bluuebunny/crossref_metadata_2025 huggingface dataset:
Galanin in Normal and Hyperplastic Anterior Pituitary Cells: From Pituitary Tumor Cell Lines to Transgenic Mice
Authors: HYDE JAMES F. , MOORE JOSEPH P. , CAI AIHUA
Published: February 2006

Abstract: Studies on the regulation of galanin expression in the epithelial cells of the anterior pituitary gland have provided a wealth of insight into the cellular and molecular biology of this unique peptide. Galanin is localized within subpopulations of specific pituitary cell types, and hypothalamic as well as gonadal factors including dopamine, somatostatin, thyrotropin‐releasing hormone, growth hormone‐releasing hormone (GHRH), estrogen, and progesterone dynamically regulate its expression and release. Galanin gene expression and peptide secretion are markedly increased in estrogen‐induced prolactinomas, wherein galanin serves as both an autocrine and paracrine hormone regulating prolactin secretion. Galanin mRNA and peptide levels are also dramatically elevated in somatotroph adenomas of human GHRH transgenic mice. Moreover, galanin secretion is increased from the hyperplastic somatotrophs of hGHRH transgenic mice. However, not all pituitary adenomas are associated with increased galanin gene expression; galanin synthesis is repressed in 131I‐induced thyrotroph adenomas. Thus, galanin acts locally to regulate pituitary hormone secretion and appears to act as a mitogenic factor to increase the proliferation of pituitary cells in a cell‐type specific manner.
Structural Organization and Chromosomal Localization of Three Human Galanin Receptor Genes
Authors: IISMAA TIINA P. , FATHI ZAHRA , HORT YVONNE J. , IBEN LAWRENCE G. , DUTTON JULIE L. , BAKER ELIZABETH , SUTHERLAND GRANT R. , SHINE JOHN
Published: February 2006

Abstract: Human galanin receptor subtypes GALR1, GALR2, and GALR3 are encoded by separate genes that are located on human chromosomes 18q23, 17q25.3, and 22q13.1, respectively. The exon:intron organization of the gene encoding GALR2 (GALNR2) and GALR3 (GALNR3) is conserved, with exon 1 encoding the NH2‐terminus to the end of transmembrane domain 3 and exon 2 encoding the remainder of the receptor, from the second intracellular loop to the COOH‐terminus. This conservation of structural organization is indicative of a common evolutionary origin for GALNR2 and GALNR3. The exon:intron organization of the gene encoding GALR1 (GALNR1) is different from that of GALNR2 and GALNR3, with exon 1 encoding the NH2‐terminus to the end of transmembrane domain 5, exon 2 encoding the third intracellular loop, and exon 3 encoding the remainder of the receptor, from transmembrane domain 6 to the COOH‐terminus. The structural organization of GALNR1 suggests convergent evolution for this gene and represents a structural organization that is unique among genes encoding G‐protein‐coupled receptors.
Galanin Causes Cl− Secretion in the Human Colon: Potential Significance of Inflammation‐Associated NF‐κB Activation on Galanin‐1 Receptor Expression and Function
Authors: BENYA RICHARD V. , MATKOWSKYJ KRISTINA A. , DANILKOVICH ALEXEY , HECHT GAIL
Published: February 2006

Abstract: Galanin is widely distributed in enteric nerves and nerve terminals throughout the gastrointestinal (GI) tract. Within the GI tract galanin is best known for its ability to alter smooth muscle contractility and regulate intestinal motility. However, recent studies also indicate that galanin can modulate epithelial ion transport. We previously showed that epithelial cells lining the human GI tract, including those of colonic origin, express Gal1 galanin receptors (Gal1‐R). We herein demonstrate that epithelial cells lining the human colon only express Gal1‐R receptors and do not express other galanin receptor subtypes. We previously showed that Gal1‐R expression was transcriptionally regulated by the transcription factor NF‐κB. Consistent with this transcription factor being activated in a number of inflammatory conditions, we show increased colonic Gal1‐R expression in patients with colitis due to a variety of causes. To further evaluate the physiology of Gal1‐R activation, we studied this receptor expressed by the human colon epithelial cell line T84. Gal1‐R activation resulted in a dose‐dependent increase in Cl− secretion; whereas infection of T84 cells with pathogens known to activate NF‐κB augmented Gal1‐R expression and Cl− secretion. Thus, galanin acts as a secretagogue in epithelial cells lining the human colon, with alterations in Gal1‐R expression possibly playing an important role in the diarrhea associated with various inflammatory processes affecting the GI tract.
Mutagenesis Study on Human Galanin Receptor GalR1 Reveals Domains Involved in Ligand Binding a
Authors: KASK KALEV , BERTHOLD MALIN , KAHL ULRIKA , JURÉUS ANDERS , NORDVALL GUNNAR , LANGEL ÜLO , BARTFAI TAMAS
Published: February 2006

Abstract: Many receptor mutants were generated and several NH2‐terminally modified galanin analogs synthesized to define the regions of hGalR1 involved in galanin binding. Ligand binding properties and functionality of mutant receptors were evaluated. The His264Ala and Phe282Ala receptor mutants, although deficient in binding in the concentration range of galanin used, remained functional albeit at least 20‐fold less efficient than the wild‐type receptor in the inhibition of stimulated cAMP production. Hence, His264 and Phe282 of hGalR1 are directly involved in galanin binding. NH2‐terminal carboxylic acid analogs of galanin (1–16) have a very low affinity for the wild‐type receptor, but substantially increased affinity for the Glu271Lys‐hGalR1, suggesting that the NH2‐terminus of galanin binds to the receptor near the transmembrane (TM) VI. Based on these findings and computer‐aided molecular modeling, we propose a binding site model for the hGalR1 receptor (possibly also for other galanin receptor subtypes): galanin binds with its NH2‐terminus to the pocket between TM III and TM VI, Trp2 of galanin interacts with His264 of the receptor, and Tyr9 is involved in an aromatic‐aromatic type of interaction with Phe282 of ECIII of GalR1.
Cloning and Evaluation of the Role of Rat GALR‐2, a Novel Subtype of Galanin Receptor, in the Control of Pain Perception
Authors: AHMAD SULTAN , O'DONNELL DAJAN , PAYZA KEM , DUCHARME JULIE , MÉNARD DANIEL , BROWN WILLIAM , SCHMIDT RALF , WAHLESTEDT CLAES , SHEN S. H. , WALKER PHILIPPE
Published: February 2006

