Rene S. Hendriksen

Effect of Abolishment of the Use of Antimicrobial Agents for Growth Promotion on Occurrence of Antimicrobial Resistance in Fecal Enterococci from Food Animals in Denmark

ABSTRACT From 1995 to 2000, a total of 673 Enterococcus faeciumand 1,088 Enterococcus faecalis isolates from pigs together with 856 E. faecium isolates from broilers were isolated and tested for susceptibility to four classes of antimicrobial agents used for growth promotion as part of the Danish program of monitoring for antimicrobial resistance. The four antimicrobials were avilamycin, erythromycin, vancomycin, and virginiamycin. Major changes in the use of antimicrobial agents for growth promotion have occurred during the last 6 years in Denmark. The government banned the use of avoparcin in 1995 and of virginiamycin in 1998. Furthermore, the producers have voluntarily stopped all use beginning in 1999. The avoparcin ban in 1995 was followed by a decrease in the occurrence of glycopeptide-resistant E. faecium (GRE) in broilers, from 72.7% in 1995 to 5.8% in 2000. The occurrence of glycopeptide resistance among isolates from pigs remained constant at around 20% from 1995 to 1997. It was shown that, in GRE from pigs, the genes encoding macrolide and glycopeptide resistance were genetically linked and that, following the decrease in the use of tylosin during 1998 and 1999, the occurrence of GRE in pigs decreased to 6.0% in 2000. From 1995 to 1997 the occurrence of erythromycin resistance among E. faecium and E. faecalis isolates from pigs was almost 90%. Use of tylosin decreased considerably during 1998 and 1999, and this decrease was followed by decreases in the occurrence of resistance to 46.7 and 28.1% among E. faecium and E. faecalis isolates from pigs, respectively. Erythromycin resistance among E. faecium isolates from broilers reached a maximum of 76.3% in 1997 but decreased to 12.7% in 2000 concomitantly with more limited use of virginiamycin. Use of virginiamycin increased from 1995 to 1997 and was followed by an increased occurrence of virginiamycin resistance among E. faecium isolates in broilers, from 27.3% in 1995 to 66.2% in 1997. In January 1998 the use of virginiamycin was banned in Denmark, and the occurrence of virginiamycin resistance decreased to 33.9% in 2000. Use of avilamycin increased from 1995 to 1996 and was followed by an increase in avilamycin resistance among E. faeciumisolates from broilers, from 63.6% in 1995 to 77.4% in 1996. Since 1996 avilamycin usage has decreased, followed by a decrease in resistance to 4.8% in 2000. Our observations show that it is possible to reduce the occurrence of antimicrobial resistance in a national population of food animals when the selective pressure is removed. Cases in which resistance to vancomycin was linked to resistance to erythromycin were exceptions. In such cases resistance did not decrease until the use of both avoparcin and tylosin was limited.

Global Monitoring of Salmonella Serovar Distribution from the World Health Organization Global Foodborne Infections Network Country Data Bank: Results of Quality Assured Laboratories from 2001 to 2007

Salmonella enterica is commonly acquired from contaminated food and is an important cause of illness worldwide. Interventions are needed to control Salmonella; subtyping Salmonella by serotyping is useful for targeting such interventions. We, therefore, analyzed the global distribution of the 15 most frequently identified serovars of Salmonella isolated from humans from 2001 to 2007 in laboratories from 37 countries that participated in World Health Organization Global Foodborne Infections Network and demonstrated serotyping proficiency in the Global Foodborne Infections Network External Quality Assurance System. In all regions throughout the study period, with the exception of the Oceania and North American regions, Salmonella serovars Enteritidis and Typhimurium ranked as the most common and second most common serovar, respectively. In the North American and Oceania (Australia and New Zealand) regions, Salmonella serovar Typhimurium was the most common serovar reported, and Salmonella serovar Enteritidis was the second most common serovar. During the study period, the proportion of Salmonella isolates reported from humans that were Salmonella serovar Enteritidis was 43.5% (range: 40.6% [2007] to 44.9% [2003]), and Salmonella serovar Typhimurium was 17.1% (range: 15% [2007] to 18.9% [2001]). Salmonella serovars Newport (mainly observed in Latin and North American and European countries), Infantis (dominating in all regions), Virchow (mainly observed in Asian, European, and Oceanic countries), Hadar (profound in European countries), and Agona (intense in Latin and North American and European countries) were also frequently isolated with an overall proportion of 3.5%, 1.8%, 1.5%, 1.5%, and 0.8%, respectively. There were large differences in the most commonly isolated serovars between regions, but lesser differences between countries within the same region. The results also highlight the complexity of the global epidemiology of Salmonella and the need and importance for improving monitoring data of those serovars of highest epidemiologic importance.

