Acetylcholinesterase activity in the brain of wild birds in Korea-2014 to 2016.

Acetylcholinesterase (AChE) activity level can be used as a diagnostic marker for anticholinesterase pesticide poisoning. In this study, we aimed to establish a baseline level of normal brain AChE activity in wild birds. AChE activity was measured in the brains of 87dead wild birds (26 species). The level of AChE activity ranged from 6.40 to 15.9 µmol/min/g of brain tissue in normal wild birds. However, the brain tissue AChE activity level in wild birds exposed to organophosphate (OP) pesticide was 48.0%-96.3% of that in the normal birds. These results may serve as reference values to facilitate routine diagnosis and monitoring of OP-poisoned wild birds.

Negative regulation of the RLR-mediated IFN signaling pathway by duck ubiquitin-specific protease 18 (USP18).

Ubiquitin-specific protease 18 (USP18) plays an important role in regulating type I interferon (IFN) signaling in innate immunity, and has a crucial impact on the IFN therapeutic effect. Although significant progress has been made in elucidating USP18 function in mammals, the role of USP18 in ducks (duUSP18) remains poorly understood. In this study, we cloned the USP18 gene from white crested ducks by reverse transcription polymerase chain reaction (RT-PCR) and rapid amplification of complementary DNA (cDNA) ends. We determined that duUSP18 cDNA contains a 52-bp 5'UTR, a 1,131-bp open reading frame and a 356-bp 3'UTR, and encodes a 376-amino acid protein. Multiple sequence alignments showed that duUSP18 shares high similarity with USP18 from other vertebrates. Overexpression of duUSP18 inhibited nuclear factor-κB (NF-κB) and interferon regulatory factor 1 (IRF1) activity, and reduced IFN-ß production following 5' triphosphate double-stranded RNA (5'ppp dsRNA) or lipopolysaccharide (LPS) stimulation. duUSP18 knockdown significantly activated 5'ppp dsRNA-induced and LPS-induced NF-κB and IRF1 activation, and induced IFN-ß expression in duck embryo fibroblasts. Furthermore, Quantitative real-time PCR (qRT-PCR) revealed that overexpression or knockdown of duUSP18 could alter the expression of genes related to the RLR-mediated IFN signaling pathway following the treatment with 5'ppp dsRNA. In addition, site-directed mutation analysis revealed that cysteine 66 (C66), histidine 313 (H313), and histidine 321 (H321) of duUSP18 were critical for inhibiting IFN-ß activity. Taken together, these results suggest that duck USP18 plays an important role in innate immune responses against double-stranded RNA viruses in the RLR-mediated IFN signaling pathway, and that further studies are warranted to elucidate its underlying mechanisms, which could provide molecular insights into the effect of the treatment of duck diseases.

Conserved Active-Site Residues Associated with OAS Enzyme Activity and Ubiquitin-Like Domains Are Not Required for the Antiviral Activity of goOASL Protein against Avian Tembusu Virus.

Interferon (IFN)-induced 2'-5'-oligoadenylate synthetase (OAS) proteins exhibit an extensive and efficient antiviral effect against flavivirus infection in mammals and birds. Only the 2'-5'-oligoadenylate synthetase-like (OASL) gene has been identified thus far in birds, except for ostrich, which has both OAS1 and OASL genes. In this study, we first investigated the antiviral activity of goose OASL (goOASL) protein against a duck-origin Tembusu virus (DTMUV) in duck embryo fibroblast cells (DEFs). To investigate the relationship of conserved amino acids that are related to OAS enzyme activity and ubiquitin-like (UBL) domains with the antiviral activity of goOASL, a series of mutant goOASL plasmids was constructed, including goOASL-S64C/D76E/D78E/D144T, goOASL∆UBLs and goOASL∆UBLs-S64C/D76E/D78E/D144T. Interestingly, all these mutant proteins significantly inhibited the replication of DTMUV in DEFs in a dose-dependent manner. Immunofluorescence analysis showed that the goOASL, goOASL-S64C/D76E/D78E/D144T, goOASL∆UBLs and goOASL∆UBLs-S64C/D76E/D78E/D144T proteins were located not only in the cytoplasm where DTMUV replicates but also in the nucleus of DEFs. However, the goOASL and goOASL mutant proteins were mainly colocalized with DTMUV in the cytoplasm of infected cells. Our data indicated that goOASL could significantly inhibit DTMUV replication in vitro, while the active-site residues S64, D76, D78 and D144, which were associated with OAS enzyme activity, the UBL domains were not required for the antiviral activity of goOASL protein.

