Radiographic evaluation of the forelimb bone development in maned wolves (Chrysocyon brachyurus).

This study aimed to assess the fusion of growth plates and the development of secondary ossification centres in the forelimb bones of maned wolves (Chrysocyon brachyurus), contrasting the findings with established data from domestic dogs. Three maned wolves, comprising one male and two females, initially aged between 3 and 4 months, were subjected to monthly radiographic evaluations until 10-11 months of age, followed by bimonthly assessments until 18-19 months of age, encompassing both forelimbs. The closure times of growth plates were observed as follows: supraglenoid tubercle (7-8 months), proximal humerus (17-19 months), distal humerus (8-9 months), medial epicondyle of the humerus (8-9 months), proximal ulna (9-10 months), proximal radius (13-15 months), distal ulna (13-15 months) and distal radius (17-19 months). Statistical analysis revealed significant differences in the areas of secondary ossification centres in the proximal epiphyses of the humerus and radius, respectively, observed from the initial evaluation at 8-9 months and 6-7 months. Conversely, the epiphyses of the supraglenoid tubercle, distal humerus, proximal ulna, distal ulna, medial epicondyle of the humerus and distal radius did not exhibit significant area differences between 3-4 months and 4-5 months, yet notable distinctions emerged at 5-6 months. In summary, while the radiographic appearance of epiphyseal growth plates and secondary ossification centres in maned wolves resembles that of domestic dogs, closure times vary. These findings contribute to understanding the dynamics of epiphyseal growth plates in this species.

Extracellular Vesicles from Scedosporium apiospermum Mycelial Cells: Implication for Fungal-Host Interplays.

The release of extracellular vesicles (EVs) has been implicated as an alternative transport mechanism for the passage of macromolecules through the fungal cell wall, a phenomenon widely reported in yeasts but poorly explored in mycelial cells. In the present work, we have purified and characterized the EVs released by mycelia of the emerging, opportunistic, widespread and multidrug-resistant filamentous fungus Scedosporium apiospermum. Transmission electron microscopy images and light scattering measurements revealed the fungal EVs, which were observed individually or grouped with heterogeneous morphology, size and electron density. The mean diameter of the EVs, evaluated by the light scattering technique, was 179.7 nm. Overall, the structural stability of S. apiospermum EVs was preserved during incubation under various storage conditions. The lipid, carbohydrate and protein contents were quantified, and the EVs' protein profile was evidenced by SDS-PAGE, revealing proteins with molecular masses ranging from 20 to 118 kDa. Through immunoblotting, ELISA and immunocytochemistry assays, antigenic molecules were evidenced in EVs using a polyclonal serum (called anti-secreted molecules) from a rabbit inoculated with conditioned cell-free supernatant obtained from S. apiospermum mycelial cells. By Western blotting, several antigenic proteins were identified. The ELISA assay confirmed that the anti-secreted molecules exhibited a positive reaction up to a serum dilution of 1:3200. Despite transporting immunogenic molecules, S. apiospermum EVs slightly induced an in vitro cytotoxicity effect after 48 h of contact with either macrophages or lung epithelial cells. Interestingly, the pretreatment of both mammalian cells with purified EVs significantly increased the association index with S. apiospermum conidia. Furthermore, EVs were highly toxic to Galleria mellonella, leading to larval death in a typically dose- and time-dependent manner. Collectively, the results represent the first report of detecting EVs in the S. apiospermum filamentous form, highlighting a possible implication in fungal pathogenesis.

Vitreous rebleeding rate after vitrectomy or phacovitrectomy in patients with proliferative diabetic retinopathy: A retrospective study.

BACKGROUND: The most common complication after vitrectomy is the rebleeding in vitreous cavity. It is important to detect the different factors that can increase the vitreous rebleeding rate in these patients. OBJECTIVE: To carry out a retrospective review of the rate of vitreous rebleeding after vitrectomy or phacovitrectomy. METHOD: Retrospective, descriptive and comparative study of patients with a diagnosis of proliferative diabetic retinopathy with phacovitrectomy or vitrectomy procedure. Personal background data, type of surgical intervention and grade of the surgeon who carried out the procedure were obtained. RESULTS: 1227 files of diabetic patients with vitrectomy or phacovitrectomy were reviewed. 65% presented systemic arterial hypertension. The average glomerular filtration rate was 63.50 (±32.36) ml/min/1.73 m2 and glycosylated hemoglobin (HBA1C) of 8% (4.6 to 15%). In the comparison of variables, a significant difference in the rate of vitreous rebleeding was obtained comparing phacovitrectomy with vitrectomy. (p = 0.003), in the relationship between vitrectomy with vitreous rebleeding, an odds ratio of 1.44 was obtained. CONCLUSION: The results obtained show a lower rate of rebleeding in patients undergoing phacovitrectomy in patients with proliferative diabetic retinopathy.

