Epidemiology of Rhodococcus equi strains on Thoroughbred horse farms

Pulsed-field gel electrophoresis of restriction endonuclease-digested genomic DNA from a large collection of clinical isolates of Rhodococcus equi, an important pathogen of foals, was used to compare strain distribution between farms and over time. Forty-four strains were found among 209 isolates, with 5 of these accounting for over half the isolates and the 22 strains isolated more than once accounting for 90% of the isolates. The average genotypic diversity on each farm and in each year was found to be less than the genotypic diversity of the isolates taken as a whole, with 5.2% of the total diversity being due to differences between farms and 5.5% to differences between years. A small number of strains on each farm were found to have caused at least half the clinical cases of disease, and these varied between farms and, to a lesser extent, years. Most strains were found on more than one farm, and some very similar restriction patterns were found among isolates from different continents, indicating that strains can be very widespread. Multiple strains were isolated in five of the six cases in which more than one isolate from a single foal was examined, indicating that disease may commonly be caused by simultaneous infection with multiple strains. It was concluded that there are a number of different strains of R. equi which carry the vapA gene, and these strains tend to be widespread, but individual farms tend to have particular strains associated with them.     

Plasmid Profiles of Virulent Rhodococcus equi Strains Isolated from Infected Foals in Poland

Rhodococcus equi is an important bacterial pathogen in foals up to 6 months old, widespread in horse farms all over the world. It was found that only virulent R. equi strains expressing 15–17 kDa virulence-associated protein (VapA) and having large virulence plasmid of 85–90 kb containing vapA gene are pathogenic for horses. To date, 12 plasmid types have been reported in VapA positive strains from horses. There are no data concerning plasmid types of Polish field R. equi strains isolated from horses and horse farm environment. The aim of the study is to determine plasmid profiles of virulent R. equi strains isolated in Poland from dead foals as well as from soil samples taken from horse breeding farms. Plasmid profiles of 10 clinical strains derived from 8 farms and 11 environmental strains from 3 farms, confirmed as virulent by PCR, were compared with 12 reference strains containing the known plasmid size and type. Plasmid DNAs were analysed by digestion with the restriction endonucleases BamHI, EcoRI, EcoT22I, and HindIII for detailed comparison and estimation of plasmid sizes. The results of RFLP analysis revealed that all except one isolates used in the study are classified as VapA 85 kb type I plasmid. One strain harboured VapA 87 kb type I plasmid. This is the first report of plasmid types of Polish field R. equi strains. The results of our preliminary investigations on horse farms located in central and eastern Poland indicate that the virulent R. equi strains thus far isolated from diseased foals and horse farms environment represent a highly uniform plasmid pattern.     

Prevalence of Clostridium botulinum in Finnish Trout Farms: Pulsed-Field Gel Electrophoresis Typing Reveals Extensive Genetic Diversity among Type E Isolates

The distribution of Clostridium botulinum serotypes A, B, E, and F in Finnish trout farms was examined. A total of 333 samples were tested with a neurotoxin-specific PCR assay. C. botulinum type E was found in 68% of the farm sediment samples, in 15% of the fish intestinal samples, and in 5% of the fish skin samples. No other serotypes were found. The spore counts determined by the most-probable-number method were considerably higher for the sediments than for the fish intestines and skin; the average values were 2,020, 166, and 310 C. botulinum type E spores kg⁻¹, respectively. The contamination rates in traditional freshwater ponds and marine net cages were high, but in concrete ponds equipped with sediment suction devices the contamination rates were significantly lower. Pulsed-field gel electrophoresis (PFGE) typing of 42 isolates obtained in this survey and 12 North American reference strains generated 28 pulsotypes upon visual inspection, suggesting that there was extensive genetic diversity and that the discriminatory power of PFGE typing in C. botulinum type E was high. A numerical analysis of SmaI-XmaI macrorestriction profiles confirmed these findings, as it divided the 54 isolates into 15 clusters at a similarity level of 76%. For this material, this level of similarity corresponded to a three-band difference in the macrorestriction profiles, which indicated that there is no genotypic proof of a close epidemiological relationship among the clusters.     