Resident Microbiome Disruption with Antibiotics Enhances Virulence of a Colonizing Pathogen.

There is growing evidence that symbiotic microbes play key roles in host defense, but less is known about how symbiotic microbes mediate pathogen-induced damage to hosts. Here, we use a natural wildlife disease system, house finches and the conjunctival bacterial pathogen Mycoplasma gallisepticum (MG), to experimentally examine the impact of the ocular microbiome on host damage and pathogen virulence factors during infection. We disrupted the ocular bacterial community of healthy finches using an antibiotic that MG is intrinsically resistant to, then inoculated antibiotic- and sham-treated birds with MG. House finches with antibiotic-disrupted ocular microbiomes had more severe MG-induced conjunctival inflammation than birds with unaltered microbiomes, even after accounting for differences in conjunctival MG load. Furthermore, MG cultures from finches with disrupted microbiomes had increased sialidase enzyme and cytadherence activity, traits associated with enhanced virulence in Mycoplasmas, relative to isolates from sham-treated birds. Variation in sialidase activity and cytadherence among isolates was tightly linked with degree of tissue inflammation in hosts, supporting the consideration of these traits as virulence factors in this system. Overall, our results suggest that microbial dysbiosis can result in enhanced virulence of colonizing pathogens, with critical implications for the health of wildlife, domestic animals, and humans.

Avian reovirus σA and σNS proteins activate the phosphatidylinositol 3-kinase-dependent Akt signalling pathway.

The present study was conducted to identify avian reovirus (ARV) proteins that can activate the phosphatidylinositol 3-kinase (PI3K)-dependent Akt pathway. Based on ARV protein amino acid sequence analysis, σA, σNS, µA, µB and µNS were identified as putative proteins capable of mediating PI3K/Akt pathway activation. The recombinant plasmids σA-pcAGEN, σNS-pcAGEN, µA-pcAGEN, µB-pcAGEN and µNS-pcAGEN were constructed and used to transfect Vero cells, and the expression levels of the corresponding genes were quantified by immunofluorescence and Western blot analysis. Phosphorylated Akt (P-Akt) levels in the transfected cells were measured by flow cytometry and Western blot analysis. The results showed that the σA, σNS, µA, µB and µNS genes were expressed in Vero cells. σA-expressing and σNS-expressing cells had higher P-Akt levels than negative control cells, pcAGEN-expressing cells and cells designed to express other proteins (i.e., µA, µB and µNS). Pre-treatment with the PI3K inhibitor LY294002 inhibited Akt phosphorylation in σA- and σNS-expressing cells. These results indicate that the σA and σNS proteins can activate the PI3K/Akt pathway.

Pigeon RIG-I Function in Innate Immunity against H9N2 IAV and IBDV.

Retinoic acid-inducible gene I (RIG-I), a cytosolic pattern recognition receptor (PRR), can sense various RNA viruses, including the avian influenza virus (AIV) and infectious bursal disease virus (IBDV), and trigger the innate immune response. Previous studies have shown that mammalian RIG-I (human and mice) and waterfowl RIG-I (ducks and geese) are essential for type I interferon (IFN) synthesis during AIV infection. Like ducks, pigeons are also susceptible to infection but are ineffective propagators and disseminators of AIVs, i.e., "dead end" hosts for AIVs and even highly pathogenic avian influenza (HPAI). Consequently, we sought to identify pigeon RIG-I and investigate its roles in the detection of A/Chicken/Shandong/ZB/2007 (H9N2) (ZB07), Gansu/Tianshui (IBDV TS) and Beijing/CJ/1980 (IBDV CJ-801) strains in chicken DF-1 fibroblasts or human 293T cells. Pigeon mRNA encoding the putative pigeon RIG-I analogs was identified. The exogenous expression of enhanced green fluorescence protein (EGFP)-tagged pigeon RIG-I and caspase activation and recruitment domains (CARDs), strongly induced antiviral gene (IFN-ß, Mx, and PKR) mRNA synthesis, decreased viral gene (M gene and VP2) mRNA expression, and reduced the viral titers of ZB07 and IBDV TS/CJ-801 virus strains in chicken DF-1 cells, but not in 293T cells. We also compared the antiviral abilities of RIG-I proteins from waterfowl (duck and goose) and pigeon. Our data indicated that waterfowl RIG-I are more effective in the induction of antiviral genes and the repression of ZB07 and IBDV TS/CJ-801 strain replication than pigeon RIG-I. Furthermore, chicken melanoma differentiation associated gene 5(MDA5)/ mitochondrial antiviral signaling (MAVS) silencing combined with RIG-I transfection suggested that pigeon RIG-I can restore the antiviral response in MDA5-silenced DF-1 cells but not in MAVS-silenced DF-1 cells. In conclusion, these results demonstrated that pigeon RIG-I and CARDs have a strong antiviral ability against AIV H9N2 and IBDV in chicken DF-1 cells but not in human 293T cells.

