Pro-Inflammatory Cytokines Enhanced In Vitro Cytotoxic Activity of Clostridioides difficile Toxin B in Enteric Glial Cells: The Achilles Heel of Clostridioides difficile Infection?

Bacterial infections are characterized by an inflammatory response, which is essential for infection containment but is also responsible for negative effects on the host. The pathogen itself may have evolved molecular mechanisms to antagonize the antimicrobial effects of an inflammatory response and to enhance its pathogenicity using inflammatory response mediators, such as cytokines. Clostridioides difficile (C. difficile) infection (CDI) causes gastrointestinal diseases with markedly increasing global incidence and mortality rates. The main C. difficile virulence factors, toxin A and B (TcdA/TcdB), cause cytopathic/cytotoxic effects and inflammation. We previously demonstrated that TcdB induces enteric glial cell (EGC) apoptosis, which is enhanced by the pro-inflammatory cytokine tumor necrosis factor alpha plus interferon gamma (CKs). However, it is unknown whether CKs-enhanced TcdB cytotoxicity (apoptosis/necrosis) is affected by the timing of the appearance of the CKs. Thus, we simulated in vitro, in our experimental model with TcdB and EGCs, three main situations of possible interactions between TcdB and the timing of CK stimulation: before TcdB infection, concomitantly with infection, or at different times after infection and persisting over time. In these experimental conditions, which all represent situations of possible interactions between C. difficile and the timing of CK stimulation, we evaluated apoptosis, necrosis, and cell cycle phases. The CKs, in all of these conditions, enhanced TcdB cytotoxicity, which from apoptosis became necrosis when CK stimulation persisted over time, and was most relevant after 48 h of TcdB:EGCs interaction. Particularly, the enhancement of apoptosis by CKs was dependent on the TcdB dose and in a less relevant manner on the CK stimulation time, while the enhancement of necrosis occurred always independently of the TcdB dose and CK stimulation time. However, since in all conditions stimulation with CKs strongly enhanced the TcdB cytotoxicity, it always had a negative impact on C. difficile pathogenicity. This study might have important implications for the treatment of CDI.

Clostridioides difficile toxin B alone and with pro-inflammatory cytokines induces apoptosis in enteric glial cells by activating three different signalling pathways mediated by caspases, calpains and cathepsin B.

Clostridioides difficile infection (CDI) causes nosocomial/antibiotic-associated gastrointestinal diseases with dramatically increasing global incidence and mortality rates. The main C. difficile virulence factors, toxins A and B (TcdA/TcdB), cause cytopathic/cytotoxic effects and inflammation. We demonstrated that TcdB induces caspase-dependent, mitochondria-independent enteric glial cell (EGC) apoptosis that is enhanced by the pro-inflammatory cytokines TNF-α and IFN-γ (CKs) by increasing caspase-3/7/9 and PARP activation. Because this cytotoxic synergism is important for CDI pathogenesis, we investigated the apoptotic pathways involved in TcdB- and TcdB + CK-induced apoptosis indepth. EGCs were pre-treated with the inhibitors BAF or Q-VD-OPh (pan-caspase), Z-DEVD-fmk (caspase-3/7), Z-IETD-fmk (caspase-8), PD150606 (calpains), and CA-074Me (cathepsin B) 1 h before TcdB exposure, while CKs were given 1.5 h after TcdB exposure, and assays were performed at 24 h. TcdB and TcdB + CKs induced apoptosis through three signalling pathways activated by calpains, caspases and cathepsins, which all are involved both in induction and execution apoptotic signalling under both conditions but to different degrees in TcdB and TcdB + CKs especially as regards to signal transduction mediated by these proteases towards downstream effects (apoptosis). Calpain activation by Ca2+ influx is the first pro-apoptotic event in TcdB- and TcdB + CK-induced EGC apoptosis and causes caspase-3, caspase-7 and PARP activation. PARP is also directly activated by calpains which are responsible of about 75% of apoptosis in TcdB and 62% in TcdB + CK which is both effector caspase-dependent and -independent. Initiator caspase-8 activation mediated by TcdB contributes to caspase-3/caspase-7 and PARP activation and is responsible of about 28% of apoptosis in both conditions. Caspase-3/caspase-7 activation is weakly responsible of apoptosis, indeed we found that it mediates 27% of apoptosis only in TcdB. Cathepsin B contributes to triggering pro-apoptotic signal and is responsible in both conditions of about 35% of apoptosis by a caspase-independent manner, and seems to regulate the caspase-3 and caspase-7 cleaved fragment levels, highlighting the complex interaction between these cysteine protease families activated during TcdB-induced apoptosis. Further a relevant difference between TcdB- and TcdB + CK-induced apoptosis is that TcdB-induced apoptosis increased slowly reaching at 72 h the value of 18.7%, while TcdB + CK-induced apoptosis increased strongly reaching at 72 h the value of 60.6%. Apoptotic signalling activation by TcdB + CKs is enriched by TNF-α-induced NF-κB signalling, inhibition of JNK activation and activation of AKT. In conclusion, the ability of C. difficile to activate three apoptotic pathways represents an important strategy to overcome resistance against its cytotoxic activity.

