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R Helmuth

Publications and source records attributed to R Helmuth.

At least 19 recordsLinked to original sources
PubMed2005

Pulsed-field gel electrophoresis, plasmid profiles and phage types for the human isolates of Salmonella enterica serovar Enteritidis obtained over 13 years in Taiwan.

AIMS: Plasmid profile, phage typing, and pulsed-field gel electrophoresis (PFGE) patterns of 124 Salmonella Enteritidis strains isolated in 1998-2002 in Taiwan were analysed and the results were compared with those of the 63 strains obtained in 1991-1997, so that molecular subtypes and epidemic strains for Salmonella Enteritidis over a 13-year period (1991-2002) could be elucidated. METHODS AND RESULTS: A total of 124 strains of Salmonella Enteritidis isolated from human in Taiwan between 1998 and 2002 were analysed by PFGE, plasmid analysis and phage typing. The results obtained were compared with those of the 63 strains obtained in 1991-1997, so that the clonal relationships for a total of 187 strains obtained over 13 years could be elucidated. For PFGE, restriction enzymes XbaI, SpeI and NotI were used for chromosomal DNA digestion. Results showed 28 PFGE pattern combinations for the 187 Salmonella strains. Of them, pattern X3S3N3 was the major subtype as 130 strains isolated from different locations during 1991-2002 showed this PFGE pattern. For all these 187 strains, the genetic similarity was higher than 80%. Plasmid analysis showed 17 distinct types, which consist of one to four plasmids and the predominant phage type of those strains was PT4 (71.6%) and PT6a (13.4%). The three methods identified different degrees of polymorphism in the following order: plasmid profile (18 types, D = 0.659) > PFGE (28 types, D = 0.512) > phage typing (13 types, D = 0.438). As PFGE patterns, phage type and plasmid profile were combined for subtyping, the 187 strains could be grouped into 46 subtypes and the discriminatory index was raised to 0.795. For these 46 subtypes, the predominant one was X3S3N3/P1/PT4, which contained 77 (41%) isolates. CONCLUSIONS: Most of the Salmonella Enteritidis strains from sporadic cases were with pattern X3S3N3. They were the prevalent and may be the epidemic strains found in Taiwan during 1991-2002. The present study suggested that the several variants were derived from a single clonal line and the genome for strains of Salmonella Enteritidis are highly conserved over a 13-year period (1991-2002). SIGNIFICANCE AND IMPACT OF THE STUDY: The results obtained here are useful for epidemiolgical study of salmonellosis caused by Salmonella Enteritidis in Taiwan. Comparing the data of the present study with those obtained for strains from other countries, the major subtypes for Salmonella Enteritidis infection in the world can be elucidated.

Bacteriophage Typing↗
PubMed2004

Characterization and localization of drug resistance determinants in multidrug-resistant, integron-carrying Salmonella enterica serotype Typhimurium strains.

The genetic background of the antimicrobial resistance of 10 selected multiresistant Salmonella serotype Typhimurium (S. Typhimurium) strains (including the emerging monophasic variant [4,5,12:i:- ]) was investigated. All strains shared class 1 integrons (with seven types of variable regions) and belonged to different lineages (L1-L6) according to their phage types, DNA polymorphisms by XbaI-pulsed-field gel electrophoresis (PFGE), integrons, and/or resistance patterns. The strains were screened for the presence and localization (chromosomal or plasmid) of 32 DNA sequences representing integron-, Tn21-like transposon-, resistance-, and virulence-plasmid genes. Strains belonging to lineage L1 (definitive phage type DT104) carried the 90-kb Salmonella virulence plasmid together with the complete or partial chromosomally located Salmonella Genomic Island 1 (SGI1). All strains belonging to the other five lineages carried their resistance determinants on various resistance plasmids. Two of these strains showed complex plasmid profiles, which included a 95 kb virulence plasmid together with two or four resistance plasmids. Two strains carried a resistance plasmid that lacked the virulence-plasmid-encoding sequences. The remaining two strains carried two different hybrid virulence-resistance plasmids. Twenty-three of the DNA sequences could be assigned to distinct XbaI genomic restriction patterns (PFGE profiles). In this way, the influence of the resistance and virulence plasmids on the PFGE profiles was determined, and several groups of resistance genes could be identified. The data obtained represent a useful epidemiological tool for tracing the emergence and distribution of multiresistant S. Typhimurium worldwide.

