleaning in place on the germination and attachment of Bacillus cereus spores

The effect of simulated cleaning in place (CIP) was determined on the structure, attachment and growth of Bacillus cereus spores isolated from raw milk and biofilms in filler nozzles from extended shelf life (ESL) milk processing lines. Simulated CIP treatment structurally affected >98% of B. cereus spores, while 0.1% remained intact. Following simulated CIP treatment, B. cereus spores were able to attach to stainless steel coupons and form biofilms. B. cereus spores were capable of germination and growth under refrigerated conditions for more than 28 days. Contamination with B. cereus spores may lead to a reduced shelf life and potentially be a safety risk in ESL milk with a prolonged shelf life.

Microbiological and process variability using biological indicators of inactivation (BIIs) based on Bacillus cereus spores of food and fish-based animal by-products to evaluate microwave heating in a pilot plant

Microwave processing can be a valid alternative to conventional heating for different types of products. It enables a more efficient heat transfer in the food matrix, resulting in higher quality products. However, for many food products a uniform temperature distribution is not possible because of heterogeneities in their physical properties and non-uniformtiy in the electric field pattern. Hence, the effectiveness of microwave inactivation treatments is influenced by both intrinsic (differences between cells) and extrinsic variability (non-uniform temperature). Interpreting the results of the process and considering its impact on microbial inactivation is essential to ensure effective and efficient processing.In this work, we quantified the variability in microbial inactivation attained in a microwave pasteurization treatment with a tunnel configuration at pilot-plant scale. The configuration of the equipment makes it impossible to measure the product temperature during treatment. For that reason, variability in microbial counts was measured using Biological Inactivation Indicators (BIIs) based on spherical particles of alginate inoculated with spores of Bacillus spp. The stability of the BIIs and the uncertainty associated to them was assessed using preliminary experiments in a thermoresistometer. Then, they were introduced in the food product to analyse the microbial inactivation in different points of the products during the microwave treatment. Experiments were made in a vegetable soup and a fish-based animal by-product (F-BP).The results show that the variation in the microbial counts was higher than expected based on the biological variability estimated in the thermoresistometer and the uncertainty of the BIIs. This is due to heterogeneities in the temperature field (measured using a thermographic camera), which were higher in the F-BP than in the vegetable soup. Therefore, for the process studied, extrinsic variability was more relevant than intrinsic variability. The methodology presented in this work can be a valid method to evaluate pasteurization treatments of foods processed by heating, providing valuable information of the microbial inactivation achieved. It can contribute to design microwave processes for different types of products and for product optimization.

Inactivation of Bacillus cereus from pork by thermal, non-thermal and single-frequency/multi-frequency thermosonication: Modelling and effects on physicochemical properties.

B cereus was extracted from pork meat and detected using conventional polymerase chain reaction, (PCR). The spores were infected in the pork and the inactivation behavior was scrutinize by thermal (80 °C and 90 °C), sole ultrasonication, US (20 kHz) and thermosonication (TS), at single frequency TSsf (20 kHz, 70 °C) and multi-frequency TSmf (20/40/60 kHz, 70 °C) using a maximum power of 300 W over a period of 25 min. The most convenient and time-saving treatment method of the infected pork were investigated. TSmf proved to be favorable treatment method as it gave a 4.16 log CFU/ml spore reduction within 5 min. The various treatments were modelled to predict the best fit for the study. Weibull model gave the best fitness and the various parameters showed the effectiveness of the treatments as follows: thermal (80 °C) < thermal (90 °C) < US (20 kHz) < TSsf (20 kHz, 70 °C) < TSmf (20/40/60 kHz, 70 °C) for the deactivation of B. cereus spores in pork. Finally, US and TSmf methods enhanced the physicochemical properties (pH, texture and color) of pork.

Visualization of Germination Proteins in Putative Bacillus cereus Germinosomes.

Bacillus cereus can survive in the form of spores for prolonged periods posing a serious problem for the manufacture of safe shelf-stable foods of optimal quality. Our study aims at increasing knowledge of B. cereus spores focusing primarily on germination mechanisms to develop novel milder food preservation strategies. Major features of B. cereus spores are a core with the genetic material encased by multiple protective layers, an important one being the spores′ inner membrane (IM), the location of many important germination proteins. To study mechanisms involved in germination of B. cereus spores, we have examined the organization of germinant receptors (GRs) in spores′ IM. Previous studies have indicated that in spores of B. cereus ATCC 14579 the L-alanine responsive GR, GerR, plays a major role in the germination process. In our study, the location of the GerR GR subunit, GerRB, in spores was examined as a C-terminal SGFP2 fusion protein expressed under the control of the gerR operon′s promoter. Our results showed that: (i) the fluorescence maxima and integrated intensity in spores with plasmid-borne expression of GerRB-SGFP2 were significantly higher than in wild-type spores; (ii) western blot analysis confirmed the expression of the GerRB-SGFP2 fusion protein in spores; and (iii) fluorescence microscopy visualized GerRB-SGFP2 specific bright foci in ~30% of individual dormant spores if only GerRB-SGFP2 was expressed, but, noticeably, in ~85% of spores upon co-expression with GerRA and GerRC. Our data corroborates the notion that co-expression of GR subunits improves their stability. Finally, all spores displayed bright fluorescent foci upon expression of GerD-mScarlet-I under the control of the gerD promoter. We termed all fluorescent foci observed germinosomes, the term used for the IM foci of GRs in Bacillus subtilis spores. Our data are the first evidence for the existence of germinosomes in B. cereus spores.

