Use of bacteriophages, which are viruses that kill bacteria, for biocontrol of pathogens and antimicrobial resistant bacteria has become increasingly important in recent years. As traditional culture-based methods are laborious and time-consuming, practicable use of bacteriophages will hinge on development of rapid and high throughput methods to analyze, characterize and screen large bacteriophage libraries. We thus established a novel method to fluorescently tag bacteriophages for virus screening and interaction studies, without the need for complicated and laborious purification procedures or genetic engineering of viruses to express fluorescent proteins. Bacteriophage PMBT14 was tagged using DNA dye Syto 13. Simply by using a membrane filter, tagged bacteriophages can be separated from non-sequestered excess dye rapidly, effortlessly, and cheaply. The procedure takes less than 30 min and makes use of simple laboratory consumables that are already commonly used for bacteriophage preparations. As proof of concept, we present here flow cytometric methods to analyze bacteriophage binding, infection and killing that are very accessible for high throughput analysis. We show that the resulting fluorescently tagged bacteriophage can be used to specifically stain its host bacterium Pseudomonas fluorescens DSM 50090. Individual fluorescent bacteriophages, their binding to and initial infection of bacteria could also be observed using confocal microscopy. The infection process was halted by the metabolic inhibitor sodium azide, suggesting a requirement of host metabolic processes for penetration by PMBT14. Flow cytometric live/dead assays was used as a complementary method to determine bacteriophage infection of its host. We made preliminary efforts to adapt the tagging method to two other bacteriophages and discuss potential pitfalls and solutions in the use of tagged phages. Fluorescent phage tagging has previously been demonstrated to facilitate analysis of bacteriophage–host interactions. The method adopted in this study makes it fast, easy as well as cost effective.

Two hundred fresh produce samples (cucumber, carrots, herbs, leaf lettuce, and ready-to-eat mixed salad leaves) were obtained from retail in northern Germany in 2015. These were investigated for microbial contamination and the presence of foodborne pathogens, including Listeria monocytogenes , Salmonella serovars, presumptive Bacillus (B.) cereus , and Shiga toxin–producing Escherichia coli using culture-dependent (enrichment, plating on selective media) and -independent (real-time polymerase chain reaction [PCR]) techniques. Overall, our results showed that the fresh produce samples generally showed high mean aerobic mesophilic bacterial counts of between 7 and 8 log ₁₀ cfu/g. However, there was also a considerable variation in total aerobic bacterial counts between different product samples. Although real-time PCR signals for pathogenic E. coli were detected in 14.0% of total samples analyzed, only one (0.5%) Shiga toxin–producing E. coli isolate of serotype O26:H11 was recovered from mixed salad leaves and contained stx 1, stx 2, and eae genes. Two L. monocytogenes isolates (1% of total samples) were recovered from packaged mixed salad leaves and belonged to PCR serogroups IIb and IVb, respectively. One Salmonella isolate (0.5%) was recovered after selective enrichment also from mixed salad leaves and it was identified as Salmonella Szentes serotype 16:k:1,2. Overall the incidence of foodborne pathogens on the northern German retail market in 2015 was very low.

In this study, nine Gram-negative, motile and rod-shaped bacteria were isolated during a Germany-wide investigation of raw milk microbiota. The strains could be differentiated from their closest relatives by phenotypic and chemotaxonomic characterization and average nucleotide identity (ANIb) values calculated from draft genome assemblies. Strains MBT-1^T, MBT-8, MBT-9, MBT-10, MBT-11 and MBT-12 were related to the Pseudomonas chlororaphis subgroup. Isolates MBT-2^T, MBT-13 and MBT-14 were closely related to Pseudomonas rhizosphaerae DSM 16299^T with an ANIb of 88.2 % and a genome-to-genome distance result of 36.0 %. The G+C content of the DNA of strains MBT-1^T and MBT-2^T was 60.84 and 62.48 mol%, respectively. The major fatty acids were C_16 : 1 ω7c (summed feature 3), C_16 : 0 and C_18 : 1 ω7c (summed feature 8). The strains were catalase-positive, while production of urease, β-galactosidase and indole were negative. Growth occurred at 4–30 °C and at pH values of pH 6.0–8.0. Based on these results, we conclude that the strains belong to two novel species, for which the names Pseudomonas kielensis sp. nov. and Pseudomonas baltica sp. nov. are proposed. The type strains are MBT-1^T (=DSM 111668^T= LMG 31954^T) and MBT-2^T (=DSM 111761^T=LMG 31955^T).

Here we present the description of a novel Pseudomonas species, designated Pseudomonas rustica sp. nov., which was isolated from raw milk samples obtained from Germany. Results of initial 16S rRNA gene sequence analysis assigned the strain into the genus Pseudomonas and showed Pseudomonas helmanticensis , Pseudomonas neuropathica and Pseudomonas atagonensis to be its closest relatives. Further studies including sequence analysis of the rpoB gene, multi-gene phylogenetic tree reconstruction, whole-genome sequence comparisons, cellular fatty acid analysis and chemotaxonomic characterization showed a clear separation from the known Pseudomonas species. Isolate MBT-4^T was closely related to Pseudomonas helmanticensis , 'Pseudomonas crudilactis' and Pseudomonas neuropathica with average nucleotide identities based on blast values of 88.8, 88.8 and 88.6%, respectively. Therefore, the strain can be classified into the Pseudomonas koreensis subgroup of the Pseudomonas fluorescens group. The G+C content of strain MBT-4^T was 58.9 mol%. The strain was catalase- and oxidase-positive, while the β-galactosidase reaction was negative. Growth occurred between 4 and 30 °C and at pH values from pH 6.0 to 8.0. In conclusion, strain MBT-4^T belongs to a novel species, for which the name Pseudomonas rustica sp. nov. is proposed. The type strain is MBT-4^T (=DSM 112348^T=LMG 32241^T) and strain MBT-17 is also a representative of this species.