Lorin D. Warnick

Emergence, distribution, and molecular and phenotypic characteristics of Salmonella enterica serotype 4,5,12:i:-.

Salmonella spp. represent one of the most common causes of bacterial foodborne illnesses around the world. The species Salmonella enterica contains more than 2500 serotypes, and emergence of new human pathogenic Salmonella strains and serotypes represents a major public health issue. Salmonella enterica subsp. enterica serotype 4,5,12:i:- represents a monophasic variant of Salmonella Typhimurium, which has rarely been identified before the mid-1990 s. The prevalence of this serotype among human salmonellosis cases has increased considerably since the mid-1990 s and Salmonella 4,5,12:i:- currently (i.e., the first decade of the 2000s) represents one of the most common serotypes among human cases in many countries around the world. This paper discusses our current knowledge of the global ecology, epidemiology, transmission, and evolution of this emerging Salmonella serotype.

Immune-mediated hemolytic anemia: 70 cases (1988-1996).

Survival times and mortality rates in dogs with idiopathic immune-mediated hemolytic anemia (IMHA) have been infrequently reported in the literature. This study evaluates survival and mortality in a large group of dogs with IMHA. The association of age, sex, and breed with IMHA was evaluated by comparing affected dogs to control dogs admitted to the hospital during the same time period. Treatment regimens were reviewed to determine the effects of different agents upon survival of dogs with IMHA during hospitalization and after discharge. Median survival times for each treatment group were 57 days (prednisone), 28 days (prednisone, cyclophosphamide), 974 days (prednisone, azathioprine), 15 days (prednisone, cyclophosphamide, azathioprine), and one day (no treatment). Overall mortality rate in the population of dogs studied was 70%. Twenty-nine (41.4%) dogs either died or were euthanized while hospitalized. Forty-one (59%) dogs were discharged from the hospital. Of the dogs discharged, 10 died within the first month, another five died within three months, and another five died within a year of discharge due to assumed complications of therapy or relapses of IMHA.

DNA Sequence-Based Subtyping and Evolutionary Analysis of Selected Salmonella enterica Serotypes

ABSTRACT While serotyping and phage typing have been used widely to characterize Salmonella isolates, sensitive subtyping methods that allow for evolutionary analyses are essential for examining Salmonella transmission, ecology, and evolution. A set of 25 Salmonella enterica isolates, representing five clinically relevant serotypes (serotypes Agona, Heidelberg, Schwarzengrund, Typhimurium, and Typhimurium var. Copenhagen) was initially used to develop a multilocus sequence typing (MLST) scheme for Salmonella targeting seven housekeeping and virulence genes (panB, fimA, aceK, mdh, icdA, manB, and spaN). A total of eight MLST types were found among the 25 isolates sequenced. A good correlation between MLST types and Salmonella serotypes was observed; only one serotype Typhimurium var. Copenhagen isolate displayed an MLST type otherwise typical for serotype Typhimurium isolates. Since manB, fimA, and mdh allowed for the highest subtype discrimination among the initial 25 isolates, we chose these three genes to perform DNA sequencing of an additional 41 Salmonella isolates representing a larger diversity of serotypes. This “three-gene sequence typing scheme” allowed discrimination of 25 sequence types (STs) among a total of 66 isolates; STs correlated well with serotypes and allowed within-serotype differentiation for 9 of the 12 serotypes characterized. Phylogenetic analyses showed that serotypes Kentucky and Newport could each be separated into two distinct, statistically well supported evolutionary lineages. Our results show that a three-gene sequence typing scheme allows for accurate serotype prediction and for limited subtype discrimination among clinically relevant serotypes of Salmonella. Three-gene sequence typing also supports the notion that Salmonella serotypes represent both monophyletic and polyphyletic lineages.

Risk factors for clinical salmonellosis in Virginia, USA cattle herds

Veterinarians working in several Virginia, USA counties noticed an increase in clinical salmonellosis in cattle herds during 1994. We conducted a case-control study to identify potential risk factors for salmonellosis in cattle herds in this region. The potential impact of exposure to poultry-related factors was of particular interest because of the close proximity of poultry and cattle operations in this part of the state. Information was collected from 23 case farms and 23 control farms matched by herd veterinarian, type of enterprise (beef vs. dairy), and location. Faecal and environmental samples were collected during herd visits for bacteriologic culture. Salmonella was isolated from 4.7% of 531 faecal, feed, water, and environmental samples collected. Salmonella Typhimurium was the serotype found most frequently in clinical cases in participating herds and from samples collected during study visits. Case herds had a higher percentage of study samples positive for Salmonella. Potential risk factors were screened using methods for pair-matched data and then analyzed in individual conditional logistic-regression models to estimate odds ratios with 95% profile likelihood confidence intervals. Significant factors included: the number of mature cows in the herd (OR=1.01; CI 1.00, 1.03), percent change in the number of mature cows during 1994 (OR=0.68; CI 0.11, 0.96), having calves usually born in a building rather than outdoors (OR=0.17; CI 0.01, 0.98), poultry manure spread on bordering property (OR=4.00; CI 1.00, 26.50), signs of rodents in cattle-housing or feed-storage areas (OR=2.75; CI 0.94, 9.92), and contact of wild geese with cattle or feed (OR=4.5; 1.16, 29.51).

