Comparative Genomic Analysis of Antimicrobial-Resistant Escherichia coli from South American Camelids in Central Germany

South American camelids (SAC) are increasingly kept in Europe in close contact with humans and other livestock species and can potentially contribute to transmission chains of epizootic, zoonotic and antimicrobial-resistant (AMR) agents from and to livestock and humans. Consequently, SAC were included as livestock species in the new European Animal Health Law. However, the knowledge on bacteria exhibiting AMR in SAC is too scarce to draft appropriate monitoring and preventive programs. During a survey of SAC holdings in central Germany, 39 Escherichia coli strains were isolated from composite fecal samples by selecting for cephalosporin or fluoroquinolone resistance and were here subjected to whole-genome sequencing. The data were bioinformatically analyzed for strain phylogeny, detection of pathovars, AMR genes and plasmids. Most (33/39) strains belonged to phylogroups A and B1. Still, the isolates were highly diverse, as evidenced by 28 multi-locus sequence types. More than half of the isolates (23/39) were genotypically classified as multidrug resistant. Genes mediating resistance to trimethoprim/sulfonamides (22/39), aminoglycosides (20/39) and tetracyclines (18/39) were frequent. The most common extended-spectrum-β-lactamase gene was bla CTX-M-1 (16/39). One strain was classified as enteropathogenic E. coli . The positive results indicate the need to include AMR bacteria in yet-to-be-established animal disease surveillance protocols for SAC

Kausale Erklärungen in der Geschichtswissenschaft - Konzepte und Fallbeispiele

Die Glücksgöttin Fortuna in Rom ist wohl eine der bekanntesten Gottheiten, die menschliche Geschicke leiten sollen. Andere Schlagwörter in diesem Zusammenhang sind Zufall, Pech, Schicksal, Glückssträhne etc. Geht man davon aus, dass Menschen zu diesen Konstrukten greifen, weil sie die Geschehnisse in der Welt und die ihnen zugrunde liegenden Ursache-Wirkung-Beziehungen nicht verstehen, so ist eine Klärung des Kausalitätsprinzips nötig. Bisher ist es nicht gelungen, Kausalgesetze in der Geschichte zu ermitteln. Nichtsdestotrotz existiert eine Vielzahl von Ansätzen zur Untersuchung ursächlicher Beziehungen. Einige ausgewählte Konzepte werden in dieser Arbeit anhand von Fallbeispielen vorgestellt

Evaluation of Operation IceBridge quick-look snow depth estimates on sea ice

We evaluate Operation IceBridge (OIB) ‘quick-look’ (QL) snow depth on sea ice retrievals using in situ measurements taken over immobile first-year ice (FYI) and multi-year ice (MYI) during March of 2014. Good agreement was found over undeformed FYI (-4.5 cm mean bias) with reduced agreement over deformed FYI (-6.6 cm mean bias). Over MYI, the mean bias was -5.7 cm but 54% of retrievals were discarded by the OIB retrieval process as compared to only 10% over FYI. Footprint scale analysis revealed a root mean square error (RMSE) of 6.2 cm over undeformed FYI with RMSE of 10.5 cm and 17.5 cm in the more complex deformed FYI and MYI environments. Correlation analysis was used to demonstrate contrasting retrieval uncertainty associated with spatial aggregation and ice surface roughness

Using unique ORFan genes as strain-specific identifiers for Escherichia coli

Background Bacterial identification at the strain level is a much-needed, but arduous and challenging task. This study aimed to develop a method for identifying and differentiating individual strains among multiple strains of the same bacterial species. The set used for testing the method consisted of 17 Escherichia coli strains picked from a collection of strains isolated in Germany, Spain, the United Kingdom and Vietnam from humans, cattle, swine, wild boars, and chickens. We targeted unique or rare ORFan genes to address the problem of selective and specific strain identification. These ORFan genes, exclusive to each strain, served as templates for developing strain-specific primers. Results Most of the experimental strains (14 out of 17) possessed unique ORFan genes that were used to develop strain-specific primers. The remaining three strains were identified by combining a PCR for a rare gene with a selection step for isolating the experimental strains. Multiplex PCR allowed the successful identification of the strains both in vitro in spiked faecal material in addition to in vivo after experimental infections of pigs and recovery of bacteria from faecal material. In addition, primers for qPCR were also developed and quantitative readout from faecal samples after experimental infection was also possible. Conclusions The method described in this manuscript using strain-specific unique genes to identify single strains in a mixture of strains proved itself efficient and reliable in detecting and following individual strains both in vitro and in vivo, representing a fast and inexpensive alternative to more costly methods.Peer Reviewe