Psychometric properties of the Brazilian version of the Canine Brief Pain Inventory (CBPI) for dogs with clinical signs of osteoarthritis and preliminary evidence of its clinical utility / Propriedades psicométricas da versão brasileira do Breve Inventário de Dor Canina (BIDC) em cães com sinais clínicos de osteoartrite e evidências preliminares da sua utilização clínica

This study aimed to determine the psychometric properties and clinical use of the Brazilian version of the Canine Brief Pain Inventory (CBPI) in dogs with hip dysplasia (HD). Forty-three dogs with HD and 16 clinically normal dogs were enrolled. The HD dogs were treated daily with 4.4mg/kg carprofen (GT = 21) or placebo (GP = 19), for four weeks. Owners completed the CBPI at two weeks (W-2) and immediately before the start of the treatment (W0), two (W2) and four (W4) weeks during treatment, and two weeks (W6) after the end of treatment. The internal structure was accessed, and the Cronbach's alpha coefficient was 0.97, indicating the high internal consistency of the instrument. Principal Component Analysis (PCA) suggested the retention of one component, which accounted for 78% of the variability. The ROC curve analysis concluded that the score 3 has an excellent performance to discriminate between normal and possible HD dogs (AUC of 0.973). There was no difference between dogs treated with carprofen versus placebo. The instrument in Portuguese showed construct and criterion validity and reliability to be used in dogs with HD.

Objetivou-se determinar as propriedades psicométricas e a utilidade clínica do Breve Inventário de Dor Canina na língua portuguesa, em cães com displasia coxofemoral (DCF). O inventário foi preenchido por tutores de 43 animais com DCF e por 16 tutores de cães saudáveis. Os animais com DCF foram tratados com carprofeno 4,4mg/kg (GT = 21) ou placebo (GP =19), administrados uma vez ao dia durante quatro semanas. As avaliações foram realizadas duas semanas e imediatamente antes do tratamento, duas e quatro semanas durante o tratamento e após duas semanas do término do tratamento. A estrutura interna calculada pelo alfa de Cronbach = 0,97 indicou alta consistência dos dados. A análise dos componentes principais identificou a retenção de apenas um componente responsável por 78% da variabilidade dos dados. A análise da curva ROC indicou que o escore 3 discrimina cães saudáveis de cães com possível DCF (ASC de 0,973). Não houve diferença entre os cães tratados com carprofeno daqueles que receberam placebo. O questionário apresentou validade de constructo e critério e confiabilidade e pode-se empregá-lo para avaliar a dor crônica em cães com osteoartrite em países de língua portuguesa.

Real-time ultrasound-guided lumbosacral epidural anaesthesia in obese or appropriate body condition score dogs: A randomized clinical trial.

Correctly identifying the puncture site and needle position in obese dogs can be challenging to achieve epidural anaesthesia. The current study aimed to evaluate a real-time ultrasound-guided technique, to perform epidural anaesthesia in obese or appropriate body condition score dogs, based on visualization of local anaesthetic flow during its injection, compared to the traditional method of palpation of anatomical landmarks. Seventy-two client-owned dogs were evaluated in a prospective, comparative, randomized clinical trial, allocated into four groups of 18 dogs. For the Palpation-guided 1 (PG1) and 2 (PG2) groups, epidural anaesthesia was based on palpating anatomical landmarks. Dogs with a body condition score (BCS) 1-5/9 were included in the PG1 (non-obese), and those with a BCS 6-9/9 in PG2 (obese) groups. In the Ultrasound-guided 1 (USG1 - BCS 1-5/9) and 2 (USG2 - BCS 6-9/9) groups, epidural anaesthesia was guided by ultrasound (US). The flow of anaesthetic through the epidural canal was observed in all dogs by US. There were fewer needle-to-bone contacts in the US-guided groups when performing epidural anaesthesia; this only occurred on the vertebral laminae, never in the vertebral canal. Ultrasound guidance enabled local anaesthetic injection into the epidural space without the need for palpation of anatomical landmarks to guide needle placement. Blood reflux occurred in 11.1% (PG1), 22.2% (PG2), 5.5% (USG1), and 0% (USG2) of the dogs. Ultrasound-guided punctures led to fewer vascular punctures. Epidural anaesthesia was effective in all animals, and no complications were observed.

Maternal plasma folate concentration is positively associated with serum total cholesterol and low-density lipoprotein across the three trimesters of pregnancy.