Genetic Diversity and Antibiotic Resistance Patterns in a Campylobacter Population Isolated from Poultry Farms in Switzerland

The diversity and genetic interrelation of Campylobacter jejuni and C. coli isolated from Swiss poultry were assessed by three independent typing methods. Samples were derived prior to slaughter from 100 randomly selected flocks (five birds per flock) raised on three different farm types. The observed flock prevalence was 54% in total, with 50% for conventional and 69% for free-range farms. Birds held on farms with a confined roaming area had the lowest prevalence of 37%. Campylobacter isolates were characterized by amplified fragment length polymorphism (AFLP), restriction fragment length polymorphism of flaA PCR fragments (flaA-RFLP), and disk diffusion testing for eight antimicrobial agents that are commonly used in veterinary or human medicine in Switzerland. Analysis of the genotypic results indicates that the Campylobacter population in Swiss poultry is genetically highly diverse. Nevertheless, occasionally, isolates with identical or nearly identical characteristics were isolated from different farms or farm types in different locations. Genetic typing by AFLP and flaA-RFLP was found to be complementary. The majority of isolates (67%) were susceptible to all tested antibiotics; however, single, double, and triple resistances were observed in 7%, 23%, and 2% of the strains, respectively. There was no correlation between genotype and antibiotic resistance. Surprisingly, sulfonamide resistance was frequently found together with streptomycin resistance. Our findings illustrate the results of common genetic exchange in the studied bacterial population.     

Longitudinal Study of Escherichia coli O157:H7 Dissemination on Four Dairy Farms in Wisconsin

A 14-month longitudinal study was conducted on four dairy farms (C, H, R, and X) in Wisconsin to ascertain the source(s) and dissemination of Escherichia coli O157:H7. A cohort of 15 heifer calves from each farm were sampled weekly by digital rectal retrieval from birth to a minimum of 7 months of age (range, 7 to 13 months). Over the 14 months of the study, the cohort heifers and other randomly selected cattle from farms C and H tested negative. Farm R had two separate periods of E. coli O157:H7 shedding lasting 4 months (November 1995 to February 1996) and 1 month (July to August 1996), while farm X had at least one positive cohort animal for a 5-month period (May to October 1996). Heifers shed O157:H7 strains in feces for 1 to 16 weeks at levels ranging from 2.0 × 10² to 8.7 × 10⁴ CFU per g. E. coli O157:H7 was also isolated from other noncohort cattle, feed, flies, a pigeon, and water associated with the cohort heifers on farms R and/or X. When present in animal drinking water, E. coli O157:H7 disseminated through the cohort cattle and other cattle that used the water source. E. coli O157:H7 was found in water at <1 to 23 CFU/ml. Genomic subtyping by pulsed-field gel electrophoresis demonstrated that a single O157:H7 strain comprised a majority of the isolates from cohort and noncohort cattle, water, and other positive samples (i.e., from feed, flies, and a pigeon, etc.) on a farm. The isolates from farm R displayed two predominant XbaI restriction endonuclease digestion profiles (REDP), REDP 3 and REDP 7, during the first and second periods of shedding, respectively. Six additional REDP that were ≥89% similar to REDP 3 or REDP 7 were identified among the farm R isolates. Additionally, the REDP of an O157:H7 isolate from a heifer on farm R in 1994 was indistinguishable from REDP 3. Farm X had one O157:H7 strain that predominated (96% of positive samples had strains with REDP 9), and the REDP of an isolate from a heifer in 1994 was indistinguishable from REDP 9. These results suggest that E. coli O157:H7 is disseminated from a common source on farms and that strains can persist in a herd for a 2-year period.     

Identification of the Most Abundant Lactobacillus Species in the Crop of 1- and 5-Week-Old Broiler Chickens

