Phage Treatment Trial to Eradicate LA-MRSA from Healthy Carrier Pigs

The increase of livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA) causes a threat to human health. LA-MRSA can be transmitted from animals to animal caretakers, which may further spread MRSA to communities and health care facilities. The objective of this work was to study the efficacy of phage treatment in the eradication of LA-MRSA from healthy carrier pigs. A total of 19 MRSA -positive weanling pigs were assigned to a test (n = 10) and a control group (n = 9). A phage cocktail containing three Staphylococcus phages, or a control buffer was administered to the nares and skin of the pigs three times every two days, after which the phage and MRSA levels in nasal and skin swab samples were monitored for a three-week period. The sensitivity of the strains isolated during the follow-up period to the phage cocktail and each phage individually was analyzed and the pig sera were tested for antibodies against the phages used in the cocktail. The phage treatment did not cause any side effects to the pigs. Phages were found in the skin and nasal samples on the days following the phage applications, but there was no reduction in the MRSA levels in the sampled animals. Phage-resistant strains or phage-specific antibodies were not detected during the experiment. The MRSA load in these healthy carrier animals was only 10–100 CFU/swab or nasal sample, which was likely below the replication threshold of phages. The effectiveness of phage treatment to eradicate MRSA from the pigs could thus not be (reliably) determined

Phage Treatment Trial to Eradicate LA-MRSA from Healthy Carrier Pigs

The increase of livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA) causes a threat to human health. LA-MRSA can be transmitted from animals to animal caretakers, which may further spread MRSA to communities and health care facilities. The objective of this work was to study the efficacy of phage treatment in the eradication of LA-MRSA from healthy carrier pigs. A total of 19 MRSA -positive weanling pigs were assigned to a test (n = 10) and a control group (n = 9). A phage cocktail containing three Staphylococcus phages, or a control buffer was administered to the nares and skin of the pigs three times every two days, after which the phage and MRSA levels in nasal and skin swab samples were monitored for a three-week period. The sensitivity of the strains isolated during the follow-up period to the phage cocktail and each phage individually was analyzed and the pig sera were tested for antibodies against the phages used in the cocktail. The phage treatment did not cause any side effects to the pigs. Phages were found in the skin and nasal samples on the days following the phage applications, but there was no reduction in the MRSA levels in the sampled animals. Phage-resistant strains or phage-specific antibodies were not detected during the experiment. The MRSA load in these healthy carrier animals was only 10–100 CFU/swab or nasal sample, which was likely below the replication threshold of phages. The effectiveness of phage treatment to eradicate MRSA from the pigs could thus not be (reliably) determined

Novel canine high-quality metagenome-assembled genomes, prophages and host-associated plasmids provided by long-read metagenomics together with Hi-C proximity ligation

The human gut microbiome has been extensively studied, yet the canine gut microbiome is still largely unknown. The availability of high-quality genomes is essential in the fields of veterinary medicine and nutrition to unravel the biological role of key microbial members in the canine gut environment. Our aim was to evaluate nanopore long-read metagenomics and Hi-C (high-throughput chromosome conformation capture) proximity ligation to provide high-quality metagenome-assembled genomes (HQ MAGs) of the canine gut environment. By combining nanopore long-read metagenomics and Hi-C proximity ligation, we retrieved 27 HQ MAGs and 7 medium-quality MAGs of a faecal sample of a healthy dog. Canine MAGs (CanMAGs) improved genome contiguity of representatives from the animal and human MAG catalogues - short-read MAGs from public datasets - for the species they represented: they were more contiguous with complete ribosomal operons and at least 18 canonical tRNAs. Both canine-specific bacterial species and gut generalists inhabit the dog's gastrointestinal environment. Most of them belonged to , followed by and . We also assembled one and one MAG. CanMAGs harboured antimicrobial-resistance genes (ARGs) and prophages and were linked to plasmids. ARGs conferring resistance to tetracycline were most predominant within CanMAGs, followed by lincosamide and macrolide ones. At the functional level, carbohydrate transport and metabolism was the most variable within the CanMAGs, and mobilome function was abundant in some MAGs. Specifically, we assigned the mobilome functions and the associated mobile genetic elements to the bacterial host. The CanMAGs harboured 50 bacteriophages, providing novel bacterial-host information for eight viral clusters, and Hi-C proximity ligation data linked the six potential plasmids to their bacterial host. Long-read metagenomics and Hi-C proximity ligation are likely to become a comprehensive approach to HQ MAG discovery and assignment of extra-chromosomal elements to their bacterial host. This will provide essential information for studying the canine gut microbiome in veterinary medicine and animal nutrition

