The Evolutionary and Historical Foundation of the Modern Horse: Lessons from Ancient Genomics.

The domestication of the horse some 5,500 years ago followed those of dogs, sheep, goats, cattle, and pigs by ∼2,500-10,000 years. By providing fast transportation and transforming warfare, the horse had an impact on human history with no equivalent in the animal kingdom. Even though the equine sport industry has considerable economic value today, the evolutionary history underlying the emergence of the modern domestic horse remains contentious. In the last decade, novel sequencing technologies have revolutionized our capacity to sequence the complete genome of organisms, including from archaeological remains. Applied to horses, these technologies have provided unprecedented levels of information and have considerably changed models of horse domestication. This review illustrates how ancient DNA, especially ancient genomes, has inspired researchers to rethink the process by which horses were first domesticated and then diversified into a variety of breeds showing a range of traits that are useful to humans.

TonB-Dependent Transport Across the Bacterial Outer Membrane.

TonB-dependent transporters (TBDTs) are present in all gram-negative bacteria and mediate energy-dependent uptake of molecules that are too scarce or large to be taken up efficiently by outer membrane (OM) diffusion channels. This process requires energy that is derived from the proton motive force and delivered to TBDTs by the TonB-ExbBD motor complex in the inner membrane. Together with the need to preserve the OM permeability barrier, this has led to an extremely complex and fascinating transport mechanism for which the fundamentals, despite decades of research, are still unclear. In this review, we describe our current understanding of the transport mechanism of TBDTs, their potential role in the delivery of novel antibiotics, and the important contributions made by TBDT-associated (lipo)proteins.

Equine herpesvirus type 1 (EHV-1) replication at the upper respiratory entry site is inhibited by neutralizing EHV-1-specific IgG1 and IgG4/7 mucosal antibodies.

Equine herpesvirus type 1 (EHV-1) is a contagious respiratory pathogen that infects the mucosa of the upper respiratory tract (URT). Mucosal immune responses at the URT provide the first line of defense against EHV-1 and are crucial for orchestrating immunity. To define host-pathogen interactions, we characterized B-cell responses, antibody isotype functions, and EHV-1 replication of susceptible (non-immune) and clinically protected (immune) horses after experimental EHV-1 infection. Nasal secretion and nasal wash samples were collected and used for the isolation of DNA, RNA, and mucosal antibodies. Shedding of infectious virus, EHV-1 copy numbers, viral RNA expression, and host B-cell activation in the URT were compared based on host immune status. Mucosal EHV-1-specific antibody responses were associated with EHV-1 shedding and viral RNA transcription. Finally, mucosal immunoglobulin G (IgG) and IgA isotypes were purified and tested for neutralizing capabilities. IgG1 and IgG4/7 neutralized EHV-1, while IgG3/5, IgG6, and IgA did not. Immune horses secreted high amounts of mucosal EHV-1-specific IgG4/7 antibodies and quickly upregulated B-cell pathway genes, while EHV-1 was undetected by virus isolation and PCR. RNA transcription analysis reinforced incomplete viral replication in immune horses. In contrast, complete viral replication with high viral copy numbers and shedding of infectious viruses was characteristic for non-immune horses, together with low or absent EHV-1-specific neutralizing antibodies during viral replication. These data confirm that pre-existing mucosal IgG1 and IgG4/7 and rapid B-cell activation upon EHV-1 infection are essential for virus neutralization, regulation of viral replication, and mucosal immunity against EHV-1.IMPORTANCEEquine herpesvirus type 1 (EHV-1) causes respiratory disease, abortion storms, and neurologic outbreaks known as equine herpes myeloencephalopathy (EHM). EHV-1 is transmitted with respiratory secretions by nose-to-nose contact or via fomites. The virus initially infects the epithelium of the upper respiratory tract (URT). Host-pathogen interactions and mucosal immunity at the viral entry site provide the first line of defense against the EHV-1. Robust mucosal immunity can be essential in protecting against EHV-1 and to reduce EHM outbreaks. It has previously been shown that immune horses do not establish cell-associated viremia, the prerequisite for EHM. Here, we demonstrate how mucosal antibodies can prevent the replication of EHV-1 at the epithelium of the URT and, thereby, the progression of the virus to the peripheral blood. The findings improve the mechanistic understanding of mucosal immunity against EHV-1 and can support the development of enhanced diagnostic tools, vaccines against EHM, and the management of EHV-1 outbreaks.