Abstract: We have identified a novel subtype of galanin receptor (GALR‐2) in rat dorsal root ganglia and spinal cord. The open reading frame of GALR‐2 is 1116 nucleotides long, encoding a protein of 372 amino acids with a theoretical molecular mass of 40.7 kD. Membranes prepared from stable pools of 293 cells expressing GALR‐2, but not wild‐type 293 cells, demonstrated high affinity galanin binding sites. Rat galanin and galanin‐related peptides M40, C7, M15, and galanin1–16 effectively competed for binding; peptide C7 demonstrated a lower affinity for rGALR‐2, and all these peptides were agonists at rGALR‐2 when assessed on a microphysiometer. Studies on the expression of GALR‐2 in various tissues by Northern and in situ hybridization analyses suggest a low abundance but wide distribution of GALR‐2 mRNA, including several discrete areas in brain and spinal cord and a high abundance in the dorsal root ganglia.
Distribution and Characterization of the Cell Types Expressing GALR2 mRNA in Brain and Pituitary Gland
Authors: DEPCZYNSKI B. , NICHOL K. , FAITHI Z. , IISMAA T. , SHINE J. , CUNNINGHAM A.
Published: February 2006

Abstract: The neuropeptide galanin mediates its activities through G‐protein‐ coupled receptors, and three receptor subtypes have been described with distinctly different patterns of regional tissue expression. GALR1 is predominantly expressed in basal forebrain, hypothalamus, as well as spinal cord. GALR2 has a wider distribution in brain and is also present in the pituitary gland and peripheral tissues. GALR3 has been found to be widely distributed at low abundance. We examined the distribution of GALR2 in rat brain and pituitary by in situ hybridization histochemistry and found it abundant in regions of hippocampus, piriform and entorhinal cortex, basal nucleus of the accessory olfactory tract, amygdala, hypothalamic nuclei, Purkinje cells, and discrete brainstem nuclei. It is also highly expressed in the intermediate and anterior lobes of the pituitary. Using combined in situ hybridization immunohistochemistry we characterized the neurotransmitter and hormonal phenotype of cells expressing GALR2 mRNA in the hypothalamus and pituitary gland. Our findings suggest GALR2 is a receptor mediating important functions of galanin in the hypothalamic‐pituitary axis and may also play a role in hippocampal and cerebellar function.
Role of Galanin in the Gastrointestinal Sphincters a
Authors: RATTAN SATISH , TAMURA WATARU
Published: February 2006

Abstract: Galanin was present and exerted potent effects in all the gastrointestinal sphincters examined. Galanin‐immunoreactive nerve fibers and neurons are present in both the myenteric and submucosal plexuses of sphincters. The neuropeptide exerts diverse effects in different sphincteric smooth muscles that may be species specific. For example, in the lower esophageal sphincter, it may cause an increase in basal tone and suppression of nonadrenergic noncholinergic (NANC) nerve‐mediated relaxation. On the contrary, in the internal anal sphincter (IAS), the predominant effect of galanin is to cause smooth muscle relaxation and augmentation of NANC nerve‐mediated relaxation. In other sphincters, galanin may either have no effect or cause either an increase or a decrease in basal tone. Most of the actions of galanin on basal smooth muscle sphincteric tone are due to its actions directly on smooth muscle cells. However, some of the relaxant actions of the peptide may also be due to activation of NANC inhibitory neurons. The basic mechanism/s responsible for sphincteric smooth muscle contraction or relaxation in response to galanin have not been investigated. The suppressive as well as the augmentatory effects of galanin on NANC nerve‐mediated sphincteric smooth muscle relaxation may be due to inhibition or facilitation, respectively, of the release of NANC inhibitory neurotransmitters such as nitric oxide and vasoactive intestinal polypeptide. Diverse effects in different gastrointestinal sphincters suggest a neuromodulatory rather than a neurotransmitter role of galanin and a significant role of the neuropeptide and putative antagonists in the pathophysiology and potential therapy of gastrointestinal motility disorders especially those affecting sphincteric function.
Differential Functions of Galanin Cell Groups in the Regulation of Eating and Body Weight a
Authors: LEIBOWITZ SARAH F.
Published: February 2006

Abstract: Evidence suggests that hypothalamic galanin (GAL) has a variety of functions related to energy and nutrient balance, reproduction, water balance, and neuroendocrine regulation. The focus of this chapter is the role of GAL in eating and body weight regulation. Findings described herein demonstrate that GAL, in a cell group of the anterior region of the paraventricular nucleus (aPVN) that projects to the median eminence, has a role in the control of fat intake, fat metabolism, and body fat. This function of aPVN GAL neurons is carried out in close relation to circulating insulin and glucose. Galanin‐expressing perikarya in the medial preoptic area (MPOA) have a similar function, although GAL here operates in association with the female steroids estrogen and progesterone. These GAL cell groups of the aPVN and MPOA contrast with those in the arcuate nucleus as well as the magnocellular vasopressin‐containing neurons of the PVN and supraoptic nucleus, which show no relation to fat balance. This evidence reveals differential functions for the distinct GAL neuronal cell groups of the hypothalamus.
Galanin‐Galanin Receptor Systems in the Hypothalamic Paraventricular and Supraoptic Nuclei: Some Recent Findings and Future Challenges aa
Authors: GUNDLACH ANDREW L. , BURAZIN TANYA C.D.
Published: February 2006

