Evaluation of different buffered peptone water (BPW) based enrichment broths for detection of Gram-negative foodborne pathogens from various food matrices

Margot, H.; Zwietering, Marcel H.; Joosten, Han; O’Mahony, E.; Stephan, Roger

doi:10.1016/j.ijfoodmicro.2015.07.033

Cited by 21 publications

(7 citation statements)

References 15 publications

(2 reference statements)

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“…This results in difficulty in the detection of pathogens using either molecular-based or culture-based methods without a pre-enrichment step (Baylis et al, 2000; Margot et al, 2015). Most of the bacteria in marketed foods are usually in an injured/stressed state, and some are in a viable but non-culturable (VBNC) state due to heat, radiation, or low temperature treatments.…”

Section: Discussionmentioning

confidence: 99%

A Multiplex RT-PCR Assay for S. aureus, L. monocytogenes, and Salmonella spp. Detection in Raw Milk with Pre-enrichment

et al. 2017

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This study firstly developed a multiplex real-time PCR (RT-PCR) technique combined with a pre-enrichment step to simultaneously detect Staphylococcus aureus (S. aureus), Listeria monocytogenes (L. monocytogenes) and Salmonella spp. in raw milk and the dairy farm environment (feces, soil, feed, water) in one reaction. Brain heart infusion (BHI) broth was selected for the enrichment step to increase the density of the target bacteria by using an incubation of 4 h before multiplex RT-PCR. The results showed that the detection limit of the multiplex real-time assay was approximately 102 CFU/mL for pure cultures and artificially contaminated milk without enrichment, while 12, 14, and 10 CFU/25 mL, respectively, for S. aureus, L. monocytogenes, and Salmonella spp. after pre-enrichment. The newly developed multiplex RT-PCR assay was applied to 46 dairy farm environmental samples and raw milk samples covering a wide variety of sample types. The results demonstrated that the multiplex RT-PCR assay coupled with the BHI enrichment broth was suitable for the simultaneous screening of S. aureus, L. monocytogenes, and Salmonella spp. in the pasture environment and in raw milk. The multiplex RT-PCR assay clearly and successfully shortened the total detection time and reduced labor compared to conventional culture-based methods for testing natural samples.

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Section: Discussionmentioning

confidence: 99%

A Multiplex RT-PCR Assay for S. aureus, L. monocytogenes, and Salmonella spp. Detection in Raw Milk with Pre-enrichment

et al. 2017

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“…Although the observation that EE-broth favours enrichment of the phylum Proteobacteria including pathogens such as STEC and Salmonella should be a desirable trait of this media, we have also previously observed that the growth of STEC spiked into mungo bean sprouts terminates prematurely thereby hindering their detection in contaminated sprouts [12]. Based on this, we concluded that this was probably due to competition and outgrowth of the STEC by non-target enterobacteria species that occur at high abundances within the mungo bean sprout background flora.…”

Section: Discussionmentioning

confidence: 99%

“…However, due to different factors such as competition with co-enriching sprout microflora, as well as differences in growth rates and presence of growth inhibitors, this enrichment does result in a biased sample [11]. In the case of sprouted seeds deliberately contaminated with STEC, we recently showed that growth of the target organisms terminates prematurely and PCR cannot reliably detect its presence even at high contamination levels [12]. Furthermore, attempts to increase the selectivity of the enrichment using different media or selective supplements did not have a significant impact on reducing the levels of co-enriching sprout background flora during enrichment [12].…”

Section: Introductionmentioning

confidence: 99%

Mungo bean sprout microbiome and changes associated with culture based enrichment protocols used in detection of Gram-negative foodborne pathogens