Bacteria from crops of 1- and 5-week-old broiler chickens fed with two brands (diets A and B) of wheat-based diets were isolated on Lactobacillus-selective medium and identified (n = 300) based on partial 16S rRNA gene sequence. The most abundant Lactobacillus species were L. reuteri (33%), L. crispatus (18.7%), and L. salivarius (13.3%). Regardless of farm and feed, L. reuteri was the most abundant species (P < 0.005) in the crops of the younger chickens. However, the amount of L. reuteri was significantly reduced in the crops of the 5-week-old chickens regardless of the feed (P = 0.016). The diversity of L. reuteri isolates was studied by fatty acid analysis, and the 94 L. reuteri isolates could be arranged into several clusters. The nisin sensitivities of the L. reuteri isolates were determined because nisin is a candidate coccidiostat. Sensitive isolates were found more frequently in younger chickens (77%) than in 5-week-old chickens (23%), whereas chickens fed with commercial feed B had a higher proportion of nisin-resistant isolates (73%) than did chickens fed with feed A (45%). Nisin-resistant strains are potential candidates for adjunct cultures for maintaining L. reuteri in its natural niche in the crop and are potential targets for genetic engineering with nisin-selectable food-grade vectors. The diversity of the L. reuteri population suggested that one should consider including several strains representing different clusters and nisin resistance phenotypes in candidate probiotic feed supplements for chickens.     

The diversity of Listeria monocytogenes strains from 10 Icelandic sheep farms

AIMS: The purpose of this study was to examine the diversity of Listeria monocytogenes strains from healthy sheep, winter feed and environment of sheep farms in Iceland. METHODS AND RESULTS: A total of 104 L. monocytogenes isolates from animals, winter feed and environment on 10 Icelandic sheep farms were compared by serotyping, ribotyping, and pulsed-field gel electrophoresis with ApaI and AscI. The isolates were divided into 24 genotypes, all identified as serovars 1/2a, 1/2b, or 4b. Nine genotypes were detected on more than one farm. On three of the farms there seemed to be a dominant strain of L. monocytogenes. Isolates from incidents of listeriosis in animals occurring on two of the farms belonged to the genotype most commonly found on the particular farm. Nine of the 24 genotypes found on the sheep farms have been associated with disease in animals and/or humans elsewhere in Iceland. CONCLUSIONS: Certain strains of L. monocytogenes seem to be widely distributed on Icelandic sheep farms. On some farms there appears to be a dominant strain of L. monocytogenes. Incidents of listeriosis in animals may tend to be associated with strains commonly found on the farm. SIGNIFICANCE AND IMPACT OF THE STUDY: This study demonstrates the diversity of L. monocytogenes present in healthy sheep and their environment.     

Genetic diversity and antibiotic resistance patterns in a campylobacter population isolated from poultry farms in Switzerland

The diversity and genetic interrelation of Campylobacter jejuni and C. coli isolated from Swiss poultry were assessed by three independent typing methods. Samples were derived prior to slaughter from 100 randomly selected flocks (five birds per flock) raised on three different farm types. The observed flock prevalence was 54% in total, with 50% for conventional and 69% for free-range farms. Birds held on farms with a confined roaming area had the lowest prevalence of 37%. Campylobacter isolates were characterized by amplified fragment length polymorphism (AFLP), restriction fragment length polymorphism of flaA PCR fragments (flaA-RFLP), and disk diffusion testing for eight antimicrobial agents that are commonly used in veterinary or human medicine in Switzerland. Analysis of the genotypic results indicates that the Campylobacter population in Swiss poultry is genetically highly diverse. Nevertheless, occasionally, isolates with identical or nearly identical characteristics were isolated from different farms or farm types in different locations. Genetic typing by AFLP and flaA-RFLP was found to be complementary. The majority of isolates (67%) were susceptible to all tested antibiotics; however, single, double, and triple resistances were observed in 7%, 23%, and 2% of the strains, respectively. There was no correlation between genotype and antibiotic resistance. Surprisingly, sulfonamide resistance was frequently found together with streptomycin resistance. Our findings illustrate the results of common genetic exchange in the studied bacterial population.     

Genotype Analysis of Escherichia coli Strains Isolated from Children and Chickens Living in Close Contact

Escherichia coli isolates from rectal swabs from 62 chickens and stools from 42 children living in close contact with chickens on the same farms in Kiambu district, Kenya, were compared for their genetic relatedness. Antibiotic susceptibility profiles broadly categorized isolates from the children and from the chickens into two separate clusters: the majority (144; 85.5%) of the E. coli isolates from children were multidrug resistant, while the majority (216; 87.1%) of the E. coli isolates from chickens were either fully susceptible or resistant only to tetracycline. Sixty- and 100- to 110-MDA plasmids were found to encode the transferable resistance to co-trimoxazole and tetracycline. HindIII restriction endonuclease digestion of the 60- and 100- to 110-MDA plasmids produced four distinct patterns for isolates from children and three distinct patterns for isolates from chickens. XbaI digestion of genomic DNA followed by pulsed-field gel electrophoresis (PFGE) analysis produced 14 distinct clusters. There were six distinct PFGE clusters among the isolates from children, while among the isolates from chickens there were seven distinct clusters. Only one PFGE cluster contained isolates from both children and chickens, with the isolates displaying an approximately 60% coefficient of similarity. This study showed that although several different genotypes of E. coli were isolated from children and chickens from the same farms, the E. coli strains from these two sources were distinct.     