Conformational epitopes of myelin oligodendrocyte glycoprotein are targets of potentially pathogenic antibody responses in multiple sclerosis

<p>Abstract</p> <p>Background</p> <p>Myelin/oligodendrocyte glycoprotein (MOG) is a putative autoantigen in multiple sclerosis (MS). Establishing the pathological relevance and validity of anti-MOG antibodies as biomarkers has yielded conflicting reports mainly due to different MOG isoforms used in different studies. Because epitope specificity may be a key factor determining anti-MOG reactivity we aimed at identifying <it>a priori </it>immunodominant MOG epitopes by monoclonal antibodies (mAbs) and at assessing clinical relevance of these epitopes in MS.</p> <p>Methods</p> <p>Sera of 325 MS patients, 69 patients with clinically isolated syndrome and 164 healthy controls were assayed by quantitative, high-throughput ELISA for reactivity to 3 different MOG isoforms, and quantitative titers correlated with clinical characteristics. mAbs defined unique immunodominant epitopes distinct to each of the isoforms.</p> <p>Results</p> <p>In the majority of human samples anti-MOG levels were skewed towards low titers. However, in 8.2% of samples high-titer anti-MOG antibodies were identified. In contrast to anti-MOG reactivity observed in a mouse model of MS, in patients with MS these never reacted with ubiquitously exposed epitopes. Moreover, in patients with relapsing-remitting MS high-titer anti-MOG IgG correlated with disability (EDSS; Spearman r = 0.574; p = 0.025).</p> <p>Conclusions</p> <p>Thus high-titer reactivity likely represents high-affinity antibodies against pathologically relevant MOG epitopes, that are only present in a small proportion of patients with MS. Our study provides valuable information about requirements of anti-MOG reactivity for being regarded as a prognostic biomarker in a subtype of MS.</p

Evidence for Contemporary Switching of the O-Antigen Gene Cluster between Shiga Toxin-Producing Escherichia coli Strains Colonizing Cattle

Shiga toxin-producing Escherichia coli (STEC) comprise a group of zoonotic enteric pathogens with ruminants, especially cattle, as the main reservoir. O-antigens are instrumental for host colonization and bacterial niche adaptation. They are highly immunogenic and, therefore, targeted by the adaptive immune system. The O-antigen is one of the most diverse bacterial cell constituents and variation not only exists between different bacterial species, but also between individual isolates/strains within a single species. We recently identified STEC persistently infecting cattle and belonging to the different serotypes O156:H25 (n = 21) and O182:H25 (n = 15) that were of the MLST sequence types ST300 or ST688. These STs differ by a single nucleotide in purA only. Fitness-, virulence-associated genome regions, and CRISPR/CAS (clustered regularly interspaced short palindromic repeats/CRISPR associated sequence) arrays of these STEC O156:H25 and O182:H25 isolates were highly similar, and identical genomic integration sites for the stx converting bacteriophages and the core LEE, identical Shiga toxin converting bacteriophage genes for stx1a, identical complete LEE loci, and identical sets of chemotaxis and flagellar genes were identified. In contrast to this genomic similarity, the nucleotide sequences of the O-antigen gene cluster (O-AGC) regions between galF and gnd and very few flanking genes differed fundamentally and were specific for the respective serotype. Sporadic aEPEC O156:H8 isolates (n = 5) were isolated in temporal and spatial proximity. While the O-AGC and the corresponding 5′ and 3′ flanking regions of these aEPEC isolates were identical to the respective region in the STEC O156:H25 isolates, the core genome, the virulence associated genome regions and the CRISPR/CAS elements differed profoundly. Our cumulative epidemiological and molecular data suggests a recent switch of the O-AGC between isolates with O156:H8 strains having served as DNA donors. Such O-antigen switches can affect the evaluation of a strain's pathogenic and virulence potential, suggesting that NGS methods might lead to a more reliable risk assessment

Clostridioides difficile in South American Camelids in Germany: First Insights into Molecular and Genetic Characteristics and Antimicrobial Resistance

