We describe the investigation of two temporally coincident illness clusters involving salmonella and Staphylococcus aureus in two states. Cases were defined as gastrointestinal illness following two meal events. Investigators interviewed ill persons. Stool, food and environmental samples underwent pathogen testing. Alabama: Eighty cases were identified. Median time from meal to illness was 5.8 h. Salmonella Heidelberg was identified from 27 of 28 stool specimens tested, and coagulase-positive S. aureus was isolated from three of 16 ill persons. Environmental investigation indicated that food handling deficiencies occurred. Colorado: Seven cases were identified. Median time from meal to illness was 4.5 h. Five persons were hospitalised, four of whom were admitted to the intensive care unit. Salmonella Heidelberg was identified in six of seven stool specimens and coagulase-positive S. aureus in three of six tested. No single food item was implicated in either outbreak. These two outbreaks were linked to infection with Salmonella Heidelberg, but additional factors, such as dual aetiology that included S. aureus or the dose of salmonella ingested may have contributed to the short incubation periods and high illness severity. The outbreaks underscore the importance of measures to prevent foodborne illness through appropriate washing, handling, preparation and storage of food.

Live poultry-associated salmonellosis is an emerging public health issue in the United States. Public and animal health officials collaborated to investigate one of the largest (356 cases, 39 states) of these outbreaks reported to date. A case was defined as illness in a person infected with the outbreak strain of Salmonella Typhimurium with illness onset between 1 March and 22 October 2013. The median patient age was seven years (range: < 1–87 years); 58% of ill persons were children ≤ 10 years, 51% were female, 25% were hospitalized; 189 (76%) of 250 patients reported live poultry exposure in the week before illness; and 149 (95%) of 157 reported purchasing live poultry from agricultural feed stores. Traceback investigations identified 18 live poultry sources, including 16 mail-order hatcheries. Environmental sampling was conducted at two mail-order hatcheries. One (2.5%) of 40 duplicate samples collected at one hatchery yielded the outbreak strain. Live poultry are an important source of human salmonellosis, particularly among children, highlighting the need for educational campaigns and comprehensive interventions at the mail-order hatchery and agricultural feed store levels. Prevention and control efforts depend on a One Health approach, involving cooperation between public and animal health officials, industry, health professionals, and consumers.

Vibrio parahaemolyticus is an aquatic halophilic bacterium that occupies estuarine and coastal marine environments, and is a leading cause of seafood-borne food poisoning cases. To investigate the environmental reservoir and potential gene flow that occurs among V. parahaemolyticus isolates, the virulence-associated gene content and genome diversity of a collection of 133 V. parahaemolyticus isolates were analyzed. Phylogenetic analysis of housekeeping genes, and pulsed-field gel electrophoresis, demonstrated that there is genetic similarity among V. parahaemolyticus clinical and environmental isolates. Whole-genome sequencing and comparative analysis of six representative V. parahaemolyticus isolates was used to identify genes that are unique to the clinical and environmental isolates examined. Comparative genomics demonstrated an O3:K6 environmental isolate, AF91, which was cultured from sediment collected in Florida in 2006, has significant genomic similarity to the post-1995 O3:K6 isolates. However, AF91 lacks the majority of the virulence-associated genes and genomic islands associated with these highly virulent post-1995 O3:K6 genomes. These findings demonstrate that although they do not contain most of the known virulence-associated regions, some V. parahaemolyticus environmental isolates exhibit significant genetic similarity to clinical isolates. This highlights the dynamic nature of the V. parahaemolyticus genome allowing them to transition between aquatic and host-pathogen states.