Abstract: Galanin and galanin receptors are widely distributed within the central nervous system, but historically much research has been focused on hypothalamic galanin systems including those in the preoptic area, paraventricular nucleus (PVN), supraoptic nucleus (SON), and median eminence. In early studies, galanin mRNA, immunoreactivity, and binding sites were detected in neurons of the SON and both the magnocellular and parvocellular regions of the PVN, all of which also contain vasopressin, oxytocin, and several other peptides. This article briefly reviews some important recent studies of the electrophysiologic effects of galanin on magnocellular neurons in vitro; regulation of galanin expression by the physiologic stimulus of lactation; the role of parvocellular galanin systems in energy balance, body weight, and obesity; and the regional and cellular localization of galanin and galanin receptor mRNAs in the PVN/SON. In relation to the latter issue, two distinct galanin receptor subtypes, GalR1 and GalR2, have now been cloned and characterized. In situ hybridization histochemical studies of rat brain by several groups have consistently demonstrated GalR1 mRNA in the SON and PVN, in the magnocellular and parvocellular regions. By contrast, our recent experiments using [35S]‐labeled oligonucleotide probes detected GalR2 mRNA enriched in the parvocellular, not the magnocellular regions of the PVN, and the transcripts were not detected in the SON, whereas studies by others using a digoxigenin‐labeled RNA probe have detected GlR2 mRNA in the SON (and PVN). Nonetheless, given the known effects of hyperosmotic stimuli, changes in metabolic status, and various hormones on galanin synthesis and release and the ability of galanin to regulate the electrical and secretory activity of magnocellular neurons, it will be of interest to determine any possible (differential) regulation of galanin receptor subtype expression and the pre‐ and postsynaptic roles of GalR1 and GalR2 receptors in magnocellular and parvocellular neurons.
Galanin in Ascending Systems: Focus on Coexistence with 5‐Hydroxytryptamine and Noradrenaline a
Authors: HÖKFELT TOMAS , XU ZHI‐QING DAVID , SHI TIE‐JUN , HOLMBERG KRISTINA , ZHANG XU
Published: February 2006

Abstract: Galanin can be synthesized in several ascending systems including cholinergic forebrain neurons, serotonergic dorsal raphe neurons, and the noradrenergic locus coeruleus system. Recent immunohistochemical studies suggest that of these three systems, the locus coeruleus neurons express the highest levels of galanin and that in cortex and hippocampus galanin peptide can only be detected in the noradrenergic projections. Electrophysiologic studies show that galanin hyperpolarizes both serotonergic dorsal raphe neurons and noradrenergic locus coeruleus neurons at fairly high concentrations (106‐10−7 M). In addition, galanin at low concentrations (10−9 M) enhances the 5‐HT‐ and noradrenaline‐induced hyperpolarization. Consequently, a galanin antagonist could attenuate an inhibitory tone on both dorsal raphe and locus coeruleus neurons and thus perhaps exert antidepressant activity.
Electrophysiologic Effects of Galanin on Neurons of the Central Nervous System a
Authors: PIERIBONE VINCENT A. , XU ZHI‐QING DAVID , ZHANG XU , HÖKFELT TOMAS
Published: February 2006

Abstract: The neuropeptidc galanin is found in a large number of neurons and nerve terminals throughout the nervous system. In nerve terminals, galanin is contained in large dense‐core vesicles and is released upon electrical stimulation. A variety of electrophysiologic studies have examined the effects of galanin application onto neurons of the central nervous system. Overall, galanin appears to have inhibitory effects in the central nervous system, causing in most cases a potassium‐mediated hyperpolarization accompanied by a decrease in input resistance. Other actions include a reduction in presynaptic excitatory inputs and an interaction with other applied neurotransmitters. These effects are robust and long lasting in most cases. Differences in the responses mediated by the various receptor subtypes have not been explored electrophysiologically. More complete analysis awaits the availability of more potent and specific receptor anatagonists.
Galanin Inhibits Performance on Rodent Memory Tasks
Authors: McDONALD MICHAEL P. , GLEASON THERESA C. , ROBINSON JOHN K. , CRAWLEY JACQUELINE N.
Published: February 2006

Abstract: Central administration of galanin produces performance deficits on a variety of rodent learning and memory tasks. Galanin impairs acquisition and/or retention of the Morris water task, delayed nonmatching to position, T‐maze delayed alternation, starburst radial maze, and passive avoidance in normal rats. A primary site of action is the ventral hippocampus, with an additional modulatory site in the medial septum‐diagonal band. The behavioral actions of galanin at rat septohippo‐campal sites mediating cognitive processes are consistent with previous reports of inhibitory actions of galanin on acetylcholine release and cholinergically activated transduction at the M1 muscarinic receptor in rat hippocampus. The peptidergic galanin receptor antagonist M40 blocks the inhibitory actions of galanin on memory tasks. Treatment combinations of M40 with an M1 agonist, TZTP, improves performance on delayed nonmatching to position, in rats with 192IgG‐saporin‐induced cholinergic lesions of basal forebrain neurons. Nonpeptide, bioavailable, subtype‐selective galanin receptor antagonists may provide tools to test the hypothesis that antagonism of endogenous galanin, which is overexpressed in the basal forebrain in Alzheimer's patients, can contribute to the alleviation of the cognitive deficits associated with Alzheimer's disease.
Galanin: A Significant Role in Depression?
Authors: WEISS JAY M. , BONSALL ROBERT W. , DEMETRIKOPOULOS MELISSA K. , EMERY MILBURN S. , WEST CHARLES H.K.
Published: February 2006

