Neurobrucellosis (Brucella ceti) in striped dolphins (Stenella coeruleoalba): Immunohistochemical studies on immune response and neuroinflammation.

Neurobrucellosis is a shared condition of cetaceans and humans. However, the pathogenesis and immune response in cetacean neurobrucellosis has not been extensively studied. In this multicentric investigation, 21 striped dolphin (Stenella coeruleoalba) neurobrucellosis (Brucella ceti) cases diagnosed over a 10-year period (2012-2022) were retrospectively evaluated. For each case, morphological changes were assessed by evaluating 21 histological parameters. Furthermore, the immunohistochemical expression of Brucella antigen, glial fibrillary acid protein (GFAP), and a selection of inflammatory cell (IBA-1, CD3, and CD20) and cytokine (tumor necrosis factor-alpha [TNF-α], interferon-gamma [IFN-γ], interleukin [IL]-1ß, IL-2, and IL-6) markers were investigated. Inflammation of the leptomeninges, ependyma, and/or choroid plexus was lymphohistiocytic, containing macrophages/microglia (IBA-1+), T-cells (CD3+), and B-cells (CD20+) in equal proportion. B-cells occasionally formed tertiary follicles. GFAP expression showed astrocytosis in most cases. Expression of TNF-α, IFN-γ, and IL-2 indicated an intense proinflammatory response, stimulating both macrophages and T-cells. Our results showed that the inflammation and neuroinflammation in neurobrucellosis of striped dolphins mimic human neurobrucellosis and in vitro and in vivo studies in laboratory animals. Cetacean disease surveillance can be exploited to expand the knowledge of the pathogenesis and immunology of infectious diseases, particularly brucellosis, under a One Health approach.

Forensic findings in urban red fox mortality in the metropolitan area of Madrid, 2014-2022.

The expanding presence of red foxes (Vulpes vulpes) in urban and suburban regions could potentially lead to increased instances of human aggression towards this species. We studied 10 deceased red foxes that were submitted by law enforcement agencies in the metropolitan area of Madrid in 2014-2022 because of suspected abuse. Forensic autopsies were performed to establish the cause and manner of death. In 4 of the 10 cases, the cause of death was deemed unnatural, involving blunt-force trauma (n = 2), asphyxia resulting from hanging (n = 1), and firearm injury (n = 1). Among the remaining cases, most had succumbed to natural causes (n = 4), often marked by severe emaciation and a high burden of parasites, primarily Sarcoptes scabiei. In 2 cases, death was undetermined given the poor preservation of the carcass. The growing prevalence of wildlife species in urban areas, particularly red foxes, may require forensic veterinary investigation of deaths potentially related to abuse.

Tertiary lymphoid organs in wild boar exposed to a low-virulent isolate of African swine fever virus.

Despite the great interest in the development of a vaccine against African swine fever (ASF) in wild boar, the immunological mechanisms that induce animal protection are still unknown. For this purpose, tertiary lymphoid organs (TLOs) of wild boar were characterised and compared with mucosa-associated lymphoid tissues (MALTs) by histopathology, histomorphometry and immunohistochemistry (CD3, CD79, PAX5, LYVE1, fibronectin). In addition, real-time polymerase chain reaction (qPCR) and immunohistochemistry (p72) were used to evaluate the presence of ASF virus (ASFV) in blood and tissues samples, respectively. TLOs were observed in animals infected with a low-virulent ASFV isolate (LVI), animals co-infected with low and high-virulent ASFV isolates (LVI-HVI) and animals infected only with the high virulence isolate (HVI). TLOs in LVI and LVI-HVI groups were located adjacent to the mucosa and presented a similar structure to MALT. Immunoexpresion of p72 observed in the inflammatory cells adjacent to TLOs/MALTs confirmed its development and reactivity generated by ASF attenuated isolates. Immunohistochemical evaluation, based on cellular composition (T and B lymphocytes), and histomorphometrical study revealed a more pronounced maturation of TLOs/MALTs in the LVI-HVI group. It is currently unclear whether these formations play a protective role by contributing to local immunity in chronic inflammatory diseases. However, the structural similarities between TLOs and MALTs and the location of TLOs close to the mucosa suggest that they may perform a similar function, facilitating a local protective response. Nevertheless, further investigations are warranted to assess the cellular and humoral dynamics of these lymphoid organs induced by attenuated isolates.