OBJECTIVE: Although amphibians are known Salmonella carriers, no such outbreaks have been reported. We investigated a nationwide outbreak of human Salmonella Typhimurium infections occurring predominantly among children from 2008 to 2011. METHODS: We conducted a matched case-control study. Cases were defined as persons with Salmonella Typhimurium infection yielding an isolate indistinguishable from the outbreak strain. Controls were persons with recent infection with Salmonella strains other than the outbreak strain and matched to cases by age and geography. Environmental samples were obtained from patients' homes; traceback investigations were conducted. RESULTS: We identified 376 cases from 44 states from January 1, 2008, to December 31, 2011; 29% (56/193) of patients were hospitalized and none died. Median patient age was 5 years (range <1-86 years); 69% were children <10 years old (253/367). Among 114 patients interviewed, 69 (61%) reported frog exposure. Of patients who knew frog type, 79% (44/56) reported African dwarf frogs (ADF), a type of aquatic frog. Among 18 cases and 29 controls, illness was significantly associated with frog exposure (67% cases versus 3% controls, matched odds ratio 12.4, 95% confidence interval 1.9-infinity). Environmental samples from aquariums containing ADFs in 8 patients' homes, 2 ADF distributors, and a day care center yielded isolates indistinguishable from the outbreak strain. Traceback investigations of ADFs from patient purchases converged to a common ADF breeding facility. Environmental samples from the breeding facility yielded the outbreak strain. CONCLUSIONS: ADFs were the source of this nationwide pediatric predominant outbreak. Pediatricians should routinely inquire about pet ownership and advise families about illness risks associated with animals.

During the 2010 cholera outbreak in Haiti, water and seafood samples were collected to detect Vibrio cholerae. The outbreak strain of toxigenic V. cholerae O1 serotype Ogawa was isolated from freshwater and seafood samples. The cholera toxin gene was detected in harbor water samples.

The PulseNet Methods Development and Validation Laboratory began a re-evaluation of the standardized pulsed-field gel electrophoresis (PFGE) protocols with the goal of optimizing their overall performance and robustness. Herein, we describe a stepwise evaluation of the PulseNet-standardized PFGE protocol for Listeria monocytogenes that led to the modification of several steps which significantly improved the overall appearance and reproducibility of the resulting PFGE data. These improvements included the following: (1) reducing the cell suspension concentration, (2) increasing lysozyme incubation temperature from 37 degrees C to 56 degrees C, and (3) decreasing the number of units of restriction enzymes AscI and ApaI. These changes were incorporated into a proposed protocol that was evaluated by 16 PulseNet participating laboratories, including 2 international participants. Results from the validation study indicated that the updated L. monocytogenes protocol is more robust than the original PulseNet-standardized protocol established in 1998 and this resulted in the official adoption of the new protocol into the PulseNet system in the spring of 2008. The modifications not only represent an improvement to the protocol but also describe procedural improvements that could be potentially applied to the PFGE analysis of other Gram-positive organisms.

Vibrio parahaemolyticus is a pathogenic marine bacterium that is the main causative agent of bacterial seafood borne gastroenteritis in the United States. An increase in the frequency of V. parahaemolyticus-related infections during the last decade has been attributed to the emergence of an O3:K6 pandemic clone in 1995. The diversity of the O3:K6 pandemic clone and serovariants has been examined using multiple molecular techniques including multilocus sequence analysis (MLSA), pulsed-field gel electrophoresis (PFGE), and group-specific PCR (GS-PCR) analysis. Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) has become a powerful tool for rapidly distinguishing between related bacterial species. In the current study we demonstrate the development of a whole-cell MALDI-TOF MS method for the identification of V. parahaemolyticus from other Vibrio spp. We identified 30 peaks that were present only in the spectra of the V. parahaemolyticus strains examined in this study that may be developed as MALDI-TOF MS biomarkers for identification of V. parahaemolyticus. We detected variation in the MALDI-TOF spectra of V. parahaemolyticus strains isolated from different geographical locations and at different times. The MALDI-TOF MS spectra of the V. parahaemolyticus strains examined were distinct from the other Vibrios examined including the closely related V. alginolyticus, V. harveyi, and V. campbellii. The results of this study demonstrate the first use of whole-cell MALDI-TOF MS analysis for the rapid identification of V. parahaemolyticus.