Cefiderocol: Clinical application and emergence of resistance.

Antibacterial drug resistance of gram-negative bacteria (GNB) results in high morbidity and mortality of GNB infection, seriously threaten human health globally. Developing new antibiotics has become the critical need for dealing with drug-resistant bacterial infections. Cefiderocol is an iron carrier cephalosporin that achieves drug accumulation through a unique "Trojan horse" strategy into the bacterial periplasm. It shows high antibacterial activity against multidrug-resistant (MDR) Enterobacteriaceae and MDR non-fermentative bacteria. The application of cefiderocol offers new hope for treating clinical drug-resistant bacterial infections. However, limited clinical data and uncertainties about its resistance mechanisms constrain the choice of its therapeutic use. This review aimed to summarize the clinical applications, drug resistance mechanisms, and co-administration of cefiderocol.

Single-cell profiling of bronchoalveolar cells reveals a Th17 signature in neutrophilic severe equine asthma.

Severe equine asthma (SEA) is a complex respiratory condition characterized by chronic airway inflammation. It shares many clinical and pathological features with human neutrophilic asthma, making it a valuable model for studying this condition. However, the immune mechanisms driving SEA have remained elusive. Although SEA has been primarily associated with a Th2 response, there have also been reports of Th1, Th17, or mixed-mediated responses. To uncover the elusive immune mechanisms driving SEA, we performed single-cell mRNA sequencing (scRNA-seq) on cryopreserved bronchoalveolar cells from 11 Warmblood horses, 5 controls and 6 with SEA. We identified six major cell types, including B cells, T cells, monocytes-macrophages, dendritic cells, neutrophils, and mast cells. All cell types exhibited significant heterogeneity, with previously identified and novel cell subtypes. Notably, we observed monocyte-lymphocyte complexes and detected a robust Th17 signature in SEA, with CXCL13 upregulation in intermediate monocytes. Asthmatic horses exhibited expansion of the B-cell population, Th17 polarization of the T-cell populations, and dysregulation of genes associated with T-cell function. Neutrophils demonstrated enhanced migratory capacity and heightened aptitude for neutrophil extracellular trap formation. These findings provide compelling evidence for a predominant Th17 immune response in neutrophilic SEA, driven by dysregulation of monocyte and T-cell genes. The dysregulated genes identified through scRNA-seq have potential as biomarkers and therapeutic targets for SEA and provide insights into human neutrophilic asthma.

Induction of in vivo-like ciliation in confluent monolayers of re-differentiated equine oviduct epithelial cells.

We recently developed re-differentiated equine oviduct epithelial cell (REOEC) monolayers demonstrating various in vivo morphological characteristics, but lacking secondary ciliation. In this study, we evaluated the effects of fetal bovine serum, reproductive steroid hormones, Wnt- and Notch ligands and inhibitors, and different EOEC seeding densities, in both conventional wells and on microporous membranes, on EOEC morphology and, in particular, secondary ciliation. REOEC monolayers were assessed by confocal microscopy after combined staining of nuclei, cilia and the cytoskeleton. Only Wnt ligands, Notch inhibitors and oviduct explant cell concentration affected EOEC morphology. Undesirable epithelial-mesenchymal transition was observed in REOEC monolayers exposed to Wnt3a containing medium and Wnt ligand CHIR 99021. With respect to secondary ciliation, only the combined effect of oviduct explant cell concentration and Notch inhibition steered REOEC monolayers to in vivo-like ciliation patterns. De-differentiated EOECs, formed 10 days after oviduct explant cell seeding, were reseeded on inserts; only at initial oviduct explant cell concentrations of 1 and 5 x106 cells per well was the formation of REOEC monolayers with a high rate of diffuse ciliation supported. Within 1 month after air-liquid interface introduction, >40% and > 20% of the REOECs showed secondary cilia, respectively. At higher oviduct explant cell seeding densities secondary ciliation was not supported after re-differentiation. Additionally, Notch inhibition helped boost secondary ciliation rates to >60% in REOEC monolayers with diffuse ciliation only. These monolayers demonstrated higher clathrin expression under follicular phase conditions. Overall, the ciliated REOEC monolayers better resemble in vivo oviduct epithelial cells than previous models.