Increased first-trimester low-density lipoprotein (LDL-C) concentration has been associated with adverse pregnancy outcomes, such as gestational diabetes. The B vitamins folate, B-6, and total B-12 are key for the methyl group-dependent endogenous synthesis of phosphatidylcholine, which is needed for lipoprotein synthesis, e.g., very low-density lipoprotein (VLDL), the precursor of circulating LDL-C. Maternal B-vitamin concentration usually declines across trimesters. Whether changes in maternal B-vitamin concentrations are associated with total cholesterol (TC), triglycerides (TG), and lipoprotein concentrations is unknown. Therefore, we explored the association between plasma folate, vitamin B-6 in the form of pyridoxal 5'-phosphate (PLP), and total B-12 with serum TC, LDL-C, HDL-C, and TG concentrations across trimesters. This secondary analysis used data of a prospective pregnancy cohort study included apparently healthy adult women (n = 179) from Rio de Janeiro, Brazil. The biomarkers were measured in fasting blood samples collected at 5-13, 20-26, and 30-36 weeks of gestation. The associations between B vitamins and lipid concentrations across trimesters were explored using linear mixed-effect models. Among B vitamins, only plasma folate was positively associated with TC (ß = 0.244, 95% CI 0.034-0.454) and LDL-C (ß = 0.193, 95% CI 0.028-0.357) concentrations. The positive relationship of maternal folate and TC and LDL-C concentrations may indicate the importance of folate as a methyl donor for lipoprotein synthesis during pregnancy.

Fenbendazole Controls In Vitro Growth, Virulence Potential, and Animal Infection in the Cryptococcus Model.

The human diseases caused by the fungal pathogens Cryptococcus neoformans and Cryptococcus gattii are associated with high indices of mortality and toxic and/or cost-prohibitive therapeutic protocols. The need for affordable antifungals to combat cryptococcal disease is unquestionable. Previous studies suggested benzimidazoles as promising anticryptococcal agents combining low cost and high antifungal efficacy, but their therapeutic potential has not been demonstrated so far. In this study, we investigated the antifungal potential of fenbendazole, the most effective anticryptococcal benzimidazole. Fenbendazole was inhibitory against 17 different isolates of C. neoformans and C. gattii at a low concentration. The mechanism of anticryptococcal activity of fenbendazole involved microtubule disorganization, as previously described for human parasites. In combination with fenbendazole, the concentrations of the standard antifungal amphotericin B required to control cryptococcal growth were lower than those required when this antifungal was used alone. Fenbendazole was not toxic to mammalian cells. During macrophage infection, the anticryptococcal effects of fenbendazole included inhibition of intracellular proliferation rates and reduced phagocytic escape through vomocytosis. Fenbendazole deeply affected the cryptococcal capsule. In a mouse model of cryptococcosis, the efficacy of fenbendazole to control animal mortality was similar to that observed for amphotericin B. These results indicate that fenbendazole is a promising candidate for the future development of an efficient and affordable therapeutic tool to combat cryptococcosis.

Radiographic evaluation of the forelimbs and hind limbs of marmosets (Callithrix spp.).

BACKGROUND: The maintenance of marmosets in captivity may trigger various disorders. The objective of this study was to provide a radiographic evaluation of the forelimbs and hind limbs of marmosets rescued from illegal wildlife trade and maintained in captivity. METHODS: Fourteen marmosets were studied. RESULTS AND CONCLUSIONS: Three marmosets were apparently normal. Of the eleven marmosets with radiographic changes in the hind limbs, five had pelvic narrowing and bilateral varus deviation of the femur, with one suggestive of rickets and another of osteomalacia. The other marmosets showed changes such as malunion of acetabular fracture, transtibial amputation, tibial antecurvatum, and knee osteoarthritis. Six marmosets showed alterations in the forelimbs such as radius fracture, scapulohumeral subluxation, and elbow joint incongruence. In conclusion, 78.57% of the marmosets had bone and/or joint changes detected by plain radiography, which were found predominantly in the hind limbs and without impediment to locomotion in captivity.

Necrohemorrhagic enteritis outbreak in a cattle feedlot in Nova Crixás, Goiás, Brazil / Surto de enterite necro-hemorrágica em um confinamento de bovinos no município de Nova Crixás, Goiás, Brasil

This study described an outbreak of necrohemorrhagic enteritis in a beef cattle feedlot in Nova Crixás, State of Goiás, Brazil, with emphasis on epidemiological, lesional, and laboratory aspects. Visits to the property were carried out and a necroscopic examination was performed on the bovine cadavers (N=57), which presented similar macroscopic alterations. Epidemiological data were collected, mainly referring to the feeding management of animals, and tissue samples were submitted to histopathological examination. Samples of feces and intestinal contents were also collected for bacterial isolation and PCR genotyping to detect the etiological agent, being confirmed Clostridium perfringens type A strains in 100% of the samples. Furthermore, 33.3% of strains isolated from intestinal contents and 40% of those isolated from feces were positive for beta-2 encoding gene. Considering the history, macroscopic and microscopic findings, as well as bacterial isolation and PCR, the diagnosis of bovine necrohemorrhagic enteritis was determined.(AU)

