Evaluation of antigen-capture ELISA and immunohistochemical methods for avian surveillance of West Nile virus.
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Accurate detection of West Nile virus (WNV) in corvids is essential for monitoring the spread of virus during the mosquito season. Viremia in corvids is very high, with titers approaching 10(8) viral particles/ml. In the presence of such marked viremia, the sensitivity of real-time reverse transcription-polymerase chain reaction (RT-PCR) analysis is unnecessary, and more cost-effective methods should be assessed. To this end, antigen-capture ELISA (ACE) and immunohistochemical (IHC) assays were evaluated. Skin, cloacal swab specimens, and feathers from corvids were tested by use of ACE, and results were compared with results obtained from use of real-time RT-PCR analysis. Of the 3 sample types, skin gave the best sensitivity (98%) and specificity (100%). Skin, brain, kidney, and spleen from corvids were analyzed by IHC, and results were compared with real-time RT-PCR results. Kidney and spleen were more often positive by use of IHC than were brain and skin tissue; however, IHC did not perform as well as ACE in the identification of virus-positive birds. Results of this study support the use of a skin sample in an ACE format as an effective surveillance method for corvids. (+info)
Sustained Bmp signaling is essential for cloaca development in zebrafish.
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Bone morphogenetic protein (Bmp) signaling has long been known to be important for the early development of the ventral mesoderm, including blood, vasculature and kidney cells. Although Bmp genes are continually expressed in the ventral cells throughout gastrulation and somitogenesis, previous studies in zebrafish have not addressed how the role of Bmp signaling changes over time to regulate ventral mesoderm development. Here, we describe the use of a transgenic inducible dominant-negative Bmp receptor line to examine the temporal roles of Bmp signaling in ventral mesoderm patterning. Surprisingly, we find that Bmp signaling from the mid-gastrula stage through early somitogenesis is important for excluding blood and vascular precursors from the extreme ventral mesoderm, and we show that this domain is normally required for development of the cloaca (the common gut and urogenital opening). Using a novel assay for cloacal function, we find that larvae with reduced mid-gastrula Bmp signaling cannot properly excrete waste. We show that the cloacal defects result from alterations in the morphogenesis of the cloaca and from changes in the expression of genes marking the excretory system. Finally, we show that HrT, a T-box transcription factor, is a Bmp-regulated gene that has an essential function in cloacal development. We conclude that sustained Bmp signaling plays an important role in specification of the zebrafish cloaca by maintaining the fate of extreme ventral cells during the course of gastrulation and early somitogenesis. Furthermore, our data suggest that alterations in Bmp signaling are one possible cause of anorectal malformations during human embryogenesis. (+info)
The immunogenicity and efficacy against H5N1 challenge of reverse genetics-derived H5N3 influenza vaccine in ducks and chickens.
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H5N1 avian influenza viruses are continuing to spread in waterfowl in Eurasia and to threaten the health of avian and mammalian species. The possibility that highly pathogenic (HP) H5N1 avian influenza is now endemic in both domestic and migratory birds in Eurasia makes it unlikely that culling alone will control H5N1 influenza. Because ducks are not uniformly killed by HP H5N1 viruses, they are considered a major contributor to virus spread. Here, we describe a reverse genetics-derived high-growth H5N3 strain containing the modified H5 of A/chicken/Vietnam/C58/04, the N3 of A/duck/Germany/1215/73, and the internal genes of A/PR/8/34. One or two doses of inactivated oil emulsion vaccine containing 0.015 to 1.2 microg of HA protein provide highly efficacious protection against lethal H5N1 challenge in ducks; only the two dose regimen has so far been tested in chickens with high protective efficacy. (+info)
Oblique pelvic osteotomy in the exstrophy/epispadias complex.
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We reviewed retrospectively 45 patients (46 procedures) with bladder exstrophy treated by bilateral oblique pelvic osteotomy in conjunction with genitourinary repair. The operative technique and post-operative management with or without external fixation are described. A total of 21 patients attended a special follow-up clinic and 24 were interviewed by telephone. The mean follow-up time was 57 months (24 to 108). Of the 45 patients, 42 reported no pain or functional disability, although six had a waddling gait and two had marked external rotation of the hip. Complications included three cases of infection and loosening of the external fixator requiring early removal with no deleterious effect. Mid-line closure failed in one neonate managed in plaster. This patient underwent a successful revision procedure several months later using repeat osteotomies and external fixation. The percentage pubic approximation was measured on anteroposterior radiographs pre-operatively, post-operatively and at final follow-up. The mean approximation was 37% (12% to 76%). It varied markedly with age and was better when external fixation was used. The wide range reflects the inability of the anterior segment to develop naturally in spite of close approximation at operation. We conclude that bilateral oblique pelvic osteotomy with or without external fixation is useful in the management of difficult primary closure in bladder exstrophy, failed primary closure and secondary reconstruction. (+info)
Impairment of testicular function in adult male Japanese quail (Coturnix japonica) after a single administration of 3-methyl-4-nitrophenol in diesel exhaust particles.