Abstract: This paper describes a hypothesis that attempts to account for how changes in noradrenergic systems in the brain can affect depression‐related behaviors and symptoms. It is hypothesized that increased activity of the locus coeruleus (LC) neurons, the principal norepinephrine (NE)‐containing cells in the brain, causes release of galanin (GAL) in the ventral tegmentum (VTA) from LC axon terminals in which GAL is colocalized with NE. It is proposed that GAL release in VTA inhibits the activity of dopaminergic cell bodies in this region whose axons project to forebrain, thereby resulting in two of the principal symptoms seen in depression, decreased motor activation and decreased appreciation of pleasurable stimuli (anhedonia). The genesis of this hypothesis, which derives from studies using an animal model of depression, is described as well as recent data consistent with the hypothesis. The formulation proposed suggests that GAL antagonists may be of therapeutic benefit in the treatment of depression.
Galanin in Somatosensory Function a
Authors: WIESENFELD‐HALLIN ZSUZSANNA , XU XIAO‐JUN
Published: February 2006

Abstract: Galanin‐like immunoreactivity and galanin receptors are found in dorsal root ganglion (DRG) cells and in dorsal horn interneurons, suggesting that this neuropeptide may have a role in sensory transmission and modulation at the spinal level. Expression of galanin or galanin receptors in the DRG and spinal cord are altered, sometimes in a dramatic fashion, by peripheral nerve injury or inflammation. Under normal conditions, galanin occurs in a small population of primary sensory neurons as well as in spinal interneurons. However, following peripheral nerve injury or inflammation, expression of galanin in primary afferents and spinal cord is upregulated. We examined the role of galanin in spinal processing of nociceptive information under normal and pathologic conditions in a large series of electrophysiologic and behavioral studies. Results suggest that under normal conditions galanin exerts tonic inhibition of nociceptive input to the central nervous system. After peripheral nerve injury the inhibitory control exerted by endogenous galanin, probably released from DRG neurons, is increased. During inflammation, galanin presumably released from dorsal horn interneurons also exerts an inhibitory function. Thus, stable galanin agonists may be useful in the treatment of inflammatory and neuro‐pathic pain.
Galanin Expression in Neuropathic Pain: Friend or Foe?a
Authors: RAMER MATTHEW S. , MA WEIMA , MURPHY PATRICIA G. , RICHARDSON PETER M. , BISBY MARK A.
Published: February 2006

Abstract: We investigated a possible link between galanin expression and evoked pain accompanying painful partial sciatic nerve lesions. Increased galanin immunoreactivity (IR) in the dorsal horn, in gracile nucleus, and in sensory neurons following chronic constriction injury (CCI) compared to complete sciatic transection suggested a facilitatory role in thermal and mechanical hypersensitivity (allodynia). We therefore investigated the effects of endogenous interleukin‐6 (IL‐6) and nerve growth factor (NGF) on allodynia and neuropeptide expression. IL‐6 knockout mice showed decreased allodynia and galanin‐IR compared to wild‐type mice, but also decreased substance P (SP)‐IR in the dorsal horn. Anti‐NGF‐treated rats with CCI also showed decreased allodynia and SP‐IR, but increased galanin‐IR in the dorsal horn. These results suggest that evoked pain is more tightly linked to SP than to galanin expression. If galanin's effects are inhibitory as the bulk of the literature suggests, its effects are subordinate to those of SP and to other changes following CCI.
Regulation of Expression of Galanin and Galanin Receptors in Dorsal Root Ganglia and Spinal Cord after Axotomy and Inflammation a
Authors: ZHANG XU , XU ZHI‐QING , SHI TIE‐JUN , LANDRY MARC , HOLMBERG KRISTINA , JU GONG , TONG YONG‐GUANG , BAO LAN , CHENG XI‐PING , WIESENFELD‐HALLIN ZSUZSANNA , LOZANO ANDRES , DOSTROVSKY JONATHAN , HÖKFELT TOMAS
Published: February 2006

Abstract: Galanin can normally be detected only in a few dorsal root ganglion (DRG) neurons, but it is dramatically upregulated after peripheral nerve injury in both rat and monkey. Galanin is stored in large dense core vesicles, which after axotomy are often found close to the membrane of afferent nerve endings in the dorsal horn. In the monkey there is an increase in galanin in many ner ve terminals in the superficial dorsal horn after axotomy, but such an inc rease is more difficult to detect in the rat. Galanin is also present in local dorsal horn neurons, where it is upregulated by peripheral inflammation. Both galanin‐R1 and galanin‐R2 receptor mRNAs are expressed in rat DRGs, mainly in, respectively, large and small DRG neurons. Galanin‐R1 receptor mRNA is downregulated in DRG neurons after axo‐tomy, and a small decrease in galanin‐R2 receptor mRNA levels can also be seen. After peripheral tissue inflammation galanin‐R1 receptor mRNA levels decrease and galanin‐R2 receptor mRNA levels increase. The present results show that galanin and galanin receptors are present in sensory and local dorsal horn neurons and are regulated by nerve injury and inflammation. Galanin may therefore be involved in processing of pain information, primarily exerting analgesic effects. Whereas local dorsal horn neurons represent a defense system against inflammatory pain, we have proposed that a second defense system, against neuropathic pain, is intrinsic to DRG neurons.
Bovine Babesiosis and Anaplasmosis Follow‐up on Cattle Relocated in an Endemic Area for Hemoparasitic Diseasesa
Authors: FIGUEROA J. V. , ALVAREZ J. A. , RAMOS J. A. , ROJAS E. E. , SANTIAGO C. , MOSQUEDA J. J. , VEGA C. A. , BUENING G. M.
Published: February 2006

