Inconspicuous post-mortem findings in rabbits from Switzerland naturally -infected with Rabbit Haemorrhagic -Disease Virus 2.

INTRODUCTION: Rabbit Haemorrhagic Disease Virus 2 (RHDV-2) emerged in France in 2010. In Switzerland, RHDV-2 was first identified in 2015 and apparently has almost replaced the classical Rabbit Haemorrhagic Disease Virus (RHDV) by now. Like RHDV, RHDV-2 causes a viral hepatitis with a peracute course and an increased mortality rate within the rabbitry. RHDV infection causes consistent gross pathological findings, especially in the liver and respiratory tract. Reports about gross pathology for animals naturally infected with RHDV-2 is scarce. The present study analysed the anamnesis and necropsy reports of 35 rabbits examined during routine diagnostics between March 2015 and May 2017. A reverse transcriptase real-time polymerase chain reaction (RT-qPCR) specific for RHDV-2 and RHDV proved a total of 25 animals to be positive for RHDV-2, while none was positive for RHDV. Additionally, histological examinations were performed on liver, lung, and kidney of 18 rabbits that had tested positive by RHDV-2 RT-qPCR. The anamnestic report more often stated an increased mortality rate in RHDV-2 positive (16/18, 89 %) compared to RHDV-2 negative rabbits (3/9, 33 %). Gross pathology did not reveal any pathognomonic changes in RHDV-2 positive animals. Histologically, the liver showed the most severe lesions followed by lung and kidney. Animals positive for RHDV-2 frequently showed signs of gastro-intestinal disease (n = 5) and/or septicaemia (n = 6) masking possible indicators of an RHDV-2 infection, such as the rather unspecific findings of an enlarged spleen or an enlarged, friable, tan-coloured liver. The authors want to raise awareness among clinicians and pathologists that in case of sudden death in commercial or pet rabbits, RHDV-2 needs to be considered as differential diagnosis and should be confirmed by laboratory diagnosis.

INTRODUCTION: Le virus 2 de la maladie hémorragique du lapin (RHDV-2) est apparu en France en 2010. En Suisse, le RHDV-2 a été identifié pour la première fois en 2015 et semble avoir presque remplacé le virus classique de la maladie hémorragique du lapin (RHDV). Comme le RHDV, le RHDV-2 provoque une hépatite virale avec une évolution suraiguë et un taux de mortalité élevé chez les lapins. L'infection par le RHDV entraîne des constatations pathologiques bruts cohérents, notamment au niveau du foie et des voies respiratoires. Les rapports sur la pathologie macroscopique des animaux naturellement infectés par le RHDV-2 sont rares. La présente étude a analysé les rapports d'anamnèse et de nécropsie de 35 lapins examinés lors de diagnostics de routine entre mars 2015 et mai 2017. Une réaction en chaîne par polymérase en temps réel à la transcriptase inverse (RT-qPCR) spécifique pour le RHDV-2 et le RHDV a prouvé qu'un total de 25 animaux étaient positifs pour le RHDV-2, tandis qu'aucun n'était positif pour le RHDV. De plus, des examens histologiques ont été réalisés sur le foie, les poumons et les reins de 18 lapins qui avaient été testés positifs par RT-qPCR pour le RHDV-2. Le rapport anamnestique faisait plus souvent état d'un taux de mortalité accru chez les lapins RHDV-2 positifs (16/18, 89 %) que chez les lapins RHDV-2 négatifs (3/9, 33 %). La pathologie macroscopique n'a révélé aucun changement pathognomonique chez les animaux RHDV-2 positifs. Sur le plan histologique, le foie présentait les lésions les plus graves, suivi des poumons et des reins. Les animaux positifs pour le RHDV-2 présentaient fréquemment des signes de maladie gastro-intestinale (n = 5) et/ou de septicémie (n = 6) masquant les indicateurs possibles d'une infection par le RHDV-2, tels que les découvertes plutôt peu spécifiques d'une rate hypertrophiée ou d'un foie hypertrophié, friable et de couleur beige. Les auteurs souhaitent sensibiliser les cliniciens et les pathologistes au fait qu'en cas de mort subite chez des lapins d'élevage ou de compagnie, le RHDV-2 doit être considéré comme un diagnostic différentiel et doit être confirmé par un diagnostic de laboratoire.