Combined effect of short-term dehydration and sublethal acute oral dicrotophos exposure confounds the diagnosis of anticholinesterase exposure in common quail (Coturnix coturnix) using plasma cholinesterase activity.

Common Quail (Coturnix coturnix) were subjected to controlled and replicated experiments in the summer of 2008 to investigate the effects of short-term dehydration on cholinesterase activity in brain and plasma and the interaction between dehydration and exposure to the organophosphorus pesticide dicrotophos in these same tissues. Our objective was to determine if dehydration could confound the diagnosis of anticholinesterase exposure using inhibition of cholinesterase activity in quail tissues. The effect of dehydration was quantified using measures of plasma osmolality and hematocrit. Dicrotophos exposure caused significant inhibition of cholinesterase activity in brain, while the effects of dehydration and interaction were not significant. Dehydration caused significant duration-dependent increases in plasma osmolality and hematocrit. Dehydration also caused a significant increase in plasma cholinesterase activity. Variation in the change in plasma cholinesterase activity in response to dehydration was significantly and positively correlated with dehydration-induced variation in both the change in plasma osmolality and the change in hematocrit. These correlations suggest that plasma cholinesterase activity in quail is not limited to plasma but occupies some larger pool of the extracellular fluid volume, and we suggest lymph is part of that pool. The effects of dehydration on plasma cholinesterase activity masked the inhibitory effects of dicrotophos. Here, the combination of dehydration and dicrotophos exposure produced plasma cholinesterase activity that was not significantly different from reference and pre-exposure values, confounding the diagnosis of anticholinesterase exposure in dehydrated, dicrotophos-exposed quail. A method to adjust plasma cholinesterase activities for the confounding effects of dehydration and enable the diagnosis of anticholinesterase exposure in dehydrated, dicrotophos-exposed quail was developed. Clinicians and practitioners responsible for the diagnosis of anticholinesterase exposure in birds are cautioned that dehydration, commonly observed in sick wildlife, may mask the effect of anticholinesterases on plasma cholinesterase activity.

Characterization of turkey inducible nitric oxide synthase and identification of its expression in the intestinal epithelium following astrovirus infection.

The inducible nitric oxide synthase (iNOS) enzyme has long been recognized as a key mediator of innate immune responses to infectious diseases across the phyla. Its role in killing or inactivating bacterial, parasitic, and viral pathogens has been documented in numerous host systems. iNOS, and its innate immune mediator NO has also been described to have negative consequence on host tissues as well; therefore understanding the pathogenesis of any infectious agent which induces iNOS expression requires a better understanding of the role iNOS and NO play in that disease. Previous studies in our laboratory and others have demonstrated evidence for increased levels of iNOS and activity of its innate immune mediator NO in the intestine of turkeys infected with astrovirus. To begin to characterize the role iNOS plays in the innate immune response to astrovirus infection, we identified, characterized, developed tkiNOS specific reagents, and demonstrated that the intestinal epithelial cells induce expression of iNOS following astrovirus infection. These data are the first to our knowledge to describe the tkiNOS gene, and demonstrate that astrovirus infection induces intestinal epithelial cells to express iNOS, suggesting these cells play a key role in the antiviral response to enteric infections.

Candida species isolated from the gastrointestinal tract of cockatiels (Nymphicus hollandicus): In vitro antifungal susceptibility profile and phospholipase activity.

