Insights into the Spectrum of Activity and Mechanism of Action of MGB-BP-3.

MGB-BP-3 is a potential first-in-class antibiotic, a Strathclyde Minor Groove Binder (S-MGB), that has successfully completed Phase IIa clinical trials for the treatment of Clostridioides difficile associated disease. Its precise mechanism of action and the origin of limited activity against Gram-negative pathogens are relatively unknown. Herein, treatment with MGB-BP-3 alone significantly inhibited the bacterial growth of the Gram-positive, but not Gram-negative, bacteria as expected. Synergy assays revealed that inefficient intracellular accumulation, through both permeation and efflux, is the likely reason for lack of Gram-negative activity. MGB-BP-3 has strong interactions with its intracellular target, DNA, in both Gram-negative and Gram-positive bacteria, revealed through ultraviolet-visible (UV-vis) thermal melting and fluorescence intercalator displacement assays. MGB-BP-3 was confirmed to bind to dsDNA as a dimer using nano-electrospray ionization mass spectrometry and nuclear magnetic resonance (NMR) spectroscopy. Type II bacterial topoisomerase inhibition assays revealed that MGB-BP-3 was able to interfere with the supercoiling action of gyrase and the relaxation and decatenation actions of topoisomerase IV of both Staphylococcus aureus and Escherichia coli. However, no evidence of stabilization of the cleavage complexes was observed, such as for fluoroquinolones, confirmed by a lack of induction of DSBs and the SOS response in E. coli reporter strains. These results highlight additional mechanisms of action of MGB-BP-3, including interference of the action of type II bacterial topoisomerases. While MGB-BP-3's lack of Gram-negative activity was confirmed, and an understanding of this presented, the recognition that MGB-BP-3 can target DNA of Gram-negative organisms will enable further iterations of design to achieve a Gram-negative active S-MGB.

The impact of the COVID-19 pandemic on a cohort of Labrador retrievers in England.

BACKGROUND: The COVID-19 pandemic is likely to have affected the welfare and health of dogs due to surges in adoptions and purchases, changes in the physical and mental health and financial status of dog owners, changes in dogs' lifestyle and routines and limited access to veterinary care. The aims of this study were to investigate whether COVID-19 restrictions were associated with differences in Labrador retrievers' lifestyle, routine care, insurance status, illness incidence or veterinary attendance with an illness, who were living in England and enrolled in Dogslife, an owner-based cohort study. Longitudinal questionnaire data from Dogslife that was relevant to the dates between the 23rd of March and the 4th of July 2020, during COVID-19 restrictions in England, were compared to data between the same dates in previous years from 2011 to 2019 using mixed regression models and adjusted chi-squared tests. RESULTS: Compared with previous years (March 23rd to July 4th, 2010 to 2019), the COVID-19 restrictions study period (March 23rd to July 4th 2020) was associated with owners reporting increases in their dogs' exercise and worming and decreases in insurance, titbit-feeding and vaccination. Odds of owners reporting that their dogs had an episode of coughing (0.20, 95% CI: 0.04-0.92) and that they took their dogs to a veterinarian with an episode of any illness (0.58, 95% CI: 0.45-0.76) were lower during the COVID-19 restrictions compared to before. During the restrictions period, owners were less likely to report that they took their dogs to a veterinarian with certain other illnesses, compared to before this period. CONCLUSIONS: Dogslife provided a unique opportunity to study prospective questionnaire data from owners already enrolled on a longitudinal cohort study. This approach minimised bias associated with recalling events prior to the pandemic and allowed a wider population of dogs to be studied than is available from primary care data. Distinctive insights into owners' decision making about their dogs' healthcare were offered. There are clear implications of the COVID-19 pandemic and associated restrictions for the lifestyle, care and health of dogs.

Mutations in SilS and CusS/OmpC represent different routes to achieve high level silver ion tolerance in Klebsiella pneumoniae.

BACKGROUND: Silver ions have potent broad-spectrum antimicrobial activity and are widely incorporated into a variety of products to limit bacterial growth. In Enterobacteriaceae, decreased silver susceptibility has been mapped to two homologous operons; the chromosomally located cus operon and the plasmid based sil operon. Here we characterised the mechanisms and clinical impact of induced silver tolerance in Klebsiella pneumoniae. RESULTS: In K. pneumoniae carriage of the sil operon alone does not give elevated silver tolerance. However, when exposed to increasing concentrations of silver nitrate (AgNO3), K. pneumoniae strains which contain the sil operon, will preferentially mutate SilS, resulting in overexpression of the genes encoding the RND efflux pump silCBA. Those strains which do not carry the sil operon also adapt upon exposure to increasing silver concentrations through mutations in another two-component regulator CusS. Secondary mutations leading to disruption of the outer membrane porin OmpC were also detected. Both routes result in a high level of silver tolerance with MIC's of >512 mg/L. When exposed to a high concentration of AgNO3 (400 mg/L), only strains that contained the sil operon were able to survive, again through mutations in SilS. The AgNO3 adapted strains were also resistant to killing by challenge with several clinical and commercial silver containing dressings. CONCLUSIONS: This study shows that K. pneumoniae has two possible pathways for development of increased silver tolerance but that the sil operon is preferentially mutated. This operon is essential when K. pneumoniae is exposed to high concentrations of silver. The potential clinical impact on wound management is shown by the increased survivability of these adapted strains when exposed to several silver impregnated dressings. This would make infections with these strains more difficult to treat and further limits our therapeutic options.