Base Sequence↗
PubMed2003

Incidence of quinolone resistance in strains of Salmonella isolated from poultry, cattle and pigs in Germany between 1998 and 2001.

This paper reports the susceptibility to the quinolone nalidixic acid and the fluoroquinolone ciprofloxacin of 14,514 strains of Salmonella isolated in Germany from poultry, cattle and pigs between 1998 and 2001. Quinolone-resistant salmonellae were most frequently isolated from poultry, with a prevalence of 10.2 to 16.8 per cent. Poultry-associated serotypes, such as Salmonella Paratyphi B (d-tartrate positive), Salmonella Hadar and Salmonella Virchow, had the highest prevalence of quinolone resistance, ranging between 35 and 74 per cent. All the nalidixic acid-resistant strains also had a reduced susceptibility to ciprofloxacin, with minimum inhibitory concentrations (MICS) of 0.125 to 2 microg/ml. A comparison of the MICS for ciprofloxacin of the strains of these poultry-associated serotypes and Salmonella Enteritidis phage type 4 isolated in 1998/99 and 2000/01 indicated that there had been a shift towards higher MIC values of up to 2 microg/ml. The quinolone resistance-determining region (QRDR) of the gyrA gene and the homologue region of the parC gene of 31 selected strains were sequenced. Several different amino acid changes were observed in gyrA of the quinolone-resistant isolates at positions 83 and 87, but no substitutions were observed in parC.

Amino Acid Sequence↗
PubMed2000

Monitoring of antibiotic resistance in bacteria of animal origin: epidemiological and microbiological methodologies.

The occurrence of antibiotic-resistant bacteria in food animals is a major public health threat. Information on the prevalence of resistance to specific drugs in both bacterial and animal species together with changes occurring over time, are necessary to understand the magnitude of the problem and to establish baselines for taking action. The aim of this paper is to define the minimum epidemiological and microbiological requirements for establishing a surveillance of antimicrobial resistance in bacteria of animal origin. Surveillance should involve different bacterial species, veterinary pathogens, zoonotic bacteria and commensal bacteria used as indicators. The collected data should be periodically updated and the reports distributed among practising veterinarians and regulatory authorities. These reports would be a useful tool for developing guidelines for the prudent use of antimicrobial agents in veterinary medicine and for action strategies.

Animal Diseases↗
PubMed2000

Salmonella in slaughter pigs of German origin: an epidemiological study.

The Salmonella prevalence in slaughter pigs of German origin was determined in seven abattoirs located in different regions of the country between February and June 1996. A total of 11,942 pigs delivered to the abattoirs in 752 batches, most of them comprised of pigs from individual finishing farms, was investigated by the bacteriological examination of faecal and gut lymph node samples, as well as of surface swabs taken from the carcasses. Salmonellae were isolated from 3.7% of the faecal samples, 3.3% of the lymph nodes and 4.7% of the surface swabs. The estimated overall prevalence of Salmonellae was 6.2% in the slaughter pigs, ranging between 1.9% and 12% in individual abattoirs. In the samples taken from carcasses, the estimated prevalence of Salmonellae reached 10.3%. 648 out of 752 batches could be included in a statistical analysis. No Salmonellae were detected in nearly 70 percent of the batches included in this analysis (n = 648). High Salmonella prevalences of more than 50 percent positive animals were detected only in 13 batches (2.0%). A statistically significant influence of the duration of the transport of slaughter pigs to the abattoirs or the waiting period in the abattoirs prior to slaughter could not be detected.

Abattoirs↗
PubMed2000

Phage types, antibiotic susceptibilities and plasmid profiles of Salmonella typhimurium and Salmonella enteritidis strains isolated in Istanbul, Turkey.