Little is known about zoonotic pathogens and their antimicrobial resistance in South American camelids (SAC) in Germany including Clostridioides (C.) difficile. The aim of this study was to investigate prevalence, molecular characteristics and antimicrobial resistance of C. difficile in SAC. Composite SAC faecal samples were collected in 43 husbandries in Central Germany and cultured for C. difficile. Toxinotyping and ribotyping was done by PCR. Whole genome sequencing was performed with Illumina® Miseq™. The genomes were screened for antimicrobial resistance determinants. Genetic relatedness of the isolates was investigated using core genome multi locus sequence typing (cgMLST) and single nucleotide polymorphism analysis. Antimicrobial susceptibility testing was done using the Etest® method. Eight C. difficile isolates were recovered from seven farms. The isolates belonged to different PCR ribotypes. All isolates were toxinogenic. cgMLST revealed a cluster containing isolates recovered from different farms. Seven isolates showed similar resistance gene patterns. Different phenotypic resistance patterns were found. Agreement between phenotypic and genotypic resistance was identified only in some cases. Consequently, SAC may act as a reservoir for C. difficile. Thus, SAC may pose a risk regarding zoonotic transmission of toxinogenic, potentially human-pathogenic and resistant C. difficile isolates

High frequency of chlamydial co-infections in clinically healthy sheep flocks

<p>Abstract</p> <p>Background</p> <p>The epidemiological situation of ovine chlamydial infections in continental Europe, especially Germany is poorly characterised. Using the German state of Thuringia as a model example, the chlamydial sero- and antigen prevalence was estimated in thirty-two randomly selected sheep flocks with an average abortion rate lower than 1%. Seven vaccinated flocks were reviewed separately.</p> <p>Results</p> <p>A wide range of samples from 32 flocks were examined. Assumption of a seroprevalence of 10% (CI 95%) at flock level, revealed that 94% of the tested flocks were serologically positive with ongoing infection (i.e. animals with seroconversion) in nearly half (47%) of the flocks. On the basis of an estimated 25% antigen prevalence (CI 95%), PCR and DNA microarray testing, together with sequencing revealed the presence of chlamydiae in 78% of the flocks. The species most frequently found was <it>Chlamydophila (C</it>.) <it>abortus </it>(50%) followed by <it>C. pecorum </it>(47%) and <it>C. psittaci </it>genotype A (25%). Mixed infections occurred in 25% of the tested flocks. Samples obtained from the vaccinated flocks revealed the presence of <it>C. abortus </it>field samples in 4/7 flocks. <it>C. pecorum </it>was isolated from 2/7 flocks and the presence of seroconversion was determined in 3/7 flocks.</p> <p>Conclusions</p> <p>The results imply that chlamydial infections occur frequently in German sheep flocks, even in the absence of elevated abortion rates. The fact that <it>C. pecorum </it>and the potentially zoonotic <it>C. psittaci </it>were found alongside the classical abortifacient agent <it>C. abortus</it>, raise questions about the significance of this reservoir for animal and human health and underline the necessity for regular monitoring. Further studies are needed to identify the possible role of <it>C. psittaci </it>infections in sheep.</p

In Vitro Antibacterial Activity of Microbial Natural Products against Bacterial Pathogens of Veterinary and Zoonotic Relevance

Antimicrobial resistance (AMR) is considered one of the greatest threats to both human and animal health. Efforts to address AMR include implementing antimicrobial stewardship programs and introducing alternative treatment options. Nevertheless, effective treatment of infectious diseases caused by bacteria will still require the identification and development of new antimicrobial agents. Eight different natural products were tested for antimicrobial activity against seven pathogenic bacterial species (Brachyspira sp., Chlamydia sp., Clostridioides sp., Mannheimia sp., Mycobacterium sp., Mycoplasma sp., Pasteurella sp.). In a first pre-screening, most compounds (five out of eight) inhibited bacterial growth only at high concentrations, but three natural products (celastramycin A [CA], closthioamide [CT], maduranic acid [MA]) displayed activity at concentrations 16 µg/mL against Mannheimia for CA, CT, and MA, respectively. CA, CT, and MA exhibited higher MIC50 and MIC90 values against Pasteurella isolates with a known AMR phenotype against commonly used therapeutic antimicrobial agents than against isolates with unknown AMR profiles. This study demonstrates the importance of whole-cell antibacterial screening of natural products to identify promising scaffolds with broad- or narrow-spectrum antimicrobial activity against important Gram-negative veterinary pathogens with zoonotic potentia