In vitro and in silico studies of enterobactin-inspired Ciprofloxacin and Fosfomycin first generation conjugates on the antibiotic resistant E. coli OQ866153.

BACKGROUND: The emergence of antimicrobial resistance in bacterial pathogens is a growing concern worldwide due to its impact on the treatment of bacterial infections. The "Trojan Horse" strategy has been proposed as a potential solution to overcome drug resistance caused by permeability issues. OBJECTIVE: The objective of our research was to investigate the bactericidal activity and mechanism of action of the "Trojan Horse" strategy using enterobactin conjugated with Ciprofloxacin and Fosfomycin against the antibiotic-resistant Escherichia coli strain OQ866153. METHODOLOGY: Enterobactin, a mixed ligand of E. coli OQ866153, was conjugated with Ciprofloxacin and Fosfomycin individually to aid active absorption via specific enterobactin binding proteins (FepABCDG). The effectiveness of the conjugates was assessed by measuring their bactericidal activity against E. coli OQ866153, as well as their ability to inhibit DNA gyrase enzyme and biofilm formation. RESULTS: The Fe+3-enterobactin-Ciprofloxacin conjugate effectively inhibited the DNA gyrase enzyme (Docking score = -8.597 kcal/mol) and resulted in a lower concentration (25 µg/ml) required to eliminate supercoiled DNA plasmids compared to the parent drug (35 µg/ml; Docking score = -6.264 kcal/mol). The Fe+3-Enterobactin-Fosfomycin conjugate showed a higher inhibition percentage (100%) of biofilm formation compared to Fosfomycin (21.58%) at a concentration of 2 mg/ml, with docking scores of -5.481 and -3.756 kcal/mol against UDP-N acetylglucosamine 1-carboxyvinyltransferase MurA. CONCLUSION: The findings of this study suggest that the "Trojan Horse" strategy using enterobactin conjugated with Ciprofloxacin and Fosfomycin can effectively overcome permeability issues caused by efflux proteins and enhance the bactericidal activity of these drugs against antibiotic-resistant strains of E. coli.

Microbiome analysis reveals the differences in gut fungal community between Dutch Warmblood and Mongolian horses.

Similar to gut bacterial community, gut fungal community are also an important part of the gut microbiota and play crucial roles in host immune regulation and metabolism. However, most studies have focused on the gut bacterial community, and research on the gut fungal community has been limited. Dutch Warmblood (DWH) and Mongolian horses (MGH) are important equine breeds, but little research has been done on their gut fungal community. Here, we assessed differences in gut fungal community between two horse species. Results showed that a total of 2159 OTUs were found in the Dutch Warmblood and Mongolian horses, of which 308 were common. Between-group analyzes of microbial diversity showed no differences in the alpha and beta diversity of gut fungal community between the two horse species. Microbiological taxonomic surveys showed that the dominant fungal phyla (Neocallimastigomycota and Ascomycota) and genera (unclassified_Neocallimastigaceae and Anaeromyces) were the same without being affected by species. Although the types of dominant fungal phyla did not change, the abundances of some fungal genera changed significantly. Results of Metastats analysis showed that there were a total of 206 fungal genera that were significantly different between the two horses, among which 78 genera showed an increase and 127 genera significantly decreased in Dutch Warmblood horses compared with Mongolian horses. In conclusion, this study investigated the composition and structure of the gut fungal community of Dutch Warmblood and Mongolian horses and found significant differences in gut fungal community between both breeds. Notably, this is the first exploration of the differences in the gut fungal community of both breeds, which may help to understand the distribution characteristics of the gut fungal community of different breeds of horses and reveal the differences in the traits of different horses.

