Cost-effective application of pulsed-field gel electrophoresis to typing of Salmonella enterica Serovar Typhimurium

Salmonella enterica serovar Typhimurium is frequently isolated from humans and animals. Phage typing is historically the first-line reference typing technique in Europe. It is rapid and convenient for laboratories with appropriate training and experience, and costs of consumables are low. Phage typing and pulsed-field gel electrophoresis (PFGE) were performed on 503 isolates of serovar Typhimurium. Twenty-nine phage types and 53 PFGE patterns were observed. Most isolates of phage types DT104, DT104b, and U310 are not distinguishable from other members of their phage type by PFGE. By contrast, PFGE of isolates of phage types DT193 and U302 shows great heterogeneity. Analysis of experience with PFGE and phage typing can facilitate the selective application of PFGE to maximize the yield of epidemiologically relevant additional information while controlling costs.     

Genetic and phenotypic variability among Salmonella enterica serovar Typhimurium isolates from California dairy cattle and humans

Fifty-six human and 24 adult dairy cattle isolates of Salmonella enterica serovar Typhimurium from a single county in California were compared using ribotyping, insertion sequence typing (IS200), pulsed-field gel electrophoresis, plasmid typing, phage typing, and antimicrobial resistance testing. The majority of the isolates fell into one of two groups which were phage types DT104 and DT193. Combining the information from all typing methods, a total of 45 different "clusters" were defined, with 35 of those including only a single isolate. The library of isolates had a high degree of variability, but antibiotic resistance and plasmid typing each defined single clusters in which human or bovine isolates predominated (chi2, P < 0.05).     

Molecular epidemiology of Salmonella enterica serovar typhimurium isolates from cattle in hokkaido, Japan: evidence of clonal replacement and characterization of the disseminated clone

The molecular epidemiology of 545 Salmonella enterica serovar Typhimurium isolates collected between 1977 and 2009 from cattle in Hokkaido, Japan, was investigated using pulsed-field gel electrophoresis (PFGE). Nine main clusters were identified from 116 PFGE patterns. Cluster I comprised 248 isolates, 243 of which possessed a sequence specific to definitive phage type 104 (DT104) or U302. The cluster I isolates were dominant in 1993 to 2003, but their numbers declined beginning in 2004. Beginning in 2002, an increase was observed in the number of cluster VII isolates, consisting of 21 PFGE patterns comprising 165 isolates. A total of 116 isolates representative of the 116 PFGE profiles were analyzed by multilocus variable-number tandem-repeat analysis (MLVA). Other than two drug-sensitive isolates, 19 isolates within cluster VII were classified in the same cluster by MLVA. Among the cluster VII isolates, an antibiotic resistance type showing resistance to ampicillin, chloramphenicol, streptomycin, sulfonamides, tetracycline, kanamycin, cefazolin, and sulfamethoxazole-trimethoprim and a resistance type showing resistance to ampicillin, streptomycin, sulfonamides, tetracycline, and kanamycin were found in 23 and 125 isolates, respectively. In the 19 isolates representative of cluster VII, the bla(TEM-1) gene was found on a Salmonella serotype Typhimurium virulence plasmid, which was transferred to Escherichia coli by electroporation along with resistance to two to four other antimicrobials. Genomic analysis by subtractive hybridization and plasmid analysis suggested that the bla(TEM-1)-carrying virulence plasmid has a mosaic structure composed of elements of different origin. These results indicate an emerging multidrug-resistant S. Typhimurium clone carrying a virulence-resistance plasmid among cattle in Hokkaido, Japan.     

Cost-Effective Application of Pulsed-Field Gel Electrophoresis to Typing of Salmonella enterica Serovar Typhimurium

Salmonella enterica serovar Typhimurium is frequently isolated from humans and animals. Phage typing is historically the first-line reference typing technique in Europe. It is rapid and convenient for laboratories with appropriate training and experience, and costs of consumables are low. Phage typing and pulsed-field gel electrophoresis (PFGE) were performed on 503 isolates of serovar Typhimurium. Twenty-nine phage types and 53 PFGE patterns were observed. Most isolates of phage types DT104, DT104b, and U310 are not distinguishable from other members of their phage type by PFGE. By contrast, PFGE of isolates of phage types DT193 and U302 shows great heterogeneity. Analysis of experience with PFGE and phage typing can facilitate the selective application of PFGE to maximize the yield of epidemiologically relevant additional information while controlling costs.     