Role of transient receptor potential ankyrin 1 receptors in rodent models of meningeal nociception - Experiments in vitro.

BACKGROUND: The TRP channel ankyrin type 1 (TRPA1) is a nonselective cation channel known to be activated by environmental irritants, cold and endogenous mediators of inflammation. Activation of TRPA1 in trigeminal afferents innervating meningeal structures has recently been suggested to be involved in the generation of headaches. METHODS: Two in vitro models of meningeal nociception were employed using the hemisected rodent head preparation, (1) recording of single meningeal afferents and (2) release of calcitonin gene-related peptide (CGRP) from the cranial dura mater. The role of TRPA1 was examined using the TRPA1 agonists acrolein and mustard oil (MO). BCTC, an inhibitor of TRP vanilloid type 1 receptor channels (TRPV1), and the TRPA1 inhibitor HC030031 as well as mice with genetically deleted TRPA1 and TRPV1 proteins, were used to differentiate between effects. RESULTS: Acrolein did not cause discharge activity in meningeal Aδ- or C-fibres but increased the electrical activation threshold. Acrolein was also effective in releasing CGRP from the dura of TRPV1-/- but not of TRPA1-/- mice. MO increased the discharge activity of afferent fibres from rat as well as C57 wild-type and TRPA1-/- but not TRPV1-/- mice. The effect was higher in C57 compared to TRPA1-/- mice. CONCLUSION: Sole TRPA1 receptor channel activation releases CGRP and increases the activation threshold of meningeal afferents but does not generate propagated activity, and so would be capable of causing local effects like vasodilatation but not pain generation. In contrast, combined TRPA1 and TRPV1 activation may be rather pronociceptive supporting headache generation. SIGNIFICANCE: Sole activation of TRPA1 receptor channels increases the activation threshold but does not cause propagated action potentials in meningeal afferents. TRPA1 agonists cause CGRP release from rodent dura mater. Peripheral TRPA1 receptors may have a pronociceptive function in trigeminal nociception only in combination with TRPV1.

From Hospital to Home: Limited Nutritional and Functional Recovery for Older Adults.

BACKGROUND: The post-hospital period may be a vulnerable time for elders recovering from acute illness. Few studies have examined nutrition outcomes of older people at nutrition risk after acute hospitalisation. OBJECTIVES: This study aims to describe a) standard nutrition care received by recently discharged older medical patients, b) change in nutritional and functional status at six weeks post-discharge and c) clinical outcomes at twelve weeks post discharge. DESIGN: Prospective cohort study. SETTING: Two metropolitan teaching hospitals in Brisbane, Australia. PARTICIPANTS: Medical patients aged ≥65 years at risk of malnutrition (Malnutrition Screening Score ≥2) and discharged to independent living in the community. MEASUREMENT: Nutritional status (Mini Nutritional Assessment (MNA), weight, lean body mass), functional status (grip strength, walk speed, activities of daily living) and health-related quality of life assessed on discharge and six weeks post-discharge. Inpatient and post-discharge nutrition intervention was recorded. Death and unplanned admissions were measured at 12 weeks. RESULTS: Of the 42 consented participants, only 14% (n=6) received post-discharge dietitian review and 19% (n=8) received practical nutrition supports at home (meal delivery, shopping assistance) as part of standard care. While there was a small improvement in MNA (18.4±4.0 to 20.1±4.2, p=0.004) and walk speed (0.7±0.3 m/s to 0.9±0.3, p=0.004) at six weeks, there was no difference in mean weight, lean body mass, grip strength or activities of daily living. Five (15%) participants lost ≥5% body weight. By twelve weeks, 17 participants (46%) had at least one unplanned hospital admission and four (10%) had died. CONCLUSIONS: Few patients at nutrition risk received nutrition-focussed care in the post-hospital period, and most did not improve nutritional or functional status at 6 weeks.