Salmonella enterica Serotype 4,5,12:i:−, an Emerging Salmonella Serotype That Represents Multiple Distinct Clones

ABSTRACT The prevalence, among human clinical cases, of Salmonella enterica serotype 4,5,12:i:−, a serotype antigenically similar to Salmonella enterica serotype Typhimurium but lacking second-phase flagellar antigens, has increased considerably over the last 10 years. To probe the evolution and ecology of this emerging serotype, we characterized 190 Salmonella isolates initially classified as Salmonella serotypes 4,5,12:i:− (n = 90) and Typhimurium (n = 100) and obtained from various sources in the United States and Spain. These isolates were characterized into six sequence types (determined by multilocus sequence typing [MLST]) and 79 pulsed-field gel electrophoresis types. The majority of Salmonella serotype 4,5,12:i:− and Typhimurium isolates (85 and 84 isolates, respectively) represented a single MLST type. Existing genome information revealed different genome deletions (which included genes responsible for phase 2 flagellum expression) in four Spanish Salmonella serotype 4,5,12:i:− isolates and one U.S. Salmonella serotype 4,5,12:i:− isolate. Fifty-nine isolates of both serotypes, representing different sources and geographical locations as well as different molecular subtypes, were thus screened for the presence of six genes and one specific region, all of which were previously found to show variable presence among Salmonella serotype 4,5,12:i:− and Typhimurium strains. All Salmonella serotype 4,5,12:i:− isolates lacked the phase 2 flagella genes fljA and fljB, which were present in all Salmonella serotype Typhimurium isolates. While all Spanish Salmonella serotype 4,5,12:i:− isolates carried the same deletion surrounding fljAB, all but two U.S. isolates showed a different genomic deletion; the two atypical U.S. isolates represented the “Spanish” deletion genotype and a unique deletion genotype. Salmonella serotype 4,5,12:i:− thus appears to represent at least two common clones, which cannot easily be differentiated with standard diagnostic procedures.

Ceftiofur-Resistant Salmonella Strains Isolated from Dairy Farms Represent Multiple Widely Distributed Subtypes That Evolved by Independent Horizontal Gene Transfer

ABSTRACT Salmonella is the leading cause of known food-borne bacterial infections in the United States, with an incidence rate of approximately 15 cases per 100,000 people. The rise of antimicrobial-resistant Salmonella subtypes, including the appearance of subtypes resistant to ceftriaxone, represents a particular concern. Ceftriaxone is used to treat invasive cases of Salmonella in children and is closely related to ceftiofur, an antibiotic commonly used to treat diseases of cattle. In order to develop a better understanding of the evolution and transmission of ceftiofur resistance in Salmonella, we characterized ceftiofur-resistant and -sensitive Salmonella isolates from seven New York dairy farms. A total of 39 isolates from these seven farms were analyzed for evolutionary relatedness (by DNA sequencing of the Salmonella genes fimA, manB, and mdh), antibiotic resistance profiles, and the presence of blaCMY-2, a beta-lactamase gene associated with resistance to cephalosporins. Our data indicate that (i) resistance to ceftriaxone and ceftiofur was highly correlated with the presence of blaCMY-2; (ii) ceftiofur-resistant Salmonella strains were geographically widespread, as shown by their isolation from farms located throughout New York State; (iii) ceftiofur-resistant Salmonella strains isolated from farms represent multiple distinct subtypes and evolutionary lineages, as determined by serotyping, DNA sequence typing, and antimicrobial-resistance profiles; and (iv) ceftiofur-resistant Salmonella strains evolved by multiple independent acquisitions of an identical blaCMY-2 allele and by clonal spread of ceftiofur-resistant subtypes.

Strategies to Analyze Milk Losses Caused by Diseases with Potential Incidence Throughout the Lactation : A Lameness Example