Non-Synonymous Substitutions in Cadherin 13, Solute Carrier Family 6 Member 4, and Monoamine Oxidase A Genes are Associated with Personality Traits in Thoroughbred Horses.

Retraining retired racehorses for various purposes can help correct behavioral issues. However, ensuring efficiency and preventing accidents present global challenges. Based on the hypothesis that a simple personality assessment could help address these challenges, the present study aimed to identify genetic markers associated with personality. Eight genes were selected from 18 personality-related candidate genes that are orthologs of human personality genes, and their association with personality was verified based on actual behavior. A total of 169 Thoroughbred horses were assessed for their tractability (questionnaire concerning tractability in 14 types of situations and 3 types of impressions) during the training process. Personality factors were extracted from the data using principal component analysis and analyzed for their association with single nucleotide variants as non-synonymous substitutions in the target genes. Three genes, CDH13, SLC6A4, and MAOA, demonstrated significant associations based on simple linear regression, marking the identification of these genes for the first time as contributors to temperament in Thoroughbred horses. All these genes, as well as the previously identified HTR1A, are involved in the serotonin neurotransmitter system, suggesting that the tractability of horses may be correlated with their social personality. Assessing the genotypes of these genes before retraining is expected to prevent problems in the development of a racehorse's second career and shorten the training period through individual customization of training methods, thereby improving racehorse welfare.

Glycoconjugate-Specific Developmental Changes in the Horse Vomeronasal Organ.

The vomeronasal organ (VNO) is a tubular pheromone-sensing organ in which the lumen is covered with sensory and non-sensory epithelia. This study used immunohistochemistry and lectin histochemistry techniques to evaluate developmental changes, specifically of the glycoconjugate profile, in the horse VNO epithelium. Immunostaining analysis revealed PGP9.5 expression in some vomeronasal non-sensory epithelium (VNSE) cells and in the vomeronasal receptor cells of the vomeronasal sensory epithelium (VSE) in fetuses, young foals, and adult horses. Olfactory marker protein expression was exclusively localized in receptor cells of the VSE in fetuses, young foals, and adult horses and absent in VNSE. To identify the glycoconjugate type, lectin histochemistry was performed using 21 lectins. Semi-quantitative analysis revealed that the intensities of glycoconjugates labeled with WGA, DSL, LEL, and RCA120 were significantly higher in adult horse VSE than those in foal VSE, whereas the intensities of glycoconjugates labeled with LCA and PSA were significantly lower in adult horse VSE. The intensities of glycoconjugates labeled with s-WGA, WGA, BSL-II, DSL, LEL, STL, ConA, LCA, PSA, DBA, SBA, SJA, RCA120, jacalin, and ECL were significantly higher in adult horse VNSE than those in foal VNSE, whereas the intensity of glycoconjugates labeled with UEA-I was lower in adult horse VNSE. Histochemical analysis of each lectin revealed that various glycoconjugates in the VSE were present in the receptor, supporting, and basal cells of foals and adult horses. A similar pattern of lectin histochemistry was also observed in the VNSE of foals and adult horses. In conclusion, these results suggest that there is an increase in the level of N-acetylglucosamine (labeled by WGA, DSL, LEL) and galactose (labeled by RCA120) in horse VSE during postnatal development, implying that they may influence the function of VNO in adult horses.

Advancing diagnostic precision: Unveiling sensitization relationships between cat, dog, and horse allergen molecules.