2016

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BackgroundFresh sprouted seeds have been associated with a number of large outbreaks caused by Salmonella and Shiga toxin-producing E. coli. However, the high number of commensal bacteria found on sprouted seeds hampers the detection of these pathogens. Knowledge about the composition of the sprout microbiome is limited. In this study, the microbiome of mungo bean sprouts and the impact of buffered peptone water (BPW) and Enterobacteriaceae enrichment broth (EE-broth)-based enrichment protocols on this microbiome were investigated.ResultsAssessments based on aerobic mesophilic colony counts showed similar increases in mungo bean sprout background flora levels independent of the enrichment protocol used. 16S rRNA sequencing revealed a mungo bean sprout microbiome dominated by Proteobacteria and Bacteroidetes. EE-broth enrichment of such samples preserved and increased Proteobacteria dominance while reducing Bacteroidetes and Firmicutes relative abundances. BPW enrichment, however, increased Firmicutes relative abundance while decreasing Proteobacteria and Bacteroidetes levels. Both enrichments also lead to various genus level changes within the Protobacteria and Firmicutes phyla.ConclusionsNew insights into the microbiome associated with mungo bean sprout and how it is influenced through BPW and EE-broth-based enrichment strategies used for detecting Gram-negative pathogens were generated. BPW enrichment leads to Firmicutes and Proteobacteria dominance, whereas EE-broth enrichment preserves Proteobacteria dominance in the mungo bean sprout samples. By increasing the relative abundance of Firmicutes, BPW also increases the abundance of Gram-positive organisms including some that might inhibit recovery of Gram-negative pathogens. The use of EE-broth, although preserving and increasing the dominance of Proteobacteria, can also hamper the detection of lowly abundant Gram-negative target pathogens due to outgrowth of such organisms by the highly abundant non-target Proteobacteria genera comprising the mungo bean sprout associated background flora.

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“…Food samples (25 g or 25 mL) were screened for S . aureus by transferring to 225 mL of buffered peptone water and mixed thoroughly 43 . The samples were incubated overnight at 37 °C for enrichment of cultures.…”

Section: Methodsmentioning

confidence: 99%

MALDI-TOF-MS based identification and molecular characterization of food associated methicillin-resistant Staphylococcus aureus

Manukumar

Umesha

2017

Sci Rep

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Food-borne methicillin resistant Staphylococcus aureus (MRSA) is involved in two-fold higher mortality rate compared to methicillin susceptible S. aureus (MSSA). Eventhough Mysuru recognized as cleanest city in the world, prevalence of food contamination is not detailed. The aim is to screen food samples from Mysuru area and to characterize MRSA strain, employing MALDI-Biotyper, multiplex PCR to distinguish between MRSA and MSSA by PCR-coupled single strand conformation polymorphism (PCR-SSCP). Of all the food-borne pathogens, S. aureus contamination accounts for 94.37 ± 0.02% (P < 0.01), strains characterized by means of nuc genes, followed by species specific identification by Coa, Eap and SpA genes and multiplex PCR to confirm the presence of three methicillin resistant staphylococcal species simultaneously using nuc and phoP genes. Amplification of mecA gene in 159 isolates confirmed that all strains are methicillin resistant, except UOM160 (MSSA) and multi-drug resistant (MDR) in 159 isolates confirmed by 22 sets of β-lactam antibiotics. MSSA and MDR-MRSA were discriminated by PCR-SSCP using nuc gene for the first time. From the present studies, compared to conventional methods MALDI-Biotyper emerged as an effective, sensitive (>99%), robust (<2 min), and alternative tool for pathogen identification, and we developed a PCR-SSCP technique for rapid detection of MSSA and MRSA strains.

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Evaluation of different buffered peptone water (BPW) based enrichment broths for detection of Gram-negative foodborne pathogens from various food matrices

Cited by 21 publications

References 15 publications

A Multiplex RT-PCR Assay for S. aureus, L. monocytogenes, and Salmonella spp. Detection in Raw Milk with Pre-enrichment

A Multiplex RT-PCR Assay for S. aureus, L. monocytogenes, and Salmonella spp. Detection in Raw Milk with Pre-enrichment

Mungo bean sprout microbiome and changes associated with culture based enrichment protocols used in detection of Gram-negative foodborne pathogens

MALDI-TOF-MS based identification and molecular characterization of food associated methicillin-resistant Staphylococcus aureus

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