Descreve-se um surto de enterite necro-hemorrágica em um confinamento de bovinos de corte no município de Nova Crixás, Estado de Goiás, Brasil, com ênfase nos aspectos epidemiológicos, lesionais e laboratoriais. Foram realizadas visitas à propriedade e todos os cadáveres bovinos (N=57) foram submetidos ao exame necroscópico, os quais apresentaram alterações macroscópicas semelhantes. Foram compilados dados epidemiológicos, sobretudo referentes ao manejo alimentar dos animais e amostras de tecido foram submetidas a exame histopatológico. Foram colhidas, também, amostras de fezes e conteúdo intestinal para isolamento bacteriano e genotipagem por PCR para detecção do agente etiológico, sendo confirmadas estirpes de Clostridium perfringens tipo A em 100% das amostras. Ainda, 33,3% das cepas de Clostridium perfringens isoladas no conteúdo intestinal e 40% daquelas isoladas nas fezes foram positivas para o gene codificador da toxina beta-2. Considerando o histórico, os achados macroscópicos e microscópicos, o isolamento bacteriano e o PCR, foi estabelecido o diagnóstico de enterite necro-hemorrágica por C. perfringens tipo A.(AU)

Necrohemorrhagic enteritis outbreak in a cattle feedlot in Nova Crixás, Goiás, Brazil / Surto de enterite necro-hemorrágica em um confinamento de bovinos no município de Nova Crixás, Goiás, Brasil

This study described an outbreak of necrohemorrhagic enteritis in a beef cattle feedlot in Nova Crixás, State of Goiás, Brazil, with emphasis on epidemiological, lesional, and laboratory aspects. Visits to the property were carried out and a necroscopic examination was performed on the bovine cadavers (N=57), which presented similar macroscopic alterations. Epidemiological data were collected, mainly referring to the feeding management of animals, and tissue samples were submitted to histopathological examination. Samples of feces and intestinal contents were also collected for bacterial isolation and PCR genotyping to detect the etiological agent, being confirmed Clostridium perfringens type A strains in 100% of the samples. Furthermore, 33.3% of strains isolated from intestinal contents and 40% of those isolated from feces were positive for beta-2 encoding gene. Considering the history, macroscopic and microscopic findings, as well as bacterial isolation and PCR, the diagnosis of bovine necrohemorrhagic enteritis was determined.(AU)

Descreve-se um surto de enterite necro-hemorrágica em um confinamento de bovinos de corte no município de Nova Crixás, Estado de Goiás, Brasil, com ênfase nos aspectos epidemiológicos, lesionais e laboratoriais. Foram realizadas visitas à propriedade e todos os cadáveres bovinos (N=57) foram submetidos ao exame necroscópico, os quais apresentaram alterações macroscópicas semelhantes. Foram compilados dados epidemiológicos, sobretudo referentes ao manejo alimentar dos animais e amostras de tecido foram submetidas a exame histopatológico. Foram colhidas, também, amostras de fezes e conteúdo intestinal para isolamento bacteriano e genotipagem por PCR para detecção do agente etiológico, sendo confirmadas estirpes de Clostridium perfringens tipo A em 100% das amostras. Ainda, 33,3% das cepas de Clostridium perfringens isoladas no conteúdo intestinal e 40% daquelas isoladas nas fezes foram positivas para o gene codificador da toxina beta-2. Considerando o histórico, os achados macroscópicos e microscópicos, o isolamento bacteriano e o PCR, foi estabelecido o diagnóstico de enterite necro-hemorrágica por C. perfringens tipo A.(AU)

Cryptococcus neoformans Glucuronoxylomannan and Sterylglucoside Are Required for Host Protection in an Animal Vaccination Model.