BACKGROUND: Recent advancements in molecular diagnostics have unveiled a multitude of allergen molecules (AMs) associated with animal sensitizations, revealing significant cross- and co-sensitization patterns among these seemingly distinct allergens. METHOD: We investigated the sensitization profiles of 120 children, sensitized to at least one of the 14 AMs from cat, dog, or horse using the Alex test, employing correlations and hierarchical clusters to explore relationship between sensitizations. RESULTS: Sensitizations to Fel d 1, Can f 4/5, and Equ c 4 differ from other cat, dog, and horse AM sensitizations, suggesting they may represent genuine sensitizations for their respective animals. High correlations were observed among various AMs, including lipocalins (Can f 1/2/6, Fel d 4/7, and Equ c 1), serum albumins (Fel d 2, Can f 3, and Equ c 3), and uteroglobins (Fel d 1 and Can f_Fd1). Hierarchical clustering of sensitizations identified two similarity clusters and one dissimilarity cluster, providing an estimation of the likelihood of cross-reactivity. Additionally, our method facilitated speculation regarding cross-, co-, or genuine sensitization. Moreover, we noted a potential increase in the number and level of sensitized animal AMs concurrent with increased sensitization to other aeroallergens with advancing age. No significant difference was detected for the presence or absence of various types of allergic comorbidities. CONCLUSION: Correlations and hierarchical clustering can unveil the extent and magnitude of cross-, co-, and genuine sensitization relationships among animal AMs. These insights can be leveraged to enhance artificial intelligence algorithms, improving diagnostic accuracy through the integration of other measures of sensitization.

Peptostreptococcus equinus sp. nov., isolated from horse faeces.

A Gram-stain-positive, non-spore-forming, and obligate anaerobic bacteria designated strain CBA3647T was isolated from a horse faecal sample in Jeju, Republic of Korea. Phylogenetic analysis based on 16S rRNA gene sequences showed that strain CBA3647T formed a distinct phyletic lineage from closely related species within the genus Peptostreptococcus. Based on comparative analysis of 16S rRNA gene sequences, Peptostreptococcus anaerobius ATCC 27337T is most closely related to strain CBA3647T with a 16S rRNA gene similarity of 98.31 %, while similarity to other type strains is below 98.0 %. The genomic DNA G+C content of strain CBA3647T was 30.0 mol%. The digital DNA-DNA hybridization values between strain CBA3647T and the six Peptostreptococcus species were equal to or less than 24 %. Cells were non-motile and oval-shaped cocci with catalase-positive and oxidase-negative activities. Growth occurred at 20-40 °C (optimum, 35 °C), pH 6-8 (optimum, pH 7), and in the presence of 0-2 % (w/v) NaCl (optimum, 1 %). Strain CBA3647T contained C14 : 0 iso and C16 : 0 as major fatty acids. Phenotypic, chemotaxonomic, and molecular properties of strain CBA3647T suggest that it represents a novel species in the genus Peptostreptococcus, which has been named Peptostreptococcus equinus sp. nov. The type strain is CBA3647T (=KACC 22891T= JCM 35846T).

A matter of differentiation: equine enteroids as a model for the in vivo intestinal epithelium.

Epithelial damage due to gastrointestinal disorders frequently causes severe disease in horses. To study the underlying pathophysiological processes, we aimed to establish equine jejunum and colon enteroids (eqJE, eqCE) mimicking the in vivo epithelium. Therefore, enteroids were cultivated in four different media for differentiation and subsequently characterized histomorphologically, on mRNA and on protein level in comparison to the native epithelium of the same donor horses to identify ideal culture conditions for an in vitro model system. With increasing enterocyte differentiation, the enteroids showed a reduced growth rate as well as a predominantly spherical morphology and less budding compared to enteroids in proliferation medium. Combined or individual withdrawal of stem cell niche pathway components resulted in lower mRNA expression levels of stem cell markers and concomitant differentiation of enterocytes, goblet cells and enteroendocrine cells. For eqCE, withdrawal of Wnt alone was sufficient for the generation of differentiated enterocytes with a close resemblance to the in vivo epithelium. Combined removal of Wnt, R-spondin and Noggin and the addition of DAPT stimulated differentiation of eqJE at a similar level as the in vivo epithelium, particularly with regard to enterocytes. In summary, we successfully defined a medium composition that promotes the formation of eqJE and eqCE consisting of multiple cell types and resembling the in vivo epithelium. Our findings emphasize the importance of adapting culture conditions to the respective species and the intestinal segment. This in vitro model will be used to investigate the pathological mechanisms underlying equine gastrointestinal disorders in future studies.

Genomic characterization of equine influenza A subtype H3N8 viruses by long read sequencing and functional analyses of the PB1-F2 virulence factor of A/equine/Paris/1/2018.