Population Genetics of Vibrio cholerae from Nepal in 2010: Evidence on the Origin of the Haitian Outbreak

ABSTRACT Cholera continues to be an important cause of human infections, and outbreaks are often observed after natural disasters, such as the one following the 2010 earthquake in Haiti. Once the cholera outbreak was confirmed, rumors spread that the disease was brought to Haiti by a battalion of Nepalese soldiers serving as United Nations peacekeepers. This possible connection has never been confirmed. We used whole-genome sequence typing (WGST), pulsed-field gel electrophoresis (PFGE), and antimicrobial susceptibility testing to characterize 24 recent Vibrio cholerae isolates from Nepal and evaluate the suggested epidemiological link with the Haitian outbreak. The isolates were obtained from 30 July to 1 November 2010 from five different districts in Nepal. We compared the 24 genomes to 10 previously sequenced V. cholerae isolates, including 3 from the Haitian outbreak (began July 2010). Antimicrobial susceptibility and PFGE patterns were consistent with an epidemiological link between the isolates from Nepal and Haiti. WGST showed that all 24 V. cholerae isolates from Nepal belonged to a single monophyletic group that also contained isolates from Bangladesh and Haiti. The Nepalese isolates were divided into four closely related clusters. One cluster contained three Nepalese isolates and three Haitian isolates that were almost identical, with only 1- or 2-bp differences. Results in this study are consistent with Nepal as the origin of the Haitian outbreak. This highlights how rapidly infectious diseases might be transmitted globally through international travel and how public health officials need advanced molecular tools along with standard epidemiological analyses to quickly determine the sources of outbreaks. IMPORTANCE Cholera is one of the ancient classical diseases and particularly prone to cause major outbreaks following major natural disasters, such as earthquakes and hurricanes, where the normal separation between sewage and drinking water is destroyed. This was the case following the 2010 earthquake in Haiti. Rumors spread that the disease was brought to Haiti by a battalion of Nepalese soldiers serving as United Nations peacekeepers. This possible connection has never been confirmed. Sequencing the genomes of bacteria can give detailed information on whether isolates from different sites share a common origin. We used this technology to sequence isolates of Vibrio cholerae from Nepal, identify single-nucleotide polymorphisms (SNPs), and compare these high-resolution genotypes to the complete genome sequences of isolates from the Haiti outbreak. We provide support for the hypothesis that the isolates were brought to Haiti from Nepal. Cholera is one of the ancient classical diseases and particularly prone to cause major outbreaks following major natural disasters, such as earthquakes and hurricanes, where the normal separation between sewage and drinking water is destroyed. This was the case following the 2010 earthquake in Haiti. Rumors spread that the disease was brought to Haiti by a battalion of Nepalese soldiers serving as United Nations peacekeepers. This possible connection has never been confirmed. Sequencing the genomes of bacteria can give detailed information on whether isolates from different sites share a common origin. We used this technology to sequence isolates of Vibrio cholerae from Nepal, identify single-nucleotide polymorphisms (SNPs), and compare these high-resolution genotypes to the complete genome sequences of isolates from the Haiti outbreak. We provide support for the hypothesis that the isolates were brought to Haiti from Nepal.

Detection of mcr-1 encoding plasmid-mediated colistin-resistant Escherichia coli isolates from human bloodstream infection and imported chicken meat, Denmark 2015.

The plasmid-mediated colistin resistance gene, mcr-1, was detected in an Escherichia coli isolate from a Danish patient with bloodstream infection and in five E. coli isolates from imported chicken meat. One isolate from chicken meat belonged to the epidemic spreading sequence type ST131. In addition to IncI2, an incX4 replicon was found to be linked to mcr-1. This report follows a recent detection of mcr-1 in E. coli from animals, food and humans in China.

Salmonella serovars from humans and other sources in Thailand, 1993-2002.

We serotyped 44,087 Salmonella isolates from humans and 26,148 from other sources from 1993 through 2002. The most common serovar causing human salmonellosis in Thailand was Salmonella enterica Weltevreden. Serovars causing human infections in Thailand differ from those in other countries and seem to be related to Salmonella serovars in different food products and reservoirs.

International Spread of an Epidemic Population of Salmonella enterica Serotype Kentucky ST198 Resistant to Ciprofloxacin

National Salmonella surveillance systems from France, England and Wales, Denmark, and the United States identified the recent emergence of multidrug-resistant isolates of Salmonella enterica serotype Kentucky displaying high-level resistance to ciprofloxacin. A total of 489 human cases were identified during the period from 2002 (3 cases) to 2008 (174 cases). These isolates belonged to a single clone defined by the multilocus sequence type ST198, the XbaI-pulsed-field gel electrophoresis cluster X1, and the presence of the Salmonella genomic island 1 variant SGI1-K. This clone was probably selected in 3 steps in Egypt during the 1990s and the early 2000s and has now spread to several countries in Africa and, more recently, in the Middle East. Poultry has been identified as a potential major vehicle for infection by this clone. Continued surveillance and appropriate control measures should be implemented by national and international authorities to limit the spread of this strain.