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The effects of 3-methyl-4-nitrophenol (PNMC), a component of diesel exhaust, on reproductive function were investigated in adult male Japanese quail. The quail were treated with a single i.m. dose of PNMC (78, 103 or 135 mg/kg body weight), and trunk blood and testes were collected 1, 2 or 4 weeks later. Various levels of testicular atrophy were observed in all groups treated with PNMC. Sperm formation, cloacal gland area, and plasma LH and testosterone concentrations were also reduced in birds with testicular atrophy. To determine the acute effect of PNMC on gonadotrophin from the pituitary, adult male quail were administrated a single i.m. injection of PNMC (25 mg/kg), and plasma concentrations of LH were measured at 1, 3 and 6 h. This dose significantly lowered plasma levels of LH at all three time points. These results suggest that PNMC acts on the hypothalamus-pituitary axis, by reducing circulating LH within a few hours of administration and subsequently reducing testosterone secretion. In addition, in order to investigate the direct effects of PNMC on the secretion of testosterone from testicular cells in quail testes, cultured interstitial cells containing Leydig cells were exposed to PNMC (10(-6), 10(-5) or 10(-4) M) for 4, 8 or 24 h. These quantities of PNMC significantly reduced the secretion of testosterone in a time- and dose-dependent manner. The present findings also suggest a direct effect of PNMC on the testis to reduce testosterone secretion. This study clearly indicates that PNMC induces reproductive toxicity at both the central and testicular levels, and disrupts testicular function in adult male quail. (+info)
Staging of intestinal development in the chick embryo.
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Comparisons between developmental studies rely on embryonic staging systems. It is important for comparison of molecular, immunohistochemical, and physiological studies of the developing chick intestine that the developmental stage of embryos is reliably determined. Good staging systems exist for the external features of the chick embryo but not for development of internal organs. To facilitate precise comparisons of chick embryo intestine development, we prepared an intestinal staging system. Embryos were fixed, other tissues dissected away, and the intestine and associated organs were then drawn to scale using a camera lucida. This produced black-and-white drawings with features of the gut clearly visible. The detailed drawings of intestine from chick embryos aged 2.5 to 10 days were correlated with age of embryos and developmental stages described in three common staging systems, Hamilton and Hamburger, Thompson and Fitzharris, and Allan and Newgreen. Descriptions of key changes in gut morphology and position are given for each stage. This staging of chick gut development will allow future studies to quote and compare development of the gut rather than external features or incubation time. This will allow much more precise reporting and comparisons in developmental studies of cell migration and gene expression. (+info)
Enhanced early West Nile virus infection in young chickens infected by mosquito bite: effect of viral dose.
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Mosquito transmission of arboviruses potentially affects the course of viral infection in the vertebrate host. Studies were performed to determine if viral infection differed in chickens infected with West Nile virus (WNV) by mosquito bite or needle inoculation. Mosquito-infected chickens exhibited levels of viremia and viral shedding that were up to 1,000 times higher at 6, 12, and 24 hours post-feeding (PF) compared with those inoculated with 10(3) PFU by needle. Follow-up studies were conducted to determine if enhanced early infection was due to a higher viral dose inoculated by mosquitoes. Needle inoculation with successively higher doses of WNV led to higher early viremia and viral shedding; a dose >or= 10(4) PFU by needle was required to attain the high early viremia observed in mosquito-infected chickens. Mosquitoes inoculated WNV at this level as estimated by feeding on a hanging drop of blood (mean: 10(2.5), range: 10(0.7)-10(4.6) PFU). These results indicate that enhanced early infection in mosquito-infected chickens may be explained by higher viral dose delivered by mosquitoes. On the other hand, chickens infected by multiple mosquitoes (N = 3-11) had viremic titers that were 25-50 times higher at 6 and 12 hours PF than in chickens infected by a single mosquito, suggesting that viral dose is not the only factor involved in enhanced early infection. The likelihood that enhanced early infection in mosquito-infected chickens is due to a higher viral dose inoculated by mosquitoes and/or other factors (saliva, inoculation location, or viral source) is discussed. (+info)
Tissue distribution of psittacid herpesviruses in latently infected parrots, repeated sampling of latently infected parrots and prevalence of latency in parrots submitted for necropsy.
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Psittacid herpesvirus-1 (PsHV-1) is the cause of an acute fatal disease in parrots and is implicated as the cause of papillomatous lesions of the digestive tract. Not all infections cause disease and some parrots are infected asymptomatically. Latently infected parrots are potential sources for virus dissemination. Tissues from parrots that died spontaneously with a history of coming from flocks where a PsHV-1 outbreak had occurred were examined for PsHV-1 DNA. Fourteen of 16 parrots examined were infected with at least 1 variant of PsHV-1; of these 13 (93%) had viral DNA in either or both the oral and cloacal mucosa, suggesting that most latently infected parrots could be detected by sampling these sites. Nine of 9 parrots shown to be infected 5 years prior to this study were positive again on repeat sampling and were infected with the same virus genotype. Opportunistic sampling of parrots submitted for diagnostic necropsy indicated that the prevalence of PsHV-1 in parrots in the sampled population was approximately 9.3%. PsHV-1 genotypes 1, 2, and 3 were found in these birds, but genotype 4 was not. Six necropsy specimens were found to be infected with two PsHV-1 genotypes and it was concluded that infection with one serotype did not protect against infection with another. Psittacid herpesvirus 2 (PsHV-2) was identified in 4 African grey parrots and a blue and gold macaw. Prior to this study PsHV-2 had only been found in African grey parrots. (+info)