Cryptococcus neoformans is an encapsulated fungal pathogen that causes meningoencephalitis. There are no prophylactic tools for cryptococcosis. Previously, our group showed that a C. neoformans mutant lacking the gene encoding sterylglucosidase (Δsgl1) induced protection in both immunocompetent and immunocompromised murine models of cryptococcosis. Since sterylglucosidase catalyzes degradation of sterylglucosides (SGs), accumulation of this glycolipid could be responsible for protective immunity. In this study, we analyzed whether the activity of SGs is sufficient for the protective effect induced by the Δsgl1 strain. We observed that the accumulation of SGs impacted several properties of the main polysaccharide that composes the fungal capsule, glucuronoxylomannan (GXM). We therefore used genetic manipulation to delete the SGL1 gene in the acapsular mutant Δcap59 to generate a double mutant (strain Δcap59/Δsgl1) that was shown to be nonpathogenic and cleared from the lung of mice within 7 days post-intranasal infection. The inflammatory immune response triggered by the Δcap59/Δsgl1 mutant in the lung differed from the response seen with the other strains. The double mutant did not induce protection in a vaccination model, suggesting that SG-related protection requires the main capsular polysaccharide. Finally, GXM-containing extracellular vesicles (EVs) enriched in SGs delayed the acute lethality of Galleria mellonella against C. neoformans infection. These studies highlighted a key role for GXM and SGs in inducing protection against a secondary cryptococcal infection, and, since EVs notoriously contain GXM, these results suggest the potential use of Δsgl1 EVs as a vaccination strategy for cryptococcosis.IMPORTANCE The number of deaths from cryptococcal meningitis is around 180,000 per year. The disease is the second leading cause of mortality among individuals with AIDS. Antifungal treatment is costly and associated with adverse effects and resistance, evidencing the urgency of development of both therapeutic and prophylactic tools. Here we demonstrate the key roles of polysaccharide- and glycolipid-containing structures in a vaccination model to prevent cryptococcosis.

Avaliação psicométrica em português do indicador de dor crônica de Helsinki em cães com sinais crônicos de osteoartrite / Psychometric testing in Portuguese of Helsinki's chronic pain index for dogs with chronic signs of osteoarthritis

Objetivou-se determinar a validade, a confiabilidade e a sensibilidade do indicador de dor crônica de Helsinki (IDCH) em português, além de estabelecer correlação entre a plataforma de pressão em cães com displasia coxofemoral (DCF). O questionário foi preenchido por tutores de 40 animais com DCF e por 16 tutores de cães saudáveis. Os animais com DCF foram tratados com carprofeno 4,4mg/kg (GT= 21) ou placebo (GP= 19), administrados uma vez ao dia, durante quatro semanas. A avaliação consistiu no preenchimento do questionário (IDCH), na escala analógica visual de dor (EAVdor) e na escala analógica visual de locomoção (EAVloc) com duas semanas (A1) e imediatamente antes do tratamento (A2), duas (S2) e quatro (S4) semanas de tratamento e após duas semanas do término do tratamento (S6). A consistência interna dos dados foi considerada excelente (alfa de Cronbach = 0,89). Houve correlação moderada entre o IDCH e a EAVdor e boa entre o IDCH e a EAVloc, porém não houve diferença entre o grupo tratado com carprofeno e placebo no IDCH. Não houve correlação entre os dados da plataforma de pressão e o IDCH. O questionário apresentou validade de constructo, critério e fidedignidade e pode-se empregá-lo em países de língua portuguesa em cães com osteoartrite.(AU)

We aimed to determine validity, reliability, and sensitivity of Helsinki's chronic pain index (HCPI) and stablish a correlation between HCPI in dogs with hip dysplasia (HD) using pressure sensitive walkway. Forty-owners of dogs with HD and 16 owners of health dogs filled a questionnaire. Dogs with HD were treated with carprofen 4.4mg/kg (GT n= 21) or with placebo (GP n= 19), both were administered once a day for 4 weeks. Evaluation was performed by the owners using the questionnaire (HCPI), the Visual Analogue Scale for pain (VASpain) and the VAS for locomotion (VASloc). The evaluation was performed 2 weeks before the treatment began (A1), immediately after treatment (A2), two (S2), four (S4) and two weeks after the end of treatment (S6) and the lameness was evaluated by pressure sensitive walkway. The internal consistency of the data was considered excellent (Cronbach´α coefficient= 0.89). There was a moderate correlation between the HCPI and VASpain. For VASloc the correlation was good. However, there was no difference between treatments, indicating low sensibility. No correlation was observed between pressure sensitive walkway and HCPI. We concluded that the questionnaire has construct and criterion validity, reliability and can be applied in dogs with osteoarthritis in Portuguese-speaking countries.(AU)

Avaliação psicométrica em português do indicador de dor crônica de Helsinki em cães com sinais crônicos de osteoartrite / Psychometric testing in Portuguese of Helsinki's chronic pain index for dogs with chronic signs of osteoarthritis