OBJECTIVE: To examine 13 Salmonella typhimurium and 22 S. enteritidis strains isolated from individual cases of gastroenteritis for their phage types, antibiotic susceptibilities and plasmid profiles. METHODS: The phage typing of S. typhimurium strains was done according to the method of Anderson et al, and the phage typing scheme of Ward et al was used for phage typing of S. enteritidis strains. Antibiotic susceptibility testing was performed by the Kirby-Bauer disk diffusion method. Extended-spectrum beta-lactamase production of the strains was determined by the three-dimensional method. Plasmid profiles of the strains were examined using the method described by Kado and Liu with some modification by Graeber et al. RESULTS: Two S. typhimurium strains were DT 193 and one was DT 22, whereas 10 strains were untypable. PT 4 was the predominant phage type among S. enteritidis strains. Four S. enteritidis strains were DT 6a, three strains were PT 1 and one strain was PT 8, whereas only one strain was untypable. Eleven of 13 S. typhimurium and three of 22 S. enteritidis strains were found to be multiresistant. Ten different resistance patterns among S. typhimurium and four different resistance patterns among S. enteritidis strains were detected. Extended-spectrum beta-lactamase production was detected in 10 of 13 S. typhimurium and in three of 22 S. enteritidis strains. All S. typhimurium strains but one were found to contain at least one plasmid, with molecular masses varying between 4 and 107 MDa, and 11 different plasmid patterns were determined. Plasmid pattern analysis permitted further differentiation of the S. enteritidis strains into nine groups. A serovar-specific virulence plasmid of 36 MDa was detected in 13 of 22 S. enteritidis strains. CONCLUSIONS: The results suggest that the majority of S. typhimurium strains were closely related.

Anti-Bacterial Agents↗
PubMed1999

Clonal relationship of Salmonella enterica serovar typhimurium phage type DT104 in Germany and Austria.

A new epidemic clone of Salmonella enterica serovar Typhimurium designated definitive phage type (DT) 104 has been emerging since 1990 to become most common type among S. Typhimurium isolates in Germany and Austria. Molecular fingerprinting (PFGE-pattern, plasmid profiles, IS200 pattern, ribotype, ERIC-type, OMP and MLE patterns) revealed the majority of the DT104 isolates to have clonal identity; they were designated as type 1 (about 95%). Moreover, clonal type 1 of DT104 was found to occur in sensitive as well as in a range of multiply drug-resistant variants and in a variety of plasmid profile types (in particular with small cryptic plasmids in the range of 1.0 to 5.0 Md). Since the clonal type 1 of DT104 has been identified among isolates from other countries, too, including such from the United Kingdom, the United Arab Emirates, the Philippines, and the Netherlands, its pandemic spread in man indicates that the import/export of this pathogen continues. About 5% of the DT104 isolates have been identified as genetically diverse indicating the independent appearance of the same multiple drug resistance and phage pattern phenotype among different Salmonella ancestor strains.

Animals↗
PubMed1999

Incidence of quinolone resistance over the period 1986 to 1998 in veterinary Salmonella isolates from Germany.

A total of 24,591 nonhuman salmonella strains isolated in Germany between 1986 and 1998 were examined for their resistance to nalidixic acid by an agar diffusion method. The rate of resistance (inhibition zone, </=13 mm) ranged from 0.2% in 1986 to a peak of 14. 8% in 1990. Between 1991 and 1998 the MICs for nalidixic acid-resistant strains ranged from more than 256 microg/ml for nalidixic acid to between 0.25 and 128 microg/ml for enrofloxacin. In the early 1990s a particularly high incidence of fluoroquinolone resistance (49.5%) was seen among isolates of Salmonella enterica serotype Typhimurium (Salmonella Typhimurium) definitive phage type 204c that mainly originated from cattle. Among isolates from poultry an increase in the incidence of nalidixic acid resistance to a peak of 14.4% was observed in 1994. This peak was due to the presence of specific resistant serotypes, mainly serotypes Hadar, Saintpaul, Paratyphi B (D-tartrate positive; formerly serotype Java) and Newport. Such strains exhibited a decreased susceptibility to enrofloxacin (MIC, 1 microg/ml). Among isolates from pigs the peak incidence of resistance was reached in 1993, with 7.5% of isolates resistant to nalidixic acid and enrofloxacin. The study demonstrates an increase in the incidence of strains that are resistant to nalidixic acid and that have decreased susceptibility to enrofloxacin after the licensing of enrofloxacin. In addition, the number of other serotypes that exhibited nalidixic acid resistance or reduced enrofloxacin susceptibility increased among the total number of isolates investigated between 1992 and 1998.