ABSTRACT: A multiplex PCR/DNA probe assay was used to monitor Babesia bovis, B. bigemina and Anaplasma marginale infection in cattle introduced to a Boophilus microplus‐infested area in Veracruz, Mexico. Eight intact, 18‐month‐old, cross‐bred beef cattle (four naive, Group A; four Babesia species‐premunized, Group B) were immediately exposed to ticks after arrival and were clinically monitored from day 6 to day 98 post‐exposure (PE) to ticks. Blood sample analysis for DNA detection by the MPCR/DNA probe assay showed that Group A animals were infected with B. bovis from day 11 up to day 22 PE, requiring treatment on days 17‐20. Group B animals were detected positive to B. bovis on days 17‐20, did not require treatment and remained persistently infected from days 70 to 84 PE. Treatment of Group A animals delayed the infection with B. bigemina. These animals became positive to the parasite on days 63‐77 PE. In contrast, Group B animals (untreated) showed B. bigemina infection on days 21‐26 and 63‐84 PE. One animal was positive for A. marginale infection on days 63‐66 PE, the rest of the animals became so on days 80‐98 PE. All infected animals required treatment with oxytetracycline. Monitoring the triple hemoparasite infection with the MPCR/DNA probe assay provided important epidemiological information. Thus, precautionary measures can be established when cattle are moved to a babesiosis/anaplasmosis risk area.
Genetic Analysis of Isolates of the Spotted Fever Group of Rickettsiae Belonging to the R. conorii Complexa
Authors: DASCH GREGORY A. , JACKSON LOUISE M.
Published: February 2006

ABSTRACT: The cytoplasmic 120 kDa antigen genes of 9 isolates of Rickettsia conorii (RC), 12 isolates of R. africae (RA), and 3 isolates of Israeli tick typhus rickettsiae (ISTT) were compared for restriction fragment length polymorphisms (RFLP) present in portions of the open reading frame amplified by polymerase chain reaction (PCR). Initially, DNAs from 13 species or serotypes of spotted fever group rickettsiae were used to select restriction enzymes (RE) that detected RFLP in gene fragments amplified with primer pairs 483WF/1514R and 764F/3409R. Among the R. conorii complex isolates, Dpn II gave RFLP differentiating all three serotypes. Unique RE patterns were obtained for RC with Bsr I and Hinf I, for RA with Mwo I, Pst I and Ssp I, and for ISTT with Hpa II. While RFLP typing of the 120 kDa gene permitted rapid separation of R. conorii complex isolates into three groups corresponding to the RC, RA, and ISTT rOmp serotypes, additional intragroup genetic variation was also detected in all three serotypes.
Use of a PCR Assay for the Specific and Sensitive Detection of Trypanosoma Spp. in Naturally Infected Dairy Cattle in Peri‐urban Kampala, Ugandaa
Authors: CLAUSEN P.‐H. , WIEMANN A. , PATZELT R. , KAKAIRE D. , POETZSCH C. , PEREGRINE A. , MEHLITZ D.
Published: February 2006

ABSTRACT: The objective of this study was to compare the sensitivity and specificity of the polymerase chain reaction (PCR) with the hematocrit centrifugation technique (HCT) and the mini‐anion‐exchange centrifugation technique (m‐AECT) for diagnosis of trypanosome infections in livestock. In a cross‐sectional study, 486 cattle from 50 randomly selected farms in Mukono County, Uganda were investigated in June 1994. The direct parasitological techniques were performed in the field, resulting in 45 (9.3%) animals positive by HCT and 78 (16%) positive by m‐AECT. The total prevalence (combined evidence of HCT and m‐AECT) was 18.9%, with 78.2%Trypanosoma brucei only, 10.9%T. vivax and 10.9% mixed (T. brucei/T. vivax) infections. Trypanosomes of the subgenus Nannomonas were not detected. DNA was prepared by lysis from 181 randomly selected blood samples and amplified by PCR using species‐specific oligonucleotide primers. Overall, the PCR gave positive results in 63 (34.8%) blood samples, with 76.2% positive only for T. brucei, 20.6% positive only for T. vivax and 3.2% positive for mixed (T. brucei/T. vivax) infections. The preliminary results from this study demonstrate that the detection rate of PCR is about two times higher than that of the direct parasitological techniques, suggesting a higher sensitivity. The higher proportion of T. vivax infections detected by PCR as compared to HCT/m‐AECT is likely to be due to false parasitological classifications which might occur under field conditions.
Microsatellite Markers for Genetic Population Studies in Glossina palpalis gambiensis (Diptera: Glossinidae)a
Authors: SOLANO P. , DUVALLET G. , DUMAS V. , CUISANCE D. , CUNY G. , TOURE S. M.
Published: February 2006

ABSTRACT: Little is known about intraspecific variability in tsetse flies and its consequences for vectorial capacity. Microsatellite markers have been developed for Glossina palpalis gambiensis. Three loci have been identified and showed size polymorphisms for insectarium samples. G. palpalis gambiensis from Burkina Faso were also subjected to PCR to investigate their genetic variability. Amplifications were observed in different species belonging to the palpalis group. These molecular markers will be useful to estimate gene flow within G. palpalis gambiensis populations and analysis could be extended to related species.
Theileria lestoquardi and T. annulata in Cattle, Sheep, and Goats: In Vitro and in Vivo Studies a
Authors: BROWN C.G. D. , ILHAN T. , KIRVAR E. , THOMAS M. , WILKIE G. , LEEMANS I. , HOOSHMAND‐RAD P.
Published: February 2006

