A nested-PCR method was used to detect the occurrence of human adenovirus in coastal waters of Southern California. Twenty-to forty-liter water samples were collected from 12 beach locations from Malibu to the border of Mexico between February and March 1999. All sampling sites were located at mouths of major rivers and creeks. Two ultrafiltration concentration methods, tangential flow filtration (TFF) and vortex flow filtration (VFF), were compared using six environmental samples. Human adenoviruses were detected in 4 of the 12 samples tested after nucleic acid extraction of VFF concentrates. The most probable number of adenoviral genomes ranged from 880 to 7,500 per liter of water. Coliphages were detected at all sites, with the concentration varying from 5.3 to 3332 PFU/liter of water. F-specific coliphages were found at 5 of the 12 sites, with the concentration ranging from 5.5 to 300 PFU/liter. The presence of human adenovirus was not significantly correlated with the concentration of coliphage (r ؍ 0.32) but was significantly correlated (r ؍ 0.99) with F-specific coliphage. The bacterial indicators (total coliforms, fecal coliforms, and enterococci) were found to exceed California recreational water quality daily limits at 5 of the 12 sites. However, this excess of bacterial indicators did not correlate with the presence of human adenoviruses in coastal waters. The results of this study call for both a reevaluation of our current recreational water quality standards to reflect the viral quality of recreational waters and monitoring of recreational waters for human viruses on a regular basis.
We constructed two collections of targeted single gene deletion (SGD) mutants and two collections of targeted multi-gene deletion (MGD) mutants in Salmonella enterica sv Typhimurium 14028s. The SGD mutant collections contain (1), 3517 mutants in which a single gene is replaced by a cassette containing a kanamycin resistance (KanR) gene oriented in the sense direction (SGD-K), and (2), 3376 mutants with a chloramphenicol resistance gene (CamR) oriented in the antisense direction (SGD-C). A combined total of 3773 individual genes were deleted across these SGD collections. The MGD collections contain mutants bearing deletions of contiguous regions of three or more genes and include (3), 198 mutants spanning 2543 genes replaced by a KanR cassette (MGD-K), and (4), 251 mutants spanning 2799 genes replaced by a CamR cassette (MGD-C). Overall, 3476 genes were deleted in at least one MGD collection. The collections with different antibiotic markers permit construction of all viable combinations of mutants in the same background. Together, the libraries allow hierarchical screening of MGDs for different phenotypic followed by screening of SGDs within the target MGD regions. The mutants of these collections are stored at BEI Resources (www.beiresources.org) and publicly available.
Human enteric pathogens, such as Salmonella spp. and verotoxigenic Escherichia coli, are increasingly recognized as causes of gastroenteritis outbreaks associated with the consumption of fruits and vegetables. Persistence in plants represents an important part of the life cycle of these pathogens. The identification of the full complement of Salmonella genes involved in the colonization of the model plant (tomato) was carried out using transposon insertion sequencing analysis. With this approach, 230,000 transposon insertions were screened in tomato pericarps to identify loci with reduction in fitness, followed by validation of the screen results using competition assays of the isogenic mutants against the wild type. A comparison with studies in animals revealed a distinct plant-associated set of genes, which only partially overlaps with the genes required to elicit disease in animals. De novo biosynthesis of amino acids was critical to persistence within tomatoes, while amino acid scavenging was prevalent in animal infections. Fitness reduction of the Salmonella amino acid synthesis mutants was generally more severe in the tomato rin mutant, which hyperaccumulates certain amino acids, suggesting that these nutrients remain unavailable to Salmonella spp. within plants. Salmonella lipopolysaccharide (LPS) was required for persistence in both animals and plants, exemplifying some shared pathogenesis-related mechanisms in animal and plant hosts. Similarly to phytopathogens, Salmonella spp. required biosynthesis of amino acids, LPS, and nucleotides to colonize tomatoes. Overall, however, it appears that while Salmonella shares some strategies with phytopathogens and taps into its animal virulence-related functions, colonization of tomatoes represents a distinct strategy, highlighting this pathogen's flexible metabolism.IMPORTANCE Outbreaks of gastroenteritis caused by human pathogens have been increasingly associated with foods of plant origin, with tomatoes being one of the common culprits. Recent studies also suggest that these human pathogens can use plants as alternate hosts as a part of their life cycle. While dual (animal/plant) lifestyles of other members of the Enterobacteriaceae family are well known, the strategies with which Salmonella colonizes plants are only partially understood. Therefore, we undertook a high-throughput characterization of the functions required for Salmonella persistence within tomatoes. The results of this study were compared with what is known about genes required for Salmonella virulence in animals and interactions of plant pathogens with their hosts to determine whether Salmonella repurposes its virulence repertoire inside plants or whether it behaves more as a phytopathogen during plant colonization. Even though Salmonella utilized some of its virulence-related genes in tomatoes, plant colonization required a distinct set of functions.KEYWORDS Salmonella, foodborne pathogens, plant-microbe interactions
Results show a significant decrease in quality of life between no pads (1.16 or pleased), a security pad and 0 or 1 pad (2.78 and 3.41 or mixed, respectively). Findings do not support defining continence with a security pad or 0 to 1 pad. Continence should be strictly defined as 0 pads.
Aims: To investigate human viral contamination in urban rivers and its impact on coastal waters of southern California, USA. Methods and Results: Three types of human viruses (adeno, entero and hepatitis A) were detected using nestedand RT-PCR from 11 rivers and creeks. Faecal indicator bacteria as well as somatic and F-specific coliphage were also tested. Approximately 50% of the sites were positive for human adenoviruses. However, there was no clear relationship between detection of human viruses and the concentration of indicator bacteria and coliphage. Both faecal indicator bacteria and human viral input at beaches near river mouths were associated with storm events. The first storm of the wet season seemed to have the greatest impact on the quality of coastal water than following storm events. Conclusions: This study provides the first direct evidence that human viruses are prevalent in southern California urban rivers. Urban run-off impacts coastal water quality most significantly during the storm season. Significance and Impact of the Study: To protect human health during water recreational activities, it is necessary to develop effective strategies to manage urban run-off during storm events.
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