Over the past years, the incidence of yeast infections, especially candidiasis, has increased. It is known that birds, including cockatiels, harbor potentially pathogenic yeasts to human beings in their gastrointestinal tract. Thus, this work aims at determining the in vitro antifungal susceptibility and phospholipase activity of Candida spp. isolated from the gastrointestinal tract and stools of cockatiels. Sixty cockatiels were assessed and samples were collected from oral cavity, crop and cloaca and stools were collected from cages where birds were kept. Yeast species were identified according to morphological and biochemical characteristics. Amphotericin B, itraconazole and fluconazole were tested against 39 C. albicans; 12 C. tropicalis; 7 C. parapsilosis and 1 C. krusei, through broth microdilution test. These same isolates were also tested for phospholipase production, on egg yolk agar. For amphotericin B, itraconazole and fluconazole, MICs were 0.25-1 µg/mL, 0.03125 to ≥16 µg/mL and 0.5 to ≥64 µg/mL, respectively, and resistance to itraconazole and fluconazole was observed in 14 (35.89%) and 4 (10.26%) C. albicans isolates, respectively. All C. albicans were positive for phospholipase production, out of which 74.36% presented high enzymatic activity. Among non-albicans Candida species, 40% produced phospholipase. The results show that cockatiels might represent a hazard to human health, as sources of infections caused by resistant Candida spp., especially to immunocompromised individuals, children and elderly.

Superoxide dismutase expression and oxidative damage in a case of myopathy in brown pelicans (Pelecanus occidentalis).

Four brown pelicans (Pelecanus occidentalis) housed at a rehabilitation facility were found dead after a 3-day history of muscle weakness and after being fed for about 2 weeks from a recent shipment of fish. The birds had pale streaking of the skeletal and heart muscles. Microscopically, the skeletal muscle, and to a lesser extent the cardiac muscle, had severe myocyte degeneration and necrosis characterized by microvacuolation with loss of cross-striations, condensation of cytoplasm, fragmentation, mineralization, and inflammatory cell infiltrates consisting of multinucleated cells, macrophages, and few heterophils. The findings were consistent with myopathy, and a nutritional myopathy caused by eating rancid fish was suspected. Immunohistochemical staining revealed abundant immunoreactive copper zinc superoxide dismutase and manganese superoxide dismutase either as diffuse homogeneous precipitates or granular aggregates in the cytoplasm of affected cells. Immunoreactivity was directly related to degree of cellular damage as estimated by light microscopic examination. We suggest that the lack of protection, despite upregulation of superoxide dismutase, is most likely attributable to supersaturation of oxidants beyond the capacity of superoxide dismutases to scavenge.

Successful treatment of capture myopathy in three wild greater sandhill cranes (Grus canadensis tabida).

Two adult and 1 juvenile free-flying greater sandhill cranes (Grus canadensis tabida) were diagnosed with capture myopathy after alpha-chloralose baiting and physical capture during a banding and feeding ecologic study. Blood samples were collected for serum biochemical analysis at the time of capture for the 2 adults, and at 24 hours postcapture, at various intervals during treatment, and at the time of release for all 3 birds. Concentrations of creatine kinase, aspartate transaminase, and lactate dehydrogenase were high within 1 hour of capture and peaked approximately 3 days after capture. By days 10-17 after capture, creatine kinase and lactate dehydrogenase concentrations both decreased to within the reference range measured for cranes at capture, but aspartate transaminase concentrations remained 2-5 times higher than the measured reference range. Treatment consisted of corticosteroids, selenium/vitamin E, parenteral fluids, and gavage feedings. Physical therapy consisted of assisting the cranes to stand and walk 2-8 times a day, massaging leg muscles, and moving limbs manually through the range of motion. The adults were released when they were able to stand up independently and were pacing in the pen. The juvenile was released 12 hours after it was able to stand independently but was returned to the pen when it fell and could not rise. It was treated supportively for an additional 3 days and then successfully released. Both adult cranes were observed on their territories with their original mates after release and returned to their territories for the subsequent 8 years, raising chicks most years. After release, the juvenile was observed in a flock of cranes near its natal territory for the next 2 days.

Activity of selected hydrolases in excretion-secretion products and extracts of adult Contracaecum rudolphii.