Forensic investigations of suspected livestock depredation by vultures: scientific tools for compensation programmes.

Human-wildlife conflicts may have devastating consequences for fauna due to targeting by humans of wildlife populations suspected to have predated livestock. Suspicion of depredation of extensively raised livestock by vultures in Europe has triggered public administration-led forensic investigations intended to distinguish between predation and scavenging in order to compensate farmers for attacks on their livestock. In this study, gross and histological analyses were carried out on suspected cases of domestic animal depredation by griffon vultures (Gyps fulvus) over a 1-year period. Fifty-eight animals were affected (n = 41, suspected depredation), including domestic cattle (75.9%), sheep (22.4%) and a goat (1.7%). All the adults affected were female and most cases of suspected depredation occurred during the peripartum period (56%). Histological investigations distinguished between post-mortem (84% of diagnostic samples) and ante-mortem (16% of diagnostic samples) cases, and gross examinations revealed significant differences (P ≤ 0.002) between ante-mortem, post-mortem and non-diagnostic samples. This study highlights the need to optimize sampling protocols to increase the success of forensic studies. The forensic investigations presented here may be applied to resolve human-wildlife conflicts involving not only vultures but other endangered carnivores and contribute to human-wildlife coexistence in rural areas by protecting the interests of both the livestock sector and endangered species.

Intrapancreatic accessory spleens in African swine fever infection of wild boar (Sus scrofa).

Intrapancreatic accessory spleen (IPAS) is one of the most frequent congenital splenic anomalies in humans; however, studies in veterinary medicine are scarce. This study aimed to describe the macroscopic, histopathological and immunohistochemical features of 11 suspected cases of IPAS in wild boar piglets of 3-4 months old. Seven of the 11 animals were immunised with a low virulence isolate of African swine fever virus (ASFV) and subsequently challenged with a highly virulent ASFV isolate (LVI-HVI group). The remaining four animals were exclusively infected with a highly virulent isolate of ASFV (HVI group). Grossly, lesions comprised focal or multifocal reddish areas of variable shape, located on the surface of the pancreatic tail or within the parenchyma. Histological and immunohistochemical studies (anti-CD79 and CD3) confirmed the presence of IPAS in eight of the 11 cases. IPAS shared the same histological structure and alterations as those observed in the original spleen. The immunohistochemical study against ASFV revealed the presence of VP72+ cells in both the spleen and IPAS of seven of the eight piglets. The results of this study describe for the first time the presence of IPAS in ASFV infection of wild boar (Sus scrofa) regardless the isolate and suggest that the infection may induce the development of ectopic splenic tissue due to an increased demand for phagocytic cells from the reticuloendothelial system. However, further studies are needed to understand the immunological mechanisms that trigger the formation of these accessory organs.

Forensic investigations of religious rituals involving poultry: a case report.

Animal victims of human cruelty are receiving increasing attention from the press and society. Veterinary pathologists and civic authorities have a duty not only to elucidate the cause, method and manner of death but also to address the motivation behind a case. Poultry are commonly used as offering to gods in Santería rituals (ebós). Only a few cases have been reported in scientific journals, in contrast with the number of cases described in the media. Here, a hen and a cockerel (Gallus domesticus) were submitted to the VISAVET Health Surveillance Centre for autopsy, which comprised macroscopic and histological studies and microbiological testing for Chlamydia psittaci, avian orthoavulavirus 1 (Newcastle disease virus), West Nile virus and avian influenza virus. The carcasses were in an active decay stage of decomposition, containing larvae of Lucilia eximia. Both animals had been decapitated and haemorrhages in the cutting region were observed histologically. Post-mortem burns were also observed. To the best of the authors' knowledge, this is the first post-mortem description of lesions in a domestic avian species due to religious rituals. European Union and Spanish forensic practitioners should be aware that these ritual sacrifices occur in some European countries. Moreover, future European and national legislation updates should consider this type of animal abuse.

An Outbreak of Aeromonas salmonicida in Juvenile Siberian Sturgeons (Acipenser baerii).