Molecular epidemiology of Salmonella enterica serovar typhimurium isolates determined by pulsed-field gel electrophoresis: comparison of isolates from avian wildlife,domestic animals,and the environment in Norway

The molecular epidemiology of 142 isolates of Salmonella enterica serovar Typhimurium from avian wildlife, domestic animals, and the environment in Norway was investigated using pulsed-field gel electrophoresis (PFGE) and computerized numerical analysis of the data. The bacterial isolates comprised 79 isolates from wild-living birds, including 46 small passerines and 26 gulls, and 63 isolates of nonavian origin, including 50 domestic animals and 13 environmental samples. Thirteen main clusters were discernible at the 90% similarity level. Most of the isolates (83%) were grouped into three main clusters. These were further divided into 20 subclusters at the 95% similarity level. Isolates from passerines, gulls, and pigeons dominated within five subclusters, whereas isolates from domestic animals and the environment belonged to many different subclusters with no predominance. The results support earlier results that passerines constitute an important source of infection to humans in Norway, whereas it is suggested that gulls and pigeons, based on PFGE analysis, represent only a minor source of human serovar Typhimurium infections. Passerines, gulls, and pigeons may also constitute a source of infection of domestic animals and feed plants or vice versa. Three isolates from cattle and a grain source, of which two were multiresistant, were confirmed as serovar Typhimurium phage type DT 104. These represent the first reported phage type DT 104 isolates from other sources than humans in Norway.     

Molecular Epidemiology of Salmonella enterica Serovar Typhimurium Isolates Determined by Pulsed-Field Gel Electrophoresis: Comparison of Isolates from Avian Wildlife, Domestic Animals, and the Environment in Norway

The molecular epidemiology of 142 isolates of Salmonella enterica serovar Typhimurium from avian wildlife, domestic animals, and the environment in Norway was investigated using pulsed-field gel electrophoresis (PFGE) and computerized numerical analysis of the data. The bacterial isolates comprised 79 isolates from wild-living birds, including 46 small passerines and 26 gulls, and 63 isolates of nonavian origin, including 50 domestic animals and 13 environmental samples. Thirteen main clusters were discernible at the 90% similarity level. Most of the isolates (83%) were grouped into three main clusters. These were further divided into 20 subclusters at the 95% similarity level. Isolates from passerines, gulls, and pigeons dominated within five subclusters, whereas isolates from domestic animals and the environment belonged to many different subclusters with no predominance. The results support earlier results that passerines constitute an important source of infection to humans in Norway, whereas it is suggested that gulls and pigeons, based on PFGE analysis, represent only a minor source of human serovar Typhimurium infections. Passerines, gulls, and pigeons may also constitute a source of infection of domestic animals and feed plants or vice versa. Three isolates from cattle and a grain source, of which two were multiresistant, were confirmed as serovar Typhimurium phage type DT 104. These represent the first reported phage type DT 104 isolates from other sources than humans in Norway.     

Epidemiological characterization of Salmonella Typhimurium DT104 prevalent among food-producing animals in the Japanese veterinary antimicrobial resistance monitoring program (1999-2001)

In the course of nationwide investigation on epidemiological characteristics in Salmonella Typhimurium isolates from food-producing animals in Japan between 1999 and 2001, fifty-seven isolates of S. Typhimurium DT104 and 104B obtained from cattle and swine at farm level in Japan between 1999 and 2001 were classified with pulsotype and antimicrobial resistance type. Most of the isolates were resistant to five or more antimicrobials and were genotyped into four groups. The present nationwide investigation shows that at least 11 types of S. Typhimurium related to DT104 are prevalent among food-producing animals across the country.     

Transmission routes of Salmonella Typhimurium DT 104 between 14 cattle and pig herds in Denmark demonstrated by molecular fingerprinting

AIMS: Salmonella Typhimurium DT 104 is generally assumed to be spread by contact between live animals, e.g. by trading. The aim of the present study was to assess the importance of other routes of transmission in the dissemination of this bacterium. METHODS AND RESULTS: An outbreak among 14 cattle and pig herds located in a geographically narrow area in Denmark was investigated. Epidemiological information and disease history of the herds was obtained through interviews. Based on this, the hypothesis for horizontal spread was proposed, and these were confirmed by comparison of the pulsed field gel electrophoresis (PFGE) and the plasmid profiles of isolates obtained by continuous sampling over a period of almost 3 years. CONCLUSIONS: The study indicated that other routes might play an important role, than the trading of live animals, in the spread of S. Typhimurium DT 104 among livestock. SIGNIFICANCE AND IMPACT OF THE STUDY: Salmonella Typhimurium DT 104 infected herd might pose a significant risk to herds located within the same geographic area. In advising on how to avoid the spread of this bacterium, factors like person contacts, sharing of equipment and contaminated slurry should be focussed on in addition to infected animals.     