The Salmonella enterica pan-genome.

Salmonella enterica is divided into four subspecies containing a large number of different serovars, several of which are important zoonotic pathogens and some show a high degree of host specificity or host preference. We compare 45 sequenced S. enterica genomes that are publicly available (22 complete and 23 draft genome sequences). Of these, 35 were found to be of sufficiently good quality to allow a detailed analysis, along with two Escherichia coli strains (K-12 substr. DH10B and the avian pathogenic E. coli (APEC O1) strain). All genomes were subjected to standardized gene finding, and the core and pan-genome of Salmonella were estimated to be around 2,800 and 10,000 gene families, respectively. The constructed pan-genomic dendrograms suggest that gene content is often, but not uniformly correlated to serotype. Any given Salmonella strain has a large stable core, whilst there is an abundance of accessory genes, including the Salmonella pathogenicity islands (SPIs), transposable elements, phages, and plasmid DNA. We visualize conservation in the genomes in relation to chromosomal location and DNA structural features and find that variation in gene content is localized in a selection of variable genomic regions or islands. These include the SPIs but also encompass phage insertion sites and transposable elements. The islands were typically well conserved in several, but not all, isolates—a difference which may have implications in, e.g., host specificity.

International Spread of Multidrug-resistant Salmonella Schwarzengrund in Food Products

This serovar was isolated from persons, food, and food animals in Thailand, Denmark, and the United States.

Identification of a novel transposon-associated phosphoethanolamine transferase gene, mcr-5, conferring colistin resistance in d-tartrate fermenting Salmonella enterica subsp. enterica serovar Paratyphi B

Objectives Plasmid-mediated mobilized colistin resistance is currently known to be caused by phosphoethanolamine transferases termed MCR-1, MCR-2, MCR-3 and MCR-4. However, this study focuses on the dissection of a novel resistance mechanism in mcr-1-, mcr-2- and mcr-3-negative d-tartrate fermenting Salmonella enterica subsp. enterica serovar Paratyphi B (Salmonella Paratyphi B dTa+) isolates with colistin MIC values >2 mg/L. Methods A selected isolate from the strain collection of the German National Reference Laboratory for Salmonella was investigated by WGS and bioinformatical analysis to identify novel phosphoethanolamine transferase genes involved in colistin resistance. Subsequently PCR screening, S1-PFGE and DNA-DNA hybridization were performed to analyse the prevalence and location of the identified mcr-5 gene. Cloning and transformation experiments in Escherichia coli DH5α and Salmonella Paratyphi B dTa+ control strains were carried out and the activity of MCR-5 was determined in vitro by MIC testing. Results In this study, we identified a novel phosphoethanolamine transferase in 14 mcr-1-, mcr-2- and mcr-3-negative Salmonella Paratyphi B dTa+ isolates with colistin MIC values >2 mg/L that were received during 2011-13. The respective gene, further termed as mcr-5 (1644 bp), is part of a 7337 bp transposon of the Tn3 family and usually located on related multi-copy ColE-type plasmids. Interestingly, in one isolate an additional subclone with a chromosomal location of the mcr-5 transposon was observed. Conclusions Our findings suggest that the transfer of colistin-resistance-mediating phosphoethanolamine transferase genes from bacterial chromosomes to mobile genetic elements has occurred in multiple independent events raising concern regarding their variety, prevalence and impact on public health.

Molecular Characterization and Antimicrobial Susceptibility of Salmonella Isolates from Infections in Humans in Henan Province, China

ABSTRACT We characterized 208 human Salmonella isolates from 2006 to 2007 and 27 human Salmonella enterica serovar Typhimurium isolates from 1987 to 1993 from Henan Province, China, by serotyping, by antimicrobial susceptibility testing, and, for the most common serovars, by pulsed-field gel electrophoresis (PFGE). The most common serovars among the 2006-2007 isolates were S. enterica serovar Typhimurium (27%), S. enterica serovar Enteritidis (17%), S. enterica serovar Derby (10%), S. enterica serovar Indiana (6%), and S. enterica serovar Litchfield (6%). A high percentage of the isolates were multiple-drug resistant, and 54% were resistant to both nalidixic acid and ciprofloxacin. Of these, 42% were resistant to a high level of ciprofloxacin (MIC > 4 μg/ml), whereas for the remaining isolates, the MICs ranged from 0.125 to 2 μg/ml. Five isolates (2%) were ceftiofur resistant and harbored blaCTX-M14 or blaCTX-M15. With the possible exception of the quinolones and cephalosporins, the 1987-1993 S. enterica serovar Typhimurium isolates were almost as resistant as the recent isolates. PFGE typing of S. enterica serovar Typhimurium showed that the most common cluster predominated over time. Two other clusters have emerged, and another cluster has disappeared.