Aeromonas salmonicida is one of the major threats to world aquaculture, causing fish furunculosis and high mortality rates in cultured fish, particularly salmonids. Although Aeromonas spp. is a thoroughly studied pathogen, little is known regarding aeromoniasis in sturgeons. After a mortality outbreak, four juvenile sturgeons (Acipenser baerii) were submitted for autopsy and tissue samples were collected for histopathological and microbiological studies. The external examination revealed size heterogenicity, skin hyperpigmentation and reduced body condition of sturgeons. Within the abdominal cavity, mild hepatomegaly and splenomegaly were observed, as well as generalized organic congestion. Histology revealed severe multifocal haemorrhagic and ulcerative dermatitis, mainly localized in the dorsal and latero-ventral areas of fish. The histological study also showed moderate to severe inflammation of gills and organic lesions compatible with septicaemia. Bacterial isolates were identified as Aeromonas salmonicida subsp. salmonicida using MALDI-TOF MS and PCR. Overall, the lesions first described here are consistent with those previously reported in other cultured fish species and contribute to a better understanding of the pathogenesis of Aeromonas salmonicida subsp. salmonicida in the Siberian sturgeon, aside from providing new diagnostic tools for bacterial diseases impacting the fast-growing industry of caviar.

Immunopathology of early and advanced epididymis lesions caused by Brucella ovis in rams.

Ovine brucellosis is an infectious disease that causes alterations in the reproductive tract in ram and abortion in ewes. Their negative economic impact in ovine production warrants a thorough understanding the interactions between B. ovis and the host. Here, epididymis lesions of rams infected by B. ovis were histopathologically staged into early and advanced. Expression by immunohistochemistry of Brucella antigens, inflammatory cell markers (CD3, CD79αcy) and cytokines (IFN-γ, TNF-α, TGF-ß1) was assessed in both stages. Early lesions were characterized by epithelial changes, interstitial inflammation, and mild fibrosis; whereas advanced lesions displayed caseous granulomas containing numerous macrophages, multinucleated giant cells, lymphocytes, and plasma cells. Expression of Brucella antigens were observed in both stages. The cellular response in B. ovis lesions were predominantly of T-cells (CD3+) whereas low numbers of B-cells and plasma cells (CD79αcy+) were present in both early and advanced lesions. IFN-γ was expressed by lymphocytes in early lesions suggesting that the adaptive immune response against B. ovis is initiated by Th1 cells, this response was also preserved in advanced stages. Expression of TNF-α was observed in neutrophils of epithelial microabscesses and intraepithelial T-cells of early lesions suggesting a promotion of neutrophil phagocytosis triggered by TNF-α. On the other hand, advanced lesions showed a reduction of TNF-α expression which may permit B. ovis persistence in granulomas. Lastly, TGF-ß1 expression (fibroblast, macrophages and less in lymphocytes) were increased with time, suggesting that B. ovis promotes TGF-ß1 secretion promoting chronicity of the lesions.

First detection of Flavobacterium psychrophilum in juvenile Siberian sturgeons (Acipenser baerii) and description of the pathological findings.

Flavobacterium psychrophilum affects many cultured fish species and is considered one of the most important bacterial pathogens causing substantial economic losses in salmonid aquaculture worldwide. Here, F. psychrophilum was identified by matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) and nested PCR as the aetiological agent causing mortality in diseased juvenile Siberian sturgeons (Acipenser baerii) reared on a freshwater fish farm. Diseased sturgeons were lethargic and displayed dark skin pigmentation, increased mucus production and the presence of skin ulcerations and haemorrhages specially on the ventral side and the base of fins. The histological examination of fish revealed proliferative branchitis, ulcerative and necrotizing dermatitis and myositis, lymphoid tissue atrophy, liver and kidney degeneration and thrombosis. To the best of our knowledge, this is the first report describing the infection of Siberian sturgeons by F. psychrophilum. The detection of F. psychrophilum in diseased Siberian sturgeons and the description of the pathological findings observed during the outbreak may contribute to a better understanding of the bacterium pathogenicity and the range of fish species susceptible to infection.

Histopathologic and immunohistochemical findings in the placentas and fetuses of domestic swine naturally infected with Brucella suis biovar 2.