Characterization of the genomes of a diverse collection of Salmonella enterica serovar Typhimurium definitive phage type 104

Salmonella enterica serovar Typhimurium definitive phage type 104 (DT104) has caused significant morbidity and mortality in humans and animals for almost three decades. We completed the full DNA sequence of one DT104 strain, NCTC13348, and showed that significant differences between the genome of this isolate and the genome of the previously sequenced strain Salmonella serovar Typhimurium LT2 are due to integrated prophage elements and Salmonella genomic island 1 encoding antibiotic resistance genes. Thirteen isolates of Salmonella serovar Typhimurium DT104 with different pulsed-field gel electrophoresis (PFGE) profiles were analyzed by using multilocus sequence typing (MLST), plasmid profiling, hybridization to a pan-Salmonella DNA microarray, and prophage-based multiplex PCR. All the isolates belonged to a single MLST type, sequence type ST19. Microarray data demonstrated that the gene contents of the 13 DT104 isolates were remarkably conserved. The PFGE DNA fragment size differences in these isolates could be explained to a great extent by differences in the prophage and plasmid contents. Thus, here the nature of variation in different Salmonella serovar Typhimurium DT104 isolates is further defined at the gene and whole-genome levels, illustrating how this phage type evolves over time.     

A multiple typing approach is recommended for the laboratory-based surveillance of salmonellosis in Romania

In Romania, Salmonella enterica serovar Typhimurium isolates are currently typed by antimicrobial resistance profiles and phage typing, as part of the national laboratory-based surveillance system of human enteric infections. The aim of the present study was to assess the added value of complementing this approach with molecular fingerprinting, namely pulsed-field gel electrophoresis (PFGE) and multiple-locus variable-number tandem-repeats analysis (MLVA). Thirty-six S. Typhimurium isolates received by the Reference Center for Human Salmonella Infections for confirmation and typing from the Microbiology Departments of three Public Health Authorities, were selected for this study. Phage typing revealed that 14 isolates (39%) were nontypeable (NT). Twenty-two isolates were assigned to 5 phage types: DT193 (11 isolates), U302 (7 isolates), DT116 (2 isolates), DT41 (1 isolate) and DT86 (1 isolate). Antimicrobial susceptibility testing showed that all the NT and DT116 isolates were multidrug resistant and extended-spectrum betalactamase producers. All the examined isolates were typeable when using the molecular approach. Both methods gave conclusive and comparable results, documenting the genetic relatedness and discriminating the outbreak isolates from sporadic cases. We conclude that in order to improve outbreak investigation and surveillance of salmonellosis in Romania, the current routine typing of Salmonella isolates should be complemented with at least one of these DNA fingerprinting methods.     

The prevalence of Salmonella spp. in bovine faecal,rumen and carcass samples at a commercial abattoir

AIMS: To determine the prevalence, serotype and antibiotic resistance profile of Salmonella isolates in cattle and on carcasses at a commercial Irish abattoir. METHODS AND RESULTS: Faecal, rumen and carcass samples were collected from a beef abattoir over a 12-month period and examined for the presence of Salmonella spp. Isolates were serotyped, phage typed (when serotype was found to be S. Typhimurium) and tested for susceptibility to a panel of antibiotics. Salmonella was isolated from 2% of faecal, 2% of rumen and 7.6% of carcass samples. Salmonella was most frequently isolated from samples taken during the period August to October. S. Dublin was isolated from 72% of positive samples. S. Agona and S. Typhimurium definitive type (DT)104 were each isolated from 14% of positive samples. All S. Typhimurium DT104 isolates were resistant to ampicillin, chloramphenicol, streptomycin, sulphafurazole and tetracycline (ACSSuT). On occasion, from a single animal, the same serotype was isolated from more than one sample (i.e. faeces and rumen; faeces and carcass; rumen and carcass; faeces, rumen and carcass). CONCLUSIONS: Salmonella is present in cattle at slaughter and on beef carcasses at an Irish abattoir, with a higher frequency of occurrence during the period August to October. Most isolates from the study are not commonly associated with human clinical infection, with the exception of S. Typhimurium DT104 (R-type ACSSuT). SIGNIFICANCE AND IMPACT OF THE STUDY: This study provides epidemiological data that is necessary for the understanding of beef as a source of human Salmonella infection.     

