A sensitive solution-hybridization assay was used to investigate the expression of genes encoding insulin-like growth factors I and II (IGF-I and -I) in the rat central nervous system (CNS). mRNAs for both IGFs are synthesized throughout the CNS of adult rats but exhibit distinct regional differences for each growth factor. IGF-I mRNA is 8-10 times more abundant in the cervical-thoracic spinal cord and in the olfactory bulb than in whole brain and is enriched 3-fold in the midbrain and cerebellum. IGF-U mRNA is minimally enriched in the medulla-pons and cerebellum but is 3-5 times less abundant in the midbrain and corpus striatum than in total brain. During CNS development the content of IGF-I and IGF-II mRNAs is highest at embryonic day 14 and declines by a factor of 3-4 at birth, to values found in adult brain. Embryonic neurons and glia synthesize IGF-I mRNA during short-term cell culture; only glia produce IGF-II mRNA. These observations show that IGF-I and IGF-il are differentially expressed in the developing and adult CNS and suggest that each growth factor may play a unique role in the mammalian nervous system. The insulin-like growth factors (IGFs), or somatomedins, are polypeptide mitogens that play fundamental roles in mammalian growth processes. IGF-I (somatomedin C), a basic peptide of 70 amino acids (1), is the major mediator through which growth hormone exerts its biological effects on postnatal growth (2-4). IGF-II, a 67-residue neutral protein (5), may perform comparable growth-promoting functions during fetal life (2, 3). Both IGFs have been found in a variety of tissues, including the brain (6-16), although the precise physiological roles of either IGF in the central nervous system (CNS) are undefined.Recent evidence suggests that both somatomedins may be involved in several aspects of neural metabolic and growth processes. Receptors for IGF-I are widely distributed throughout the mammalian CNS (17)(18)(19) MATERUILS AND METHODSAnimals. Female rats of the Sprague-Dawley strain were obtained after timed matings from commercial suppliers. At specific times during gestation, rats were killed by cervical dislocation. The uterine horns were excised and the embryos were dissected from maternal tissues. Brain tissue was isolated and pooled for RNA isolation or harvested for cell culture. Neonatal, juvenile, and adult male Sprague-Dawley rats were killed by cervical dislocation, and specific organs were quickly excised and placed on ice. The following regions were dissected from adult animals (age, 60-90 (28). Morphological appearance and celltype-specific staining for glial fibrillary acidic protein indicated that these cultures were >80% astroglia.Molecular Cloning. The gene encoding rat IGF-I was isolated as a series of overlapping clones from two rat chromosomal libraries in bacteriophage X as described (29). The gene specifying rat IGF-II was isolated similarly from a chromosomal library (30) and was characterized by restricAbbreviations: IGF, insulin-like growth factor; CNS, central n...

No abstract

Results are presented from four studies between 2002 and 2011 into the feasibility of routinely monitoring Marek's disease virus serotype 1 (MDV-1) in broiler house dust using real-time quantitative PCR (qPCR) measurement. Study 1 on two farms showed that detection of MDV-1 occurred earlier on average in dust samples tested using qPCR than standard PCR and in spleen samples from five birds per shed assayed for MDV-1 by qPCR or standard PCR. DNA quality following extraction from dust had no effect on detection of MDV-1. Study 2 demonstrated that herpesvirus of turkeys (HVT) and MDV serotype 2 (MDV-2) in addition to MDV-1 could be readily amplified from commercial farm dust samples, often in mixtures. MDV-2 was detected in 11 of 20 samples despite the absence of vaccination with this serotype. Study 3 investigated the reproducibility and sensitivity of the qPCR test and the presence of inhibitors in the samples. Samples extracted and amplified in triplicate showed a high level of reproducibility except at very low levels of virus near the limit of detection. Mixing of samples prior to extraction provided results consistent with the proportions in the mixture. Tests for inhibition showed that if the template contained DNA in the range 0.5-20 ng/microl no inhibition of the reaction was detectable. The sensitivity of the tests in terms of viral copy number (VCN) per milligram of dust was calculated to be in the range 24-600 VCN/mg for MDV-1, 48-1200 VCN/mg for MDV-2, and 182-4560 VCN/mg for HVT. In study 4 the results of 1976 commercial tests carried out for one company were analyzed. Overall 23.1% of samples were positive for MDV-1, 26.1% in unvaccinated and 16.4% in vaccinated chickens. There was marked regional and temporal variation in the proportion of positive samples and the MDV-1 load. The tests were useful in formulating Marek's disease vaccination strategies. The number of samples submitted has increased recently, as has the incidence of positive samples. These studies provide strong evidence that detection and quantitation of MDV-1, HVT, and MDV-2 in poultry house dust using qPCR is robust, sensitive, reproducible, and meaningful, both biologically and commercially. Tactical vaccination based on monitoring of MDV-1 rather than routine vaccination may reduce selection pressure for increased virulence in MDV-1.

Pathotyping results in broiler chickens with maternal antibody broadly agreed with those in specific-pathogen-free chickens in other studies, with some important differences. MDV load in the spleen at 7 dpc and in isolator dust at both 14 and 21 dpc was a powerful early predictor of subsequent MD incidence.

No abstract