Primary leiomyosarcoma in the heart of a guinea pig.

Leiomyosarcoma was diagnosed in the heart of a 3-year-old female guinea pig. Neoplastic tissue was located in the ventricular septum extending into the right ventricular lumen, but was not obvious grossly. Microscopically, the mass was nodular and infiltrative and was composed of streams and bundles of spindle cells. Immunohistochemically, the neoplastic cells expressed vimentin and α-smooth muscle actin. There was no cross striation of the muscle cells. This is the first report of cardiac leiomyosarcoma in a guinea pig.

Gonadal activity in male and female captive fossas (Cryptoprocta ferox) during the mating season.

The fossa is an endangered, mongoose-like carnivore species endemic to Madagascar with a breeding season (BS) in the southern hemisphere spring. For the present study, faecal samples of captive fossas were collected for over 1 year for five males and four non-pregnant females, and for two pregnant animals during the reproductive period. The goal was to assess gonadal activity using non-invasive hormone measurements of faecal testosterone (T) and gestagen metabolites using assays previously established in our laboratory and further validated in this study. All study animals were housed in northern hemisphere zoos. In males, the seasonal T metabolite profile revealed a peak in spring (March). High performance liquid chromatography (HPLC) analysis was used to characterize the faecal hormone metabolite composition. The highest immunoreactivity was detected in the position of dihydrotestosterone, whereas native T was not detected. In the two pregnant females, gestagen metabolite concentrations increased 4-9 days after the initial matings and remained elevated throughout gestation with concentrations dramatically higher than those of the non-pregnant females during the BS. In these females, gestagen metabolite analyses did not reveal a seasonal pattern similar to the males. The HPLC-analysis revealed that the major proportion of immunoreactivity was associated with an unknown metabolite, whereas native progesterone was undetectable. The seasonal hormone pattern of male fossas gives proof of the reproductive seasonality of this species. The elevated 5alpha-pregnan-3beta-ol-20-one levels in pregnant animals allows for the characterization of pregnancy in the fossa based on analysis of faecal steroid metabolite concentrations.

The in vitro anti-denaturation effects induced by natural products and non-steroidal compounds in heat treated (immunogenic) bovine serum albumin is proposed as a screening assay for the detection of anti-inflammatory compounds, without the use of animals, in the early stages of the drug discovery process / Los efectos de la anti-desnaturalización in vitro inducida por productos naturales y compuestos no esteroidales en la albúmina sérica bovina (inmunogénica) tratada con calor, se proponen aquí como ensayo de pesquizaje para la detección de compuestos inflamatorios sin el uso de animales en las etapas tempranas del proceso de descubrimiento del medicamento

There are emerging ethical issues with regards to the use of animals in the early stages of drug discovery for anti-inflammatory and degenerative diseases from natural products using the activity-directed isolation pathways when many compounds (eg > 100) are present in the crude extract or fraction and are to be tested. The above-mentioned is the main reason for proposing the use of the in vitro anti-denaturation (stabilization) effects of heat treated (immunogenic) bovine serum albumin (BSA) as an assay. Current methods used for detecting and isolating a wide range of anti-inflammatory compounds in the early stages of the drug discovery process utilize a large number of animals. When BSA is heated and is undergoing denaturation, it expresses antigens associated to Type III hyper-sensitive reaction and which are related to diseases such as serum sickness, glomerulonephritis, rheumatoid arthritis and systemic lupus erythematosus. Thus, the assay that is being proposed should be applicable to the discovery of drugs for treating the above mentioned diseases and others, once the compounds stabilize the denaturation process.