Equine influenza virus (EIV) remains a threat to horses, despite the availability of vaccines. Strategies to monitor the virus and prevent potential vaccine failure revolve around serological assays, RT-qPCR amplification, and sequencing the viral hemagglutinin (HA) and neuraminidase (NA) genes. These approaches overlook the contribution of other viral proteins in driving virulence. This study assesses the potential of long-read nanopore sequencing for fast and precise sequencing of circulating equine influenza viruses. Therefore, two French Florida Clade 1 strains, including the one circulating in winter 2018-2019 exhibiting more pronounced pathogenicity than usual, as well as the two currently OIE-recommended vaccine strains, were sequenced. Our results demonstrated the reliability of this sequencing method in generating accurate sequences. Sequence analysis of HA revealed a subtle antigenic drift in the French EIV strains, with specific substitutions, such as T163I in A/equine/Paris/1/2018 and the N188T mutation in post-2015 strains; both substitutions were in antigenic site B. Antigenic site E exhibited modifications in post-2018 strains, with the N63D substitution. Segment 2 sequencing also revealed that the A/equine/Paris/1/2018 strain encodes a longer variant of the PB1-F2 protein when compared to other Florida clade 1 strains (90 amino acids long versus 81 amino acids long). Further biological and biochemistry assays demonstrated that this PB1-F2 variant has enhanced abilities to abolish the mitochondrial membrane potential ΔΨm and permeabilize synthetic membranes. Altogether, our results highlight the interest in rapidly characterizing the complete genome of circulating strains with next-generation sequencing technologies to adapt vaccines and identify specific virulence markers of EIV.

Corelease of Genotoxic Polycyclic Aromatic Hydrocarbons and Nanoparticles from a Commercial Aircraft Jet Engine - Dependence on Fuel and Thrust.

Jet engines are important contributors to global CO2 emissions and release enormous numbers of ultrafine particles into different layers of the atmosphere. As a result, aviation emissions are affecting atmospheric chemistry and promote contrail and cloud formation with impacts on earth's radiative balance and climate. Furthermore, the corelease of nanoparticles together with carcinogenic polycyclic aromatic hydrocarbons (PAHs) affects air quality at airports. We studied exhausts of a widely used turbofan engine (CFM56-7B26) operated at five static thrust levels (idle, 7, 30, 65, and 85%) with conventional Jet A-1 fuel and a biofuel blend composed of hydro-processed esters and fatty acids (HEFA). The particles released, the chemical composition of condensable material, and the genotoxic potential of these exhausts were studied. At ground operation, particle number emissions of 3.5 and 0.5 × 1014 particles/kg fuel were observed with highest genotoxic potentials of 41300 and 8800 ng toxicity equivalents (TEQ)/kg fuel at idle and 7% thrust, respectively. Blending jet fuel with HEFA lowered PAH and particle emissions by 7-34% and 65-67% at idle and 7% thrust, respectively, indicating that the use of paraffin-rich biofuels is an effective measure to reduce the exposure of airport personnel to nanoparticles coated with genotoxic PAHs (Trojan horse effect).

US-wide equine strongylid egg count data demonstrate seasonal and regional trends.

Equine strongylid parasites are ubiquitous around the world and are main targets of parasite control programmes. In recent years, automated fecal egg counting systems based on image analysis have become available allowing for collection and analysis of large-scale egg count data. This study aimed to evaluate equine strongylid fecal egg count (FEC) data generated with an automated system over three years in the US with specific attention to seasonal and regional trends in egg count magnitude and sampling activity. Five US regions were defined; North East, South East, North Central, South Central and West. The data set included state, region and zip code for each FEC. The number of FECs falling in each of the following categories were recorded: (1) 0 eggs per gram (EPG), (2) 1 ⩽ 200 EPG, (3) 201 ⩽ 500 EPG and (4) >500 EPG. The data included 58 329 FECs. A fixed effects model was constructed fitting the number of samples analysed per month, year and region, and a mixed effects model was constructed to fit the number of FECs falling in each of the 4 egg count categories defined above. The overall proportion of horses responsible for 80% of the total FEC output was 18.1%, and this was consistent across years, months and all regions except West, where the proportion was closer to 12%. Statistical analyses showed significant seasonal trends and regional differences of sampling frequency and FEC category. The data demonstrated that veterinarians tended to follow a biphasic pattern when monitoring strongylid FECs in horses, regardless of location.