Effect of pulsed electric field with moderate heat (80°C) on inactivation, thermal resistance and differential gene expression inB. cereusspores

Bacillus cereus spores pose a challenge to the food industry owing to their ability to germinate in food during storage and produce toxins. The objective of the current study was to investigate the effect of pulsed electric field (PEF) at low electric field strength (i.e., 7.3, 8.1, 8.8 and 9.4 kV/cm) and moderate temperatures (65, 70, 75 and 80°C) on B. cereus spores. PEF treatment at 9.4 kV/cm at 80°C (pulse width of 20 μs and frequency of 300 Hz) led to a 3.1 log CFU/ml reduction in the number of B. cereus spores and importantly, the D88°C values of the surviving spores were reduced by 12 min. Differential gene expression revealed the upregulation of the gene (BC1768) encoding for chitooligosaccharide deacetylase involved in peptidoglycan degradation in PEF treated spores. The gene (BC2729) encoding for a penicillin-binding protein required for septation during sporulation was downregulated in PEF treated spores. Practical applications PEF treatment is an emerging food processing technology that has gained attention owing to its potential to provide high-quality foods. However, to date the inactivation of bacterial spores by PEF has only been partially successful and then only when combined with high heat. The effect of PEF on the thermal stability of spores has not previously been reported. The current work bridges these gaps by providing insight into differential gene expression associated with the PEF treatment of B. cereus spores. The study indicates the potential for PEF to be incorporated into a hurdle regime targeting spore inactivation and provides data on specific genes that may help to provide a better understanding of the stress induced in B. cereus spores by PEF treatment.

Bacillus cereus, a geobiological marker for gold prospecting isolated from soil from the Jiaodong Gold Mine

An efficient method for new gold deposits exploration is of great interest as the existing deposits depleted. In this paper, we investigated the possibility of using B. cereus in the soil from Jiaodong Gold Mine as a biogeochemical indicator of gold. Cell counting, growth curve assay, SEM/TEM-EDS and X-ray Absorption Near Edge Spectroscopy (XANES) techniques were used to investigate the sporulation characteristics, reducing capability and detoxification mechanisms of the B. cereus strain in the environment containing gold. The results showed that: (1) The frequencies of B. cereus spores were higher in areas where elevated concentrations of Au were detected, which was not observed for other elements, such as As, Cu, Zn, Cd and Pb. (2) B.cereus has the ability to facilitated the solubilization of Au. The dissolved gold subsequently stimulated B.cereus spores formation, and mother cells release the contents of their cytoplasm to ameliorate gold toxicity. (3) Toxic Au (III)-complexe was reduced to nontoxic Au (0) nanoparticles by B. cereus. Before reduced to Au (0), cellular Au was coupled with the formation of AuS intermediate species. The results showed that B.cereus spores were confirmed to exist in the gold mine soil, and the reduction and detoxification of gold were mediated by the cytoplasm released by the mother cells, which led to the formation of gold nanoparticles.

Hurdle enhancement of acidic electrolyzed water antimicrobial efficacy on Bacillus cereus spores using ultrasonication.

This study evaluated the inactivation effect of ultrasonic treatment combined with acidic electrolyzed water (AEW) on Bacillus cereus spores. AEW treatment reduced the spores by 1.05-1.37 log CFU/mL while the sporicidal effect of ultrasound was minor. More strikingly, simultaneous ultrasonic and AEW treatments for 30min led to 2.29 log CFU/mL reduction and thus, considered a synergistic effect. Flow cytometry combined with SYTO/PI staining analysis revealed that ultrasound hydrolyzed the cortex while the AEW partially damaged the integrity of the inner membrane. Scanning and transmission electron microscopies were used to characterize the ultrastructural changes. The detachment of the exosporium induced by ultrasound was the most apparent difference compared with the control group, and the electron density of spores appeared to be heterogeneous after treatment with AEW. These results indicated that combining ultrasound with AEW is a promising decontamination technology with potential uses in the food industry and environmental remediation.

Bacillus cereus spores and Staphylococcus aureus sub. aureus vegetative cells inactivation in human milk by high-pressure processing

The effect of high hydrostatic pressure (HHP) on pathogen microorganisms is influenced by an important number of variables like treatment conditions, microorganism type and food matrix, among others. This technology could be an alternative to pasteurization in Human Milk Banks. In this study, a response surface methodology with a central composite design (CCD) was applied to determine the HHP conditions (pressure intensity and holding time) for the maximum inactivation of Staphylococcus aureus sub. aureus and Bacillus cereus in human milk. Precision and bias factors were calculated in order to validate the acceptance of predictions. These were considered within the acceptable range for vegetative cells models. The conditions to reach a maximum bacterial inactivation (within the ranges of this assay) were a pressure intensity of 593.96 MPa for 233 s. These conditions reached a decrease of 5.81 and 6.93 log cfu mL−1 in S. aureus sub. aureus and vegetative B. cereus, respectively. On the other hand, the decrease of B. cereus spores followed a trend that did not fit the model under study. Probably, the application of a second HHP treatment and/or the combination with higher temperature of processing would enhance the inactivation of this pathogen. This model could be a useful tool to increase milk safety by applying HHP treatments in human milk banks.