ABSTRACT: Theileria annulata, causing bovine tropical theileriosis, and T. lestoquardi (syn T. hirci), the agent of malignant ovine theileriosis, are both transmitted by ticks of the genus Hyalomma. Their distribution is thus very similar and, should these parasites infect more than one ruminant species, the difficulty in interpreting epidemiological studies is magnified considerably. A pilot series of experiments was thus conducted in which cattle, sheep and goats were infected with sporozoites of a single stock of each of T. annulata and T. lestoquardi from a laboratory colony of H.a.anatolicum. Reciprocal cross‐immunity and serological studies and in vitro culture isolations in mononuclear cells of each ruminant species illustrated both the similarity of these organisms and their differences. The importance of these findings in discriminating parasites in epidemiological studies and the control of these diseases with cell culture vaccines is emphasized.
Characterization and Discrimination of Three Theileria parva Stabilates Involved in East Coast Fever Vaccinationa a
Authors: SPARAGANO O. A. E. , ZANAA O. , AMBROSE N.
Published: February 2006

ABSTRACT: Three vaccine stabilates of Theileria parva, of which sporozoites are being used against East Coast fever, were characterized by immunological and molecular biology techniques before being used for a national vaccination campaign in Kenya. T. parva Marikebuni stabilates 316 and 3014, and T. parva Lanet were used in this study and were discriminated from other Kenyan field Theileria isolates. IFAT results showed that all the animals were producing antibodies regardless of the stock used. Primers designed on the TPR1 gene sequence were used for PCR and Decamers were used for RAPD. Specific DNA band patterns (1,877 bp; 1,059 bp, and 443 bp) for the three vaccine stocks were observed. These molecular markers could be used to trace vaccinated animals in Kenya and to identify which isolates are responsible for reactions in animals.
Comparative Evaluation of 16S, map1 and pCS20 Probes for the Detection of Cowdria and Ehrlichia Species in Ticks a
Authors: ALLSOPP M. T. E. P. , HATTINGH C. M. , VOGEL S. W. , ALLSOPP B. A.
Published: February 2006

ABSTRACT: We have developed a panel of 16S ribosomal RNA gene probes for heartwater epidemiology; five of these detect different Cowdria genotypes (Ball3, Senegal, Omatjenne, Crystal Springs, and Mara 87/7); one detects all five of these genotypes; one detects any Group III Ehrlichia species other than Cowdria; one detects any Group II Ehrlichia species. We have used these probes on PCR‐amplified rickettsial 16S rRNA genes from over 200 Amblyomma ticks. Control ticks were laboratory‐reared and either uninfected or fed on sheep experimentally infected with different Cowdria isolates, field ticks were harvested from animals in heartwater‐endemic and heartwater‐free areas. All the samples were also examined by PCR amplification and probing for two other Cowdria genes (map1 and pCS20) which have been used for heartwater epidemiology. This paper describes the first direct comparison of all the currently available DNA probes for heartwater‐associated organisms.
PCR Detection of Cowdria ruminantium Infection in Ticks and Animals from Heartwater‐Endemic Regions of Zimbabwe
Authors: MAHAN S. M. , PETER T. F. , SIMBI B. H. , BURRIDGE M. J.
Published: February 2006

ABSTRACT: The development of a PCR assay for the detection of Cowdria ruminantium infection in ticks has been previously described. Here we report a further evaluation of this assay by comparison with a DNA probe and with the mouse inoculation assay (MIA). Application of the PCR assay in determining the prevalence of infection in ruminants and ticks from heartwater‐endemic areas of Zimbabwe is also described. One hundred uninfected and 120 infected Amblyomma hebraeum ticks were analyzed by PCR, DNA probe and the MIA. These tests detected infection in 92%, 77% and 8% of the infected ticks, respectively, showing PCR to be the most sensitive assay. None of the uninfected ticks were positive by any of the 3 tests. The PCR assay detected infection rates of 10.5%, 12.5% and 3.2% in 200 male, 241 female and 95 nymphal A. hebraeum ticks, respectively, which were collected from a heartwater‐endemic farm. The PCR test was also applied to cattle of different age groups and goats from the same heartwater‐endemic farm. In a cross‐sectional survey, the assay detected infection in 3.3% to 26.7% of the cattle and in 23.3% of the goats. The implications of these findings and the potential for the application of PCR in heartwater diagnosis is discussed.
Detection and Differentiation of Theileria annulata and Theileria parva Using Macroschizont‐derived DNA Probes
Authors: SHAYAN P. , BIERMANN REINHILD , SCHEIN E. , GERDES J. , AHMED J. S.
Published: February 2006

ABSTRACT: A λgt11 expression library based on T. annulata‐infected cells was screened with an antiserum raised in rabbits against partially purified schizonts of T.annulata. Two clones were detected, sequenced and designated as SA288 and SB288 (Shayan et al., submitted for publication). From the sequences of these two genes oligonucleotide primers were designed for specific amplification of parasite DNA by polymerase chain reaction (PCR). We could show that these genes are of parasitic origin and do occur in all T. annulata stocks tested in the present study. In addition, a target sequence for SA288 could also be identified in T.parva‐schizonts. None of them reacted with genomic DNA of different Babesia spp. A third primer pair was designed from the DNA‐sequence of a gene encoding for the T.parva‐specific casein kinase II‐alpha subunit. Using this primer pair, a target sequence could only be detected in T.parva. Taken together, the primers described here can be used as molecular tools in PCR for the detection of Theileria parasites and to distinguish T.annulata from T.parva.
Theileria annulata: Carrier State and Immunitya
Authors: ILHAN T. , WILLIAMSON S. , KIRVAR E. , SHIELS B. , BROWN C.G. D.
Published: February 2006

