Further monitoring for Salmonella species and Escherichia coli O86 at a bird table in south-west Scotland

Pooled faeces collected from a garden bird table were screened for Salmonella species and Escherichia coli O86, two recognised causes of garden bird mortality. Dead birds found at the site were also screened for these organisms, and bird numbers and meteorological data were recorded. In the first year of the study, 48.5 per cent of the samples were positive for Salmonella Typhimurium DT56 (variant), decreasing to 38.9 per cent in the second year and 12.8 per cent in the third year. E. coli O86 was not recovered from any of the 288 samples of pooled faeces tested. S. Typhimurium DT56 (variant) was recovered from the carcases of eight birds found dead at the site, and E. coli O86 was isolated from three carcases. The degree of contamination of the bird table with S. Typhimurium DT56 (variant) was positively correlated with the number of house sparrows observed in the garden, but not with the number of blackbirds, chaffinches or greenfinches.     

Epidemiology and characterization of animal Salmonella enterica subspecies Enterica serotype typhimurium DT104 in Hungary

Reports on the internationally emerging significance of multiresistant zoonotic Salmonella in animals and man prompted studies to estimate the significance of multiresistant Salmonella enterica subspecies enterica serotype Typhimurium (S. Typhimurium) phage type DT104 of animal origin in Hungary. A collection of 231 strains (primarily of goose, turkey, poultry and porcine origin from the years 1997-1998) was tested for resistance against 7 selected antibiotics (ampicillin, chloramphenicol, enrofloxacin, nalidixic acid, streptomycin, tetracycline and sulphamethoxazole). Strains with resistance against 3 or more were defined as multiresistant. All strains were phage typed using Felix-Callow's S. Typhimurium phage typing system, and 91 of them (suspect DT104) were also typed according to Anderson's definitive typing (DT) system. In this study, 14% of animal strains from 1997-1998 was classified as DT104, for which turkey, pig and duck seemed to be the main carriers, and the multiresistant non-DT104 strains represented a further 6% of this collection. The prevalence of DT104 was highest among strains of turkey origin (50%), followed by strains of pig (29%), chicken (25%), duck (19%), and goose (3%) origin. The other DT104 related phage types (DT12 and U302) were only detected in the case of 4 strains (2 of porcine, and one each of turkey and of goose origin). The DT104 corresponded to the Felix-Callow types 2/3 or 2c/3 in each case, except in the case of 3 turkey strains where they corresponded to type 35/3. Nalidixic acid resistance was detected in all multiresistant turkey strains and in some of other animal origin but none of these strains were resistant to enrofloxacin. A retrospective analysis (based on the above relationship) indicated that S. Typhimurium strains corresponding to DT104 could be present and increase in the Hungarian farm animal population from about 2% to 20% between 1985 and 1990, in a manner similar to the emergence of human DT104, as reported elsewhere (Pászti et al., 2000). The 91 suspect DT104 strains were also tested for plasmid profile and for spvC gene indicating the presence of the large serotype specific plasmid (Ssp). No characteristic plasmid profile could be attributed to S. Typhimurium DT104. The serovar-specific large plasmid was detected by PCR for spvC in 100% of DT104 strains and in 77% of the non-DT104 strains. The virulence of two DT104 strains was tested in orally infected day-old chicks and compared with virulence of 4 non-DT104 strains. Higher colonizing virulence of DT104 strains could be established as compared to the other strains.     

