Whole-genome analysis of vancomycin-resistant Enterococcus faecium causing nosocomial outbreaks suggests the occurrence of few endemic clonal lineages in Bavaria, Germany.

OBJECTIVES: Infections caused by vancomycin-resistant Enterococcus faecium (VREfm) represent a major public health concern due to limited treatment options. Among invasive isolates of VREfm, ST117, ST80 and ST78 represent the most frequently detected STs by MLST in Germany. In this study, we investigated the genetic diversity of isolates of VREfm recovered from different nosocomial outbreaks in Bavaria, Germany, by WGS. METHODS: Between January 2018 and April 2019, 99 non-replicate isolates of VREfm originating from nosocomial outbreaks at eight different hospitals in Bavaria were investigated for genetic diversity by WGS. In detail, complex types (CTs) were identified by core-genome MLST. Furthermore, an SNP analysis was performed for all VREfm strains. RESULTS: Most of the isolates of this study (76%) belonged to three major clonal groups, which occurred in at least three hospitals: ST80/CT1065 vanB (n = 45; six hospitals), ST117/CT71 vanB (n = 11; four hospitals) and ST78/CT894like vanA (n = 19; three hospitals). Moreover, isolates of the predominant lineage ST80/CT1065 vanB showed a maximum difference of 36 SNPs as revealed by SNP analysis. CONCLUSIONS: Whole-genome analysis of VREfm causing nosocomial outbreaks suggests the occurrence of few endemic clonal lineages in Bavarian hospital settings, namely ST80/CT1065 vanB, ST117/CT71 vanB and ST78/CT894like vanA. Further studies are needed for a better understanding of the factors affecting the successful spread of the above-mentioned lineages.

Seroprevalence of Brucella Infection in Wild Boars (Sus scrofa) of Bavaria, Germany, 2019 to 2021 and Associated Genome Analysis of Five B. suis Biovar 2 Isolates.

Brucella species are highly pathogenic zoonotic agents and are found in vertebrates all over the world. To date, Germany is officially declared free from brucellosis and continuous surveillance is currently limited to farm ruminants. However, porcine brucellosis, mostly caused by B. suis biovar 2, is still found in wild boars and hares. In the present study, a three-year monitoring program was conducted focusing on the wild boar population in the state of Bavaria. Serologic screening of 11,956 animals and a direct pathogen detection approach, including a subset of 681 tissue samples, was carried out. The serologic incidence was 17.9%, which is in approximate accordance with previously published results from various European countries. Applying comparative whole genome analysis, five isolated B. suis biovar 2 strains from Bavaria could be assigned to three known European genetic lineages. One isolate was closely related to another strain recovered in Germany in 2006. Concluding, porcine brucellosis is endemic in Bavaria and the wild boar population represents a reservoir for genetically distinct B. suis biovar 2 strains. However, the transmission risk of swine brucellosis to humans and farm animals is still regarded as minor due to low zoonotic potential, awareness, and biosafety measures.

Emergence of novel ST1299 vanA lineages as possible cause for the striking rise of vancomycin resistance among invasive strains of Enterococcus faecium at a German university hospital.

IMPORTANCE: The proportion of VREfm among all Enterococcus faecium isolated from blood cultures in German hospitals has increased in the period 2015-2020 from 11.9% to 22.3% with a country-wide spread of the clonal lineage ST117/CT71 vanB. In this study, we provided useful information about the genetic diversity of invasive strains of E. faecium. Moreover, our findings confirm the nosocomial spread of novel ST1299 vanA lineages, which recently had a rapid expansion in Austria and the south-eastern part of Germany.

Whole-Genome Investigation of Salmonella Dublin Considering Mountain Pastures as Reservoirs in Southern Bavaria, Germany.

Worldwide, Salmonella Dublin (S. Dublin) is responsible for clinical disease in cattle and also in humans. In Southern Bavaria, Germany, the serovar was identified as a causative agent for 54 animal disease outbreaks in herds between 2017 and 2021. Most of these emerged from cattle herds (n = 50). Two occurred in pig farms and two in bovine herds other than cattle. Genomic analysis of 88 S. Dublin strains isolated during these animal disease outbreaks revealed 7 clusters with 3 different MLST-based sequence types and 16 subordinate cgMLST-based complex types. Antimicrobial susceptibility investigation revealed one resistant and three intermediate strains. Furthermore, only a few genes coding for bacterial virulence were found among the isolates. Genome analysis enables pathogen identification and antimicrobial susceptibility, serotyping, phylogeny, and follow-up traceback analysis. Mountain pastures turned out to be the most likely locations for transmission between cattle of different herd origins, as indicated by epidemiological data and genomic traceback analyses. In this context, S. Dublin shedding was also detected in asymptomatic herding dogs. Due to the high prevalence of S. Dublin in Upper Bavaria over the years, we suggest referring to this administrative region as "endemic". Consequently, cattle should be screened for salmonellosis before and after mountain pasturing.

Combined Clinical, Epidemiological, and Genome-Based Analysis Identified a Nationwide Outbreak of Burkholderia cepacia Complex Infections Caused by Contaminated Mouthwash Solutions.