Smart plant disease net: Adaptive Dense Hybrid Convolution network with attention mechanism for IoT-based plant disease detection by improved optimization approach.

Plant diseases are rising nowadays. Plant diseases lead to high economic losses. Internet of Things (IoT) technology has found its application in various sectors. This led to the introduction of smart farming, in which IoT has been utilized to help identify the exact spot of the diseased affected region on the leaf from the vast farmland in a well-organized and automated manner. Thus, the main focus of this task is the introduction of a novel plant disease detection model that relies on IoT technology. The collected images are given to the Image Transmission phase. Here, the encryption task is performed by employing the Advanced Encryption Standard (AES) and also the decrypted plant images are fed to the pre-processing stage. The Mask Regions with Convolutional Neural Networks (R-CNN) are used to segment the pre-processed images. Then, the segmented images are given to the detection phase in which the Adaptive Dense Hybrid Convolution Network with Attention Mechanism (ADHCN-AM) approach is utilized to perform the detection of plant disease. From the ADHCN-AM, the final detected plant disease outcomes are obtained. Throughout the entire validation, the offered model shows 95% enhancement in terms of MCC showcasing its effectiveness over the existing approaches.

Putting the cart before the horse: mixed-methods participatory investigation of working equid harnessing practices in three selected towns of the Oromia national regional state in Ethiopia.

BACKGROUND: Millions of working equids provide socio-economic support for many low-income communities worldwide. With the prevalence of harness-related wounds reported as higher than 60%, this study aims to describe the equipment used by working equids in three locations of the Ethiopian national regional state of Oromia (Fiche, Bishoftu and Shashamene), and the attitudes and practices of equid owners, users and harness makers regarding work equipment. This mixed-methods study consists of cross-sectional surveying of working equids used for taxi or transport of goods or water, as well as cart-driver questionnaires and focus groups (FG) with working equid stakeholders. Activities conducted with FG included participatory ranking of equipment attributes and equipment drawing exercises. Indicators of equipment design and assembly, as well as cart-driver attitudes and practices were described quantitatively. Associations between equipment characteristics and species, work-type and cart-driver indicators were investigated through univariable logistic regression models, whereas focus group discussions were transcribed and analysed thematically. RESULTS: In total, 368 working equid surveys and cart-driver questionnaires were completed and 87 participants took part in nine FG. Equipment composition and characteristics differed considerably from ideal animal draught and harnessing principles described in the literature, with none of the observed harnesses adhering to all principles and thus not considered fully adequate. Various harness compositions were used, with only saddles and breast collars present in all. Donkey equipment had fewer components than that of horses, such as swingle trees (OR 0.02; 95% CI 0.01-0.06; p < 0.001) or girths (OR 0.09; 95%CI 0.02-0.4; p = 0.002). Horse equipment was more likely to have functional elements such as breeching (OR 7.8; 95% CI 2.9-20.9; p < 0.001). Of all equipment attributes, FG participants ranked cost, design and impact on animal wellbeing as having the highest importance. Thematic analysis identified motivations and priorities regarding equipment; awareness and knowledge of design and function; barriers to using ideal equipment; and consequences of equipment design and practices as key themes. CONCLUSIONS: Various weaknesses of design, assembly and use of equid work equipment were identified. Promoting behavioural change through awareness and training could lead to a sustainable improvement of work equipment quality, access, and prevention of equipment-related problems.

Assessment of fecal bacterial viability and diversity in fresh and frozen fecal microbiota transplant (FMT) product in horses.