Comparison of Salmonella enterica subspecies enterica serovar Typhimurium isolates from dairy cattle and humans using in vitro assays of virulence

Salmonella enterica subspecies enterica serovar Typhimurium (Salmonella typhimurium) can infect and cause disease in a wide range of host species however there have been suggestions that this serovar may have genes involved with host range or specificity [Tsolis, R.M., Townsend, S.M., Miao, E.A., Miller, S.I., Ficht, T.A., Adams, L.G., Baumler, A.J., 1999. Identification of a putative S. enterica serotype Typhimurium host range factor with homology to IpaH and YopM by signature-tagged mutagenesis. Infect. Immun. 67 (12), 6385-6393]. Our goal in this study was to determine if in vitro virulence assays would support this suggestion. Twelve human and 10 bovine isolates of S. typhimurium from a single county in California were evaluated using in vitro virulence assays of adhesion and invasion. The resulting data was combined with results from previously reported genotypic and phenotypic testing of the isolates and statistical analysis performed using multivariate general linear models. Human isolates had higher adhesion values in each of the statistical models tested (p<0.05) but no statistical differences were found in the invasion values of human and bovine source isolates. Both adhesion and invasion values differed between the two largest groups of isolates segregated on the basis of pulsed-field gel patterns. The findings suggest there may be genetically defined in vitro virulence attributes in S. typhimurium that are associated with host species.     

Effects of microcin 24-producing Escherichia coli on shedding and multiple-antimicrobial resistance of Salmonella enterica serotype typhimurium in pigs

OBJECTIVE: To investigate the effect of an Escherichia that produced microcin 24 (Mcc24) on shedding of of Salmonella enterica serotypeTyphimurium in swine and evaluate evidence of in vivo activation of the Mcc24-mediated, multiple-antibiotic resistance (mar) operon. ANIMALS: 36 crossbred weaned pigs. PROCEDURE: 24 pigs were allocated to 2 groups (12 pigs/group). Pigs in 1 group received daily oral administration of an Mcc24-producing E coli, whereas the other group received a non-Mcc24-producing E coli. All pigs were challenge exposed with Salmonella Typhimurium chi4232. A third group of 6 pigs received Mcc24-producing E coli and was challenge exposed with an Mcc24-sensitive, marA-deleted strain of Salmonella Typhimurium 4232. After challenge exposure, fecal samples from all pigs were cultured to detect shedding of Salmonella Typhimurium and Salmonella Typhimurium isolates were screened for resistance to ciprofloxacin. Fecal samples were collected throughout the study, and tissue samples were collected during necropsy. RESULTS: Differences in shedding of Salmonella Typhimurium were not detected between groups receiving Mcc24-producing or non-Mcc24-producing E coli. No significant differences were found in quantitative analysis between groups receiving Mcc24-producing and non-Mcc24-producing E coli. Evidence of mar activation was not detected. CONCLUSIONS AND CLINICAL RELEVANCE: Microcin-producing E coli did not exert an effect on shedding of SalmonellaTyphimurium or mar activation in pigs. It may be difficult or impractical to create the conditions required for Mcc24 to be an effective part of a food safety intervention to reduce shedding of Salmonella Typhimurium.     

Prevalence of Escherichia coli O157:H7 prophage-like sequences among German Salmonella enterica serotype Typhimurium phage types and their use in detection of phage type DT104 by the polymerase chain reaction

A 1.6kb DNA fragment identified by random amplifiable polymorphic DNA differentiation (RAPD) from a Salmonella enterica serotype Typhimurium phage type DT104 isolate was used to investigate the prevalence of the region in 160 DT104 isolates, 83 other epidemiological important S. Typhimurium phage types and 20 strains selected from 17 other Salmonella serotypes. PCR screening tests using two different primer-sets derived from the RAPD fragment's nucleotide sequence showed that 76% of the 160 DT104 isolates investigated, including subtypes DT104A, DT104B, DT104B low, DT104H and DT104L, reacted positively. High sensitivity was shown for DT104 strains expressing at least the penta-resistance pattern ACSSuT (97% of 104 strains tested). DT104 susceptible strains showed only a sensitivity of 35% (17 strains tested). In contrast, 83% of the 83 strains from the other S. Typhimurium phage types reacted negatively. Strains from five out of the 17 other serotypes showed a positive signal with one primer-set. The other primer-set exhibited only a positive reaction with one S. Dublin isolate. The analysis of a 2415bp extended sequence revealed homologies to genes encoded by Escherichia coli O157:H7 prophages, suggesting that the described region contains genes of a prophage specific for DT104 and related phage types.     

