Biomedical subjects

T C Mettenleiter

Publications and source records attributed to T C Mettenleiter.

At least 19 recordsLinked to original sources

PubMed2005

Fish rhabdoviruses: molecular epidemiology and evolution.

Rhabdoviruses may cause serious diseases in wild and farmed fish. Within the Rhabdoviridae six genera have been established: Ephemerovirus, Cytorhabdovirus, Nucleorhabdovirus, Lyssavirus, Vesiculovirus, and Novirhabdovirus. Viruses that infect fish are official or tentative members of the genera Vesiculovirus and Novirhabdovirus, or are listed as unassigned rhabdoviruses. In this report, we summarize and discuss published and our own unpublished data on the molecular epidemiology and phylogeography of fish rhabdoviruses including intrapopulational differences and subgrouping of fish rhabdoviruses, in particular the species spring viremia of carp virus (SVCV), infectious hematopoietic necrosis virus (IHNV) and viral hemorrhagic septicemia virus (VHSV).

Animals↗

PubMed2004

[Zoonoses in working- and wild animals and their significance in Germany. An overview].

The control of infectious diseases transmitted from animals to humans (zoonoses) was recently put on a new basis in the European Union when a new Zoonoses Directive entered into force. Brucellosis, campylobacteriosis, echinococcosis, listeriosis, salmonellosis, trichinosis, and the respective causative agents, tuberculosis due to Mycobacterium bovis, and verotoxigenic Escherichia coli must be included in monitoring. Additional zoonoses and zoonotic agents are to be monitored according to the epidemiological situation. Against this background, the current knowledge on important zoonoses transmitted from livestock and some wildlife animals to humans as well as the epidemiological situation in Germany with regard to these diseases is summarized.

Animals↗

PubMed2003

Orexin neurons project to diverse sympathetic outflow systems.

The viral transneuronal labeling method was used to demonstrate that orexin-containing neurons of the lateral hypothalamic area (LHA) are linked via multisynaptic connections to different sympathetic outflow systems. Two different types of transneuronal tracing experiments were performed: single- and double-virus studies. In the first series of experiments, Bartha pseudorabies virus (PRV), a retrograde transneuronal tracer, was injected into single sympathetic targets, viz., stellate ganglion, adrenal gland, celiac ganglion, and kidney. Six to 7 days post-injection, orexin (hypocretin) neurons were transneuronally labeled. In a second set of experiments, the double-virus tracing method was used to determine whether single orexin LHA neurons are linked to two different sympathetic outflow systems. Two isogenic forms of Bartha PRV were used that differed by a single gene. beta-Galactosidase Bartha PRV was injected into the stellate ganglion and green fluorescent protein Bartha PRV into the adrenal gland of the same rat. The reverse placement of viral injections was made in another set of rats. In both paradigms, some orexin LHA neurons were transneuronally labeled with both viruses, indicating that they are capable of modulating multiple sympathetic outflow systems. These findings raise the possibility that orexin LHA neurons regulate general sympathetic functions, such as those that occur during arousal or the fight-or-flight response.

Adrenergic Fibers↗

PubMed2003

Single CNS neurons link both central motor and cardiosympathetic systems: a double-virus tracing study.