Phenotypic and Genotypic Characterization of Non-Starter Lactic Acid Bacteria in Mature Cheddar Cheese

Non-starter lactic acid bacteria were isolated from 14 premium-quality and 3 sensorially defective mature Irish Cheddar cheeses, obtained from six manufacturers. From countable plates of Lactobacillus-selective agar, 20 single isolated colonies were randomly picked per cheese. All 331 viable isolates were biochemically characterized as mesophilic (i.e., group II) Lactobacillus spp. Phenotypically, the isolates comprised 96.4% L. paracasei, 2.1% L. plantarum, 0.3% L. curvatus, 0.3% L. brevis, and 0.9% unidentified species. Randomly amplified polymorphic DNA (RAPD) analysis was used to rapidly identify the dominant strain groups in nine cheeses from three of the factories, and through clustering by the unweighted pair group method with arithmetic averages, an average of seven strains were found per cheese. In general, strains isolated from cheese produced at the same factory clustered together. The majority of isolates associated with premium-quality cheese grouped together and apart from clusters of strains from defective-quality cheese. No correlation was found between the isomer of lactate produced and RAPD profiles, although isolates which did not ferment ribose clustered together. The phenotypic and genotypic methods employed were validated with a selection of 31 type and reference strains of mesophilic Lactobacillus spp. commonly found in Cheddar cheese. RAPD analysis was found to be a useful and rapid method for identifying isolates to the species level. The low homology exhibited between RAPD banding profiles for cheese isolates and collection strains demonstrated the heterogeneity of the L. paracasei complex.     

Silage Collected from Dairy Farms Harbors an Abundance of Listeriaphages with Considerable Host Range and Genome Size Diversity

Since the food-borne pathogen Listeria monocytogenes is common in dairy farm environments, it is likely that phages infecting this bacterium (“listeriaphages”) are abundant on dairy farms. To better understand the ecology and diversity of listeriaphages on dairy farms and to develop a diverse phage collection for further studies, silage samples collected on two dairy farms were screened for L. monocytogenes and listeriaphages. While only 4.5% of silage samples tested positive for L. monocytogenes, 47.8% of samples were positive for listeriaphages, containing up to >1.5 × 10⁴ PFU/g. Host range characterization of the 114 phage isolates obtained, with a reference set of 13 L. monocytogenes strains representing the nine major serotypes and four lineages, revealed considerable host range diversity; phage isolates were classified into nine lysis groups. While one serotype 3c strain was not lysed by any phage isolates, serotype 4 strains were highly susceptible to phages and were lysed by 63.2 to 88.6% of phages tested. Overall, 12.3% of phage isolates showed a narrow host range (lysing 1 to 5 strains), while 28.9% of phages represented broad host range (lysing ≥11 strains). Genome sizes of the phage isolates were estimated to range from approximately 26 to 140 kb. The extensive host range and genomic diversity of phages observed here suggest an important role of phages in the ecology of L. monocytogenes on dairy farms. In addition, the phage collection developed here has the potential to facilitate further development of phage-based biocontrol strategies (e.g., in silage) and other phage-based tools.     

