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Escherichia coliO157:H7 is a common cause of foodborne illness in the United States. Beef ground at establishments regulated by the U.S. Department of Agriculture, Food Safety and Inspection Service is routinely tested for E. coliO157:H7. Prior to December 2013, boxed beef product (wholesale cuts of beef, such as beef loin, packaged into bags and boxed for shipping) was not always tested for this pathogen. Downstream processors or retailers may grind the product; and, if the ground beef is not cooked to the recommended temperature, pathogens on the exterior of the beef introduced to the interior through grinding may survive. On 18 October 2013, the Allegheny County Health Department identified two E. coliO157:H7 cases, both of whom were food handlers at restaurant A, a restaurant that ground locally produced boxed beef for hamburgers on site. Case finding was conducted through public messaging, employee surveys, and disease surveillance. All potential cases were interviewed using a standard questionnaire. A confirmed case was defined as laboratory-confirmed E. coliO157:H7 with exposure to restaurant A. A probable case was defined as a patient with compatible symptoms and exposure to restaurant A but without laboratory confirmation. All human and food isolates were characterized by pulsed-field gel electrophoresis and multilocus variable-number tandem repeat analysis. The analysis identified 14 confirmed and 10 probable cases of E. coli; 18 nonintact ground beef samples tested positive for E. coliO157:H7. Nine confirmed cases were restaurant A employees. All confirmed cases recalled eating a restaurant A hamburger in the 10 days before illness onset; most cases reported consuming medium to rare hamburgers. Multiple pulsed-field gel electrophoresis and multilocus variable-number tandem repeat analysis patterns were identified among both the human and ground beef isolates, and the patient isolates matched those found in ground beef samples. Restaurant A voluntarily closed for 1.5 days, changed beef suppliers, ceased grinding beef in-house, and has had no new cases since reopening.

ABSTRACTSalmonella entericasubsp. entericaserovar Newport (S. Newport) is the third most prevalent cause of food-borne salmonellosis. Rapid, efficient, and accurate methods for identification are required to track specific strains of S. Newport during outbreaks. By exploiting the hypervariable nature of virulence genes and clustered regularly interspaced short palindromic repeats (CRISPRs), we previously developed a sequence-based subtyping approach, designated CRISPR–multi-virulence-locus sequence typing (CRISPR-MVLST). To demonstrate the applicability of this approach, we analyzed a broad set of S. Newport isolates collected over a 5-year period by using CRISPR-MVLST and pulsed-field gel electrophoresis (PFGE). Among 84 isolates, we defined 38 S. Newport sequence types (NSTs), all of which were novel compared to our previous analyses, and 62 different PFGE patterns. Our data suggest that both subtyping approaches have high discriminatory abilities (>0.95) with a potential for clustering cases with common exposures. Importantly, we found that isolates from closely related NSTs were often similar by PFGE profile as well, further corroborating the applicability of CRISPR-MVLST. In the first full application of CRISPR-MVLST, we analyzed isolates from a recent S. Newport outbreak. In this blinded study, we confirmed the utility of CRISPR-MVLST and were able to distinguish the 10 outbreak isolates, as defined by PFGE and epidemiological data, from a collection of 20 S. Newport isolates. Together, our data show that CRISPR-MVLST could be a complementary approach to PFGE subtyping for S. Newport.

AbstractThe aim of this study was to identify Salmonellaserotypes infecting cattle in Pennsylvania, to compare infection rates for the predominant serotype, Salmonella entericaserotype Cerro, with the infection rates for the same serotype in humans, and to study the clonal diversity and antimicrobial resistance for this serotype in cattle from 2005 to 2010. Clonal diversity among the selected isolates was studied using pulsed-field gel electrophoresis (PFGE) and repetitive (rep)–polymerase chain reaction (PCR). SalmonellaCerro showed the single largest increase as a cause of cattle infections over the study period. The proportional distribution of SalmonellaCerro serotype among laboratory-submitted Salmonellapositive cases in cattle was 36.1% in the year 2010 compared to 14.3% in 2005. A simultaneous decrease in serotype Newport infections was also observed in cattle (25% in 2005, to 10.1% in 2010). Studies of clonal diversity for cattle and human isolates revealed a predominant PFGE type but showed some variability. All tested isolates (n=60) were susceptible to sulfamethoxazole-trimethoprim, but 2% of cattle isolates (n=1/50) and 20% of human isolates (n=2/10) showed resistance to tetracycline and sulfisoxazole. One human isolate showed additional resistance to ampicillin and gentamicin. This study suggests an increase in SalmonellaCerro infections in the cattle population and a decrease in SalmonellaNewport infections. The increase in Cerro infections appears to be restricted to the cattle population, but occasional human infections occur.