Objetivou-se determinar a validade, a confiabilidade e a sensibilidade do indicador de dor crônica de Helsinki (IDCH) em português, além de estabelecer correlação entre a plataforma de pressão em cães com displasia coxofemoral (DCF). O questionário foi preenchido por tutores de 40 animais com DCF e por 16 tutores de cães saudáveis. Os animais com DCF foram tratados com carprofeno 4,4mg/kg (GT= 21) ou placebo (GP= 19), administrados uma vez ao dia, durante quatro semanas. A avaliação consistiu no preenchimento do questionário (IDCH), na escala analógica visual de dor (EAVdor) e na escala analógica visual de locomoção (EAVloc) com duas semanas (A1) e imediatamente antes do tratamento (A2), duas (S2) e quatro (S4) semanas de tratamento e após duas semanas do término do tratamento (S6). A consistência interna dos dados foi considerada excelente (alfa de Cronbach = 0,89). Houve correlação moderada entre o IDCH e a EAVdor e boa entre o IDCH e a EAVloc, porém não houve diferença entre o grupo tratado com carprofeno e placebo no IDCH. Não houve correlação entre os dados da plataforma de pressão e o IDCH. O questionário apresentou validade de constructo, critério e fidedignidade e pode-se empregá-lo em países de língua portuguesa em cães com osteoartrite.(AU)

We aimed to determine validity, reliability, and sensitivity of Helsinki's chronic pain index (HCPI) and stablish a correlation between HCPI in dogs with hip dysplasia (HD) using pressure sensitive walkway. Forty-owners of dogs with HD and 16 owners of health dogs filled a questionnaire. Dogs with HD were treated with carprofen 4.4mg/kg (GT n= 21) or with placebo (GP n= 19), both were administered once a day for 4 weeks. Evaluation was performed by the owners using the questionnaire (HCPI), the Visual Analogue Scale for pain (VASpain) and the VAS for locomotion (VASloc). The evaluation was performed 2 weeks before the treatment began (A1), immediately after treatment (A2), two (S2), four (S4) and two weeks after the end of treatment (S6) and the lameness was evaluated by pressure sensitive walkway. The internal consistency of the data was considered excellent (Cronbach´α coefficient= 0.89). There was a moderate correlation between the HCPI and VASpain. For VASloc the correlation was good. However, there was no difference between treatments, indicating low sensibility. No correlation was observed between pressure sensitive walkway and HCPI. We concluded that the questionnaire has construct and criterion validity, reliability and can be applied in dogs with osteoarthritis in Portuguese-speaking countries.(AU)

Cytoglobin Attenuates Neuroinflammation in Lipopolysaccharide-Activated Primary Preoptic Area Cells via NF-κB Pathway Inhibition.

Cytoglobin (Cygb) is a hexacoordinate protein, associated with the transport of oxygen, nitric oxide scavenging, tumor suppression and protection against oxidative stress and inflammation. This protein is expressed in brain areas including the preoptic area (POA) of the anterior hypothalamus, the region responsible for the regulation of body temperature. In this study, we show that Cygb is upregulated in the rat hypothalamus 2.5 h and 5 h after intravenous administration of lipopolysaccharide (LPS). We investigated the effect of treatment with Cygb in POA primary cultures stimulated with LPS for 4 h. The levels of tumor necrosis factor-alpha (TNF-α) and interleukin-6 (IL-6) were measured and the results showed that Cygb reduced the concentrations of both cytokines. We further observed a decrease in immunoreactivity of the inflammatory transcription factor nuclear factor-κB (NF-κB), but not NF-IL6 and STAT3, in the nucleus of Cygb-treated POA cells. These findings suggest that Cygb attenuates the secretion of IL-6 and TNF-α in LPS-stimulated POA primary cultures via inhibition of the NF-κB signaling pathway, indicating that this protein might play an important role in the control of neuroinflammation and fever.

Characterisation of the in vivo interactions between detomidine and methadone in horses: Pharmacokinetic and pharmacodynamic modelling.

BACKGROUND: Pharmacokinetic (PK)/pharmacodynamic (PD) modelling offers new insights to design protocols for sedation and analgesia in standing horses. OBJECTIVES: To evaluate the parameters and interactions between detomidine and methadone when given alone or combined in standing horses. STUDY DESIGN: Randomised, placebo-controlled, blinded, crossover. METHODS: Eight adult healthy horses were given six treatments intravenously: saline (SAL); detomidine (5 µg/kg bwt; DET); methadone (0.2 mg/kg bwt; MET) alone or combined with detomidine (2.5 [MLD], 5 [MMD] or 10 [MHD] µg/kg bwt). Venous blood samples were obtained at predetermined times between 0 and 360 min after drug administration. Plasma detomidine and methadone were measured using a single, liquid/liquid extraction technique by liquid chromatography coupled with a triple quadrupole mass spectrometer (LC-MS/MS). Sequential PK/PD modelling compared rival models, with and without PK and PD interaction between drugs, to fit the PD data including height of the head above the ground (HHAG), a visual analogue scale for sedation (VAS), electrical (ET), thermal (TT) and mechanical (MT) nociceptive thresholds and gastrointestinal motility (GIM) [1]. RESULTS: Two and three compartment models best described the PK of detomidine and methadone, respectively. Detomidine decreased its own clearance as well as the clearance of methadone. The interaction of methadone on the effect of detomidine revealed an infra-additive (partial antagonism) effect for HHAG (α = -1.33), VAS (α = -0.98) and GIM (α = -1.05), a positive potentiation for ET (pot = 0.0041) and TT (pot = 0.133) and a synergistic to additive effect for MT (α = 0.78). MAIN LIMITATIONS: This is a small experimental study. CONCLUSIONS: Different PK/PD interactions were demonstrated for each PD parameter and could be modelled in vivo. The modelling of our data will allow us to simulate and predict the effect of constant rate infusions of both drugs for future investigations.