Porcine brucellosis, which is caused by Brucella suis biovar (bv) 2, is a re-emerging disease that causes reproductive problems in pigs in Europe. The pathogenesis and lesions of B. suis intrauterine infection are poorly characterized; characterization could facilitate the diagnosis and investigation of porcine brucellosis. We collected samples of placentas and fetuses for histologic and microbiologic studies during an outbreak of abortions on a pig-breeding farm in Spain. Brucella was cultured from the vaginal swabs obtained from sows that had aborted, some placentas, and fetal tissues (spleen, liver, lung, gastric content); molecular testing confirmed B. suis bv 2 infection. Histologically, there was necrotizing and hemorrhagic placentitis; suppurative hepatitis; lymphoid depletion and sinusoidal histiocytosis in the spleen, lymph nodes, and thymus; and bronchointerstitial pneumonia. Hemorrhages were observed in the umbilical cord, heart, kidneys, and brain. We detected Brucella by immunohistochemistry (IHC) in all of the placentas and fetal organs studied, specifically in the trophoblasts of the chorionic epithelium, in the cytoplasm of macrophages in the chorionic stroma, and extracellularly in necrotic debris. Furthermore, we assessed the lymphocyte population in the placentas through the use of IHC (anti-CD3, anti-Pax5 antibodies), revealing that the lymphocytic response was composed of T cells but not B cells.

Characterization of the Fecal and Mucosa-Associated Microbiota in Dogs with Chronic Inflammatory Enteropathy.

Canine chronic inflammatory enteropathy implicates multifactorial pathogenesis where immunological dysregulation and gut microbiota changes have a central role. Most sequencing-based taxonomic studies have been focused on the fecal microbiota. However, the analysis of these samples does not provide complete information regarding the composition of the small intestine affected by this canine disease. Therefore, in this study, we aimed to characterize the intestinal bacterial microbiota in dogs with inflammatory bowel disease (IBD) (n = 34) by means of duodenal biopsies and fecal samples collected at the time of the diagnosis and to compare those to a group of healthy dogs (n = 12) using the 16S ribosomal RNA (16S rRNA) gene-targeted sequencing (Illumina MiSeq platform). Our study showed that IBD dogs presented differences in the fecal bacterial communities when compared with healthy dogs, with a lower relative abundance of Prevotellaceae (p = 0.005), Prevotella (p = 0.002), and Prevotellaceae Ga6A1 group (0.006); Erysipelotrichales (p = 0.019), Candidatus Stoquefichus (p < 0.001), Erysipelotrichaceae (p = 0.011), and Allobaculum (p = 0.003); Lachnospiraceae NK4A136 group (p = 0.015), Sellimonas (p = 0.042), Oscillospirales (p = 0.037), Oscillospiraceae UCG-005 (p < 0.001), Faecalibacterium (p = 0.028), and Fournierella (p = 0.034); Acidaminococcales, Acidaminococcaceae, and Phascolarctobacterium (p = 0.001); Aeromonadales (p = 0.026), Succinivibrionaceae (p = 0.037), and Succinivibrio (p = 0.031). On the other hand, a higher relative abundance of Enterococcaceae (Enterococcus; p = 0.003), Streptococcaceae (Streptococcus, p = 0.021), Enterobacterales (p = 0.027), Enterobacteriaceae (p = 0.008), and Escherichia-Shigella (p = 0.011) was detected. Moreover, when evaluating α-diversity, the dogs with IBD showed lower diversity in terms of richness and abundance of species (observed species [p = 0.031] and Shannon index [p = 0.039]). Furthermore, fecal microbiota in dogs with IBD was significantly different from healthy dogs (p = 0.006). However, only a few taxa relative abundance shifts (lower Rubrobacteria, Rubrobacterales, Rubrobacteriaceae, and Rubrobacter [p = 0.002]; Cyanobacteria [p = 0.010], Vampirivibrionia, Obscuribacterales, and Obscuribacteraceae [p = 0.005]; Neisseriaceae [p = 0.004] and Conchiformibius [p = 0.003]) were observed when assessing duodenal-associated microbiota of dogs with IBD. Thus, even if the bowel inflammation mainly affects the small intestine in the IBD-affected dogs of the study, fecal specimens may constitute a better sample due not only to their easy availability but also in terms of searching for bacterial taxa as biomarkers for canine IBD. The use of different diets in the study can also have a partial influence on the microbiota composition. Future studies encompassing multi-omics approaches should evaluate the functionality in both levels to unravel the pathophysiology of canine IBD.