Genetic Diversity of Listeria monocytogenes Strains from a High-Prevalence Dairy Farm

Listeria monocytogenes is a significant food-borne human and veterinary pathogen. Contaminated silage commonly leads to disease in livestock, but the pervasive nature of the bacterium can make it difficult to identify the source of infection. An investigation of bovine listeriosis that occurred on a Pacific Northwest dairy farm (“farm A”) revealed that the clinical strain was closely related to fecal strains from asymptomatic cows, and that farm environment was heavily contaminated with a diversity of L. monocytogenes strains. In addition, the farm A clinical strain was closely related to clinical and environmental strains obtained 1 year prior from a second Northwest dairy farm (“farm B”). To investigate the source(s) of contamination on farm A, environmental samples were collected from farm A at two time points. Pulsed-field gel electrophoresis characterization of 538 isolates obtained from that farm identified 57 different AscI pulsovars. Fecal isolates obtained from individual cows were the most genetically diverse, with up to 94% of fecal samples containing more than one pulsovar. The maximum numbers of pulsovars and serotypes isolated from a fecal sample of one cow were 6 and 4, respectively. Serotype 1/2a was isolated most frequently at both time points. Microarray genotyping of bovine listeriosis, fecal, and silage strains from both farms identified four probes that differentiated listeriosis strains from environmental strains; however, no probe was common to both bovine listeriosis strains.     

Use of Pulsed-Field Gel Electrophoresis and Flagellin Gene Typing in Identifying Clonal Groups of Campylobacter jejuni and Campylobacter coli in Farm and Clinical Environments

Although campylobacters have been isolated from a wide range of animal hosts, the association between campylobacters isolated from humans and animals in the farm environment is unclear. We used flagellin gene typing and pulsed-field gel electrophoresis (PFGE) to investigate the genetic diversity among isolates from animals (cattle, sheep, and turkey) in farm environments and sporadic cases of campylobacteriosis in the same geographical area. Forty-eight combined fla types were seen among the 315 Campylobacter isolates studied. Six were found in isolates from all four hosts and represented 50% of the total number of isolates. Seventy-one different SmaI PFGE macrorestriction profiles (mrps) were observed, with 86% of isolates assigned to one of 29 different mrps. Fifty-seven isolates from diverse hosts, times, and sources had an identical SmaI mrp and combined fla type. Conversely, a number of genotypes were unique to a particular host. We provide molecular evidence which suggests a link between campylobacters in the farm environment with those causing disease in the community.     

An Ecological Study of Lactococci Isolated from Raw Milk in the Camembert Cheese Registered Designation of Origin Area

The genetic diversity of lactococci isolated from raw milk in the Camembert cheese Registered Designation of Origin area was studied. Two seasonal samples (winter and summer) of raw milk were obtained from six farms in two areas (Bessin and Bocage Falaisien) of Normandy. All of the strains analyzed had a Lactococcus lactis subsp. lactis phenotype, whereas the randomly amplified polymorphic DNA (RAPD) technique genotypically identified the strains as members of L. lactis subsp. lactis or L. lactis subsp. cremoris. The genotypes were confirmed by performing standard PCR with primers corresponding to a region of the histidine biosynthesis operon. The geographic distribution of each subspecies of L. lactis was determined; 80% of the Bocage Falaisien strains were members of L. lactis subsp. lactis, and 30.5% of the Bessin strains were members of L. lactis subsp. lactis. A dendrogram was produced from a computer analysis of the RAPD profiles in order to evaluate the diversity of the lactococci below the subspecies level. The coefficient of similarity for 117 of the 139 strains identified as members of L. lactis subsp. cremoris was as high as 66%. The L. lactis subsp. lactis strains were more heterogeneous and formed 10 separate clusters (the level of similarity among the clusters was 18%). Reference strains of L. lactis subsp. lactis fell into 2 of these 10 clusters, demonstrating that lactococcal isolates are clearly different. As determined by the RAPD profiles, some L. lactis subsp. lactis strains were specific to the farms from which they originated and were recovered throughout the year (in both summer and winter). Therefore, the typicality of L. lactis subsp. lactis strains was linked to the farm of origin rather than the area. These findings emphasize the significance of designation of origin and the specificity of “Camembert de Normandie” cheese.     

Diversity of isolates of Rhodococcus equi from Australian Thoroughbred horse farms

Pulsed field gel electrophoresis of restriction endonuclease digested genomic DNA from a collection of clinical isolates of Rhodococcus equi was used to compare strain diversity on different Thoroughbred horse farms over time. Restricted diversity was found among the isolates tested, as the same strains were detected on multiple farms and in multiple years. Marked variation occurred in strain prevalence with some strains being represented by single isolates, and the most prevalent by 26 isolates. There were dominant strains on some farms and the prevalence of some strains differed between farms. Infection with multiple strains was noted in some cases where multiple isolates from a single foal were examined.     