Animals↗
PubMed1998

Comparative studies of diagnostic bacteriological methods for the recovery of Salmonella from faecal samples from flocks of layers.

Several conventional culture procedures were compared for the recovery of Salmonella from faecal samples of layer flocks. The cultural media employed consisted of Rappaport-Vassiliadis and tetrathionate broth for selective enrichment and three different plating media (Brilliant Green, Xylose-Lysine-Desoxycholate and modified semisolid Rappaport-Vassiliadis medium). The initial enrichment and plating procedures were repeated in a four-stage secondary enrichment. Selective enrichment in Rappaport-Vassiliadis or tetrathionate broth followed by plating on modified semisolid Rappaport-Vassiliadis medium (selective motility enrichment) resulted in a significantly higher isolation rate for Salmonella from faecal samples of layers compared to Brilliant Green and Xylose-Lysine-Desoxycholate medium. The selective enrichment yielded additional Salmonella isolates up to the second stage. Further enrichment did not increase the isolation rate.

Animals↗
PubMed1998

Studies of the presence of the virulence factors, adhesion, invasion, intracellular multiplication and toxin formation in salmonellas of different origin.

Salmonellas of different origin were classified into two groups (11 strains of common serovars which had been isolated from organs of calves having died from salmonellosis and 18 strains belonging to rare serovars which showed uncommon metabolic characteristics and had been isolated from spices and spiced foods). The strains were examined with regard to different virulence parameters. All salmonellas investigated possessed the genetic information on invasion (invA) and toxin formation (stn). They adhered equally well to epithelial cells, could penetrate into these and survive and multiply inside the cells. The formation of toxic substances could be detected in all strains after co-cultivation with epithelial cells in the CHO-K1 test. Significant differences between the groups of strains could be demonstrated only for the invasion of epithelial cell monolayers. Since adhesion, invasion and the ability of intracellular survival and multiplication as well as toxin formation constitute virulence parameters of salmonellas, it must be assumed that also the Salmonella serotypes studied which have been rarely observed epidemiologically constitute a risk for humans.

Animals↗
PubMed1997

Detection of the induction of Salmonella enterotoxin gene expression by contact with epithelial cells with RT-PCR.

All strains of Salmonella enterica investigated were found to carry the Salmonella enterotoxin gene (stn) as determined by PCR and hybridization studies. However, when using CHO-K1 cells for testing the toxicity of the strains, not all strains showed a toxic effect (cell elongation) on the cells or did so only at a low level. The cultivation of Salmonella in contact with epithelial cells (IEC-6) led to an increase in the production of toxin. The stn gene expression was detectable with the help of the RT-PCR after 3 h of incubation. The RNA of the strains was isolated, transcribed into cDNA (with MMLV-reverse transcriptase) and amplified using PCR. The PCR products were separated electrophoretically using a polyacrylamide gel and detected by silver staining.

Animals↗
PubMed1997

Importance of the serovar-specific plasmid for virulence of salmonella strains in calves.

To evaluate the influence of serovar-specific plasmids on salmonella virulence in calves, experiments were performed involving infection, by the oral route, with mixtures of strains containing equal counts of a plasmid-carrying and a plasmid-free strain of the same serovar. The concentration ratio between the plasmid-carrying and the plasmid-free strain which had developed in the organs of the infected animals was used for a comparative evaluation of virulence and pathogenetic behaviour of the strains. While in the S. typhimurium strains studied, the presence of the plasmid was accompanied by a significantly increased colonization and multiplication of the agent in the host's body, examination of S. enteritidis and S. dublin revealed that the plasmid-free strains exhibited identical or even significantly higher bacterial counts than the plasmid-carrying strains in organs. The fact that plasmid-free salmonella strains with a high virulence for calves have been found demonstrates that the presence of a serovar-specific plasmid is not an indispensable requirement for the development of salmonellosis in calves.