Epidemiology of Salmonella enterica serovars enteritidis and Typhimurium in animals and people in Scotland between 1990 and 2001

Two serovars of salmonella which are currently of particular importance in both human and animal infections are Salmonella enterica serovars Enteritidis phage type 4 (PT4) and Typhimurium definitive type 104 (DT104). This paper describes the trends in the relationships between the levels of infection of people and a range of farm animal species with these two serovars and explores some of the reasons behind them. In 1996, there was a peak of 520 reports of S Typhimurium DT104 infection in people in Scotland, but the number has decreased every year since, to 96 in 2001. In cattle the incidence of S Typhimurium DT104 also peaked in 1996, with 138 incidents, and it has similarly decreased every year to 2001 when there were 10 reported incidents. Similar declines have been observed in its incidence in sheep and pigs. In people the number of reports of S Enteritidis PT4 peaked in 1997 at 1684 and then declined to 457 in 2001. In chickens, the number of reports of S Enteritidis PT4 peaked in 1998 at 34 incidents, but no incidents were reported in the following three years.     

Integron content of Salmonella enterica serotype Typhimurium strains isolated in Hungary in the years 1997-1999

The integron content of 52 DT104/U302 phage type strains and 53 non-DT104/U302 strains of Salmonella enterica serotype Typhimurium (S. Typhimurium) was studied in PCR experiments using a 5'-CS/3'-CS primer pair (Lévesque et al., 1995). Forty-three out of 44 streptomycin- and/or ampicillin-resistant DT104 and related phage type strains were found to carry a 1 kb and/or 1.2 kb long integron. The other resistance markers did not affect the number and size of integrons; no integron-free multidrug-resistant (MDR) DT104 strains were found. The two large groups of DT104 strains (Felix-Callow's phage types 2 and 2c) proved to be identical in respect of integron patterns (IPs), supporting the views of those authors who consider DT104 a single clone. Strains of human and animal origin did not differ from each other in their IPs. Within the non-DT104 phage types, ampicillin- and/or streptomycin-resistant, integron-free MDR strains were also found. Based on amplicons varying between 290 and 3500 bp an IP system was suggested. The commonest amplicon sizes in non-DT104 strains were 1450 and 2050 bp. The IPs of DT104 strains and of non-DT104 strains containing an integron of 1 and 1.2 kb size were stable. In contrast, the IPs of other non-DT104 strains showed a varying degree of instability. Integron loss was frequently associated with spontaneous plasmid elimination and changes of R-type among the descendants of a given strain.     

Antimicrobial susceptibility and multi-drug resistance of Salmonella enterica subspecies enterica serovars in Sudan

This study was undertaken to determine the antimicrobial resistance patterns of Salmonella enterica subspecies enterica recovered from human, food, water, and animal samples collected in Khartoum State, Sudan. A total of 64 Salmonella isolates belonging to 28 different serovars were tested for their susceptibility to 13 antimicrobial agents. The majority of isolates (98.4 %) were resistant to at least one antimicrobial agent. Isolates were frequently resistant to ampicillin (90.6 %), cephalexin (50.0 %), nalidixic acid (25.0 %), streptomycin (21.9 %), kanamycin (18.8 %), gentamicin (17.2 %), and co-trimoxazole and trimethoprim (12.5 %). The most common pattern of multiple drug resistance included resistance to ampicillin and cephalexin. Most isolates were sensitive to chloramphenicol (98.4 %), ciprofloxacin (93.8 %), and norfloxacin (90.6 %). Two chicken- and the two human-origin S. Kentucky isolates were resistant to both ciprofloxacin and norfloxacin. All S. Kentucky isolates and the one S. Rissen isolate demonstrated multi-drug resistance. The results indicate the significance of multi-drug-resistant Salmonella serovars isolated from chickens and other animals and foods as sources for multi-drug-resistant Salmonella in humans in Sudan.     