Characterization of Salmonella enterica serovar typhimurium from marine environments in coastal waters of Galicia (Spain)

Twenty-three Salmonella enterica serovar Typhimurium isolates from marine environments were characterized by phage typing, pulsed-field gel electrophoresis (PFGE) analysis, plasmid analysis, and antibiotic resistance, and the distribution of the different types in the coastal waters were subsequently analyzed. Five phage types were identified among the isolates (PT41, PT135, PT99, DT104, and DT193). PT135 isolates were exclusively detected during the winter months from 1998 to 2000, whereas DT104 and PT41 isolates were detected exclusively in the summer months from 2000 to 2002. XbaI PFGE analysis revealed 9 PFGE types, and plasmid profiling identified 8 plasmid types (with 1 to 6 plasmids) among the isolates. Only three isolates presented multidrug resistance to antibiotics. Two DT104 isolates were resistant to 8 and 7 antibiotics (profiles ACCeFNaSSuT and ACeFNeSSuT), whereas a PT193 isolate presented resistance to 6 antibiotics (profile ACFSSu). In addition, four PT41 isolates were resistant to a single antibiotic. The detection of multidrug-resistant phage types DT104 and DT193 in shellfish emphasizes the importance of monitoring the presence of Salmonella in routine surveillance of live bivalve molluscs.     

Heterogeneity in expression of lipopolysaccharide by strains of Salmonella enterica serotype Typhimurium definitive phage type 104 and related phage types

AIMS: To investigate lipopolysaccharide (LPS) expression in Salmonella enterica serotype Typhimurium definitive phage type 104 (Salmonella Typhimurium DT104) and related phage types. METHODS AND RESULTS: Isolates were examined for the expression of LPS by SDS-PAGE and silver staining and subtyped by Pulsed Field Gel Electrophoresis (PFGE). The 100 isolates expressed one of two LPS profiles designated A (72%) and B (28%). LPS profiling was able to discriminate between isolates of identical PFGE type. Among 10 groups of outbreak isolates examined, each group was of a single LPS profile: A, 8/10 and B, 2/10. All 10 outbreaks were identical by PFGE analysis. CONCLUSIONS: Isolates of Salmonella Typhimurium DT104 and related phage types expressed one of two distinct LPS profiles. The two LPS profiles appear similar but shifted and in phase with one another, suggesting that the heterogeneity is due to changes in the LPS core region rather than among the repeating oligosaccharide units of the long-chain LPS. SIGNIFICANCE AND IMPACT OF THE SUTDY: LPS profiling provides a useful adjunct to PFGE and other molecular methods for the subtyping of this group of bacteria in epidemiological investigations.     

Molecular characterization of Irish Salmonella enterica serotype typhimurium: detection of class I integrons and assessment of genetic relationships by DNA amplification fingerprinting

Salmonella enterica is among the principal etiological agents of food-borne illness in humans. Increasing antimicrobial resistance in S. enterica is a cause for worldwide concern. There is concern at present in relation to the increasing incidence of human infection with antimicrobial agent-resistant strains of S. enterica serotype Typhimurium, in particular of phage type DT104. Integrons appear to play an important role in the dissemination of antimicrobial resistance genes in many Enterobacteriaceae including S. enterica. In this study the antimicrobial susceptibilities and phage types of 74 randomly collected strains of S. enterica serotype Typhimurium from the Cork region of southern Ireland, obtained from human, animal (clinical), and food sources, were determined. Each strain was examined for integrons and typed by DNA amplification fingerprinting (DAF). Phage type DT104 predominated (n = 48). Phage types DT104b (n = 3), -193 (n = 9), -195 (n = 6), -208 (n = 3), -204a (n = 2), PT U302 (n = 1), and two nontypeable strains accounted for the remainder. All S. enterica serotype Typhimurium DT104 strains were resistant to ampicillin, chloramphenicol, streptomycin, Sulfonamide Duplex, and tetracycline, and one strain was additionally resistant to trimethoprim. All DT104 strains but one were of a uniform DAF type (designated DAF-I) and showed a uniform pattern of integrons (designated IP-I). The DT104b and PT U302 strains also exhibited the same resistance phenotype, and both had the DAF-I and IP-I patterns. The DAF-I pattern was also observed in a single DT193 strain in which no integrons were detectable. Greater diversity of antibiograms and DAF and IP patterns among non-DT104 phage types was observed. These data indicate a remarkable degree of homogeneity at a molecular level among contemporary isolates of S. enterica serotype Typhimurium DT104 from animal, human, and food sources in this region.     