Intestinal T-cell lymphoma in an Asian small-clawed otter: case report and literature review of lymphoma in the subfamily Lutrinae.

The Asian small-clawed otter (Aonyx cinereus) is an endangered species that is common in zoologic collections. A 17-y-old female Asian small-clawed otter under human care, with a clinical history of chronic renal disease, was euthanized because of deteriorating health. Histologically, the jejunal wall was infiltrated by a monomorphic population of small neoplastic lymphocytes that expanded the lamina propria of the villi and crypts, and on rare occasions invaded the submucosa. The tumor was composed of T cells (CD3+) with a proliferation index of 16%. Based on the World Health Organization (WHO) Classification of Hematopoietic Neoplasms in Domestic Animals, this lymphoma was classified as an enteropathy-associated T-cell lymphoma (EATL) type 2. We also present here a review of the literature on intestinal lymphoma in the subfamily Lutrinae (otters).

Extracellular matrix proteins (fibronectin, collagen III, and collagen I) immunoexpression in goat tuberculous granulomas (Mycobacterium caprae).

The lesion resulting from the interaction between Mycobacterium and the host immune response is the tuberculous granuloma. Tuberculous granulomas, except in incipient stages, are partially or totally encapsulated by connective tissue. The aim of this study was to assess the immunoexpression of the extracellular matrix proteins fibronectin, collagen III, and collagen I in granulomas caused by Mycobacterium caprae in goats (Capra aegagrus hircus) to understand capsule development at different granuloma stages. For this purpose, a retrospective study of 56 samples of tuberculous granulomas in lung (n = 30) and mediastinal lymph node (n = 26) from 17 goats naturally infected with M. caprae in stages I (n = 15), II (n = 14) and III (n = 27) was carried out. Fibronectin immunoreaction was extracellular, fibrillar-reticular in the center of stage I, II and III granulomas and peripheral in stages II and III granulomas. Collagen III immunoexpression was extracellular and fibrillar in the center of stages I, II and III tuberculous granulomas in lung and mediastinal lymph node, and progressive expression was observed in the periphery of stages II and III granulomas. Finally, collagen I immunoexpression was extracellular and fibrillar, showing a progressive loss of central expression and an increase in peripheral expression in stage III granulomas compared to stage I granulomas. Immunoexpression of these extracellular matrix proteins could help understand fibrogenesis and dating in tuberculous granuloma in both animal models and humans.

Environment and Offspring Surveillance in Porcine Brucellosis.

Porcine brucellosis, caused by Brucella suis (B. suis), is a notifiable disease causing significant economic losses in production systems. Most infected pigs may act as carriers and shed B. suis even if asymptomatic. This can contribute to environmental persistence, thus hindering control efforts. Here, the environment and the offspring were investigated during and after a B. suis outbreak at a sow breeding farm. The diagnosis of B. suis in sows (n = 1,140) was performed by culture and polymerase chain reaction (PCR) from vaginal swabs, indirect enzyme-linked immunosorbent assay (I-ELISA) from sera, and brucellin skin test (BST). B. suis diagnosis in post-weaning pigs (n = 899) was performed by I-ELISA in sera and BST. The environmental surveillance programme was implemented by placing gauze sponges (n = 175) pre-hydrated in a surfactant and inactivating liquid for Brucella DNA detection by PCR in different farm areas. Our results showed that the offspring of infected sows reacted to in vivo techniques for B. suis. Furthermore, the offspring born during the outbreak displayed higher seropositivity (I-ELISA) and reactivity (BST) than those pigs born after. Brucella DNA was detected in pregnant sow areas, boxes, boots, and post-weaning pig areas. In addition, Brucella DNA environmental detection was higher during the B. suis outbreak than the post B. suis outbreak. The environmental approach has proven to be a simple, practical, valuable, and safe method to detect and monitor B. suis. These results suggest a role of the environment and the offspring that should be considered in porcine brucellosis surveillance and control programmes.

Feline Gastrointestinal Eosinophilic Sclerosing Fibroplasia-Extracellular Matrix Proteins and TGF-ß1 Immunoexpression.