Disorder predispositions and protections of Labrador Retrievers in the UK.

The Labrador Retriever is one of the most popular dog breeds worldwide, therefore it is important to have reliable evidence on the general health issues of the breed. Using anonymised veterinary clinical data from the VetCompass Programme, this study aimed to explore the relative risk to common disorders in the Labrador Retriever. The clinical records of a random sample of dogs were reviewed to extract the most definitive diagnoses for all disorders recorded during 2016. A list of disorders was generated, including the 30 most common disorders in Labrador Retrievers and the 30 most common disorders in non-Labrador Retrievers. Multivariable logistic regression was used to report the odds of each of these disorders in 1462 (6.6%) Labrador Retrievers compared with 20,786 (93.4%) non-Labrador Retrievers. At a specific-level of diagnostic precision, after accounting for confounding, Labrador Retrievers had significantly increased odds of 12/35 (34.3%) disorders compared to non-Labrador Retrievers; osteoarthritis (OR 2.83) had the highest odds. Conversely, Labrador Retrievers had reduced odds of 7/35 (20.0%) disorders; patellar luxation (OR 0.18) had the lowest odds. This study provides useful information about breed-specific disorder predispositions and protections, which future research could evaluate further to produce definitive guidance for Labrador Retriever breeders and owners.

Surveillance of a vomiting outbreak in dogs in the UK using owner-derived and internet search data.

BACKGROUND: In early 2020, the Small Animal Veterinary Surveillance Network reported evidence of an outbreak of acute prolific vomiting in dogs in the UK. The aims of this study were to investigate whether there was evidence for a vomiting outbreak in Dogslife and Google Trends data and to describe its characteristics. METHODS: Incidence of Dogslife vomiting reports and the Google search index for 'dog vomiting' and 'puppy vomiting' between December 2019 and March 2020 was compared to the respective data from the same months in previous years. Risks for dogs vomiting and factors influencing veterinary attendance in Dogslife were identified using multivariable logistic regression. RESULTS: This study confirmed a vomiting outbreak was evident in UK dogs between December 2019 and March 2020 using data from Dogslife and Google Trends. The odds of a vomiting incident being reported to Dogslife was 1.51 (95% CI: 1.24-1.84) in comparison to previous years. Dogslife data identified differences in owner-decision making when seeking veterinary attention and identified factors associated with dogs at higher odds of experiencing a vomiting episode. CONCLUSION: Owner-derived data including questionnaires and internet search queries should be considered a valid, valuable source of information for veterinary population health surveillance.

Development of photoactivable phenanthroline-based manganese(I) CO-Releasing molecules (PhotoCORMs) active against ESKAPE bacteria and bacterial biofilms.

The synthesis and biological evaluation of a series of phenanthroline-based visible-light-activated manganese(I) carbon-monoxide-releasing molecules (PhotoCORMs) against ESKAPE bacteria and bacterial biofilms is reported. Four carbonyl compounds of general formula fac-[Mn(N∧N)(CO)3(L)] have been synthesized and characterized. Despite being thermally stable in the absence of light, these PhotoCORMs readily release CO upon blue (435-450 nm) LED light irradiation as confirmed by spectrophotometric CO releasing experiments (Mb Assay). The antibacterial activity of the four PhotoCORMs has been investigated against a panel of ESKAPE bacteria. The compounds 1-3 were found to be effective antibacterials at low concentrations against multidrug-resistant Klebsiella pneumoniae and Acinetobacter baumannii when photoactivated with blue-light. In addition, the PhotoCORMs 1-2 were found to inhibit the formation of Klebsiella pneumoniae and Acinetobacter baumannii bacterial biofilms at low concentrations (MIC = 4-8 µg/mL), turning out to be promising candidates to combat antimicrobial resistance. The antibacterial and biofilm inhibitory effect of the PhotoCORMs is plausibly due to the release of CO as well as the formation of phenanthroline photo-by-products as revealed by spectroscopy and microbiology experiments.

Is it time to stop sweeping data cleaning under the carpet? A novel algorithm for outlier management in growth data.

All data are prone to error and require data cleaning prior to analysis. An important example is longitudinal growth data, for which there are no universally agreed standard methods for identifying and removing implausible values and many existing methods have limitations that restrict their usage across different domains. A decision-making algorithm that modified or deleted growth measurements based on a combination of pre-defined cut-offs and logic rules was designed. Five data cleaning methods for growth were tested with and without the addition of the algorithm and applied to five different longitudinal growth datasets: four uncleaned canine weight or height datasets and one pre-cleaned human weight dataset with randomly simulated errors. Prior to the addition of the algorithm, data cleaning based on non-linear mixed effects models was the most effective in all datasets and had on average a minimum of 26.00% higher sensitivity and 0.12% higher specificity than other methods. Data cleaning methods using the algorithm had improved data preservation and were capable of correcting simulated errors according to the gold standard; returning a value to its original state prior to error simulation. The algorithm improved the performance of all data cleaning methods and increased the average sensitivity and specificity of the non-linear mixed effects model method by 7.68% and 0.42% respectively. Using non-linear mixed effects models combined with the algorithm to clean data allows individual growth trajectories to vary from the population by using repeated longitudinal measurements, identifies consecutive errors or those within the first data entry, avoids the requirement for a minimum number of data entries, preserves data where possible by correcting errors rather than deleting them and removes duplications intelligently. This algorithm is broadly applicable to data cleaning anthropometric data in different mammalian species and could be adapted for use in a range of other domains.