Clostridioides difficile Toxin B Induced Senescence: A New Pathologic Player for Colorectal Cancer?

Clostridioides difficile (C. difficile) is responsible for a high percentage of gastrointestinal infections and its pathological activity is due to toxins A and B. C. difficile infection (CDI) is increasing worldwide due to the unstoppable spread of C. difficile in the anthropized environment and the progressive human colonization. The ability of C. difficile toxin B to induce senescent cells and the direct correlation between CDI, irritable bowel syndrome (IBS), and inflammatory bowel diseases (IBD) could cause an accumulation of senescent cells with important functional consequences. Furthermore, these senescent cells characterized by long survival could push pre-neoplastic cells originating in the colon towards the complete neoplastic transformation in colorectal cancer (CRC) by the senescence-associated secretory phenotype (SASP). Pre-neoplastic cells could appear as a result of various pro-carcinogenic events, among which, are infections with bacteria that produce genotoxins that generate cells with high genetic instability. Therefore, subjects who develop IBS and/or IBD after CDI should be monitored, especially if they then have further CDI relapses, waiting for the availability of senolytic and anti-SASP therapies to resolve the pro-carcinogenic risk due to accumulation of senescent cells after CDI followed by IBS and/or IBD.

Use of nutraceuticals in the stallion: Effects on semen quality and preservation.

Nutritional supplements are widely used in the equine industry with the aim of improving horse health, sports or reproductive performances. Over the years, a number of studies have focused on investigating the effects of several dietary compounds on the quality and preservation of stallion semen. This paper reviews the literature available on the use of nutritional supplementation for the improvement of reproductive performance and semen quality in equine species, critically appraising the benefits and negative effects of several compounds found in complementary feeds such as PUFAs from different sources, vitamins and antioxidants, carnitine and botanical extracts. Different nutraceuticals have been highlighted to improve stallion fertility by providing optimal levels of antioxidants, with the most promising results obtained by the combination of PUFAs and antioxidants that resulted to be essential for the maintenance of normal reproductive functions and the reduction of cryodamage in cooled and frozen equine semen.

Management of otitis externa with an led-illuminated gel: a randomized controlled clinical trial in dogs.