Feline gastrointestinal eosinophilic sclerosing fibroplasia (FGESF) has been described as an inflammatory disorder with an eosinophilic component with etiopathogenesis that is still unknown. Sixteen intestinal samples from two veterinary diagnostic services (2014-2017) were included in the study. A histopathological criterion classified the cases into three grades (mild, moderate, and severe) according to the distribution of the lesions and the course. An immunohistochemical study of collagen I, collagen III, fibronectin, and transforming growth factor ß1 (TGF-ß1) was performed in each case. An immunohistochemical study of mild grades shows greater collagen III immunoexpression, compared to collagen I and fibronectin, which suggests an "early" stage of fibrosis. In more intense grades, an increased immunoexpression of collagen I, compared to collagen III, suggests a "late" stage of fibrosis. Otherwise, the highest expression of TGF-ß1 was observed in the moderate phase, due to the high proliferation of reactive fibroblast and intense inflammation. The results suggest that the inflammatory infiltrate is the trigger for the elevation in TGF-ß1, altering the collagen type III:I ratio. In conclusion, immunohistochemical studies can be a very useful method in diagnosing cases of FGESF of mild grades and could help to apply a differential diagnosis regarding feline eosinophilic chronic enteritis (CEE) in the context of inflammatory bowel disease (IBD).

Bcl-2 Immunoexpression in Feline Epitheliotropic Intestinal T-Cell Lymphomas.

Lymphoma is the most common malignant hematopoietic neoplasm in domestic felines. Twenty-two cases of feline epitheliotropic duodenal T-cell lymphoma were characterized morphologically and immunohistochemically (CD3, Pax5, Ki-67), and Bcl-2 immunoexpression was established. Most cases were in domestic shorthair cats (88.2%), with a mean age of 11.2 years. All lymphomas were CD3+, with a low-to-moderate expression of Ki-67 (<30%). A correlation between the tumoral pattern of infiltration in the lamina propria and the intraepithelial distribution of the neoplastic lymphocytes was established (p = 0.0155). Intraepithelial nests of neoplastic lymphocytes were predominantly observed in lymphomas with a patchy distribution in the lamina propria, whereas intraepithelial plaques were seen in lymphomas with an obliteration pattern. Bcl-2 was expressed in neoplastic cells in all cases, and a higher expression was associated with increased villous stunting (p = 0.0221), and tended to be present in those cases with increased epithelial damage. The expression of Bcl-2 and the degree of epitheliotropism were correlated with neoplastic progression in epitheliotropic intestinal T-cell lymphomas; those displaying high Bcl-2 immunoexpression showed increased villous stunting and epithelial damage, suggesting that Bcl-2 is overexpressed in advanced tumor stages, and may be used as a predictor of tumoral behavior in feline epitheliotropic intestinal T-cell lymphomas. This entity showed many similarities with human MEITL, so the latter entity should be considered in further lymphoma classifications of domestic animals.

Isolation and immunophenotyping by flow cytometry of canine peripheral blood and intraepithelial and lamina propria duodenal T lymphocytes.

The gut associated lymphoid tissue (GALT) effector sites play a crucial role on the pathogenesis of many immune-mediated gastrointestinal diseases. The lymphocytes at these effector sites are principally T cells which present important morphological, phenotypical and functional differences. Flow cytometry (FC) is one of the most commonly used techniques to characterize intestinal lymphocytes in human and animal models. Published studies with a focus on dogs for intraepithelial lymphocytes (IEL) immunophenotyping exist in very limited numbers. Moreover, no lamina propria lymphocytes (LPL) isolation protocols in the canine species have been described for FC evaluation. In addition to immune intestinal dysregulation, imbalances in the peripheral blood immune system have been described in both human and animal gastrointestinal disorders. The aim of this study was to provide a protocol for canine IEL and LPL isolation for FC immunophenotyping of T cells subsets. Specifically, T helper, T cytotoxic, activated Th and Tc lymphocytes, regulatory, double negative, double positive, IFN-γ and IL-4 producing T cells, and to compare their respective populations between these effector sites and with the blood stream compartment in healthy dogs. The potential relationship of these cells distributions with age, sex and breed was also evaluated. This study included sixteen healthy dogs of different sexes and breeds with a mean age of 4.55 ± 2.93 years old. The selected protocols for the three immune compartments showed proper cell yield, purity, viability, and the absence of phenotypic and functional disturbances. Histologically, an adequate separation of the duodenal epithelium from the lamina propria was also observed. All the proposed T cells subsets were identified in the three immune compartments studied, showing some statistically significant differences in their distributions at these locations that highlight the importance of their individual evaluation. This study provides an adequate method for canine small intestine IEL and LPL isolation for FC immunophenotyping and is key for future studies on the gastrointestinal immune system associated with different canine diseases.