Molecular basis of quinolone resistance in Escherichia coli from wild birds

Nine quinolone resistant (minimal inhibitory concentration [MIC] was > 32 microg/mL for nalidixic acid, > 1 microg/mL for ciprofloxacin) isolates of Escherichia coli have been found in wild birds with septicemia. All of the isolates were aerobactin positive. The mechanisms of resistance were characterised by sequencing the quinolone resistance-determining region (QRDR) of the gyrA, gyrB, parC, and parE genes. Sequence analysis of the gyrA gene in all isolates identified only 1 nucleotide substitution at codon Serine-83 for Leucine-83. Sequence analysis of the gyrB, parC, and parE QRDR genes revealed no mutations in any of the isolates. This study was conducted to determine the importance of these genes in the susceptibility of E. coli strains isolated from wild birds to quinolones.     

Concentration and prevalence of Escherichia coli O157 and Salmonella serotypes in sheep during slaughter at two Australian abattoirs

Objective This study aimed to determine the presence and concentration of Escherichia coli O157 and Salmonella spp. on fleece, faeces and carcases of sheep during slaughter. Procedure Faeces, fleece and pre-chill carcase samples were collected from 164 sheep slaughtered at two Australian abattoirs. The presence of E. coli O157 and Salmonella spp. were determined by use of automated immunomagnetic separation (AIMS) with enumeration by use of the ‘most probable number’ (MPN) method. Results Escherichia coli O157 was isolated from 5% of faeces, 3% of fleeces and 0.6% of pre-chill carcases. The mean log₁₀ count of E. coli O157 positive faecal samples was 2.32 MPN/g, but counts on fleeces and carcases were below the countable limit (−1 log₁₀ MPN/cm²). Salmonella spp. were isolated from 20% of faeces, 13% of fleeces and 1.3% of pre-chill carcases. The mean log₁₀ count of Salmonella spp. in faeces was 1.43 MPN/g and on fleece was −0.24 MPN/cm², but counts on carcases were below the countable limit (−1 log₁₀ MPN/cm²). Conclusion The prevalence and concentration of pathogens were low in the sheep tested in this study, indicating a low risk of human infection from products derived from these animals.     

Characterization of Escherichia coli O86 O-antigen gene cluster and identification of O86-specific genes

Escherichia coli O86 belongs to the enteropathogenic E. coli (EPEC) group, some strains of which are pathogens of humans, wild birds and farm animals. The O-antigen gene cluster of E. coli O86 was amplified by long-range PCR using primers based on the housekeeping genes galF and gnd, and then sequenced. Genes involved in GDP-Fuc and N-acetyl-galactosamine (GalNAc) synthesis and genes encoding glycosyltransferases, O-unit flippase and O-antigen polymerase were identified on the basis of homology. By screening against 186 E. coli and Shigella-type strains, two genes specific to E. coli O86 were identified. A polymerase chain reaction (PCR) assay, based on the specific O-antigen genes identified here, could be used for the rapid detection of E. coli O86 in environmental and clinical samples. The relationship between E. coli O86 and O127 was also determined by comparing the two O-antigen gene clusters.     

An epidemic of salmonellosis caused by Salmonella Typhimurium DT160 in wild birds and humans in New Zealand