Background: In September 2018, Burkholderia cepacia complex (BCC) infections in 3 patients associated with exposure to a mouthwash solution (MWS) were reported to the Robert Koch Institute (RKI). As the product was still on the market and the scale of the outbreak was unclear, a nation-wide investigation was initiated. Methods: We aimed to investigate BCC infections/colonizations associated with MWS. Hospitals, laboratories, and public health services were informed that BCC isolates should be sent to the RKI. These isolates were typed by pulsed-field gel electrophoresis (PFGE) and whole-genome sequencing (WGS) including development of an ad hoc core genome MLST (cgMLST) scheme. Results: In total, 36 patients from 6 hospitals met the case definition, the last patient in November 2018. Twenty-nine isolates from 26 of these patients were available for typing. WGS analysis revealed 2 distinct cgMLST clusters. Cluster 1 (Burkholderia arboris) contained isolates from patients and MWS obtained from 4 hospitals and isolates provided by the manufacturer. Patient and MWS isolates from another hospital were assigned to cluster 2 (B. cepacia). Conclusions: The combined clinical, epidemiological, and microbiological investigation, including whole-genome analysis, allowed for uncovering a supraregional BCC outbreak in health care settings. Strains of B. arboris and B. cepacia were identified as contaminating species of MWS bottles and subsequent colonization and putative infection of patients in several hospitals. Despite a recall of the product by the manufacturer in August 2018, the outbreak lasted until December 2018. Reporting of contaminated medical products and recalls should be optimized to protect patients.

First report of the new emerging global clone ST1193 among clinical isolates of extended-spectrum ß-lactamase (ESBL)-producing Escherichia coli from Germany.

OBJECTIVES: Sequence type 1193 (ST1193) is a new emerging global clone of Escherichia coli. The main goal of this study was to determine the prevalence and molecular characteristics of ST1193 among clinical isolates of extended-spectrum ß-lactamase (ESBL)-producing E. coli from University Hospital of Erlangen, Germany. METHODS: Between November 2015 and February 2016, all consecutive non-duplicate clinical E. coli isolates showing resistance to cefotaxime or ceftazidime were further analysed for ESBL production by the combined disk method. ESBL genes were identified by PCR and sequencing. Bacterial strain typing was performed by PCR-based phylogrouping, MLST and whole-genome sequencing. RESULTS: ESBL production was confirmed in 51 isolates. The globally dominant ST131 occurred at a frequency of 37.3% (n=19). Major non-ST131 sequence types were ST38 (n=4; 7.8%), ST10 (n=3; 5.9%) and ST1193 (n=3; 5.9%). Among the ESBL-producing E. coli ST1193, two expressed CTX-M-14 and one expressed CTX-M-15 ESBL type. All three ST1193 isolates belonged to serogroup O75:H5, phylogroup B2, and harboured IncFIA and IncFIB plasmids and the virulence factors genes iha, sat, gad, vat and senB. Moreover, they showed ciprofloxacin resistance and exhibited a set of four conserved mutations defining quinolone resistance (gyrA S83L, gyrA D87N, parC S80I and parC L416F). CONCLUSIONS: This study revealed for the first time in Germany the occurrence of ST1193 among clinical isolates of ESBL-producing E. coli. Further national or regional multicentre studies are needed to assess the effective relevance of ESBL-producing E. coli ST1193 as a nosocomial pathogen in Germany.

Molecular Characterization of Extended-Spectrum ß-Lactamase-Producing Escherichia coli Isolates from Milk Samples of Dairy Cows with Mastitis in Bavaria, Germany.

The aim of this study was to determine the rate of extended-spectrum ß-lactamase (ESBL)-producing microorganisms among Escherichia coli isolates causing bovine mastitis, including molecular characterization of these isolates. Therefore, a total of 490 bovine E. coli isolates from milk samples of dairy cows with mastitis were investigated for ESBL production by antimicrobial susceptibility testing, PCR-based detection, and sequencing of ESBL encoding genes, which were identified in 22 isolates (4.5%). Moreover, resistance to the fluoroquinolones enrofloxacin and marbofloxacin occurred in 15 of 22 ESBL-producing isolates (68.2%). All ESBL-producing isolates carried a blaCTX-M-like gene, with blaCTX-M-14 (n = 10) as the most prevalent type. Seven isolates producing CTX-M-14 and belonging to phylogenetic group A were further investigated for genetic relatedness by multilocus sequence typing. Five of them could be assigned to four different sequence types (STs): ST10 (n = 2), ST167 (n = 1), ST410 (n = 1), and ST744 (n = 1), whereas the remaining two isolates could not be assigned. To conclude, the rate of ESBL-producing E. coli associated with cattle mastitis was 4.5%. Furthermore, a high proportion of fluoroquinolone coresistance could be detected. Therefore, careful and continuous surveillance of ESBL-producing E. coli in cattle and consequent implementation of prevention measures are needed to avoid a further spread of these multidrug-resistant bacteria.