Actualmente surgen problemas éticos en relación con el uso de animales en las etapas tempranas del descubrimiento de medicamentos anti-inflamatorios y contra enfermedades degenerativas, a partir de productos naturales, usando vías de aislamiento dirigido por actividad, cuando muchos compuestos están presentes (p.ej. > 100) en la fracción o extracto crudo, y deben ser probados. Lo anterior representa la razón principal para proponer el uso de los efectos de la anti-desnaturalización (estabilización) in vitro de la albúmina sérica bovina (inmunogénica) tratada con calor (ASB) como ensayo. Los métodos corrientes usados para detectar y aislar una amplia gama de compuestos anti-inflamatorios en las etapas tempranas del proceso de descubrimiento del medicamento, utilizan un gran número de animales. Cuando la ASB es calentada y sometida a un proceso de desnaturalización, expresa antígenos en relación con la reacción hipersensitiva de tipo III, relacionada a su vez con enfermedades tales como la enfermedad del suero, la glomerulonefritis, la artritis reumatoide, y el lupus sistémico y eritematoso. De este modo, el ensayo que aquí proponemos debe ser aplicable al descubrimiento de medicamentos para el tratamiento de las enfermedades anteriormente mencionadas y otras, una vez que los compuestos estabilicen el proceso de desnaturalización.

The in vitro anti-denaturation effects induced by natural products and non-steroidal compounds in heat treated (immunogenic) bovine serum albumin is proposed as a screening assay for the detection of anti-inflammatory compounds, without the use of animals, in the early stages of the drug discovery process.

There are emerging ethical issues with regards to the use of animals in the early stages of drug discovery for anti-inflammatory and degenerative diseases from natural products using the activity-directed isolation pathways when many compounds (eg > 100) are present in the crude extract or fraction and are to be tested The above-mentioned is the main reason for proposing the use of the in vitro anti-denaturation (stabilization) effects of heat treated (immunogenic) bovine serum albumin (BSA) as an assay. Current methods used for detecting and isolating a wide range of anti-inflammatory compounds in the early stages of the drug discovery process utilize a large number of animals. When BSA is heated and is undergoing denaturation, it expresses antigens associated to Type III hypersensitive reaction and which are related to diseases such as serum sickness, glomerulonephritis, rheumatoid arthritis and systemic lupus erythematosus. Thus, the assay that is being proposed should be applicable to the discovery of drugs for treating the above mentioned diseases and others, once the compounds stabilize the denaturation process.

In vitro anti-proliferation/cytotoxic activity of epingaione and its derivatives on the human SH-SY5Y neuroblastoma and TE-671 sarcoma cells.

Epingaione (4-Methyl-1-(5-methyl-2, 3,4,5-tetrahydro-[2,3']bifuranyl-5-yl)-pentan-2-one) was isolated as one of the major lipophilic secondary metabolites from the leaves and stems of Bontia daphnoides L. The compound gave 79.24% and 50.83% anti-proliferation/cytotoxic activity on the human SH-SY5Y neuroblastoma and TE-671 sarcoma cells in vitro at 50 pg/mL, respectively. Epingaione was transformed into eleven derivatives under laboratory conditions using ethanol, some gave greater anti-proliferation/cytotoxic activity on the cancer cell lines tested. One of the derivatives (compound 2) with enhanced cytotoxic activity was elucidated as 5'-Ethoxy-5-methyl-5-(4-methyl-2-oxo-pentyl)-2,3,4,5-tetrahydro-5'H-[2,3']bifuranyl-2'-one. Both epingaione and compound 2 caused an accumulation of arrested or dead SH-SY5Y neuroblastoma in the m-phase of the cell cycle as revealed by the m-phase specific marker KE 67.