Whole-Genome Sequence Analysis of an Extensively Drug-Resistant Salmonella enterica Serovar Agona Isolate from an Australian Silver Gull (Chroicocephalus novaehollandiae) Reveals the Acquisition of Multidrug Resistance Plasmids.

Although most of the approximately 94 million annual human cases of gastroenteritis due to Salmonella enterica resolve without medical intervention, antimicrobial therapy is recommended for patients with severe disease. Wild birds can be natural hosts of Salmonella that pose a threat to human health; however, multiple-drug-resistant serovars of S. enterica have rarely been described. In 2012, silver gull (Chroicocephalus novaehollandiae) chicks at a major breeding colony were shown to host Salmonella, most isolates of which were susceptible to antibiotics. However, multiple-drug-resistant (MDR) Escherichia coli with resistance to carbapenems, ceftazidime, and fluoroquinolones was reported from this breeding colony. In this paper, we describe a novel MDR Salmonella strain subsequently isolated from the same breeding colony. SG17-135, an isolate of S. enterica with phenotypic resistance to 12 individual antibiotics but only nine antibiotic classes including penicillins, cephalosporins, monobactams, macrolides, fluoroquinolones, aminoglycosides, dihydrofolate reductase inhibitors (trimethoprim), sulfonamides, and glycylcyclines was recovered from a gull chick in 2017. Whole-genome sequence (WGS) analysis of SG17-135 identified it as Salmonella enterica serovar Agona (S Agona) with a chromosome comprising 4,813,284 bp, an IncHI2 ST2 plasmid (pSG17-135-HI2) of 311,615 bp, and an IncX1 plasmid (pSG17-135-X) of 27,511 bp. pSG17-135-HI2 housed a complex resistance region comprising 16 antimicrobial resistance genes including blaCTX-M-55 The acquisition of MDR plasmids by S. enterica described here poses a serious threat to human health. Our study highlights the importance of taking a One Health approach to identify environmental reservoirs of drug-resistant pathogens and MDR plasmids.IMPORTANCE Defining environmental reservoirs hosting mobile genetic elements that shuttle critically important antibiotic resistance genes is key to understanding antimicrobial resistance (AMR) from a One Health perspective. Gulls frequent public amenities, parklands, and sewage and other waste disposal sites and carry drug-resistant Escherichia coli Here, we report on SG17-135, a strain of Salmonella enterica serovar Agona isolated from the cloaca of a silver gull chick nesting on an island in geographic proximity to the greater metropolitan area of Sydney, Australia. SG17-135 is closely related to pathogenic strains of S Agona, displays resistance to nine antimicrobial classes, and carries important virulence gene cargo. Most of the antibiotic resistance genes hosted by SG17-135 are clustered on a large IncHI2 plasmid and are flanked by copies of IS26 Wild birds represent an important link in the evolution and transmission of resistance plasmids, and an understanding of their behavior is needed to expose the interplay between clinical and environmental microbial communities

Comparative genomics of dairy-associated Staphylococcus aureus from selected sub-Saharan African regions reveals milk as reservoir for human-and animal-derived strains and identifies a putative animal-related clade with presumptive novel siderophore