Antimicrobial susceptibility, phage types, and pulse types of Salmonella Typhimurium, in São Paulo, Brazil

A total of 283 Salmonella Typhimurium strains isolated from cases of human infections and non human sources, were examined for antimicrobial susceptibility and the incidence of resistance was 38% and multiple resistance (to three or more antimicrobials) was 15%. All 43 multidrug-resistant strains (MDR) and 13 susceptible ones were characterized by phage typing and pulsed- field gel electrophoresis (PFGE). The strains encompassed 14 definitive phage types (DT), three were untypable (UT), and 18 atypicals or reaction does not conform (RDNC), which belonged to 21 PFGE patterns, A1-A21. The predominant phage types were DT49, DT193, and RDNC and two strains belonging to DT 104 and 104b were also identified. The most common PFGE patterns were A1 and A8. Analysis by PFGE and phage typing demonstrated that the most of the MDR were multiclonal and association among multiresistance, phage typing, and PFGE patterns was not so significant.     

National survey for Salmonella in pigs, cattle and sheep at slaughter in Great Britain (1999-2000)

AIMS: The objective of these surveys was to estimate the prevalence of faecal carriage of Salmonella in healthy pigs, cattle and sheep at slaughter, and of pig carcase contamination with Salmonella. These data can be used as a baseline against which future change in Salmonella prevalence in these species at slaughter can be monitored. METHODS AND RESULTS: In this first randomized National Survey for faecal carriage of Salmonella in slaughter pigs, cattle and sheep in Great Britain, 2509 pigs, 891 cattle and 973 sheep were sampled in 34 pig abattoirs and 117 red meat abattoirs in England, Scotland and Wales. Carriage of Salmonella in 25 g caecal contents was identified in 578 (23.0% pigs) but in only 134 (5.3%) of carcase swabs. The predominant Salmonella serovars found in both types of sample were S. Typhimurium (11.1% caeca, 2.1% carcases) and S. Derby (6.3% caeca, 1.6% carcases). The main definitive phage types (DT) of S. Typhimurium found were DT104 (21.9% of caecal S. Typhimurium isolates), DT193 (18.7%), untypable strains (17.6%), DT208 (13.3%) and U302 (13.3%). Three isolates of S. Enteritidis (PTs 13A and 4) and one enrofloxacin-resistant S. Choleraesuis were also isolated. A positive 'meat-juice ELISA' was obtained from 15.2% of pigs at 40% optical density (O.D.) cut-off level and 35.7% at 10% cut-off. There was poor correlation between positive ELISA results or carcase contamination and the caecal carriage of Salmonella. The ratio of carcase contamination to caecal carriage rates was highest in abattoirs from the midland region of England and in smaller abattoirs. In cattle and sheep 1 g samples of rectal faeces were tested. Two isolates (i.e. 0.2%) were recovered from cattle, one each of S. Typhimurium, DT193 and DT12. One sheep sample (0.1%) contained a Salmonella, S. Typhimurium DT41. In a small subsidiary validation exercise using 25 g of rectal faeces from 174 cattle samples, three (1.7%) isolates of Salmonella (S. Typhimurium DT104, S. Agama, S. Derby) were found. CONCLUSIONS: The carriage rate of Salmonella in prime slaughter cattle and sheep in Great Britain was very low compared with pigs. This suggests that future control measures should be focused on reduction of Salmonella infection on pig farms and minimizing contamination of carcases at slaughter. SIGNIFICANCE AND IMPACT OF THE STUDY: This work has set baseline figures for Salmonella carriage in these species slaughtered for human consumption in Great Britain. These figures were collected in a representative way, which enables them to be used for monitoring trends and setting control targets.     

