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Abstract Background Obesity is a risk factor for diabetes mellitus, which commonly coexists with pancreatitis in cats. However, obesity has not previously been associated with pancreatitis in cats. Objectives To evaluate factors affecting serum concentrations of pancreatic lipase immunoreactivity (fPLI), trypsin-like immunoreactivity (fTLI), cobalamin and folate in clinically healthy lean, overweight and obese, or diabetic cats. Animals Seventy-nine client-owned cats (27 healthy lean [LN, BCS 4-5/9], 30 healthy overweight and obese [OW, BCS 7-9/9], and 22 diabetic [DM]) were included. Methods Cross-sectional study. The cats underwent physical examination, and blood tests. Linear regression models compared differences in fPLI, fTLI, cobalamin, and folate concentrations. Fisher's exact test assessed the proportions of cats with fPLI and fTLI indicative of pancreatitis, and hypocobalaminemia. A random forest algorithm identified explanatory variables for cats having fPLI levels indicative of pancreatitis. Results No LN cats, while 6/30 (20%) of OW and 10/22 (45%) of DM cats had fPLI concentrations indicative of pancreatitis. Body condition score (P = .02) and body weight (P = .002) were positively associated with fPLI levels in LN and OW cats. Higher fPLI, and lower cobalamin concentrations were associated with higher age across groups. Conclusions and Clinical Importance Body condition score and body weight were associated with higher fPLI levels in nondiabetic cats. A larger proportion of OW and DM cats had fPLI concentrations indicative of pancreatitis compared to LN cats. Whether this indicates subclinical pancreatitis remains to be determined. Hypocobalaminemia was less frequent in OW compared to DM cats.

Abstract Background Concurrent exocrine pancreatic dysfunction and decreased pancreatic organ size are common findings in various stages of human type 1 diabetes mellitus (DM). Exocrine pancreatic insufficiency (EPI) is incompletely described in diabetic dogs. Objective To compare canine trypsin-like immunoreactivity (cTLI) of diabetic dogs with that of healthy controls. A secondary aim was to evaluate the correlation between duration of DM and cTLI. Animals Thirty client-owned diabetic dogs and thirty client-owned control dogs. Methods Cross-sectional study. Diabetic and healthy control dogs were included if they had no clinical evidence of pancreatitis and if serum samples obtained after food was withheld were available. Serum cTLI was measured at a reference laboratory and compared between groups. Canine pancreatic lipase immunoreactivity (cPLI) was analyzed concurrently as an indicator of pancreatitis. Results The median cTLI concentration in all diabetic dogs (36.4 μg/L [range, 7.0-288 μg/L]) did not differ from control dogs (28.7 μg/L [range, 12.8-58.6 μg/L]) (P = .07; difference −7.8 μg/L [95% Confidence Interval (CI), −23.5 to 0.6 μg/L]). There was still no difference in cTLI between groups after exclusion of dogs with cPLI consistent with pancreatitis (n = 8 diabetic dogs). There was no correlation between cTLI and DM duration in all diabetic dogs (r = −0.07, [95% CI, −0.43 to 0.3], P = .7). Conclusions and Clinical Importance There was no evidence of EPI as evaluated using cTLI in this cohort of diabetic dogs, but concurrent increases in cPLI suggest cTLI might not be the optimal indicator of exocrine pancreatic dysfunction in dogs with DM.