BACKGROUND: Enzymes contained in excretion-secretion (ES) products of parasites released to the environment play multiple roles: they facilitate hatching and moulting of larvae, enable a parasite to migrate within tissues, inhibit blood coagulation, defend the parasite from host's immunological response, and enhance feeding and nutrition. The aim of the study was to determine hydrolase activity in ES products and extracts from adult Contracaecum rudolphii. MATERIALS AND METHODS: Adult nematodes were isolated from intestines of black cormorants nesting at Katy Rybackie (the Vistula Lagoon). Nematode batches of 30 individuals each were placed in 5 ml portions of antibiotic-enriched physiological salt solution and incubated for 24 hours at 37 degrees C. After incubation, the solutions containing ES products were collected and dialysed for 24 h at 4 degrees C against distilled water. Extracts were obtained by homogenising the nematodes with the physiological salt solution (0.9% NaCl). The homogenate was centrifuged for 10 minutes at 3000xg. Enzyme activities were assayed in the supernatant. The enzymatic activity in ES products and homogenates was determined with the API ZYM kit (Bio Merieux S.A., Lyon, France). Hydrolase activities were expressed in volumetric units (nmol) of the hydrolysed substrate. RESULTS: The nematode ES products showed 10 hydrolases to be active. The highest activity was that of esterases, except for lipase the activity of which was not detected. Among glucosidases, the highest activity was shown by alpha-glucosidase, much lower activities being typical of beta-galactosidase and N-acetyl-beta-glucosaminidase. The remaining glucosidases proved inactive. Among proteases, leucine arylamidase and valine arylamidase were found to be active only. The nematode extracts revealed activities of 15 hydrolases. The highest activity was typical of esterases. Among glucosidases, the highest activity was typical of alpha-glucosidase, beta-glucuronidase and N-acetyl-beta-glucosaminidase. Activities of the remaining glucosidases were much lower. No activity of alpha-galactosidase was detected. Among proteolytic enzymes, leucine arylamidase proved the most active, while activities of valine arylamidase and chymotrypsin were much lower. The remaining proteases revealed no detectable activity. CONCLUSIONS: The ES products of adult C. rudolphii were found to contain active hydrolases which may damage the epithelium lining the host's alimentary tract. Activities of almost all glucosidases in the parasite's extracts suggests that, like in most nematodes, the parasite's main energy source is derived from carbohydrate metabolism.

Avian mortality events in the United States caused by anticholinesterase pesticides: a retrospective summary of National Wildlife Health Center records from 1980 to 2000.

We reviewed the U.S. Geological Survey National Wildlife Health Center (NWHC) mortality database from 1980 to 2000 to identify cases of poisoning caused by organophosphorus and carbamate pesticides. From the 35,022 cases from which one or more avian carcasses were submitted to the NWHC for necropsy, we identified 335 mortality events attributed to anticholinesterase poisoning, 119 of which have been included in earlier reports. Poisoning events were classified as confirmed (n = 205) when supported by findings of > or =50% inhibition of cholinesterase (ChE) activity in brain tissue and the detection of a specific pesticide in the gastrointestinal contents of one or more carcasses. Suspected poisonings (n = 130) were defined as cases where brain ChE activity was > or =50% inhibited or a specific pesticide was identified in gastrointestinal contents. The 335 avian mortality events occurred in 42 states. Washington, Virginia, and Ohio had the highest frequency of events, with 24 (7.2%), 21 (6.3%), and 20 (6.0%) events, respectively. A total of 8877 carcasses of 103 avian species in 12 orders was recovered. Because carcass counts underestimate total mortality, this represents the minimum actual mortality. Of 24 different pesticides identified, the most frequent were famphur (n = 59: 18%), carbofuran (n = 52; 15%), diazinon (n = 40; 12%), and fenthion (n = 17; 5.1%). Falconiformes were reported killed most frequently (49% of all die-offs) but Anseriformes were found dead in the greatest numbers (64% of 8877 found dead). The majority of birds reported killed by famphur were Passeriformes and Falconiformes, with the latter found dead in 90% of famphur-related poisoning events. Carbofuran and famphur were involved in mortality of the greatest variety of species (45 and 33, respectively). Most of the mortality events caused by diazinon involved waterfowl.

Differential toxicities of organophosphate and carbamate insecticides in the nestling European starling (Sturnus vulgaris).