Pharmacological inhibition of pigmentation in Cryptococcus.

Melanin formation is a promising target for antifungal development. We screened a collection of 727 compounds that were previously approved for clinical use in humans for inhibition of pigmentation in Cryptococcus gattii, a lethal fungal pathogen that causes damage to both immunocompetent and immunocompromised hosts. The pyrimidine analogues flucytosine (5-fluorocytosine [5-FC]), 5-fluorouracil (5-FU) and carmofur were identified as efficient inhibitors of pigmentation in the C. gattii model. Since melanin synthesis is enzymatically catalyzed by laccase in Cryptococcus, we investigated whether inhibition of pigmentation by the pyrimidine analogues was laccase-mediated. Enzyme activity and expression of LAC genes were not involved in the effects of the pyrimidine analogues, suggesting alternative cellular targets for inhibition of pigmentation. To address this hypothesis, we screened a collection of approximately 8000 mutants of C. gattii that were produced by insertional mutation after incubation with Agrobacterium tumefaciens and identified a gene product required for the anti-pigmentation activity of 5-FC as a beta-DNA polymerase. Reduced expression of this gene affected capsule formation and urease activity, suggesting essential roles in the cryptococcal physiology. These results demonstrate a previously unknown antifungal activity of 5-FC and reveal a promising target for the development of novel antifungals.

Sedative and antinociceptive effects of different detomidine constant rate infusions, with or without methadone in standing horses.

BACKGROUND: Standing surgery avoids the risks of general anaesthesia in horses. OBJECTIVES: To assess sedation, antinociception and gastrointestinal motility in standing horses after a detomidine loading dose and 2-h constant rate intravenous (i.v.) infusion, with or without methadone. STUDY DESIGN: Blinded, randomised, crossover with seven healthy adult cross-bred horses, three geldings and four females (404 ± 22 kg). METHODS: Five i.v. treatments were administered to all horses with 1-week washout period: saline (SAL), detomidine low (2.5 µg/kg bwt + 6.25 µg/kg bwt/h) (DL) and high doses (5 µg/kg bwt + 12.5 µg/kg bwt/h) (DH) alone or combined with methadone (0.2 mg/kg bwt + 0.05 mg/kg bwt/h), (DLM) and (DHM), respectively. Height of head above the ground (HHAG), electrical (ET), thermal (TT) and mechanical (MT) nociceptive thresholds and gastrointestinal motility were evaluated at predetermined times between 5 and 240 min. A mixed effect model and Kruskal-Wallis test were used to analyse normally and non-normally distributed data, respectively. RESULTS: Sedation (<50% basal HHAG) was achieved for the duration of the infusion, and for an additional 15 min in DH and DHM groups. Nociceptive thresholds were higher than baseline, to the greatest degree and the longest duration, with DHM (ET and TT for 135 min and MT for 150 min). After DH, TT was significantly higher than baseline from 30 to 120 min and MT from 15 to 135 min. After DLM, ET was increased at 90 min, TT at 30 min and MT for 120 min. Gastrointestinal motility was reduced for up to 135 min after DL, 150 min after DLM and 210 min after DH and DHM. MAIN LIMITATIONS: Nociceptive thresholds are not equivalent to surgical stimuli. CONCLUSION: Methadone with the highest detomidine dose (DHM) may provide sufficient sedation and analgesia for standing surgical procedures and warrants further investigation.