BACKGROUND: Canine otitis externa is a painful condition which can be challenging to treat due to difficulties in the administration of otic medication. This can be due to lack of owner compliance in the application of ear drops or due to the resentment that some dogs demonstrate when attempts are made to administer topical medication into a sensitive ear canal. The aim of the study was to assess the efficacy of a topical LED-illuminated gel (LIG) in canine otitis externa in comparison to standard of care therapy. Dogs with spontaneous otitis externa were randomly allocated in three groups: groups QW received LIG once weekly; BW received LIG twice weekly; group C received enrofloxacin and silver sulfadiazine twice daily. LIG consists of a topical application of a gel containing chromophores that, when illuminated by a LED lamp, re-emit fluorescent light which can stimulate physiological responses, promoting healing and controlling bacteria. The evaluation protocol (T0 to T5) considered clinical assessment (OTIS-3-index-scoring-system; pruritus-severity-scale; pain-severity-score; aural temperature), cytological scoring system, quali-quantitative bacteriologic assessment. RESULTS: All groups (QW, n = 21; BW, n = 23; C, n = 20) showed improvement during the study (QW: P < 0.02 for cytological and pain scores, P < 0.003 for bacteriologic assessment, P < 10- 4 for pruritus, total OTIS-3 and temperature assessments; BW: P < 10- 4 for all clinical, cytological and bacteriologic assessments; C: P < 0.02 for all clinical and cytological assessments, P < 10- 4 for bacteriologic assessment). The highest clinical score reduction occurred in Group BW (P < 0.014 in T3; P < 0.001 in T4 and P < 10- 4 in T5). BW reached the clinically relevant effect level at T3 (- 3.26 ± 1.21 levels), QW reached it at T4 (- 3.24 ± 0.99), C did not reach it. No differences between groups were seen in the reduction of CFU/mL (T0-T5). CONCLUSIONS: All treatment groups showed a positive clinical effect. LIG administered twice-a-week was the most favourable protocol of the study. LIG may be considered beneficial in the management of canine otitis externa; it seems to be effective in controlling the clinical condition, including the signs of inflammation and local pain, the bacterial growth, and it may help increasing treatment compliance.

Management of canine perianal fistula with fluorescence light energy: preliminary findings.

BACKGROUND: Canine perianal fistula (CPF) is a severe, painful, debilitating skin condition affecting the perianal skin. It often interferes with the quality of life of both dog and owner. Conventional medical treatment involves the use of immunosuppressive therapy; however, the successful resolution of lesions can be limited by poor owner compliance, adverse drug effects and dependence on costly therapies. HYPOTHESIS/OBJECTIVES: The present study aimed to assess the potential benefits of fluorescent light energy (FLE) on cases of CPF. ANIMALS: Four dogs with active perianal fistulas METHODS AND MATERIALS: FLE was applied as sole management therapy once a week with two consecutive applications in the same session for each dog until clinical signs had significantly improved, with weekly assessments for a six week period. Dogs were assessed by measuring the size of lesions at the start of the study and then weekly for six weeks, using planimetry software. Owners recorded vocalization and straining frequency scores during their pet's defaecation, and perianal licking frequency on a 0-5 point scale to assess the response to therapy. RESULTS: All dogs improved with FLE, achieving a significant reduction in vocalization, straining and licking after two weeks (P = 0.002). After five weeks of FLE therapy, lesional areas had significantly decreased (P = 0.04). Only one dog required more than seven applications. No adverse events were reported. CONCLUSION AND CLINICAL IMPORTANCE: FLE may be a promising alternative therapy for CPF.

Preoperative topical liposomal ozone dispersion to reduce bacterial colonization in conjunctival sac and periocular skin: Preliminary study in dogs.

Prophylaxis represents a keystone to reduce periocular skin and ocular conjunctiva bacterial load before surgical procedures. Despite many prophylactic agents are available the preferred perioperative ocular surface antimicrobial is still unknown. The purpose of this study was to assess the effectiveness of preoperative liposomal ozone dispersion in reducing bacterial colonization from the conjunctival sac and periocular skin in dogs, in comparison with povidone-iodine and fluoroquinolone. Twenty-two owned dogs consisting with 44 eyes in total scheduled for ophthalmic surgical procedure were enrolled for the study and divided in four groups receiving either ozone dispersion or povidone iodine in eyelid and conjunctiva, fluoroquinolone or placebo. A swab was taken before and after the antisepsis protocol evaluating total microbial count, coagulase positive and negative staphylococci. Statistical analysis revealed a significant decrease in colony forming units (CFU) for total microbial count, coagulase positive and negative staphylococci both for liposomal ozone dispersion and povidone iodine. No statistical differences were detected in median CFU for both one-day placebo and fluoroquinolone preoperative prophylactic topical therapy. The results of this preliminary study demonstrate that liposomal ozone-dispersion is as effective as povidone iodine to reduce preoperative bacterial load in ocular surface.

Fecal Proteomic Analysis in Healthy Dogs and in Dogs Suffering from Food Responsive Diarrhea.