Two anatomical experiments were performed to test the hypothesis that single CNS neurons link the central areas that regulate the somatomotor and sympathetic systems. First, the retrograde neuronal tracer cholera toxin beta-subunit was injected into the lateral parafascicular thalamic nucleus, a region that projects to both the motor cortex and striatum. Several days later, a second injection of the retrograde transneuronal tracer, pseudorabies virus (PRV), was made in the same rats in the stellate ganglion, which provides the main sympathetic supply to the heart. Using immunohistochemical methods, we demonstrate that the cholinergic neurons of the pedunculopontine tegmental nucleus (PPN) are connected to both systems. The second experiment used two isogenic strains of Bartha PRV as double transneuronal tracers. One virus contained the unique gene for green fluorescent protein (GFP) and the other had the unique gene for beta-galactosidase (beta-gal). GFP-PRV was injected in the stellate ganglion and beta-gal-PRV was injected into the primary motor cortex. Double-labeled neurons were found in the lateral hypothalamic area (50% contained orexin) and PPN (approximately 95% were cholinergic). Other double-labeled neurons were identified in the deep temporal lobe (viz., amygdalohippocampal zone and lateral entorhinal cortex), posterior hypothalamus, ventral tuberomammillary nucleus, locus coeruleus, laterodorsal tegmental nucleus, periaqueductal gray matter, dorsal raphe nucleus, and nucleus tractus solitarius. These results suggest these putative command neurons integrate the somatomotor and cardiosympathetic functions and may affect different behaviors (viz., arousal, sleep, and/or locomotion).

Animals↗

PubMed2003

Eradication of Aujeszky's disease in Germany.

Aujeszky's disease (AD) manifested itself in both German states in 1960. Owing to the historical development, in the subsequent two decades, the development of the disease and of its control in the Western and Eastern parts of Germany went different ways. This article describes differences and particularities in the development of AD in Germany leading to the establishment of a national AD eradication programme after re-unification of the two German states at the beginning of the last decade. The basic principles of the German AD eradication programme are described, and the results of 10 years of efforts to control the disease are presented and discussed. Without any doubt, as in other European countries, implementation of the national eradication programme resulted in a considerable progress in the eradication of AD. Since the eradication programme has been established in 1989, particularly in West Germany, the number of AD outbreaks has decreased steadily from about 2000 cases in 1987 to 0 cases recorded in 2001. Recently, Germany has been declared as officially AD-free by the European Commission.

Animals↗

PubMed2002

CNS inputs to the suprachiasmatic nucleus of the rat.

The neural circuits that modulate the suprachiasmatic nucleus (SCN) of the rat were studied with the retrograde transneuronal tracer--pseudorabies virus. First-order afferents were also identified using cholera toxin beta subunit. Olfactory processing regions (viz., main olfactory bulb, anterior olfactory nucleus, taenia tecta, endopiriform nucleus, medial amygdaloid nucleus, piriform cortex, and posteriomedial cortical amygdaloid nucleus) were virally labeled. The subfornical organ directly innervates SCN; two other circumventricular organs: organum vasculosum of the lamina terminalis and area postrema provide multisynaptic inputs. Direct limbic afferents arise from lateral septum, bed nucleus of the stria terminalis, amygdalohippocampal zone, and ventral subiculum; multineuronal connections come from the basolateral and basomedial amygdaloid nuclei, ventral hippocampus, amygdalopiriform area, as well as lateral entorhinal, perirhinal, and ectorhinal cortices. Most preoptic regions project directly to SCN. Multisynaptic inputs come from the lateral preoptic region. Hypothalamic inputs originate from the anterior, arcuate, dorsal, dorsomedial, lateral, paraventricular, posterior, periventricular posterior, retrochiasmatic, subparaventricular, ventromedial and tuberomammillary nuclei. Paraventricular thalamic nucleus, intergeniculate leaflet and zona incerta directly innervate SCN. Polyneuronal inputs arise from the subparafascicular parvicellular thalamic nucleus. Brainstem afferents originate from the pretectum, superior colliculus, periaqueductal gray matter, parabrachial nucleus, pedunculopontine nucleus, raphe system, locus coeruleus, nucleus incertus and reticular formation. Nucleus tractus solitarius, C3 catecholamine region, rostral ventrolateral medulla and spinal trigeminal nucleus provide indirect inputs. We propose that the SCN receives feedback primarily from interoceptive systems such as the circumventricular, autonomic, and neuroendocrine systems that are important in the central regulation of glucose metabolism (e.g., insulin and glucocorticoids).

Afferent Pathways↗

PubMed2001

Protection of chickens from lethal avian influenza A virus infection by live-virus vaccination with infectious laryngotracheitis virus recombinants expressing the hemagglutinin (H5) gene.

