Construction of Species-Specific PCR Primers for Detection of Coccidia Parasites in Captive-Reared Northern Bobwhites

Captive rearing and subsequent release of game birds, including northern bobwhites (Colinus virginianus), has become common in certain areas. In this practice, bobwhites are often raised in confinement to ‘flight ready’ and subsequently released for hunting. It is estimated that 30–40 million bobwhites are raised in captivity annually and some farms in the USA produce upwards of 1 million birds annually for this market. Raising game birds in these densities greatly facilitates the transmission of pathogenic organisms. Coccidiosis has been previously identified as an important disease in captive bobwhites and infection can lead to weight loss, diarrhea, poor feather growth, dehydration and, in severe cases, death. Eimeria lettyae, E. colini, and E. dispersa are the three described coccidia species from bobwhites. We investigated the prevalence and distribution of species of coccidia in captive bobwhite facilities throughout the United States. We collected litter or intestinal samples from 31captive bobwhite facilities originating from 13 states. Species-specific PCR primers were constructed against the internal transcribed spacer region 1 (ITS-1) of the ribosomal RNA gene of the various Eimeria spp. to aid in parasite detection and distinction. Primers were used to detect the specific Eimeria spp. in the collected samples. All 31 samples were positive for coccidia. Results of the primer survey disclosed E. lettyae, E. dispersa, and an unidentified Eimeria sp. in 20 (64.5%), 22 (72%), and 29 (93.5%) of the samples, respectively. Thirteen (41.9%) samples had 3 Eimeria spp. detected, 14 (45.2%) samples had 2 spp. detected, and 4 (12.9%) samples had 1 sp. detected. Flock age or geographical location was not associated with the presence of any particular Eimeria spp. To our knowledge, this is the first study of coccidia in captive bobwhites. Previous studies of Eimeria spp. in wild northern bobwhite are rare and disclosed variable prevalence rates ranging from 0 to 36%; no efforts were made to distinguish the coccidia species in these studies It would be helpful to use the species-specific primers constructed in this study to examine the prevalence and distribution of the Eimeria spp. in wild bobwhites from throughout their range to investigate the potential for captive-raised bobwhites to be a source of coccidiosis for wild bobwhites

HIF- and Non-HIF-Regulated Hypoxic Responses Require the Estrogen-Related Receptor in Drosophila melanogaster

Low-oxygen tolerance is supported by an adaptive response that includes a coordinate shift in metabolism and the activation of a transcriptional program that is driven by the hypoxia-inducible factor (HIF) pathway. The precise contribution of HIF-1a in the adaptive response, however, has not been determined. Here, we investigate how HIF influences hypoxic adaptation throughout Drosophila melanogaster development. We find that hypoxic-induced transcriptional changes are comprised of HIF-dependent and HIF-independent pathways that are distinct and separable. We show that normoxic set-points of carbohydrate metabolites are significantly altered in sima mutants and that these animals are unable to mobilize glycogen in hypoxia. Furthermore, we find that the estrogen-related receptor (dERR), which is a global regulator of aerobic glycolysis in larvae, is required for a competent hypoxic response. dERR binds to dHIFa and participates in the HIF-dependent transcriptional program in hypoxia. In addition, dERR acts in the absence of dHIFa in hypoxia and a significant portion of HIF-independent transcriptional responses can be attributed to dERR actions, including upregulation of glycolytic transcripts. These results indicate that competent hypoxic responses arise from complex interactions between HIF-dependent and -independent mechanisms, and that dERR plays a central role in both of these programs

Prothoracicotropic Hormone Regulates Developmental Timing and Body Size in Drosophila

In insects, control of body size is intimately linked to nutritional quality as well as environmental and genetic cues that regulate the timing of developmental transitions. Prothoracicotropic hormone (PTTH) has been proposed to play an essential role in regulating the production and/or release of ecdysone, a steroid hormone that stimulates molting and metamorphosis. In this report we examine the consequences on Drosophila development of ablating the PTTH-producing neurons. Surprisingly, PTTH production is not essential for molting or metamorphosis. Instead, loss of PTTH results in delayed larval development and eclosion of larger flies with more cells. Prolonged feeding, without changing the rate of growth, causes the developmental delay and is a consequence of low ecdysteroid titers. These results indicate that final body size in insects is determined by a balance between growth rate regulators such as insulin and developmental timing cues such as PTTH that set the duration of the feeding interval

Prothoracicotropic Hormone Regulates Developmental Timing and Body Size in Drosophila

In insects, control of body size is intimately linked to nutritional quality as well as environmental and genetic cues that regulate the timing of developmental transitions. Prothoracicotropic hormone (PTTH) has been proposed to play an essential role in regulating the production and/or release of ecdysone, a steroid hormone that stimulates molting and metamorphosis. In this report we examine the consequences on Drosophila development of ablating the PTTH-producing neurons. Surprisingly, PTTH production is not essential for molting or metamorphosis. Instead, loss of PTTH results in delayed larval development and eclosion of larger flies with more cells. Prolonged feeding, without changing the rate of growth, causes the developmental delay and is a consequence of low ecdysteroid titers. These results indicate that final body size in insects is determined by a balance between growth rate regulators such as insulin and developmental timing cues such as PTTH that set the duration of the feeding interval