Characterization of Salmonella enterica Serovar Typhimurium from Marine Environments in Coastal Waters of Galicia (Spain)

Twenty-three Salmonella enterica serovar Typhimurium isolates from marine environments were characterized by phage typing, pulsed-field gel electrophoresis (PFGE) analysis, plasmid analysis, and antibiotic resistance, and the distribution of the different types in the coastal waters were subsequently analyzed. Five phage types were identified among the isolates (PT41, PT135, PT99, DT104, and DT193). PT135 isolates were exclusively detected during the winter months from 1998 to 2000, whereas DT104 and PT41 isolates were detected exclusively in the summer months from 2000 to 2002. XbaI PFGE analysis revealed 9 PFGE types, and plasmid profiling identified 8 plasmid types (with 1 to 6 plasmids) among the isolates. Only three isolates presented multidrug resistance to antibiotics. Two DT104 isolates were resistant to 8 and 7 antibiotics (profiles ACCeFNaSSuT and ACeFNeSSuT), whereas a PT193 isolate presented resistance to 6 antibiotics (profile ACFSSu). In addition, four PT41 isolates were resistant to a single antibiotic. The detection of multidrug-resistant phage types DT104 and DT193 in shellfish emphasizes the importance of monitoring the presence of Salmonella in routine surveillance of live bivalve molluscs.     

Genetic and Phenotypic Variability among Salmonella enterica Serovar Typhimurium Isolates from California Dairy Cattle and Humans

Fifty-six human and 24 adult dairy cattle isolates of Salmonella enterica serovar Typhimurium from a single county in California were compared using ribotyping, insertion sequence typing (IS200), pulsed-field gel electrophoresis, plasmid typing, phage typing, and antimicrobial resistance testing. The majority of the isolates fell into one of two groups which were phage types DT104 and DT193. Combining the information from all typing methods, a total of 45 different “clusters” were defined, with 35 of those including only a single isolate. The library of isolates had a high degree of variability, but antibiotic resistance and plasmid typing each defined single clusters in which human or bovine isolates predominated (χ², P < 0.05).     

Identification of novel Salmonella enterica serovar Typhimurium DT104-specific prophage and nonprophage chromosomal sequences among serovar Typhimurium isolates by genomic subtractive hybridization

Genomic subtractive hybridization was performed between Salmonella enterica serovar Typhimurium LT2 and DT104 to search for novel Salmonella serovar Typhimurium DT104-specific sequences. The subtraction resulted mainly in the isolation of DNA fragments with sequence similarity to phages. Two fragments identified were associated with possible virulence factors. One fragment was identical to irsA of Salmonella serovar Typhimurium ATCC 14028, which is suggested to be involved in macrophage survival. The other fragment was homologous to HldD, an Escherichia coli O157:H7 lipopolysaccharide assembly-related protein. Five selected DNA fragments-irsA, the HldD homologue, and three fragments with sequence similarity to prophages-were tested for their presence in 17 Salmonella serovar Typhimurium DT104 isolates and 27 non-DT104 isolates by PCR. All five selected DNA fragments were Salmonella serovar Typhimurium DT104 specific among the serovar Typhimurium isolates tested. These DNA fragments can be useful for better detection and typing of Salmonella serovar Typhimurium DT104.     

Immunoassays to detect the serum antibody response of cattle to infection with Salmonella Typhimurium definitive type 104 and following vaccination with Bovivac S

AIMS: To use ELISA and immunoblotting assays to examine the serum antibody response of cattle infected with Salmonella Typhimurium DT104 and following vaccination with Bovivac S. METHODS AND RESULTS: Three hundred and twenty-nine cattle, including 16 shedding multiresistant Salmonella Typhimurium DT104, were screened for serum antibodies binding to O=1, 4, 5, 12 lipopolysaccharide (LPS) antigens before and after vaccination with Bovivac S. Sera with an ELISA reading of 0.9A405 or above were shown to contain antibodies, of the IgG-class only, to the LPS of Salmonella Typhimurium using immunoblotting. Prior to vaccination, only 11 cattle had serum IgG-class antibodies to the O=4, 5 LPS antigens, and of these one also had antibodies to outer membrane proteins and H=i flagellar antigens. Following vaccination, 87 out of 315 cattle developed serum antibodies to the LPS of Salmonella Typhimurium. CONCLUSIONS: Evidence of infection of cattle with Salmonella Typhimurium was readily obtained with an LPS-based ELISA in association with an immunoblotting procedure, supplementing existing bacteriological procedures. This enabled the detection of an increase in the number of cattle with serum antibodies to Salmonella Typhimurium LPS following vaccination with Bovivac S. SIGNIFICANCE AND IMPACT OF THE STUDY: The immunoassays described provided evidence of infection with Salmonella Typhimurium and served as a valuable adjunct to established bacteriology.