ABSTRACT: Recovery from primary infection of Theileria annulata results in the development of a persistent carrier state in the vertebrate host. The carrier state is of great importance in the maintenance of the life cycle by alternate tick/cattle challenge and both contributes to and may be necessary for maintenance of immunity. Therefore, an accurate determination of carrier animals could be useful in determining immune status and may allow the necessary control measures to be implemented. Detailed information on the carrier state of animals following immunization with attenuated cell lines is lacking. In this study, relationship between immune response, persistence of the parasite, and the antibody response has been investigated. Calves were infected with T. annulata sporozoites, low passage (non‐attenuated) or high passage (attenuated, vaccine) cell lines and later challenged with a lethal dose of heterologous sporozoites. The presence and persistence of the parasite were monitored by PCR using primers derived from genes coding for ssrRNA and a 30 kDa major merozoite surface protein, by Giemsa stained blood smears to detect the presence of piroplasms and also by attempting to establish infected mononuclear cell cultures from venous blood. Antibody responses were measured by indirect ELISA using a merozoite recombinant antigen and IFAT using piroplasm and macroschizont antigens. Results showed that there was an evident relationship between the persistence of carrier status, antibody response in ELISA and immune response to challenge.
Development of a Non‐radioactive DNA Probe andin SituHybridization for Detection ofAnaplasma marginalein Ticks and Cattlea
Authors: KOCAN KATHERINE M. , GE NIE‐LIN , BLOUIN EDMOUR F. , MURPHY GEORGE L.
Published: February 2006

ABSTRACT:A non‐radioactive DNA probe was developed for detection ofAnaplasma marginalein ticks and cattle. The probe was labeled with digoxigenin 11‐dUTP by polymerase chain reaction. The probe was tested on bovine blood and was found to be a sensitive and specific detection method forA. marginalein cattle. The DNA probe was then adapted forin situhybridization (ISH) ofA. marginaleinDermacentor andersoniandD. variabilisticks infected either as nymphs or adults. One‐half of each tick was studied with ISH while the other half was examined with light and electron microscopy. In male ticks infected as adults, tick gut cells first became infected withA. marginalewhile ticks fed on an infected calf, and they remained infected as they transmission fed on a second, susceptible calf. At the onset of transmission feeding, salivary glands became infected withA. marginale. During transmission feeding infection was also observed in interstitial, reproductive, skeletal muscle, fat body and Malpighian tubule tissue, resulting in a generalizedA. marginaleinfection. When adult ticks that acquired infection as nymphs were examined with ISH and microscopy, gut tissues of bothD. andersoniandD. variabilisbecame infected withA. marginale. However, salivary gland infection was seen only inD. variabilis, even though both species of ticks transmittedA. marginaleto susceptible calves.A. marginalewas not seen with ISH or microscopy in hemocytes collected from both species of ticks and, thus, hemocytes do not appear to play a role in the development ofA. marginalein ticks.
Vaccination against Contagious Bovine Pleuropneumonia and the Use of Molecular Tools in Epidemiology
Authors: THIAUCOURT F. , LORENZON S. , DAVID A. , TULASNE J. J. , DOMENECH J.
Published: February 2006

ABSTRACT: Contagious bovine pleuropneumonia is a serious threat to cattle not only in Africa but also in southern Europe and possibly Asia. It is now present in countries that had been free of the disease for many years, giving rise to doubts about the efficiency of the control strategies. In Africa CBPP is controlled mainly by a vaccination policy that uses variant strains of Mycoplasma mycoides subsp mycoides biotype SC, called T1/44 or T1sr. Until recently, it was not possible to differentiate the various strains within the biotype and consequently to identify the vaccine strains. Restriction analysis of mycoplasma DNA with HindIII and Pst1 has been applied to 24 strains of African origin and one European strain. Each enzyme gave rise to different restriction profiles and the combination of the results permitted subdivision of these strains into 9 groups. Interestingly, some profiles of pathogenic strains seem to be restricted to certain geographical areas. The profile of the poorly immunogenic vaccinal strain KH3J is also very peculiar, and it is easily distinguished from that of the other vaccine strains originating from T1. This technique is simple once the strains are isolated. Efforts are now under way to use molecular tools based on PCR products to alleviate the difficulty of isolation.
Analysis of Cellular Responses to Native and Recombinant Proteins of Cowdria ruminantiumaa
Authors: TOTTÉ PHILIPPE , McKEEVER DECLAN , JONGEJAN FRANS , BARBET ANTONY , MAHAN SUMAN M. , MWANGI DUNCAN , BENSAID ALBERT
Published: February 2006

ABSTRACT: The response of PBMC collected from cattle immunized with killed Cowdria to MAP1, MAP1‐B, MAP2 and to a lysate of Cowdria was characterized in vitro. CD4+ T lymphocytes responded strongly to Cowdria lysates while B cells and γδ T cells proliferated in response to recombinant proteins. Short‐term cell lines derived from PBMC using Cowdria lysates were more than 90% CD4+ and did not respond to any of the recombinant proteins. Restimulation of PBMC with MAP1, MAP1‐B, and MAP2 proved difficult, and cultures usually stopped proliferating after 2 to 3 weeks. Only one cell line could be generated using MAP1 and it was composed of 85%γδ T cells. This study indicates that in animals immunized with killed Cowdria, proteins other than the serologically immunodominant MAP1 and MAP2 are strongly immunogenic for T lymphocytes.
Inhibitory Effect of Cowdria ruminantium on the Expression of MHC Class I and Class II Molecules on Bovine Endothelial Cellsa
Authors: VACHIÉRY N. , TRAP I. , TOTTÉ P. , MARTINEZ D. , BENSAID A.
Published: February 2006

ABSTRACT: Bovine endothelial cells constitutively express MHC class I molecules, whereas MHC class II molecules can be induced by interferon gamma (IFNγ) treatment. Endothelial cells may play a role as antigen presenting cells (APC), but their role in the immune response against Cowdria ruminantium is unknown. We have studied the effect of C. ruminantium infection on the expression of MHC class I and class II molecules on the surface of bovine endothelial cells. Bovine brain endothelial cells (BBEC) from the microvasculature and bovine umbilical endothelial cells (BUEC) from the macrovasculature were cultured in vitro, infected with different concentration of Cowdria, and treated or not with IFNγ. We observed that Cowdria induced a reduction of MHC molecules expression on the surface of BUEC and BBEC. This inhibitory effect of Cowdria on MHC molecules could affect the capacity of endothelial cells to act as efficient antigen‐presenting cells.
Note on Transferrin, Hemoglobin Types, and Packed Cell Volume in Senegalese Trypanotolerant Djallonke Sheep
Authors: MISSOHOU A. , NGUYEN T. C. , DORCHIES PH. , GUEYE A. , SOW R. S.
Published: February 2006