Genotypic and Phenotypic Diversity of phlD-Containing Pseudomonas Strains Isolated from the Rhizosphere of Wheat

Production of 2,4-diacetylphloroglucinol (2,4-DAPG) in the rhizosphere by strains of fluorescent Pseudomonas spp. results in the suppression of root diseases caused by certain fungal plant pathogens. In this study, fluorescent Pseudomonas strains containing phlD, which is directly involved in the biosynthesis of 2,4-DAPG, were isolated from the rhizosphere of wheat grown in soils from wheat-growing regions of the United States and The Netherlands. To assess the genotypic and phenotypic diversity present in this collection, 138 isolates were compared to 4 previously described 2,4-DAPG producers. Thirteen distinct genotypes, one of which represented over 30% of the isolates, were differentiated by whole-cell BOX-PCR. Representatives of this group were isolated from eight different soils taken from four different geographic locations. ERIC-PCR gave similar results overall, differentiating 15 distinct genotypes among all of the isolates. In most cases, a single genotype predominated among isolates obtained from each soil. Thirty isolates, representing all of the distinct genotypes and geographic locations, were further characterized. Restriction analysis of amplified 16S rRNA gene sequences revealed only three distinct phylogenetic groups, one of which accounted for 87% of the isolates. Phenotypic analyses based on carbon source utilization profiles revealed that all of the strains utilized 49 substrates and were unable to grow on 12 others. Individually, strains could utilize about two-thirds of the 95 substrates present in Biolog SF-N plates. Multivariate analyses of utilization profiles revealed phenotypic groupings consistent with those defined by the genotypic analyses.     

Multilocus Sequence Typing, Pulsed-Field Gel Electrophoresis, and fla Short Variable Region Typing of Clonal Complexes of Campylobacter jejuni Strains of Human, Bovine, and Poultry Origins in Luxembourg

Campylobacter jejuni is the most common cause of bacterial gastroenteritis in Luxembourg, with a marked seasonal peak during summer. The majority of these infections are thought to be sporadic, and the relative contribution of potential sources and reservoirs is still poorly understood. We monitored human cases from June to September 2006 (n = 124) by molecular characterization of isolates with the aim of rapidly detecting temporally related cases. In addition, isolates from poultry meat (n = 36) and cattle cecal contents (n = 48) were genotyped for comparison and identification of common clusters between veterinary and human C. jejuni populations. A total of 208 isolates were typed by sequencing the fla short variable region, macrorestriction analysis resolved by pulsed-field gel electrophoresis (PFGE), and multilocus sequence typing (MLST). We observed a high diversity of human strains during a given summer season. Poultry and human isolates had a higher diversity of sequence types than isolates of bovine origin, for which clonal complexes CC21 (41.6%) and CC61 (18.7%) were predominant. CC21 was also the most common complex found among human isolates (21.8%). The substantial concordance between PFGE and MLST results for this last group of strains suggests that they are clonally related. Our study indicates that while poultry remains an important source, cattle could be an underestimated reservoir of human C. jejuni cases. Transmission mechanisms of cattle-specific strains warrant further investigation.     

Temporal Dynamics and Impact of Manure Storage on Antibiotic Resistance Patterns and Population Structure of Escherichia coli Isolates from a Commercial Swine Farm

Many confined-livestock farms store their wastes for several months prior to use as a fertilizer. Storing manure for extended periods could significantly bias the composition of enteric bacterial populations subsequently released into the environment. Here, we compared populations of Escherichia coli isolated from fresh feces and from the manure-holding tank (stored manure) of a commercial swine farm, each sampled monthly for 6 months. The 4,668 confirmed E. coli isolates were evaluated for resistance to amikacin, ampicillin, cephalothin, chloramphenicol, kanamycin, nalidixic acid, streptomycin, sulfamethoxazole, tetracycline, trimethoprim, and trimethoprim plus sulfamethoxazole. A subset of 1,687 isolates was fingerprinted by repetitive extragenic palindromic PCR (rep-PCR) with the BOXA1R primer to evaluate the diversity and the population structure of the collection. The population in the stored manure was generally more diverse than that in the fresh feces. Half of the genotypes detected in the stored manure were never detected in the fresh fecal material, and only 16% were detected only in the fresh feces. But the majority of the isolates (84%) were assigned to the 34% of genotypes shared between the two environments. The structure of the E. coli population showed important monthly variations both in the extent and distribution of the diversity of the observed genotypes. The frequency of detection of resistance to specific antibiotics was not significantly different between the two collections and varied importantly between monthly samples. Resistance to multiple antibiotics was much more temporally dynamic in the fresh feces than in the stored manure. There was no relationship between the distribution of rep-PCR fingerprints and the distribution of antibiotic resistance profiles, suggesting that specific antibiotic resistance determinants were dynamically distributed within the population.     