Different laboratory markers are routinely used in the diagnosis and management of gastrointestinal (GI) disease in dogs. In the present study, starting from feces from both healthy dogs and dogs suffering from food responsive diarrhea (FRD), we tried to find proteins differently expressed in the two groups of dogs, by using a proteomic approach. Interestingly, we found that the immunoglobulin J-chain isoform 1 (species: Canis lupus familiaris) was identified only in diseased dogs (not in healthy). J-chain combines especially IgA monomers to IgA dimers and plays a crucial role for their secretions into mucosal interface. Being the first study of that kind in the dog, it is only possible to hypothesize that their presence could be likely due to an increased activation of the immune system or to a mucosal damage or both in FRD patients. Similarly, it is still impossible to assess whether this protein could be used as diagnostic/prognostic marker of GI disease; however, this study represents a promising first step toward fecal proteomics in canine GI disorders.

Fluorescence biomodulation in the management of canine interdigital pyoderma cases: a prospective, single-blinded, randomized and controlled clinical study.

BACKGROUND: Interdigital pyoderma is a common multifactorial, inflammatory disease of the canine interdigital skin. Lesions commonly become infected secondarily. In addition to management of the underlying cause, management of the chronic inflammatory changes in the interdigital skin created by secondary infection and by the release of keratin into deep tissues is required. Fluorescence biomodulation appears to modulate the inflammatory process in dermatological disorders and has shown promise in preliminary studies evaluating its use in superficial and deep pyoderma in dogs. HYPOTHESIS/OBJECTIVES: To evaluate the effect of a fluorescence biomodulation (FB) system used in conjunction with systemic antibiotic on clinical manifestations of canine interdigital pyoderma (CIP), compared to dogs treated with antibiotic alone. ANIMALS: Thirty-six dogs diagnosed with CIP. METHODS AND MATERIALS: Dogs were randomly allocated to treatment groups of either antibiotic alone (Group A) or antibiotic plus twice-weekly FB application (Group B). Dogs were scored over a 12 week period on the basis of two measured parameters: a global lesion score composed of four different lesions types and neutrophil engulfing bacterial scores. RESULTS: A statistically significant decrease was seen by Week 3 in both measured parameters for Group B compared to Group A. The mean time-to-resolution of lesions was 4.3 weeks in Group B and 10.4 weeks in Group A. CONCLUSION AND CLINICAL IMPORTANCE: The FB system shows promise as an adjunct therapy to systemic antibiotic use in the management of CIP.

Clinical investigation on Theileria equi and Babesia caballi infections in Italian donkeys.

BACKGROUND: Interest in the welfare and diseases of donkeys is constantly increasing in several countries. Despite this, clinical research into donkeys needs to be in continual development since they show different reactions compared to horses in many conditions, including infectious diseases, and need specific clinical and therapeutic approaches. No reports are currently available on clinical and clinical pathology data regarding donkeys with natural piroplasms infection. RESULTS: Venous blood samples were taken from one hundred and thirty eight donkeys and underwent indirect fluorescent antibody test (IFAT) to detect IgG antibodies against Theileria equi and Babesia caballi and real-time polimerase chain reaction (PCR) to detect Babesia spp. and Theileria spp. Clinical examinations, haematological analyses and serum bilirubin evaluation were also performed and compared with positive or negative status. A seroprevalence of 40.6% and 47.8% was found for T. equi and B. caballi, respectively; double positivity was detected in 19.6% of the animals. PCR results showed that 17.4% of the animals tested positive for T.equi and 3.6% for B. caballi with no double positivity. Twelve donkeys (8.7%) had clinical signs consistent with chronic forms of the disease and no acute forms were detected. Fifty-eight donkeys had haematological and serum bilirubin alterations and 56 (96.6%) of them were IFAT and/or PCR positive. Changes in erythrocyte number, packed cell volume, hemoglobin concentration, mean corpuscular hemoglobin, platelets number and total bilirubin were significantly associated with positive and symptomatic animals. CONCLUSION: Nonspecific clinical presentation seems to be very common in donkeys and several clinical pathology alterations persist after natural infection. Therefore, apparently healthy donkeys can have masked but severe clinical pathology alterations. Acute forms are very seldom observed in donkeys. Clinical monitoring of chronically infected donkeys is recommended since such animals represent a risk both for transmission to other animals and for their own health; furthermore, their production performances could be reduced. The study should also be intended as a contribution for veterinary practitioners because it describes the most usual clinical presentations and laboratory findings of equine piroplasmosis in naturally infected donkeys in endemic areas.