AbstractAim:To examine the prevalence of antimicrobial-resistant Salmonellain chicken meat and correlate with isolates from ill humans.Methods:We isolated Salmonellafrom raw chicken purchased from a randomly selected sample of retail outlets in central Pennsylvania during 2006–2007. Salmonellaisolates from meat were compared, using pulsed-field gel electrophoresis, to isolates in the PulseNet database of Salmonellarecovered from humans.Results:Of 378 chicken meat samples, 84 (22%) contained Salmonella. Twenty-six (31%) of the Salmonellaisolates were resistant to ≥3 antimicrobials and 18 (21%) were resistant to ceftiofur. All ceftiofur-resistant isolates exhibited reduced susceptibility (minimum inhibitory concentration >2 μg/mL) to ceftriaxone and carried a blaCMYgene, as detected by polymerase chain reaction. Among the 28 Salmonellaserovar Typhimurium isolates, 20 (71.4%) were resistant to ≥3 antimicrobials and 12 (42.9%) were resistant to ceftiofur. One ceftiofur-resistant Salmonellaserovar Typhimurium poultry isolate exhibited a rare pulsed-field gel electrophoresis pattern indistinguishable from a human isolate in PulseNet; both isolates carried the blaCMY-2gene.Conclusions:These data demonstrate the presence of multidrug-resistant Salmonellain poultry meat, including blaCMYplasmid-mediated genes that confer resistance to both ceftiofur, used in poultry, and ceftriaxone, used for treating salmonellosis in humans. This study illustrates the potential for molecular subtyping databases to identify related Salmonellaisolates from meat and ill humans, and suggests that chicken could be a source for multidrug-resistant salmonellosis in humans.

ABSTRACTFive different serotypes of Salmonella entericawere implicated in a large outbreak linked to fresh Roma tomatoes served at gas station deli counters in Pennsylvania and nearby states during July 2004: S. entericaserotypes Javiana, Anatum, Thompson, Typhimurium, and Muenchen. One of these serotypes, Anatum, was isolated from both tomatoes and patients. Pulsed-field gel electrophoresis (PFGE) played a key role in identifying the outbreak-associated isolates and distinguishing them from unrelated sporadic isolates. It also demonstrated that the genetic fingerprints of serotype Anatum isolates derived from patients were indistinguishable from those derived from tomatoes. Rapid communication of PFGE fingerprints with other public health laboratories through the Centers for Disease Control and Prevention's PulseNet USA national molecular surveillance network for bacterial food-borne pathogens facilitated the tracking of this outbreak in other states. The work described in this report emphasizes the laboratory's role in core public health functions and services, thereby providing a highly visible example of public health in action.

ABSTRACTCertain Escherichia colistrains bind the Fc fragment of immunoglobulin G (IgG) at the bacterial cell surface. Previous work established that this nonimmune Ig binding depends on several large proteins with apparent molecular masses that can exceed 200 kDa. ForE. colistrain ECOR-9, four distinct genes (designatedeibA, eibC, eibD, andeibE) are responsible for Ig binding. Two eibgenes are linked to eaagenes, which are homologous to genes for the autotransporter family of secreted proteins. With reference to the E. coliK-12 chromosome, theeibA-eaaAcluster is adjacent to trpA(min 28.3) while the eibC-eaaCcluster is adjacent toaspS(min 42.0). Sequence adjacent to theeibA-eaaAcluster converges with that of strain K-12 precisely as observed for the Atlas family of prophages, suggesting that eibAis part of one of these. All four eibgenes, when cloned into plasmid vectors, impart IgG binding to E. coliK-12 strains, and three impart IgA binding also. The IgG binding occurs at the bacterial cell surface, and its expression increases survival in serum by up to 3 orders of magnitude. Theeibsequences predict a C-terminal peptide motif that is characteristic of outer membrane proteins, and the protein sequences show significant similarity near the C terminus to both the YadA virulence factor of Yersiniaspecies and the universal surface protein A II of Moraxella catarrhalis. The sizes predicted for Eib proteins from DNA sequence are much smaller than their apparent sizes on sodium dodecyl sulfate-polyacrylamide gel electrophoresis, possibly reflecting stable oligomerization.