Staphylococcus aureus infection is considered to be a neglected tropical disease with huge impact on human and animal health alike. Dairy production in sub-Saharan Africa (SSA) relies heavily on various animals such as cows, goats, and camels, depending on the region. S. aureus causes mastitis and exhibits high prevalence in raw milk. The population structure including genotypic and phenotypic traits of dairy S. aureus in relation to animal and human isolates is, however, unknown for SSA. In this work, 20 S. aureus dairy isolates from East and West Africa were selected for comparative genomics and phenotypic analysis. Comparing their population structure revealed a large diversity of different origins suggesting milk to be a reservoir for human and animal strains alike. Furthermore, a novel putative siderophore was detected in multiple strains in a distinct animal-clade with strains of global origin. This putative siderophore shares a high genetic identity with that from Streptococcus equi suggesting possible horizontal gene transfer. These findings combined with the virulence genes harbored by these dairy-derived strains such as pvl, human evasion factor scn, various enterotoxin, leucocidin and antibiotic resistance genes, stresses the need for an integrative One Health approach to tackle the problem of S. aureus infections in animals and humans in sub-Saharan Africa

Prevalence and Population Diversity of Listeria monocytogenes Isolated from Dairy Cattle Farms in the Cantabria Region of Spain

Listeria monocytogenes is an opportunistic pathogen that is widely distributed in the environment. Here we show the prevalence and transmission of L. monocytogenes in dairy farms in the Cantabria region, on the northern coast of Spain. A total of 424 samples was collected from 14 dairy farms (5 organic and 9 conventional) and 211 L. monocytogenes isolates were recovered following conventional microbiological methods. There were no statistically significant differences in antimicrobial resistance ratios between organic and conventional farms. A clonal relationship among the isolates was assessed by pulsed field gel electrophoresis (PFGE) analysis and 64 different pulsotypes were obtained. Most isolates (89%, n = 187) were classified as PCR serogroup IVb by using a multiplex PCR assay. In this case, 45 isolates of PCR serogroup IVb were whole genome-sequenced to perform a further analysis at genomic level. In silico MLST analysis showed the presence of 12 sequence types (ST), of which ST1, ST54 and ST666 were the most common. Our data indicate that the environment of cattle farms retains a high incidence of L. monocytogenes, including subtypes involved in human listeriosis reports and outbreaks. This pathogen is shed in the feces and could easily colonize dairy products, as a result of fecal contamination. Effective herd and manure management are needed in order to prevent possible outbreaks.This work was supported by Research Project grants RTA08-099, RTA2008-00080-C02, RTA2014-00045-C03-01 (INIA and FEDER) from the Spanish Ministry of Economy and Competitiveness and RTI2018-098267-R-C31 (INIA and FEDER) from the Spanish Ministry of Science and Innovation.S

A living inquiry: activating and integrating the multiple identities of dramatherapist, researcher, artist and teacher

The critical narrative explores published and creative works as a performer and how this has informed the author’s development as researcher, artist, dramatherapist and teacher in Higher Education. Through clinical work as a dramatherapist in different settings and practice as a dramatherapy educator the author explores the integrating of therapy and teaching approaches such as meaning making, deep learning and student-centred learning. The research is a living inquiry that is an unfolding of the researcher’s experience and deepening of understanding over a fourteen-year period as a clinician and teacher. The author examines the encounter and interrelationship dynamic between the performer and audience; therapist and client; teacher and student; and researcher and research participant as a means for co-construction and co-creation of meaning, knowledge and raising awareness. The author explores the importance of a research approach that is congruent with a dramatherapy and teacher identity emphasising non-verbal exploration, the senses, the imagination and play as a means for deepening understanding of dramatherapy and teaching practice. Through the creative works of a solo performer the author explores how engagement as an artist informs and strengthens their identity as a dramatherapist, researcher and teacher. Different research methodologies including heuristic inquiry, autoethnography and a/r/tography are considered for their suitability for a living inquiry that addresses the multiplicity of roles.N/