Blue-green endoscopy in a dog presenting chronic vomiting-regurgitation.

A 2-year-old male Maremma sheepdog presenting with chronic vomiting-regurgitation was examined at the University Veterinary Teaching Hospital, Camerino University. An oesophagogastroscopy with a single blue + green (BG) filter restricting wavelengths from 400 to 550 nm was carried out. A conventional white light endoscopy showed a dilated oesophagus with mildly diffuse erythematous mucosa (more accentuated proximal to the cardia); some portions of the gastric mucosa were covered with fluids and appeared only slightly erythematous. A blue green endoscopy highlighted the oesophageal lesions in dark blue, which made them appear more clearly defined from the remaining mucosa. In the gastric antrum, a small, slightly darker blue roundish area was visible. This area did not show up under the white light endoscopy. A histopathological assessment of biopsy specimens from the distal oesophagus, antrum (including the area highlighted only by BG endoscopy) and gastric body showed chronic-active hyperplastic esophagitis and superficial squamous epithelial dysplasia, while gastric samples showed severe diffuse hyperaemic gastritis of the antrum and superficial diffuse atrophy of the gastric body. The authors believe that the use of a BG endoscopy could be useful in veterinary medicine to increase the diagnostic potential of endoscopic assessment in animals.

Nuclear Magnetic Resonance (NMR) Metabolomics: Current Applications in Equine Health Assessment.

Metabolomics can allow for the comprehensive identification of metabolites within biological systems, at given time points, in physiological and pathological conditions. In the last few years, metabolomic analysis has gained popularity both in human and in veterinary medicine, showing great potential for novel applications in clinical activity. The aim of applying metabolomics in clinical practice is understanding the mechanisms underlying pathological conditions and the influence of certain stimuli (i.e., drugs, nutrition, exercise) on body systems, in the attempt of identifying biomarkers that can help in the diagnosis of diseases. Proton Nuclear Magnetic Resonance spectroscopy (1H-NMR) is well tailored to be used as an analytical platform for metabolites' detection at the base of metabolomics studies, due to minimal sample preparation and high reproducibility. In this mini-review article, the scientific production of NMR metabolomic applications to equine medicine is examined. The research works are very different in methodology and difficult to compare. Studies are mainly focused on exercise, reproduction, and nutrition, other than respiratory and musculoskeletal diseases. The available information on this topic is still scant, but a greater collection of data could allow researchers to define new reliable markers to be used in clinical practice for diagnostic and therapeutical purposes.

A Prospective, Blinded, Open-Label Clinical Trial to Assess the Ability of Fluorescent Light Energy to Enhance Wound Healing after Mastectomy in Female Dogs.

Mammary gland tumors represent the most frequently diagnosed malignant neoplasm in intact female dogs, and surgical removal represents the current gold standard treatment. To promote wound healing and prevent possible bacterial contamination, perioperative antimicrobials are commonly used in clinical practice, even though there are no publications establishing guidelines for the use of such drugs in canine mastectomy. The aim of the present study was to evaluate the ameliorative effect of fluorescent light energy on the quality of the healing process after mastectomy surgery in female dogs, in the absence of perioperative antimicrobial administration. Nine female dogs received a multiple-gland mastectomy due to gland tumors and received FLE application immediately after surgery and then five days after. The surgical incisions were evaluated by a blind investigator over time using the Modified Hollander Cosmesis and Modified Draize Wound Healing Score systems. Statistical analysis revealed a significant ameliorative effect of FLE in the control of step-off borders, contour irregularities, and excessive distortion. In addition, erythema, edema, and serous discharge were lower for those wounds managed with FLE. These results underscore the advantageous impact of FLE on the healing of post-mastectomy wounds in female dogs, offering the dual benefits of reducing potential infection risks and lessening the home care burden for pet owners.

Genome-wide association study for morphological and hunting-behavior traits in Braque Français Type Pyrénées dogs: A preliminary study.