The concept of B-esterase buffering against anti-cholinesterase (ChE) insecticide toxicity has been extensively researched in mammalian species. Presumably due to relatively low levels of anti-ChE detoxifying enzyme activity in birds, however, avian species are often more susceptible to the toxic effects of these compounds. We quantified B-esterase buffering of organophosphate (diazinon and methyl parathion) and carbamate (aldicarb and oxamyl) toxicity in nestling European starlings (Sturnus vulgaris). The differential toxicities were studied using mortality, behavioral observation, and inhibitor affinity data. The toxicities of diazinon, methyl parathion, and oxamyl were affected by the removal of butyrylcholinesterase (BChE) using the specific inhibitor tetraisopropylpyrophosphoramide (iso-OMPA). When BChE was absent, aldicarb toxicity was not affected. Theoretically, compounds affected by BChE removal would have a higher affinity for BChE or carboxylesterase (CaE) than acetylcholinesterase (AChE). However, this was only the case for diazoxon, which had a 1,000-fold higher affinity for plasma BChE and CaE than AChE. Methyl paraoxon and aldicarb had a higher affinity for plasma AChE than for BChE or CaE. Oxamyl had similar IC50 values for all three enzymes studied. The generation of IC50 curves for each inhibitor revealed the presence of nonsensitive forms of CaE in both the plasma and brain. Based on the results of this research, there appears to be no strict correlation between mortality data and inhibitor affinities for each esterase that alone can explain the differential toxicities of these compounds.

[Plasma cholesterol determination in birds--a diagnostic tool for detection of organophosphate and carbamate intoxication]. / Plasma-Cholinesterase-Bestimmung bei Vögeln--ein diagnostisches Hilfsmittel zum Nachweis von Intoxikationen mit Organophosphaten und Carbamaten.

An investigation was done on the clinical usefulness of the dry chemistry analyzer Vitros DT 60 II for determination of avian plasma cholinesterase. The analytical reliability of the method, evaluated by precision and accuracy, proved to be high for plasma of numerous pet and wild birds. Values of normal plasma-cholinesterase activity were established for different psittacine and European wild birds. Significant differences in physiologic plasma-cholinesterase activity were noted between closely related species as well as between juvenile and adult birds. These findings emphasize the necessity to use control values of the same species and age group for comparison. Dry chemistry plasma-cholinesterase determination can be used as a diagnostic tool for detection of organophosphate and carbamate poisonings in the majority of investigated birds.

Transfection of avian LMH-2A hepatoma cells with cationic lipids.

LMH-2A is an estrogen-responsive avian hepatoma cell line whose susceptibility to cationic-lipid-mediated transfection is poorly described. 3 beta[N-N',N'-dimethylaminoethane)-carbamoyl] cholesterol (DCC) requires a one-step synthesis, and can be used to formulate transfection-grade liposomes when combined with dioleoylphosphatidyl-ethanolamine (DOPE) 1/1 (wt/wt). Luciferase activities in LMH-2A cells were 8.5-fold and 87.5-fold greater than those in HepG2 and FTO2B cells, respectively, following DCC-liposome-mediated transfection with a reporter consisting of the human cytomegalovirus immediate-early promoter (CMV), joined to Photinus pyralis luciferase (L) cDNA, designated pCMVL. Using pCMVL, N-(2-bromoethyl)-N,N-dimethyl-2,3-bis(9-octadecenyloxy)-propana minimun bromide) (BMOP)/DOPE 1/1 (wt/wt), at a 7.5:1 ratio with DNA, produced luciferase activities that were 2.9-fold higher than those of DCC-liposomes, at an optimal 10:1 lipid:DNA ratio. At optimal lipid:DNA ratios, commercially available liposomes, Transfectam, Lipofectamine, and Lipofectin, produced luciferase activities that were 1.39, 1.03, and 0.47-fold those of DCC-liposomes. The effect of 0, 10, 100, or 500 nM/L 17 beta-estradiol on the expression of pCMVL and a second luciferase reporter containing the -593/+48 promoter region of the estrogen-responsive avian apo VLDL-II gene, designated pApoL, was tested in cells cultured in the presence or absence of 10% chicken serum. The CMV promoter supported a high level of expression in LMH-2A cells that was unaffected by serum alone, but was weakly responsive to estrogen. Estrogen responses of both reporters reached a plateau at 10 nM/L. Estrogen increased the expression of pApoL 24-fold and 79-fold in the absence and presence of serum, respectively. The -593/+48 region of the apo VLDL-II promoter may not contain previously reported negative insulin response elements, but chicken serum contains factors that enhance estrogen responsiveness of this region.

Effects of the intestinal flagellate, Cochlosoma anatis, on intestinal mucosal morphology and disaccharidase activity in Muscovy ducklings.