Population Health of Spotted Salamanders (Ambystoma maculatum) in Created Vernal Pools: an Integrative Approach

Habitat creation is an important tool for conservation to counteract habitat loss and degradation. Vernal pools are susceptible to destruction due to limited detection, protection, and regulation. These wetlands provide fishless breeding habitat for many amphibian species including spotted salamanders (Ambystoma maculatum) in eastern North America. Determining whether created vernal pool habitat is successful is often determined by demographic data of colonizing populations. I suggest that hormone levels, population genetics, and disease prevalence can improve our understanding of population health in created habitat. The goal of this dissertation was to assess the health of spotted salamander larvae in created vernal pools quantified by corticosterone levels, genetic diversity and structure, and ranavirus prevalence. I also wanted to determine if there were habitat characteristics impacting these parameters to inform future construction techniques and produce habitat conducive to healthy colonizing amphibian populations. I examined whether habitat traits influenced corticosterone levels of salamander larvae in created vernal pools. The strongest model predicting corticosterone levels included larval total length, pool-water temperature, year sampled, and pool diameter. Annual variation in corticosterone levels and habitat characteristics, and positive associations with water temperature and salamander body size highlighted the importance of controlling for external influences. The negative association between pool diameter and corticosterone indicated that larvae in larger pools (up to 12.75-m maximum diameter) were less stressed and potentially healthier. Water-borne hormone sampling is relatively new to amphibians, so I attempted to biologically validate the method for spotted salamanders by comparing water-borne and plasma corticosterone concentrations of the same individuals. There were differences in corticosterone concentrations between larvae, metamorphs, and adults, but there were no correlations between water-borne and plasma concentrations for any of the age groups. The two sampling methods have different units of measurement which might affect the association between the two. I evaluated the genetic structure and genetic diversity of spotted salamander larvae in created vernal pools. I also examined whether local habitat characteristics at the pool level influenced effective number of breeders, relatedness, or genetic diversity. The youngest pools exhibited genetic differentiation, founder’s effect, and low effective number of breeders. Effective number of breeders was positively associated with pool age, vegetation cover, pool diameter, and sample size. Pool cover and vegetation cover was also negatively associated with relatedness. Allelic richness and expected heterozygosity did not have strong environmental predictors. I surveyed spotted salamander larvae in created vernal pools for ranavirus prevalence and measured viral load in individual larvae. I tested associations between ranavirus prevalence and viral load and habitat characteristics, genetic diversity, corticosterone levels, and body size. I detected ranavirus in 62% of pools in 84 of 1,128 larvae (7%). Prevalence at pools ranged from 0–63%. Salamanders infected with ranavirus had greater total length, which was also positively correlated with viral load. There were no associations between ranavirus prevalence or viral load and habitat characteristics, salamander genetic diversity, relatedness, effective number of breeders, or corticosterone levels. These results indicate larger pool diameter (up to 12.75 m) and greater vegetation cover are important habitat traits to consider when creating vernal pool habitat for spotted salamanders. This research demonstrates that effective number of breeders can increase and genetic differentiation can decrease within 4–5 years of pool creation, a sign of rapid colonization and potential population establishment. The widespread occurrence of ranavirus in created pools illustrates the risk of disease exposure in newly created habitat; even with the generally low prevalence among salamanders in the majority of pools. The correlation between ranavirus and salamander total length could be a reflection of differences in susceptibility through developmental stages or increased risk of exposure over time. This study contributes to the growing body of knowledge on the association between habitat quality and corticosterone levels and genetic diversity of amphibians. The disease surveillance will add a species and county record for West Virginia, contribute to current distribution maps, and document disease prevalence levels in created habitat. The results of this dissertation will help inform current management and future site selection and habitat creation by highlighting important habitat characteristics for spotted salamanders. Finally, this research illustrates the value of incorporating interdisciplinary approaches to assess the success of habitat creation and colonizing wildlife populations