In vitro anti-proliferation/cytotoxic activity of epingaione and its derivatives on the human SH-SY5Y neuroblastoma and TE-671 sarcoma cells / Actividad citotóxica antiproliferativa in vitro de la epingaiona y sus derivados, sobre el neuroblastoma humano sh-sy5y y las células del sarcoma te-671

Epingaione (4-Methyl-1-(5-methyl-2, 3,4,5-tetrahydro-[2,3']bifuranyl-5-yl)-pentan-2-one) was isolated as one of the major lipophilic secondary metabolites from the leaves and stems of Bontia daphnoides L. The compound gave 79.24and 50.83anti-proliferation/cytotoxic activity on the human SH-SY5Y neuroblastoma and TE-671 sarcoma cells in vitro at 50 pg/mL, respectively. Epingaione was transformed into eleven derivatives under laboratory conditions using ethanol, some gave greater anti-proliferation/cytotoxic activity on the cancer cell lines tested. One of the derivatives (compound 2) with enhanced cytotoxic activity was elucidated as 5'-Ethoxy-5-methyl-5-(4-methyl-2-oxo-pentyl)-2,3,4,5-tetrahydro-5'H-[2,3']bifuranyl-2'-one. Both epingaione and compound 2 caused an accumulation of arrested or dead SH-SY5Y neuroblastoma in the m-phase of the cell cycle as revealed by the m-phase specific marker KE 67.

La epingaiona (4-Metil-1-(5-metil-2,3,4,5-tetrahidro-[2,3']bifuranil-5-il)-pentan-2-uno) fue aislada como uno de los principales metabolitos lipofilicos secundarios de las hojas y tallos de Bontia daphnoides L. El compuesto produjo 79.24 % y 50.83 % de actividad citotóxica/anti-proliferación sobre el neuroblastoma humano SH-SY5Y y las células del sarcoma TE-671 in vitro a 50 µg/mL, respectivamente. La epingaiona fue transformada en once derivados en condiciones de laboratorio, utilizando etanol. Algunos produjeron mayor actividad citotóxica y antiproliferativa sobre las líneas celulares cancerosas sometidas a ensayo. Uno de los derivados (compuesto 2) de elevada actividad citotóxica fue identificado como 5'-Etoxi-5-metil-5-(4-metil-2-oxo-pentil)-2,3,4,5-tetrahidro-5'H- [2,3']bifuranil-2'-uno. Tanto la epingaiona como el compuesto 22 causaron una acumulación de neuroblastomas SH-SY5Y muertos o detenidos en la fase m del ciclo celular, según lo revela el marcador KE 67 específico de la fase m.

HPLC-coupled spectroscopic techniques (UV, MS, NMR) for the structure elucidation of phthalides in Ligusticum chuanxiong.

Ligusticum chuanxiong Hort., a plant that is frequently used in traditional Chinese medicine, has been studied using HPLC-coupled spectroscopic techniques such as HPLC-UV, HPLC-MS as well as HPLC-NMR. With the aid of these modern spectroscopic techniques, the main constituents, namely senkyunolide A, butylphthalide, neocnidilide and Z-ligustilide, have been characterized and identified. Phthalide dimers, present in smaller amounts, have been identified by HPLC-UV and HPLC-MS analysis and compared with reference compounds. Stereochemical features of some phthalide monomers have been determined by detailed spectroscopic studies for the first time.

Scaphopetalone and scaphopetalumate, a lignan and a triterpene ester from Scaphopetalum thonneri.

From the methanol extract of the stem bark of Scaphopetalum thonneri, two new compounds, including one lignan, named scaphopetalone, one new ester of ferulic acid, named scaphopetalumate were isolated together with three known compounds including: two coumarins (scopoletin and scopolin), and one pentacyclic triterpene (oleanolic acid). The structure of the new compounds were elucidated by means of spectroscopic analyses.

Escuside, a new coumarin-secoiridoid from Fraxinus ornus bark.

A new coumarin-secoiridoid, named escuside, was isolated from the bark of Fraxinus ornus and its structure determined by spectroscopic methods.

Isolation from Cussonia barteri of 1'-O-chlorogenoylchlorogenic acid and 1'-O-chlorogenoylneochlorogenic acid, a new type of quinic acid esters.