BACKGROUND: Currently, lack of standardization for fecal microbiota transplantation (FMT) in equine practice has resulted in highly variable techniques, and there is no data on the bacterial metabolic activity or viability of the administered product. The objectives of this study were to compare the total and potentially metabolically active bacterial populations in equine FMT, and assess the effect of different frozen storage times, buffers, and temperatures on an equine FMT product. Fresh feces collected from three healthy adult horses was subjected to different storage methods. This included different preservation solutions (saline plus glycerol or saline only), temperature (-20 °C or -80 °C), and time (fresh, 30, 60, or 90 days). Samples underwent DNA extraction to assess total bacterial populations (both live and dead combined) and RNA extraction followed by reverse transcription to cDNA as a proxy to assess viable bacteria, then 16s rRNA gene amplicon sequencing using the V1-V2 region. RESULTS: The largest difference in population indices and taxonomic composition at the genus level was seen when evaluating the results of DNA-based (total) and cDNA-based (potentially metabolically active) extraction method. At the community level, alpha diversity (observed species, Shannon diversity) was significantly decreased in frozen samples for DNA-based analysis (P < 0.05), with less difference seen for cDNA-based sequencing. Using DNA-based analysis, length of storage had a significant impact (P < 0.05) on the bacterial community profiles. For potentially metabolically active populations, storage overall had less of an effect on the bacterial community composition, with a significant effect of buffer (P < 0.05). Individual horse had the most significant effect within both DNA and cDNA bacterial communities. CONCLUSIONS: Frozen storage of equine FMT material can preserve potentially metabolically active bacteria of the equine fecal microbiome, with saline plus glycerol preservation more effective than saline alone. Larger studies are needed to determine if these findings apply to other individual horses. The ability to freeze FMT material for use in equine patients could allow for easier clinical use of fecal transplant in horses with disturbances in their intestinal microbiome.

Preovulatory follicular fluid secretome added to in vitro maturation medium influences the metabolism of equine cumulus-oocyte complexes.

BACKGROUND: In vitro embryo production is a highly demanded reproductive technology in horses, which requires the recovery (in vivo or post-mortem) and in vitro maturation (IVM) of oocytes. Oocytes subjected to IVM exhibit poor developmental competence compared to their in vivo counterparts, being this related to a suboptimal composition of commercial maturation media. The objective of this work was to study the effect of different concentrations of secretome obtained from equine preovulatory follicular fluid (FF) on cumulus-oocyte complexes (COCs) during IVM. COCs retrieved in vivo by ovum pick up (OPU) or post-mortem from a slaughterhouse (SLA) were subjected to IVM in the presence or absence of secretome (Control: 0 µg/ml, S20: 20 µg/ml or S40: 40 µg/ml). After IVM, the metabolome of the medium used for oocyte maturation prior (Pre-IVM) and after IVM (Post-IVM), COCs mRNA expression, and oocyte meiotic competence were analysed. RESULTS: IVM leads to lactic acid production and an acetic acid consumption in COCs obtained from OPU and SLA. However, glucose consumption after IVM was higher in COCs from OPU when S40 was added (Control Pre-IVM vs. S40 Post-IVM: 117.24 ± 7.72 vs. 82.69 ± 4.24; Mean µM ± SEM; p < 0.05), while this was not observed in COCs from SLA. Likewise, secretome enhanced uptake of threonine (Control Pre-IVM vs. S20 Post-IVM vs. S40 Post-IVM: 4.93 ± 0.33 vs. 3.04 ± 0.25 vs. 2.84 ± 0.27; Mean µM ± SEM; p < 0.05) in COCs recovered by OPU. Regarding the relative mRNA expression of candidate genes related to metabolism, Lactate dehydrogenase A (LDHA) expression was significantly downregulated when secretome was added during IVM at 20-40 µg/ml in OPU-derived COCs (Control vs. S20 vs. S40: 1.77 ± 0.14 vs. 1 ± 0.25 vs. 1.23 ± 0.14; fold change ± SEM; p < 0.05), but not in SLA COCs. CONCLUSIONS: The addition of secretome during in vitro maturation (IVM) affects the gene expression of LDHA, glucose metabolism, and amino acid turnover in equine cumulus-oocyte complexes (COCs), with diverging outcomes observed between COCs retrieved using ovum pick up (OPU) and slaughterhouse-derived COCs (SLA).