The fungal microbiota (mycobiota) is an integral part of the complex multikingdom microbial community colonizing the mammalian gastrointestinal tract and has an important role in immune regulation 1 – 6 . Although aberrant changes in the mycobiota have been linked to several diseases, including inflammatory bowel disease 3 – 9 , it is currently unknown whether fungal species captured by deep sequencing represent living organisms and whether specific fungi have functional consequences for disease development in affected individuals. Here we developed a translational platform for the functional analysis of the mycobiome at the fungal-strain- and patient-specific level. Combining high-resolution mycobiota sequencing, fungal culturomics and genomics, a CRISPR–Cas9-based fungal strain editing system, in vitro functional immunoreactivity assays and in vivo models, this platform enables the examination of host–fungal crosstalk in the human gut. We discovered a rich genetic diversity of opportunistic Candida albicans strains that dominate the colonic mucosa of patients with inflammatory bowel disease. Among these human-gut-derived isolates, strains with high immune-cell-damaging capacity (HD strains) reflect the disease features of individual patients with ulcerative colitis and aggravated intestinal inflammation in vivo through IL-1β-dependent mechanisms. Niche-specific inflammatory immunity and interleukin-17A-producing T helper cell (T H 17 cell) antifungal responses by HD strains in the gut were dependent on the C. albicans -secreted peptide toxin candidalysin during the transition from a benign commensal to a pathobiont state. These findings reveal the strain-specific nature of host–fungal interactions in the human gut and highlight new diagnostic and therapeutic targets for diseases of inflammatory origin. Genetically diverse Candida albicans strains in patients with inflammatory bowel disease secrete a toxin and aggravate IL-1β-dependent intestinal inflammation.

Maternal deprivation (MD) induces behavioral changes and impacts brain circuits that could be associated with the pathophysiology of depression. This study investigated the markers of microglia and astrocyte activation as well as indoleamine 2,3-dioxygenase (IDO) expression in developmental programming after early life MD (on postnatal days (PNDs) 20, 30, 40, and 60). On PND 60, the rats that were subjected to MD displayed depressive-like behavior. On PND 10, it was found that there was a decrease in the level of glial fibrillary acidic protein (GFAP) immunopositive cells, a decrease in the level of IDO expression, and an increase in the level of Iba-1 (microglial marker) in the hippocampus of rats that were subjected to MD. On PND 20, levels of GFAP were also found to have decreased in the hippocampus, and there was an increase in the level of Iba-1 in the hippocampus. AIF-1 (microglial marker) expression was observed in the PFC following MD. On PND 30, the levels of Iba-1 remained elevated. On PND 40, the levels of GFAP were found to have increased in the hippocampus of rats that were subjected to MD. On PND 60, the levels of GFAP and AIF-1 remained elevated following MD. These results suggest that early life stress induces negative developmental programming in rats, as demonstrated by depressive-like behavior in adult life. Moreover, MD increases microglial activation in both early and late developmental phases. The levels of GFAP and IDO decreased in the early stages but were found to be higher in later developmental periods. These findings suggest that MD could differentially affect the expression of the IDO enzyme, astrocytes, and microglial activation depending on the neurodevelopmental period. The onset of an inflammatory state from resident brain cells could be associated with the activation of the kynurenine pathway and the development of depressive behavior in adulthood.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has infected 78 million individuals and is responsible for over 1.7 million deaths to date. Infection is associated with the development of variable levels of antibodies with neutralizing activity, which can protect against infection in animal models 1 , 2 . Antibody levels decrease with time, but, to our knowledge, the nature and quality of the memory B cells that would be required to produce antibodies upon reinfection has not been examined. Here we report on the humoral memory response in a cohort of 87 individuals assessed at 1.3 and 6.2 months after infection with SARS-CoV-2. We find that titres of IgM and IgG antibodies against the receptor-binding domain (RBD) of the spike protein of SARS-CoV-2 decrease significantly over this time period, with IgA being less affected. Concurrently, neutralizing activity in plasma decreases by fivefold in pseudotype virus assays. By contrast, the number of RBD-specific memory B cells remains unchanged at 6.2 months after infection. Memory B cells display clonal turnover after 6.2 months, and the antibodies that they express have greater somatic hypermutation, resistance to RBD mutations and increased potency, indicative of continued evolution of the humoral response. Immunofluorescence and PCR analyses of intestinal biopsies obtained from asymptomatic individuals at 4 months after the onset of coronavirus disease 2019 (COVID-19) revealed the persistence of SARS-CoV-2 nucleic acids and immunoreactivity in the small bowel of 7 out of 14 individuals. We conclude that the memory B cell response to SARS-CoV-2 evolves between 1.3 and 6.2 months after infection in a manner that is consistent with antigen persistence. In a cohort of 87 individuals with COVID-19, the memory B cell response at 6.2 months after the onset of disease evolves in a manner that is consistent with the persistence of SARS-CoV-2 antigen.

The Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) antibody neutralization response and its evasion by emerging viral variants and variant of concern (VOC) are unknown, but critical to understand reinfection risk and breakthrough infection following vaccination. Antibody immunoreactivity against SARS-CoV-2 antigens and Spike variants, inhibition of Spike-driven virus–cell fusion, and infectious SARS-CoV-2 neutralization were characterized in 807 serial samples from 233 reverse transcription polymerase chain reaction (RT-PCR)–confirmed Coronavirus Disease 2019 (COVID-19) individuals with detailed demographics and followed up to 7 months. A broad and sustained polyantigenic immunoreactivity against SARS-CoV-2 Spike, Membrane, and Nucleocapsid proteins, along with high viral neutralization, was associated with COVID-19 severity. A subgroup of “high responders” maintained high neutralizing responses over time, representing ideal convalescent plasma donors. Antibodies generated against SARS-CoV-2 during the first COVID-19 wave had reduced immunoreactivity and neutralization potency to emerging Spike variants and VOC. Accurate monitoring of SARS-CoV-2 antibody responses would be essential for selection of optimal responders and vaccine monitoring and design.

Coeliac disease is an autoimmune disorder triggered in genetically predisposed individuals by the ingestion of gluten proteins from wheat, barley and rye. The a-gliadin gene family of wheat contains four highly stimulatory peptides, of which the 33-mer is the main immunodominant peptide in patients with coeliac. We designed two sgRNAs to target a conserved region adjacent to the coding sequence for the 33-mer in the a-gliadin genes. Twenty-one mutant lines were generated, all showing strong reduction in a-gliadins. Up to 35 different genes were mutated in one of the lines of the 45 different genes identified in the wild type, while immunoreactivity was reduced by 85%. Transgene-free lines were identified, and no off-target mutations have been detected in any of the potential targets. The low-gluten, transgene-free wheat lines described here could be used to produce low-gluten foodstuff and serve as source material to introgress this trait into elite wheat varieties.

Outer membrane protein K (OmpK) has been proven to be an ideal vaccine candidate for broad-spectrum cross-prevention against Vibriosis. However, due to the extensive biological and genetic diversity of Vibrio species, current OmpK subunit vaccines can only target different strains of the same bacterial species or closely related species and have difficulty providing promising cross-immunoprotection against more diverse Vibrio infections. In recent years, the development of epitope-focused vaccines has been described as the latest stage in the development of vaccine formulations, providing new ideas for the development of broad-spectrum Vibrio vaccines. Interestingly, a cryptic epitope (K7) was identified in OmpK from Vibrio species, which is itself immunogenic but is not involved in the immune response to intact OmpK. Epitope K7 is a 15-residue hairpin structure in OmpK predicted to contain a 6-residue extracellular turn region. Interestingly, unlike other highly variable extracellular long loops, epitope K7 is the only conserved extracellular short turn in OmpK, with a similarity of 33 % to 93 %. K7 homologous peptides stimulated the production of specific antibodies, confirming their high immunogenicity. Cross-immunoreactivity between K7 homologous and K7-induced antibodies was evaluated by peptide-based ELISA, western blot, and cell-based ELISA. Flow cytometry and immunofluorescence assay further confirmed that the native epitope K7 in OmpK is surface-exposed and therefore an extracellular target that binds to antibodies. Moreover, an antibody-dependent and complement-mediated serum bactericidal assay suggested that epitope K7-induced antibodies have vibriocidal activity. In conclusion, we identified a conserved cryptic epitope with cross-immunoreactivity in OmpK from Vibrio species. Our results suggest that epitope K7 could be an ideal candidate for the design of epitope-focused vaccines against diverse Vibrio infections.