Newly hatched female Muscovy ducklings were randomly separated into 2 groups of 12 and 1 group of 13. Ducklings in the first 2 groups were each orally inoculated with 0.5 ml of sterile normal saline containing 0 and 3 x 10(6) trophozoites of Cochlosoma anatis, respectively. Birds in the third group were each orally inoculated with 3 x 10(6) trophozoites for 5 consecutive days. Birds were weighed daily for the first 5 days and then on days 7, 14 and 21 post-inoculation (p.i.). On days 6, 7, 8, 13, 14, 15, 20, 21 and 22 p.i., 1 bird from each group was killed and samples of intestine at 7 levels were taken for trophozoite counts, mucosal disaccharidase analyses and morphometric analysis. Body weights did not differ among treatment groups at any time during the experiment. Trophozoite numbers did not change over the period 6-22 days p.i. Trophozoite numbers were lowest in the anterior small intestine and increased distally, but very few were observed in the caecum. Crypt depth was greater in all regions of the small intestine in inoculated groups compared to uninoculated controls, and was significantly increased in the duodenum, proximal jejunum and mid-jejunum (P < 0.05). Villus height was greater in inoculated groups compared to controls at all levels of the intestine and was significantly increased in the duodenum, proximal jejunum and ileum (P < 0.05). Mucosal palatinase and maltase activity in the small intestine were reduced in inoculated groups compared to uninoculated controls; palatinase activities were significantly reduced in the proximal and mid-jejunum and maltase activities were significantly reduced in the mid-jejunum (P < 0.05). Sucrase activities were significantly increased at all levels of the small intestine in inoculated ducklings compared to uninoculated controls (P < 0.05). Although no clinical signs were evident, Cochlosoma infection significantly altered intestinal morphometrics and mucosal enzyme concentrations in ducklings, in several cases in a counter-intuitive direction.

Gangliosidosis in emus (Dromaius novaehollandiae).

A 6-month-old female emu (Dromaius novaehollandiae) died following acute central nervous system signs. Hematoxylin-and-eosin-stained sections revealed that neurons of the brain were distended with nonstaining 1-to-2-microns vacuoles. Ultrastructural examination of the affected neurons revealed numerous membranous cytoplasmic bodies (MCBs) similar in appearance to the MCBs seen in mammalian gangliosidoses. A full sibling of this emu was donated for study. This 7-month-old female emu was stunted compared with hatchmates. Neurologic examination revealed hypermetric gait, persistent head tremor, and mild ataxia. No gross lesions were evident at postmortem. Histopathologic and electron microscopic findings were similar to those in the index case in that swollen, pale neurons were present in the cerebrum, pons, medulla, cerebellum, spinal cord, spinal ganglia, autonomic ganglia, myenteric plexus, and ganglion cell layer of the retina. Analysis of brain gangliosides of the affected 7-month-old emu revealed 14- and 25-fold increases of GM1 and GM3 gangliosides, respectively, compared with control emus. The total brain ganglioside sialic acids were, on a wet weight basis, 519 micrograms/g (control A), 658 micrograms/g (control B), and 1800 micrograms/g (affected emu). The familial association seen with this condition suggests that emus are affected by an inherited disorder similar to mammalian gangliosidoses.

Serum enzymes as indicators of capture myopathy in mallards (Anas platyrhynchos).

Serum concentrations of the enzymes creatine kinase (CK) and aspartate aminotransferase (AST) were measured in captive and wild mallards (Anas platyrhynchos) as indicators of muscle damage. Baseline values for both enzymes were determined from six captive male mallards. During winter 1990 to 1991, six diets (including controls) representative of food available in the Mississippi alluvial valley were fed to captive female mallards housed in an outdoor aviary at the White River National Wildlife Refuge, Arkansas County, Arkansas (USA). Controlled handling of penned mallards resulted in elevated serum CK (means = 1,352 IU/liter; SD = 1,212) and AST (means = 101 IU/liter; SD = 95) concentrations consistent with myopathies. These serum enzyme elevations were not affected (P > 0.3) by dietary selenium concentrations in the six diets or by energy malnutrition suffered by birds fed soybeans. Capture of wild mallards with an entanglement type rocket net resulted in serum CK and AST concentrations (means = 12,035 and 330 IU/liter; SD = 8,125 and 171, respectively) that were higher (P < 0.001) than those reported after capture with an enveloping type rocket net. Baseline values, controlled handling values, and entanglement rocket net values for serum CK and AST all differed (P < 0.0001).