AIM: This study reports an outbreak of salmonellosis due to S. Typhimurium DT160 which caused extensive mortality in wild birds and enteric disease in humans in New Zealand during the winter and spring months of the year 2000. METHODS: Necropsies were performed and microbiological examinations undertaken on wild birds from populations in which mass mortality was reported, and on captive indigenous birds which died suddenly during the winter and spring of 2000. Affected tissues were examined histologically and isolates of S. Typhimurium were phage typed and examined using pulsedfield gel electrophoresis (PFGE). Isolates of S. Typhimurium obtained from cases of human enteric disease which occurred during these months were phage typed, examined using PFGE and compared with the bird isolates. RESULTS: Central and northern areas of the South Island and the southern North Island were worst affected with die-offs of several hundreds of sparrows and other birds reported in rural areas. Mortalities reached a peak in winter (July-August) 2000 and decreased to small numbers during the spring and early summer. The birds usually died of an acute septicaemia with multifocal necrotising lesions in the liver and spleen. Human cases throughout the country increased gradually over the same period. Isolates from birds, livestock and humans examined using PFGE were indistinguishable from one another. CONCLUSION: This strain of Salmonellahas emerged as a major cause of septicaemia in wild birds in New Zealand. Because of the close association between house sparrows (Passer domesticus) and humans, the organism also poses a serious zoonotic risk. The possibility that the infection may spread to involve indigenous species needs investigation.     

Isolation of different serovars of Salmonella enterica from wild birds in Great Britain between 1995 and 2003

Postmortem examinations were carried out on the carcases of 779 wild birds. Salmonellosis was a common cause of death in greenfinches (Carduelis chloris), house sparrows (Passer domesticus) and chaffinches (Fringilla coelebs), and was also responsible for the deaths of other birds such as goldfinches (Carduelis carduelis), feral pigeons and different species of gulls. Most cases of salmonellosis in finches occurred between January and March, whereas salmonellosis in house sparrows tended to occur between October and March. Salmonella Typhimurium DT40 and DT56 (variant) predominated in finches and sparrows, DT41 and DT195 were the most common strains isolated from gulls, and DT2 and DT99 were recovered from feral pigeons. These "wild bird" strains of Salmonella made up less than 0.5 per cent of the isolates of Salmonella recovered from cattle, sheep, pigs, chickens or turkeys in Great Britain over the same period, but they made up nearly 3 per cent of the isolates from more extensively reared avian livestock such as gamebirds, ducks and geese.     

Feedstuffs as a vehicle of cattle exposure to Escherichia coli O157:H7 and Salmonella enterica

Feed has been reported as a vehicle for transmission of Salmonella enterica in cattle and several lines of evidence suggest that feed can be a vehicle for transmitting Escherichia coli O157:H7 as well. To show whether microbial contamination of feeds could contribute to the populations of S. enterica and E. coli O157:H7 on a farm, we compared isolates from feed samples to bovine fecal isolates from the same farm using pulsed-field gel electrophoresis (PFGE). Four of 2365 component feed samples (0.2%) and 1 of 226 feed mill samples (0.4%) were positive for E. coli O157:H7. Twenty of 2405 (0.8%) component feed samples and none of 226 feed mill samples were positive for Salmonella. PFGE profiles from E. coli O157:H7 isolated from a component feed sample closely resembled that from a fecal isolate collected later from the same farm, and a similar observation was made of a Salmonella Tyhpimurium isolate from component feed on another farm. There were indistinguishable PFGE profiles from component feed Salmonella Tyhpimurium DT104 isolates and fecal isolates from the same farm. These results provide evidence for a role of cattle feed in transmission of E. coli O157:H7; S. enterica; cattle-bacteria.     

Salmonella in poultry flocks and humans--S. enterica subspecies enterica serovar Enteritidis in the past