Colistin Selection of the Mcr-1 Gene in Broiler Chicken Intestinal Microbiota.

Colistin has a long story of safe use in animals for the treatment and prevention of certain bacterial diseases. Nevertheless, the first description of the mcr-1 gene showed that colistin resistance can spread by horizontal gene transfer and changed the landscape. This study aimed to assess the effect of colistin administration on the dispersion of resistance in the microbiota of day-old broiler chicks and how the presence of mcr-1 genes influences the spread of colistin resistance determinants. In this study, 100 one-day-old chicks were divided into four groups of 25 animals (G1, G2, G3, and G4). Animals from G3/G4 were challenged with mcr-1-carrying Salmonella (day 7), while colistin (600 mg/L) was administered daily to G2/G4 animals through drinking water (from day 8 to day 15). Two quantitative PCR assays were performed to compare the amount of Salmonella and mcr-1 that were present in the caecal samples. We observed that levels of mcr-1 were higher in G3/G4 animals, especially G4, due to the spread of mcr-1-carrying Salmonella. On day 21, Salmonella levels decreased in G4, reaching similar values as those for G3, but mcr-1 levels remained significantly higher, suggesting that colistin may accelerate the spreading process when mcr-1-carrying bacteria reach the gut.

Effect of Biscuit Flour and Fermented Defatted "Alperujo" Co-Administration on Intestinal Mucosa Morphology and Productive Performance in Laying Hens.

In this study, the effects of co-administration with biscuit flour and fermented defatted "alperujo" (FDA) on gut health were evaluated in a batch of laying hens (Hy-Line 2015) on a commercial farm. Animals were divided into two groups: control group and treatment group; and histological and morphometric analyses of all sections of the intestine (duodenum, jejunum, ileum, cecum and rectum) were performed at 10, 18, 25, 50 and 75 weeks of age. During the whole productive period, a decrease in the mortality rate (p = 0.01) was observed in treated hens, as well as an increase in the number of eggs produced (p < 0.001), their size (p < 0.025), and weight (p < 0.024). In the early and late stages of production (10, 18 and 50 weeks), a significant increase (p < 0.001) in the height and depth of the intestinal villi was observed in the treatment group. Villi height was also significantly higher (p < 0.001) in the treatment group up to week 50 in the cecum, and at weeks 18 and 50 in the rectum. We concluded that an economical and sustainable feeding system with less environmental impact, such as co-supplementation with biscuit flour and FDA, could maintain gut health without negatively impacting laying hens' productive performance.

Dietary supplementation with fermented defatted "alperujo" induces modifications of the intestinal mucosa and cecal microbiota of broiler chickens.

Nutraceuticals are not only nutritionally beneficial for animals but also their use as feed supplements may reduce environmental contamination. The effect of fermented defatted "alperujo," an olive oil by-product, supplementation on the intestinal health of broiler chickens was assessed by analyzing the intestinal mucosal morphology of the duodenum and the cecum. The microbiota of the cecum was also characterized by analyzing the V3-V4 region of the 16S rRNA gene on days 7, 14, 21, 28, 35, and 42. Supplemented broilers from 14 to 35 D of age showed an increase in villus height in the duodenum. This increase likely improved digestibility and absorption capacity during growth, leading to the observed increase in BW at day 35 of life. A progressive increase in crypt depth in both the duodenum and the cecum was also observed. This modification likely enhanced epithelial renewal, thus safeguarding the turnover capacity of the intestinal mucosa. Our molecular analysis of cecal microbiota suggests that this dietary supplement may favor the growth of certain bacteria and may control the spread of pathogenic bacteria by means of competitive exclusion.