High-throughput genotyping offers great potential to increase our understanding of the genomic basis of canid variation. Braque Français Type Pyrénées (BRA) are smart, agile, and friendly dogs originally developed for tracking, hunting, and retrieving feathered game. On a population of 44 unrelated BRA dogs, single nucleotide polymorphism (SNP) genotype data from the CanineHD Whole-Genome Genotyping BeadChip and evaluation scores for 12 traits related to morphology and hunting performance were available. After quality filtering, 95,859 SNPs on the 38 dog autosomes (CFA) were retained. Phenotypic scores were expressed on a scale from 1 (worst) to 6 (best) and were mostly poorly to moderately correlated except for some morphological traits (e.g. r = 0.81 between the conformation of the head and that of the eye). From GWAS, a total of 378 SNP-phenotype associations with posterior odds of association > 1 have been detected. The strongest associations were found for the eye conformation, for the skull/muzzle ratio, and for connection to the hunter. These included both new and previously identified markers and genes potentially involved with type and behavior traits in BRA. Six of the significant markers mapped within SETDB2, a gene known to be related to pointing behavior in dogs. These results advance our understanding of the genetic basis for morphology and hunting behavior in dogs and identify new variants which are potential targets for further research.

PhosphoLipidome Alteration Induced by Clostridioides difficile Toxin B in Enteric Glial Cells.

Clostridioides difficile (C. difficile) is responsible for a spectrum of nosocomial/antibiotic-associated gastrointestinal diseases that are increasing in global incidence and mortality rates. The C. difficile pathogenesis is due to toxin A and B (TcdA/TcdB), both causing cytopathic and cytotoxic effects and inflammation. Recently, we demonstrated that TcdB induces cytopathic and cytotoxic (apoptosis and necrosis) effects in enteric glial cells (EGCs) in a dose/time-dependent manner and described the underlying signaling. Despite the role played by lipids in host processes activated by pathogens, to counter infection and/or induce cell death, to date no studies have investigated lipid changes induced by TcdB/TcdA. Here, we evaluated the modification of lipid composition in our in vitro model of TcdB infection. Apoptosis, cell cycle, cell viability, and lipidomic profiles were evaluated in EGCs treated for 24 h with two concentrations of TcdB (0.1 ng/mL; 10 ng/mL). In EGCs treated with the highest concentration of TcdB, not only an increased content of total lipids was observed, but also lipidome changes, allowing the separation of TcdB-treated cells and controls into different clusters. The statistical analyses also allowed us to ascertain which lipid classes and lipid molecular species determine the clusterization. Changes in lipid species containing inositol as polar head and plasmalogen phosphatidylethanolamine emerged as key indicators of altered lipid metabolism in TcdB-treated EGCs. These results not only provide a picture of the phospholipid profile changes but also give information regarding the lipid metabolism pathways altered by TcdB, and this might represent an important step for developing strategies against C. difficile infection.

Characterization of different 5'-untranslated exons of the ASIP gene in black-and-tan Doberman Pinscher and brindle Boxer dogs.

Differential expression of the ASIP gene and its interaction with MC1R have provided basic insight into pigment-type switching in mammals. Here, we report the characterization of a specific red-haired skin transcript and a specific black-haired skin transcript in the ASIP gene in the black-and-tan Doberman Pinscher. It is also shown that the brindle-haired skin of the Boxer exhibits a deregulated expression resulting in various 5'-untranslated exons. Comparative sequence analysis revealed a short interspersed element and a poly(A) stretch inserted within the promoter region of the ASIP in the Boxer. Genotyping studies have shown that both insertions are also present in brindle and fawn animals of the Boxer and Great Dane breeds. Furthermore, we genotyped MC1R and K loci for their known variants that affect coat color in dogs. As expected, all animals were homozygotes (E(M) /E(M) ) for the mask mutation, and fawn animals were k(y) /k(y) . Unexpectedly, we found that all brindle animals were heterozygotes k(B) /k(y) . Our results suggest that differential expression of ASIP determine pigment-type switching in a MC1R and K allele-dependent manner in dogs.

