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Rapid and robust detection assays for Salmonella Enteritidis (SE) in shell eggs are essential to enable a quick testing turnaround time (TAT) at the earliest checkpoint and to ensure effective food safety control. Real-time polymerase chain reaction (qPCR) assays provide a workaround for the protracted lead times associated with conventional Salmonella diagnostic testing. However, DNA-based analysis cannot reliably discriminate between signals from viable and dead bacteria. We developed a strategy based on an SE qPCR assay that can be integrated into system testing to accelerate the detection of viable SE in egg-enriched cultures and verify the yielded SE isolates. The specificity of the assay was evaluated against 89 Salmonella strains, and SE was accurately identified in every instance. To define the indicator for a viable bacteria readout, viable or heat-inactivated SE were spiked into shell egg contents to generate post-enriched, artificially contaminated cultures to establish the quantification cycle (Cq) for viable SE. Our study has demonstrated that this technique could potentially be applied to accurately identify viable SE during the screening stage of naturally contaminated shell eggs following enrichment to provide an early alert, and that it consistently identified the serotypes of SE isolates in a shorter time than conventional testing. [ABSTRACT FROM AUTHOR]

Background: Bacterial communities are responsible for biological nutrient removal and flocculation in engineered systems such as activated floccular sludge. Predators such as bacteriophage and protozoa exert significant predation pressure and cause bacterial mortality within these communities. However, the roles of bacteriophage and protozoan predation in impacting granulation process remain limited. Recent studies hypothesised that protozoa, particularly sessile ciliates, could have an important role in granulation as these ciliates were often observed in high abundance on surfaces of granules. Bacteriophages were hypothesized to contribute to granular stability through bacteriophage-mediated extracellular DNA release by lysing bacterial cells. This current study investigated the bacteriophage and protozoan communities throughout the granulation process. In addition, the importance of protozoan predation during granulation was also determined through chemical killing of protozoa in the floccular sludge. Results: Four independent bioreactors seeded with activated floccular sludge were operated for aerobic granulation for 11 weeks. Changes in the phage, protozoa and bacterial communities were characterized throughout the granulation process. The filamentous phage, Inoviridae, increased in abundance at the initiation phase of granulation. However, the abundance shifted towards lytic phages during the maturation phase. In contrast, the abundance and diversity of protozoa decreased initially, possibly due to the reduction in settling time and subsequent washout. Upon the formation of granules, ciliated protozoa from the class Oligohymenophorea were the dominant group of protozoa based on metacommunity analysis. These protozoa had a strong, positive-correlation with the initial formation of compact aggregates prior to granule development. Furthermore, chemical inhibition of these ciliates in the floccular sludge delayed the initiation of granule formation. Analysis of the bacterial communities in the thiram treated sludge demonstrated that the recovery of 'Candidatus Accumulibacter' was positively correlated with the formation of compact aggregates and granules. Conclusion: Predation by bacteriophage and protozoa were positively correlated with the formation of aerobic granules. Increases in Inoviridae abundance suggested that filamentous phages may promote the structural formation of granules. Initiation of granules formation was delayed due to an absence of protozoa after chemical treatment. The presence of 'Candidatus Accumulibacter' was necessary for the formation of granules in the absence of protozoa. [ABSTRACT FROM AUTHOR]