Türkiye'de etlik piliçlerde görülen Eimeria türlerinin moleküler olarak belirlenmesi]

The objective of this study was to determine whether seven Eimeria species involved in chicken coccidiosis are present in Turkey and to assess their prevelance in commercial flocks. Litter and faecal samples were collected from 1110 broiler flocks housed in 817 farms (about 12% of all broiler farms in Turkey) between September 2006 and September 2007. Coccidian oocysts were found in 624 (56.2 %) of the samples examined. Species-specific polymerase chain reaction (PCR) and nested PCR tests targeting the internal transcribed spacer-1 (ITS-1) sequences of the genomic rDNA were performed for all seven Eimeria species. The results of species-specific PCR assays confirmed the presence of E. maxima, E. tenella, K acervulina and E. praecox, and nested PCR results showed the presence of E mitis and E. brunetti. Although morphologic and morphometric observations revealed the presence of oocysts resembling E. necatrix; it was not confirmed by species specific or nested PCR. Nucleotide sequences of Turkish Eimeria isolates obtained by sequencing of the PCR products from 6 Eimeria species have been entered into the GenBank sequence database under accession numbers HQ680469 through HQ680474.Ankara University Office of Scientific Research ProjectsAnkara University [2005K120140-9-6]This research was supported by a grant of Ankara University Office of Scientific Research Projects (project number: 2005K120140-9-6) and summarized from Esin Guven's Ph.D thesis

Functional interactions between the Moses corepressor and DHR78 nuclear receptor regulate growth in Drosophila

Expression of the Drosophila orphan nuclear receptor DHR78 is regulated by the steroid hormone ecdysone and is required for growth and viability during larval stages. In contrast to our understanding of its biological functions, however, relatively little is known about how DHR78 acts as a transcription factor. Here we show that DHR78 is an obligate partner for Moses (Middleman of seventy-eight signaling), a SAM (sterile α motif) domain-containing cofactor that requires DHR78 for its stability. Unlike other nuclear receptor cofactors, Moses has no obvious interaction domains and displays a unique binding specificity for DHR78. Moses acts as a corepressor, inhibiting DHR78 transcriptional activity independently of histone deacetylation. Consistent with their close association, DHR78 and Moses proteins are coexpressed during development and colocalize to specific genomic targets in chromatin. Moses mutants progress normally through early larval stages, like DHR78 mutants, but display an opposite overgrowth phenotype, with hypertrophy of adult tissues. Genetic interactions between DHR78 and moses result in a similar phenotype, suggesting that the relative dose of Moses and DHR78 regulates growth and prevents cancer. The tight functional association between DHR78 and Moses provides a new paradigm for understanding the molecular mechanisms by which cofactors modulate nuclear receptor signaling pathways

The influence of dERR and dHIFa on hypoxic transcripts.

<p>(A) HIF-independent (HI), HIF-dependent (HD), ERR-dependent (ED), and ERR&HIF-dependent (DM) gene sets identified by microarray schemes outlined in Figures S1 and S2. Circles are scaled to size by number of transcripts in each set. (B) A Venn diagram demonstrating the overlap of the HI/HD/ED H-genes sets. Note, HI and HD genes sets are, by definition, mutually exclusive. The asterisks indicate that the overlap is significant (<i>p</i>-value<0.05), as determined by hypergeometric probability. (C) A Venn diagram demonstrating the overlap of the HD/ED/DM H-genes sets. qRT-PCR analysis of hypoxia-regulated genes falling into specific Venn overlaps, as indicated by arrows. (D) The top ten affected transcripts, as assessed by the H-responses measured in the control background, for each of the seven Venn categories shown in <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1003230#pgen-1003230-g007" target="_blank">Figure 7C</a>. Hypoxic expression for each transcript in the different mutant backgrounds (compared to <i>w¹¹¹⁸</i>) is reported as fold-change difference. Additionally shown is the N/H ratio obtained for <i>w¹¹¹⁸</i> animals. (E) Normoxic and hypoxic expression of each of the six genes (<i>Pfk, fatiga, spermine oxidase, NMNAT, LDH, ALAS</i>) was determined using RNA collected from animals of the indicated genotypes at late-L3. Samples were collected in triplicate and are independent from those used in the microarrays. Values are normalized to <i>rp49</i> expression and are reported relative to the value obtained for <i>w¹¹¹⁸</i> in normoxia.</p

List of 20 top transcripts whose expression changes in response to hypoxic challenge in a dHIF-dependent or -independent fashion.

<p>For HIF-dependent genes (taken from the top 20 down-regulated transcripts in the HD H-genes set), transcripts are sorted according to normalized microarray values obtained when comparing <i>w¹¹¹⁸</i> hypoxia samples with <i>sima</i> hypoxia samples. For the HIF-independent genes (taken from the top 20 up-regulated HI H-genes set), transcripts are sorted according to the normalized microarray values obtained when comparing <i>sima</i> normoxia with <i>sima</i> hypoxia samples. For comparative purposes, the respective hypoxic changes observed in <i>w¹¹¹⁸</i> control animals are reported in the last column. Additionally, the first four columns show the Affymetrix probe set ID, CG number, gene title, and the putative process/function of the encoded protein.</p