ABSTRACT: In this study we examined transferrin (Tf) and hemoglobin (Hb) types and frequencies and their relationship with Packed Cell Volume (PCV), which is considered as a selection criterion for the trypanotolerance trait. Blood samples were collected from 96 sheep and were typed for Tf and Hb. The frequencies of the alleles TfA, TfB, TfC and TfD were respectively 0.276, 0.005, 0.109, and 0.609. At the locus Hb, all animals were monomorphic B. The lowest PCV value was observed in animals homozygous for TfC, while the highest value was found in heterozygous (CD) animals; however, the difference was not significant.
Epidemiology and Control of East Coast Fever in Zambia: A Field Trial with Traditionally Managed Sanga Cattlea
Authors: MINJAUW B. , OTTE M. J. , JAMES A. D.
Published: February 2006

ABSTRACT: The main objective of the reported field trial was to compare different East Coast Fever (ECF) control strategies for their efficacy, effect on cattle productivity and cost‐effectiveness. Five strategies were tested in groups of traditionally managed Sanga cattle over a period of 2.5 years. Two groups were under intensive tick control, one group immunized by the infection and treatment method and the other non‐immunized. Two groups were under no tick control, one group immunized and the other non‐immunized (the control group). The fifth group was under strategic tick control and was immunized against ECF.

All ECF control methods tested significantly reduced mortality, but no marked differences to the control group were seen in other production parameters. No difference in mortality was observed between animals protected from ECF by immunization or by tick control. The most cost‐effective method of controlling the disease was by immunization. A financial analysis showed that under the prevailing conditions the break‐even price for immunization ranged from US$21.5 to US$25.7 depending of the proportion of reactors. The carrier state induced by immunization did not lead to a persistent high incidence of ECF in non‐immunized animals using the same grazing area.
Prospective Study for the Detection of Anaplasma marginale Theiler, 1911 (Rickettsiales: Anaplasmataceae) in Costa Ricaa
Authors: HERRERO M. V. , PEREZ E. , GOFF W. L. , DE ECHAIDE S. TORIONI , KNOWLES D. P. , McELWAIN T. F. , ALVAREZ V. , ALVAREZ A. , BUENING G. M.
Published: February 2006

ABSTRACT: A prospective study was conducted to assess the dynamics of the infection and host response to Anaplasma marginale in one closed herd in the dry tropical forest of Costa Rica. The study subjects were the dams and their calves born during 1 breeding season (1995‐1996). All cows were sampled at 3 month intervals for antibody detection using a competitive ELISA (cELISA) and for antigen detection using PCR/nonradioactive probe assay. All 24 calves born during the study were individually identified at birth and subsequently sampled each month for PCR and cELISA. Ticks were identified from all animals throughout the entire study period. The results from this study confirmed that the cELISA is a reliable assay for identifying new and carrier infections and that carrier infections can exist at levels below that detectable by PCR. In addition, it was demonstrated that calves born in this region will most likely be exposed to Anaplasma within the first 6 months of age.
A Partially Protective Clone from Cowdria ruminantium Identified by Using a Salmonella Vaccine Delivery Systema
Authors: BRAYTON K. A. , VAN DER WALT M. , VOGEL S. W. , ALLSOPP B. A.
Published: February 2006

ABSTRACT: A strategy has been developed to screen the Cowdria genome, using a Salmonella vaccine delivery system, to identify genes that code for protection‐stimulating proteins. We have cloned mini‐libraries of Cowdria into this Salmonella system, used the recombinant bacteria to immunize outbred mice, and then challenged them after two weeks with a lethal dose of Cowdria. When one of these mini‐libraries was tested in a group of 5 mice, one mouse lived much longer than the others. The experiment was repeated with each of the clones from the mini‐library being tested individually in 10 mice, and one mouse survived the challenge. This clone has been tested repeatedly in larger groups of mice and is proven to protect 14% of outbred mice against a lethal Cowdria challenge.
Evaluation of the MAP‐1B ELISA for Cowdriosis with Field Sera from Livestock in Zimbabwea
Authors: MAHAN S. M. , SEMU S. M. , PETER T. F. , JONGEJAN F.
Published: February 2006

ABSTRACT: The Map 1B (Senegal) antigen‐based indirect ELISA was evaluated in Zimbabwe with field sera from heartwater‐free and heartwater‐endemic areas. Of 205 sheep sera samples from a heartwater‐free area, 34 were negative and 171 were positive by immunoblotting. These 171 samples were classified as false positives. Of the same 205 samples, 199 were negative and only 6 were positive by the MAP 1B ELISA. Of 72 cattle samples tested from a similar area, 71 were negative, with only 1 sample positive by MAP 1B ELISA. By immunoblotting, 43 of 72 cattle sera were negative and 29 were positive. Of the 46 goat samples tested from a heartwater‐free area, only 2 were positive by the MAP 1B ELISA. Based on these results, the MAP 1B ELISA was more specific for heartwater than the immunoblotting assay. Of 96 and 282 cattle sera analyzed from heartwater‐endemic farms in the lowveld and highveld of Zimbabwe, respectively, approximately 33% were positive by the MAP 1B ELISA. Goat sera from the same farms had a higher sero‐prevalence (>90%). The implications of these results for serodiagnosis of heartwater using the MAP 1B ELISA will be discussed.