The objective of this study was to test the hypothesis that lameness incidence in lactating dairy cows decreases milk production compared with controls and to determine the best method of the 5 combinations of 2 study designs (retrospective cohort and retrospective matched cohort) and 3 repeated measures statistical methods: ANOVA (with binary classification of lameness); ANOVA with a lameness index (LAMIX) as the variable of interest; and analysis of covariance (ANCOVA, controlling for mean milk production for the first 3 wk of lactation and with a binary classification of lameness). Data were from 1 dairy farm located near Ithaca, NY, and from cows that calved between January 1, 2004 and January 1, 2007. The LAMIX was defined as -1, -2, -3, -4, and -5 = 1, 2, 3, 4, and > or =5 mo before lameness was detected, respectively; 1, 2, 3, 4, and 5 = 1, 2, 3, 4, and 5 mo after diagnosis, respectively; and 0 = cows that were never lame. With the binary lameness classification analyzed by repeated measures ANOVA, there was no effect of lameness on milk yield. The model was biased because lame cows had higher milk yields before lameness compared with nonlame cows. When the LAMIX was used, milk production before lameness was greater than after lameness (3.1 +/- 0.28 kg/d). Yet, point estimates generated for LAMIX were inaccurate because of the multicollinearity detected between LAMIX and week of lactation and because of the inability of adjusting the least squares means for the interaction of LAMIX and week of lactation. Therefore, the most appropriate models were the ANCOVA models (both for the matched and nonmatched retrospective-cohort designs). The estimated losses associated with lameness were 314 and 424 kg/cow per 305-d lactation, respectively, for the matched and nonmatched designs. Furthermore, high milk yield in the beginning of the lactation was a risk factor for lameness.

Antimicrobial resistance in nontyphoidal Salmonella.

Salmonella is one of the leading causes of foodborne illness in countries around the world. Treatment of Salmonella infections, in both animals and humans has become more difficult with the emergence of multidrug-resistant (MDR) Salmonella strains. Foodborne infections and outbreaks with MDR Salmonella are also increasingly reported. To better monitor and control the spread of MDR Salmonella, it is important to understand the mechanisms responsible for drug resistance and how drug resistance is transmitted to and between Salmonella strains. This review summarizes current knowledge on antimicrobial drugs used to treat Salmonella infections and provides an overview of MDR Salmonella in the United States and a discussion of the genetics of Salmonella drug resistance, including the mechanisms responsible for the transmission of drug-resistance genes in Salmonella, using data from the United States and other countries.

Multilocus Sequence Typing Supports the Hypothesis that Cow- and Human-Associated Salmonella Isolates Represent Distinct and Overlapping Populations

ABSTRACT A collection of 179 human and 156 bovine clinical Salmonella isolates obtained from across New York state over the course of 1 year was characterized using serotyping and a multilocus sequence typing (MLST) scheme based on the sequencing of three genes (fimA, manB, and mdh). The 335 isolates were differentiated into 52 serotypes and 72 sequence types (STs). Analyses of bovine isolates collected on different farms over time indicated that specific subtypes can persist over time on a given farm; in particular, a number of farms showed evidence for the persistence of a specific Salmonella enterica serotype Newport sequence type. Serotypes and STs were not randomly distributed among human and bovine isolates, and selected serotypes and STs were associated exclusively with either human or bovine sources. A number of common STs were geographically widespread. For example, ST6, which includes isolates representing serotype Typhimurium as well as the emerging serotype 4,5,12:i:-, was found among human and bovine isolates in a number of counties in New York state. Phylogenetic analyses supported the possibility that serotype 4,5,12:i:- is closely related to Salmonella serotype Typhimurium. Salmonella serotype Newport was found to represent two distinct evolutionary lineages that differ in their frequencies among human and bovine isolates. A number of Salmonella isolates carried two copies of manB (33 isolates) or showed small deletion events in fimA (nine isolates); these duplication and deletion events may provide mechanisms for the rapid diversification of Salmonella surface molecules. We conclude that the combined use of an economical three-gene MLST scheme and serotyping can provide considerable new insights into the evolution and transmission of Salmonella.

Evaluation of herd sampling for Salmonella isolation on midwest and northeast US dairy farms

Epidemiologic investigations of Salmonella infections in dairy cattle often rely on testing fecal samples from individual animals or samples from other farm sources to determine herd infection status. The objectives of this project were to evaluate the effect of sampling frequency on Salmonella isolation and to compare Salmonella isolation and serogroup classification among sample sources on 12 US dairy farms sampled weekly for 7-8 weeks. Three herds per state were enrolled from Michigan, Minnesota, New York and Wisconsin based upon predefined herd-size criteria. Weekly samples were obtained from cattle, bulk tank milk, milk filters, water and feed sources and environmental sites. Samples were submitted to a central laboratory for isolation of Salmonella using standard laboratory procedures. The herd average number of cattle fecal samples collected ranged from 26 to 58 per week. Salmonella was isolated from 9.3% of 4049 fecal samples collected from cattle and 12.9% of 811 samples from other sources. Serogroup C1 was found in more than half of the samples and multiple serogroups were identified among isolates from the same samples and farms. The percentage of herd visits with at least one Salmonella isolate from cattle fecal samples increased with overall herd prevalence of fecal shedding. Only the three herds with an average fecal shedding prevalence of more than 15% had over 85% of weekly visits with at least one positive fecal sample. The prevalence of fecal shedding from different groups of cattle varied widely among herds showing that herds with infected cattle may be classified incorrectly if only one age group is tested. Testing environmental sample sources was more efficient for identifying infected premises than using individual cattle fecal samples.