Food Sensing: Detection of Bacillus cereus Spores in Dairy Products.

Milk is a source of essential nutrients for infants and adults, and its production has increased worldwide over the past years. Despite developments in the dairy industry, premature spoilage of milk due to the contamination by Bacillus cereus continues to be a problem and causes considerable economic losses. B. cereus is ubiquitously present in nature and can contaminate milk through a variety of means from the farm to the processing plant, during transport or distribution. There is a need to detect and quantify spores directly in food samples, because B. cereus might be present in food only in the sporulated form. Traditional microbiological detection methods used in dairy industries to detect spores show limits of time (they are time consuming), efficiency and sensitivity. The low level of B. cereus spores in milk implies that highly sensitive detection methods should be applied for dairy products screening for spore contamination. This review describes the advantages and disadvantages of classical microbiological methods used to detect B. cereus spores in milk and milk products, related to novel methods based on molecular biology, biosensors and nanotechnology.

Inactivation kinetics of Bacillus cereus spores by Plasma activated water (PAW)

In recent years, plasma activated water has attracted more attention as a new disinfectant. The purpose of this study was to explore impact of variation of different treatment conditions on the inactivation kinetics of Bacillus cereus spores by PAW. All survival curves showed that the number of spores has decreased rapidly at first, followed by tailing results from the reduction inactivation rate. A linear and two nonlinear models (Weibull and Log-logistic model) were fitted to these data, and Log-logistic model fitted the inactivation of the B. cereus spores best. B. cereus spores in 106 CFU/mL was reduced by 1.62–2.96 log CFU/mL by PAW at 55 °C due to the reactive species generated in PAW. Elevated temperature, lower initial spore concentration, lower bovine serum albumin content, and smaller activation volume of PAW considerably enhanced PAW inactivation of B. cereus spores. These results provide an approach to evaluate the inactivation efficacy of different treatment conditions for PAW.

Performance Testing of a Venturi-Based Backpack Spray Decontamination System.

The performance of 2 disinfectant chemicals, peracetic acid (PAA) and hypochlorous acid (HOCl), was evaluated using a Venturi-nozzle-based light decontamination system (LDS) for delivery. The atomization equipment combined low-pressure air and disinfectant via a handheld lance, producing a fine, dense aerosol. A range of microorganisms, including Bacillus cereus and Bacillus anthracis (Vollum) spores, were used as test challenges to evaluate chemicals and equipment. The tests undertaken included assessments over fixed and variable exposure times, use of multiple surface materials, and a live agent challenge. Over a fixed-time exposure of 60 minutes, aerosolized PAA gave 7- to 8-log reductions of all test challenges, but HOCl was less effective. Material tests showed extensive kill on most surfaces using PAA (≥6-log kill), but HOCl showed more variation (4- to 6-log). Testing using B. anthracis showed measurable PAA induced spore kill inside 5 minutes and >6-log kill at 5 minutes or over. HOCl was less effective. The results demonstrate the importance of testing decontamination systems against a range of relevant microbiological challenges. Disinfectant efficacy may vary depending on product choice, types of challenge microorganisms, and their position in a treated area. The most effective disinfectants demonstrate biocidal efficacy despite these factors. The data confirmed PAA as an effective disinfectant capable of rapidly killing a range of microorganisms, including spores. HOCl was less effective. The LDS system successfully delivered PAA and HOCl over a wide area and could be suitable for a range of frontline biosecurity applications.

Impact of vaccination during pregnancy and staphylococci concentration on the presence of Bacillus cereus in raw human milk.

ObjectiveThis study aimed to determine whether vaccination during pregnancy, prematurity, and staphylococci concentration influenced the presence of B. cereus or staphylococcal enterotoxins (SEs) in raw human milk from healthy mothers.Study designHuman milk samples were collected from 152 healthy women. B. cereus, S. aureus and coagulase-negative staphylococci (CNS) were enumerated using selective agar culture media. The detection of B. cereus spores and SEs were determined using ELISA.ResultsCNS and B. cereus concentrations in milk from non-vaccinated mothers were higher than that from mothers vaccinated during pregnancy, but S. aureus did not differ. Prematurity did not affect B. cereus or staphylococci in human milk. S. aureus and CNS concentrations in human milk with the presence of B. cereus were higher than that with the absence of B. cereus. Viable B. cereus was present in 9.2% of raw human milk samples whereas SEs were not detected in any samples.ConclusionsVaccination during pregnancy and low concentration of staphylococci could reduce the risk of B. cereus in raw human milk. The screening of B. cereus in raw human milk must be performed before pasteurization to reduce the risk of B. cereus infection in preterm infants.