After importing of breeder lines for laying flocks from Canada into the former GDR in 1966 the egg industry in this country was completely isolated from that in Western Germany or other Western European countries until opening the border in Germany in 1989. Because of this isolation from other countries, an analysis of the clonal diversity of Salmonella (S.) Enteritidis isolates originated from humans, chickens and food in the former GDR during the 1980s would provide a unique opportunity to obtain new insight into factors that may have triggered the S. Enteritidis epidemic. While isolates had previously been typed by the phage typing scheme of Lalko and Laszlo we applied for the first time the extended phage typing scheme by Ward for the retrospective analysis of the S. Enteritidis strains. Furthermore, isolates of phage type (PT) 4/6 (Ward / Lalko and Laszlo) from different livestocks were investigated by ribotyping. Although in total the PT4/6 dominated between 1986 and 1989 in poultry, other phage types have prevailed in the early 1980s and represented a considerable fraction of isolates until 1989. For instance, PT8/7 was isolated from one large layer farm (district Halle) from 1988 until 1989. During that time in another farm (district Cottbus) only PT8/7 was detected too. PT4/6 was isolated from neither of these two laying hen farms. The strains of PT4/6 could be distinguished by ribotyping in 19 different subtypes. The strains from the northern farms were distinct from those isolated in the southern regions. As farms which were harbouring either PT4/6 or PT8/7 had obtained laying hens from the same sources (breeder lines in Deersheim and Spreenhagen) it is highly probable that S. Enteritidis infection was acquired from the environment at each individual farm. This conclusion is also supported by the presence of different PT4/6 ribotypes in different farms. The presence of different phage types or PT4/6 ribotypes at different farms of laying hens suggests that in each case the S. Enteritidis strains present in the environment were able to enter chicken flocks.     

Spatial distribution of Escherichia coli O157-positive farms in Scotland

Using a sample of 949 Scottish farms with finishing cattle, the spatial distribution of Escherichia coli O157-positive farms was investigated using disease mapping models. The overall prevalence of E. coli O157-positive farms was estimated as 22%. The regions used in this study were the 16 postcode areas of Scotland. For each region, the posterior relative risk (RR) was estimated as a model-based alternative to the saturated standardized morbidity ratio (SMR), i.e., the ratio between observed and expected cases in a region. Three Bayesian hierarchical models with generalized linear modeling of the area-specific risks were used to estimate the posterior relative risk of E. coli O157-positive farms in the postcode areas: a random-effects model incorporating only spatially uncorrelated heterogeneity; a model incorporating both spatially correlated and uncorrelated heterogeneity; and a pseudo-mixture model with unstructured correlation and a weighted mix of two variance components representing the spatial correlation and a jump structure. None of the models identified any areas with a significant increase or decrease in risk. The deviance information criteria slightly favored the simplest model (RR range: 0.92--1.09). However, this model appeared to smooth out more of the variation in the RR compared to the pseudo-mixture model, which gave a more informative pattern of the posterior relative risks (range: 0.81--1.22).     

Transduction of bla(CMY-2), tet(A), and tet(B) from Salmonella enterica subspecies enterica serovar Heidelberg to S. Typhimurium

Antimicrobial resistance of Salmonella spp., especially resistance mediated by extended spectrum β-lactamases (ESBL), is a growing public health concern. Understanding the mechanisms through which Salmomella spp. acquire the resistance genes can lead to the development of intervention and mitigation strategies. Thirty-one Salmonella isolates of bovine origin were analyzed by serotyping, antimicrobial susceptibility testing, phage induction and bacterial host range determination, and phage transduction of antimicrobial resistance. The Salmonella isolates consisted of 12 serovars. Resistance to 1 or more antibiotics was detected in 12 isolates. Inducible phages were recovered from 19 Salmonella (61%) by spot lysis assay. Of the 19 phage samples, 13 were able to multiply in 2 or more Salmonella of various serovars. A cross-serovar transduction of antimicrobial resistance was observed from S. Heidelberg to S. Typhimurium. Transfection of an antimicrobial-susceptible strain of S. Typhimurium with a phage propagated in a S. Heidelberg resistant to multiple β-lactam antibiotics and tetracycline resulted in independent acquisition of bla_CMY-2, tet(A), and tet(B). These data indicate that inducible phages are common in Salmonella of bovine origin, many of them demonstrate a broad host range, and wild-type phage mediated transduction may contribute to the dissemination of antimicrobial resistance, including the resistance mediated by ESBL.