Necrohemorrhagic enteritis outbreak in a cattle feedlot in Nova Crixás, Goiás, Brazil

ABSTRACT: This study described an outbreak of necrohemorrhagic enteritis in a beef cattle feedlot in Nova Crixás, State of Goiás, Brazil, with emphasis on epidemiological, lesional, and laboratory aspects. Visits to the property were carried out and a necroscopic examination was performed on the bovine cadavers (N=57), which presented similar macroscopic alterations. Epidemiological data were collected, mainly referring to the feeding management of animals, and tissue samples were submitted to histopathological examination. Samples of feces and intestinal contents were also collected for bacterial isolation and PCR genotyping to detect the etiological agent, being confirmed Clostridium perfringens type A strains in 100% of the samples. Furthermore, 33.3% of strains isolated from intestinal contents and 40% of those isolated from feces were positive for beta-2 encoding gene. Considering the history, macroscopic and microscopic findings, as well as bacterial isolation and PCR, the diagnosis of bovine necrohemorrhagic enteritis was determined.

RESUMO: Descreve-se um surto de enterite necro-hemorrágica em um confinamento de bovinos de corte no município de Nova Crixás, Estado de Goiás, Brasil, com ênfase nos aspectos epidemiológicos, lesionais e laboratoriais. Foram realizadas visitas à propriedade e todos os cadáveres bovinos (N=57) foram submetidos ao exame necroscópico, os quais apresentaram alterações macroscópicas semelhantes. Foram compilados dados epidemiológicos, sobretudo referentes ao manejo alimentar dos animais e amostras de tecido foram submetidas a exame histopatológico. Foram colhidas, também, amostras de fezes e conteúdo intestinal para isolamento bacteriano e genotipagem por PCR para detecção do agente etiológico, sendo confirmadas estirpes de Clostridium perfringens tipo A em 100% das amostras. Ainda, 33,3% das cepas de Clostridium perfringens isoladas no conteúdo intestinal e 40% daquelas isoladas nas fezes foram positivas para o gene codificador da toxina beta-2. Considerando o histórico, os achados macroscópicos e microscópicos, o isolamento bacteriano e o PCR, foi estabelecido o diagnóstico de enterite necro-hemorrágica por C. perfringens tipo A.

A glucuronoxylomannan-like glycan produced by Trichosporon mucoides.

Trichosporon asahii shares with Cryptococcus species the ability to produce glucuronoxylomannan (GXM), an immunomodulatory fungal polysaccharide. The ability of other opportunistic species of Trichosporon to produce GXM-like polysaccharides is unknown. In this study, we observed that T. mucoides was less pathogenic than T. asahii in an infection model of Galleria mellonella and asked whether this difference was related to the characteristics of GXM-like molecules. Compositional analysis of samples obtained from both pathogens indicated that the components of GXM (mannose, xylose and glucuronic acid) were, in fact, detected in T. mucoides and T. asahii glycans. The identification of the T. mucoides glycan as a GXM-like molecule was confirmed by its reactivity with a monoclonal antibody raised to cryptococcal GXM and incorporation of the glycan into the cell surface of an acapsular mutant of C. neoformans. T. mucoides and T. asahii glycans differed in molecular dimensions. The antibody to cryptococcal GXM recognized T. mucoides yeast forms less efficiently than T. asahii cells. Experiments with animal cells revealed that the T. mucoides glycan manifested antiphagocytic properties. Comparative phagocytosis assays revealed that T. mucoides and T. asahii were similarly recognized by macrophages. However, fungal association with the phagocytes did not depend on the typical receptors of cryptococcal GXM, as concluded from assays using macrophages obtained from Tlr2-/- and Cd14-/- knockout mice. These results add T. mucoides to the list of fungal pathogens producing GXM-like glycans, but also indicate a high functional diversity of this major fungal immunogen.

Golgi Reassembly and Stacking Protein (GRASP) Participates in Vesicle-Mediated RNA Export in Cryptococcus Neoformans.

Golgi reassembly and stacking protein (GRASP) is required for polysaccharide secretion and virulence in Cryptococcus neoformans. In fungal species, extracellular vesicles (EVs) participate in the export of polysaccharides, proteins and RNA. In the present work, we investigated if EV-mediated RNA export is functionally connected with GRASP in C. neoformans using a graspΔ mutant. Since GRASP-mediated unconventional secretion involves autophagosome formation in yeast, we included the atg7Δ mutant with defective autophagic mechanisms in our analysis. All fungal strains exported EVs but deletion of GRASP or ATG7 profoundly affected vesicular dimensions. The mRNA content of the graspΔ EVs differed substantially from that of the other two strains. The transcripts associated to the endoplasmic reticulum were highly abundant transcripts in graspΔ EVs. Among non-coding RNAs (ncRNAs), tRNA fragments were the most abundant in both mutant EVs but graspΔ EVs alone concentrated 22 exclusive sequences. In general, our results showed that the EV RNA content from atg7Δ and WT were more related than the RNA content of graspΔ, suggesting that GRASP, but not the autophagy regulator Atg7, is involved in the EV export of RNA. This is a previously unknown function for a key regulator of unconventional secretion in eukaryotic cells.