1'-O-Chlorogenoylchlorogenic acid and 1'-O-chlorogenoylneochlorogenic acid, a new type of quinic acid esters, have been isolated, in addition to six known quinic acid esters, rutin, and a mixture of saponins, from the methanol extract of Cussonia barteri Seemann (Araliaceae) leaves collected in Cameroon. Structure determination was achieved by NMR, mass, IR, and UV spectroscopy. All compounds were tested for inhibitory activity on 5-lipoxygenase and cyclooxygenase-1, for antimicrobial activity against Bacillus subtilis, Pseudomonas fluorescens, and Cladosporium cucumerinum, and for haemolytic activity.

Isolation and identification of N-mercapto-4-formylcarbostyril, an antibiotic produced by Pseudomonas fluorescens.

Pseudomonas fluorescens strain G308 isolated from barley leaves produces a novel antibiotic substance that was purified by preparative TLC and HPLC and identified as N-mercapto-4-formylcarbostyril (Cbs) by LC/DAD, IR, LC-ES(+)/MS, LC-ES(-)/MS, GC-EI/MS, LC-HRES(+)/MS, mass isotope ratios analysis, 1H NMR and 13C NMR analysis. The purified new antibiotic compound is effective against many phytopathogenic fungi in vitro. The compound inhibited at 25 ppm spore germination and germ tube growth of the following fungi; Fusarium oxysporum f. sp. lycopersici, Fusarium culmorum, Cladosporium cucumerinum and Colletotrichum lagenarium. At concentrations up to 125 ppm, the compound did not interfere with release of zoospores from sporangia and germination of encysted zoospores of Phytophthora infestans.

Vanillic acid 4-O-beta-D-(6'-O-galloyl) glucopyranoside and other constituents from the bark of Terminalia macroptera Guill.et Perr.

A new phenolic glucoside gallate, vanillic acid 4-O-beta-D-(6'-O-galloyl) glucopyranoside (1) was isolated from the bark of Terminalia macroptera Guill.et Perr., together with 3,3',4'-tri-O-methylellagic acid (2) and two triterpene glucopyranosyl esters, 24-deoxysericoside (3) and chebuloside II (4). Compounds 2-4, not described previously for this plant, showed antimicrobial activities against Bacillus subtilis, while 3 and 4 possessed haemolytic properties. In both assays 1 was found to be inactive.

Diterpenoid and limonoids from the stem of Pterorhachis zenkeri.

One new diterpenoid, methyl 3alpha-hydroxy-7-oxo-dehydroabietate (1), two new limonoids, 3alpha-deacetyl-amoorastatin (2) and 9beta-amoorastatin (3), and the known limonoid amoorastatin (4) were isolated from the stem of Pterorhachis zenkeri.

Sesquiterpene lactones and a myoinositol from glandular trichomes of Viguiera quinqueremis (Heliantheae; Asteraceae).

The extract of the floral parts of Viguiera quinqueremis afforded, in addition to known compounds, six new sesquiterpene lactones as well as a new myoinositol derivative. All compounds were detected in glandular trichomes which were collected micromechanically from the anther appendages and were analyzed by HPLC. Structure identification was performed by 1H NMR measurements including LC NMR and LC MS experiments.

Isolation and frontier molecular orbital investigation of bioactive quinone-methide triterpenoids from the bark of Salacia petenensis.

The crude dichloromethane bark extract of Salacia petenensis (Hippocrateaceae) from Monteverde, Costa Rica, shows antibacterial and cytotoxic activity. Bioactivity-directed separation led to the isolation of tingenone and netzahualcoyonol as the biologically active materials. Also isolated from the extract were 3-methoxyfriedel-2-en-1-one (a new natural product) and 29-hydroxyfriedelan-3-one. The structures of these compounds were elucidated on the basis of NMR spectral analysis. Molecular orbital calculations have been carried out using the semi-empirical PM3 and Hartee-Fock 3-21G ab initio techniques on the quinone-methide nortriterpenoids tingenone and netzahualcoyonol, as well as on the nucleotide bases adenine, guanine, cytosine, and thymine. The molecular orbital calculations suggest that a possible mode of cytotoxic action of quinone-methide triterpenoids involves quasi-intercalative interaction of the compounds with DNA followed by nucleophilic addition of the DNA base to carbon-6 of the triterpenoid.