Exploring fluorescent light energy as management option for canine superficial bacterial folliculitis.

Superficial bacterial folliculitis (SBF) represents a common dermatological diagnosis in dogs that can be successfully managed with either topical and/or systemic treatments. In the present study we evaluated the efficacy of a fluorescent light energy (FLE) device as sole management for SBF. The same FLE device has been shown, as adjunct therapy to systemic antibiotic or alone, to effectively control clinical manifestation of interdigital furunculosis. Twenty dogs were randomized to receive FLE once (six dogs) or twice (six dogs) weekly in comparison with oral anti-biotic (eight dogs) until complete healing. FLE regimen was able to significantly reduce the time needed to clinical resolution for oral antibiotic, supporting owners' compliance and welfare of dogs.

Sensitivity and specificity of in vivo COVID-19 screening by detection dogs: Results of the C19-Screendog multicenter study.

Trained dogs can recognize the volatile organic compounds contained in biological samples of patients with COVID-19 infection. We assessed the sensitivity and specificity of in vivo SARS-CoV-2 screening by trained dogs. We recruited five dog-handler dyads. In the operant conditioning phase, the dogs were taught to distinguish between positive and negative sweat samples collected from volunteers' underarms in polymeric tubes. The conditioning was validated by tests involving 16 positive and 48 negative samples held or worn in such a way that the samples were invisible to the dog and handler. In the screening phase the dogs were led by their handlers to a drive-through facility for in vivo screening of volunteers who had just received a nasopharyngeal swab from nursing staff. Each volunteer who had already swabbed was subsequently tested by two dogs, whose responses were recorded as positive, negative, or inconclusive. The dogs' behavior was constantly monitored for attentiveness and wellbeing. All the dogs passed the conditioning phase, their responses showing a sensitivity of 83-100% and a specificity of 94-100%. The in vivo screening phase involved 1251 subjects, of whom 205 had a COVID-19 positive swab and two dogs per each subject to be screened. Screening sensitivity and specificity were respectively 91.6-97.6% and 96.3-100% when only one dog was involved, whereas combined screening by two dogs provided a higher sensitivity. Dog wellbeing was also analyzed: monitoring of stress and fatigue suggested that the screening activity did not adversely impact the dogs' wellbeing. This work, by screening a large number of subjects, strengthen recent findings that trained dogs can discriminate between COVID-19 infected and healthy human subjects and introduce two novel research aspects: i) assessement of signs of fatigue and stress in dogs during training and testing, and ii) combining screening by two dogs to improve detection sensitivity and specificity. Using some precautions to reduce the risk of infection and spillover, in vivo COVID-19 screening by a dog-handler dyad can be suitable to quickly screen large numbers of people: it is rapid, non-invasive and economical, since it does not involve actual sampling, lab resources or waste management, and is suitable to screen large numbers of people.

Efficacy of an enterovaccine in recurrent episodes of diarrhea in the dog: a pilot study.

Recurrent episodes of self-limiting diarrhea in the dog, due to sudden dietary changes and to stressful or exciting situations, are conditions sometimes difficult to treat. Colifagina(®), a commercially available bacterial enterovaccine, showed, in previous studies performed on experimentally induced colitis in mice, to be able to improve both disease activity index and histological appearance, increase colonic secretion of IgA, and reduce inflammatory chemokine secretion. In the present study Colifagina(®) was administered to five dogs presenting recurrent episodes of self-limiting diarrhea and to one dog presenting chronic diarrhea. During the follow-up period, almost all patients decreased the number of episodes of abnormal defecation and the fecal score of such episodes improved in five out of six dogs. Even if further studies are needed to understand the exact potential of the compound, in dogs presenting recurrent episodes of self-limiting diarrhea due to sudden dietary changes and/or stressing or exciting situations, Colifagina(®) seems to be helpful in managing most of these patients.

Chronic hepatitis in man and in dog: a comparative update.

Chronic hepatitis is a frequent pathologic condition encountered in both dogs and humans; however, in the latter etiologic factors are usually searched and found that allow targeted therapeutic approaches, whereas in dogs this is less frequent. This review will take into consideration chronic hepatitis in dogs, and discuss differences and similarities between the two species with respect to this disease.