Animals↗
PubMed1997

How to modify conditions limiting resistance in bacteria in animals and other reservoirs.

Antimicrobial agents in veterinary medicine are used for three purposes: therapy, prophylaxis, and nutrition. The major public health risk is that selection pressure leads to an increase in the pool of resistance genes. Since 1987, the nutritional use of antimicrobials in Europe has been regulated by a council directive, which demands special investigations into the potential of antimicrobials to increase rates of drug resistance. However, the prophylactic and therapeutic use of antimicrobials has sometimes led to the emergence of resistant bacteria. For example, the selective effect of the prophylactic use of gentamicin and the therapeutic use of quinolones led to the emergence of resistant salmonellae. To prevent the spread of resistant microorganisms from animals to humans, it should be recognized that antibiotics are not suitable as a compensation for poor hygiene standards or for the eradication of a pathogen from a certain environment. They should be used only by doctors or veterinarians.

Animals↗
PubMed1996

[Immunization with potential Salmonella enteritidis mutants--1. Production and in vitro characterization].

Production and in vitro characterization of potential vaccine strains are the first steps leading to an efficient Salmonella Enteritidis oral live vaccine for homologous immunization of poultry. The paper presents the results of the production of adenine-amino acid auxotrophic mutants using N-methyl-N'-nitro-N-nitrosoguanidine mutagenesis. The mutant strains were characterized using the following properties: auxotrophy, stability of mutation, reversion rate, generation time, metabolic properties, serotype, motility, plasmid content, phage type, SDS-PAGE patterns, as well as cell culture adhesion and invasion. Ten S. Enteritidis double auxotrophic mutants were obtained which are stable auxotrophically and where the risk of reversion was minimal. All strains were found to be plasmid-free. 5 mutants were selected for further investigations concerning their attenuation and immunological value.

Animals↗
PubMed1995

Molecular marker analysis of Salmonella typhimurium from surface waters, humans, and animals.

Salmonella contamination of North Sea water was detected for the first time in 1988 in Germany during routine examinations of bathing areas. Since then, subsequent isolations along the coast have been reported regularly. To define the source of contamination, strains isolated from seawater and rivers were studied by molecular marker methods. Their properties were compared with those of strains originating from possible sources of contamination such as humans, cattle, and sewage treatment plant water. Plasmid profile analysis of whole bacterial populations and the determination of antibiotic resistance patterns demonstrated, that contamination through the surrounding cattle industry could be excluded. Cattle isolates belonged to a widespread clone of phage type 204c which was multiresistant and exhibited an unique plasmid pattern which was never found in sea water isolates. Outer membrane protein and lipopolysaccharide analysis failed to demonstrate differences among the Salmonella populations and proved in this case insufficient for molecular marker discrimination.

Animals↗
PubMed1994

Salmonella enteritidis phage types in Germany.

In order to monitor the epidemiological situation of S. enteritidis in Germany, in 1990-91 1138 isolates from more than 180 locations in West Germany were phage typed. 1124 strains (98.8%) from all sources were typeable, belonging to 21 different phage types (PT). PT4 strains were isolated most frequently (70.8%). In addition, PT7, 25, 34 and 8 were of epidemiological relevance with incidences of 7.2 to 4.5%. The comparison of data shows that in Germany, like in other parts of Europe, PT4 predominates. This phage type is, however, infrequent in North America, where PT8 has the highest incidence.

Animals↗
PubMed1994

Molecular typing methods for S. enteritidis.

The predominance of certain phage types of Salmonella enteritidis in various countries makes further epidemiological subgrouping necessary. Today this is achieved by using molecular typing methods. For various bacterial species, plasmid profiling, the pattern of outer membrane proteins and lipopolysaccharides, the fingerprinting of total genomic DNA including ribotyping, and multilocus enzyme electrophoretic typing, have proven very useful. When such methods have been applied to S. enteritidis, they revealed a homogeneous, clonal structure in contemporary PT4 isolates. Furthermore they indicate that the clone observed today emerged from a heterogeneous population before the onset of the epidemic.

Animals↗