Genetic Diversity and Biological Control Activity of Novel Species of Closely Related Pseudomonads Isolated from Wheat Field Soils in South Australia

Rhizobacteria closely related to two recently described species of pseudomonads, Pseudomonas brassicacearum and Pseudomonas thivervalensis, were isolated from two geographically distinct wheat field soils in South Australia. Isolation was undertaken by either selective plating or immunotrapping utilizing a polyclonal antibody raised against P. brassicacearum. A subset of 42 isolates were characterized by amplified 16S ribosomal DNA restriction analysis (ARDRA), BIOLOG analysis, and gas chromatography-fatty acid methyl ester (GC-FAME) analysis and separated into closely related phenetic groups. More than 75% of isolates tested by ARDRA were found to have >95% similarity to either Pseudomonas corrugata or P. brassicacearum-P. thivervalensis type strains, and all isolates had >90% similarity to either type strain. BIOLOG and GC-FAME clustering showed a >70% match to ARDRA profiles. Strains representing different ARDRA groups were tested in two soil types for biological control activity against the soilborne plant pathogen Gaeumannomyces graminis var. tritici, the causative agent of take-all of wheat and barley. Three isolates out of 11 significantly reduced take-all-induced root lesions on wheat plants grown in a red-brown earth soil. Only one strain, K208, was consistent in reducing disease symptoms in both the acidic red-brown earth and a calcareous sandy loam. Results from this study indicate that P. brassicacearum and P. thivervalensis are present in Australian soils and that a level of genetic diversity exists within these two novel species but that this diversity does not appear to be related to geographic distribution. The result of the glasshouse pot trial suggests that some isolates of these species may have potential as biological control agents for plant disease.     

Antimicrobial Resistance in Salmonella enterica Serovar Heidelberg Isolates from Retail Meats, Including Poultry, from 2002 to 2006

Salmonella enterica serovar Heidelberg frequently causes food-borne illness in humans. There are few data on the prevalence, antimicrobial susceptibility, and genetic diversity of Salmonella serovar Heidelberg isolates in retail meats. We compared the prevalences of Salmonella serovar Heidelberg in a sampling of 20,295 meats, including chicken breast (n = 5,075), ground turkey (n = 5,044), ground beef (n = 5,100), and pork chops (n = 5,076), collected during 2002 to 2006. Isolates were analyzed for antimicrobial susceptibility and compared genetically using pulsed-field gel electrophoresis (PFGE) and PCR for the bla_CMY gene. A total of 298 Salmonella serovar Heidelberg isolates were recovered, representing 21.6% of all Salmonella serovars from retail meats. One hundred seventy-eight (59.7%) were from ground turkey, 110 (36.9%) were from chicken breast, and 10 (3.4%) were from pork chops; none was found in ground beef. One hundred ninety-eight isolates (66.4%) were resistant to at least one compound, and 49 (16.4%) were resistant to at least five compounds. Six isolates (2.0%), all from ground turkey, were resistant to at least nine antimicrobials. The highest resistance in poultry isolates was to tetracycline (39.9%), followed by streptomycin (37.8%), sulfamethoxazole (27.7%), gentamicin (25.7%), kanamycin (21.5%), ampicillin (19.8%), amoxicillin-clavulanic acid (10.4%), and ceftiofur (9.0%). All isolates were susceptible to ceftriaxone and ciprofloxacin. All ceftiofur-resistant strains carried bla_CMY. PFGE using XbaI and BlnI showed that certain clones were widely dispersed in different types of meats and meat brands from different store chains in all five sampling years. These data indicate that Salmonella serovar Heidelberg is a common serovar in retail poultry meats and includes widespread clones of multidrug-resistant strains.