The H5 hemagglutinin (HA) gene of a highly pathogenic avian influenza virus (AIV) isolate (A/chicken/Italy/8/98) was cloned and sequenced, and inserted at the non-essential UL50 (dUTPase) gene locus of a virulent strain of infectious laryngotracheitis virus (ILTV). Northern and Western blot analyses of the obtained ILTV recombinants demonstrated stable expression of the HA gene under control of the human cytomegalovirus immediate-early gene promoter. In vitro replication of the HA-expressing ILTV mutants was not affected, and infection of chickens revealed a reduced but still considerable virulence, similar to that of a UL50 gene deletion mutant without foreign gene insertion. The immunized animals produced specific antibodies against ILTV and AIV HA, and were protected against challenge infections with either virulent ILTV, or two different highly pathogenic AIV strains (A/chicken/Italy/8/98, A/chicken/Scotland/59). After challenge, no ILTV could be reisolated from protected animals, and shedding of AIV was considerably reduced. Thus, although attenuation remains to be improved, genetically engineered ILTV live-virus vaccines might be used as vectors to protect chickens also against other pathogens.

Animals↗

PubMed2001

Synaptic and neurotransmitter activation of cardiac vagal neurons in the nucleus ambiguus.

Cardiac vagal neurons play a critical role in the control of heart rate and cardiac function. These neurons, which are primarily located in the nucleus ambiguus (NA) and the dorsal motor nucleus of the vagus (DMNX), dominate the neural control of heart rate under normal conditions. Cardiac vagal activity is diminished and unresponsive in many disease states, while restoration of parasympathetic activity to the heart lessens ischemia and arrhythmias and decreases the risk of sudden death. Recent work has demonstrated that cardiac vagal neurons are intrinsically silent and therefore rely on synaptic input to control their firing. To date, three major synaptic inputs to cardiac vagal neurons have been identified. Stimulation of the nucleus tractus solitarius evokes a glutamatergic pathway that activates both NMDA and non-NMDA glutamatergic postsynaptic currents in cardiac vagal neurons. Acetylcholine excites cardiac vagal neurons via three mechanisms, activating a direct ligand-gated postsynaptic nicotinic receptor, enhancing postsynaptic non-NMDA currents, and presynaptically by facilitating transmitter release. This enhancement by nicotine is dependent upon activation of pre- and postsynaptic P-type voltage-gated calcium channels. Additionally, there is a GABAergic innervation of cardiac vagal neurons. The transsynaptic pseudorabies virus that expresses GFP (PRV-GFP) has been used to identify, for subsequent electrophysiologic study, neurons that project to cardiac vagal neurons. Bartha PRV-GFP-labeled neurons retain their normal electrophysiological properties, and the labeled baroreflex pathways that control heart rate are unaltered by the virus.

Animals↗

PubMed2001

Pseudorabies virus UL37 gene product is involved in secondary envelopment.

Herpesvirus envelopment is a two-step process which includes acquisition of a primary envelope resulting from budding of intranuclear capsids through the inner nuclear membrane. Fusion with the outer leaflet of the nuclear membrane releases nucleocapsids into the cytoplasm, which then gain their final envelope by budding into trans-Golgi vesicles. It has been shown that the UL34 gene product is required for primary envelopment of the alphaherpesvirus pseudorabies virus (PrV) (B. G. Klupp, H. Granzow, and T. C. Mettenleiter, J. Virol. 74:10063-10073, 2000). For secondary envelopment, several virus-encoded PrV proteins are necessary, including glycoproteins E, I, and M (A. R. Brack, J. M. Dijkstra, H. Granzow, B. G. Klupp, and T. C. Mettenleiter, J. Virol. 73:5364-5372, 1999). We show here that the product of the UL37 gene of PrV, which is a constituent of mature virions, is involved in secondary envelopment. Replication of a UL37 deletion mutant, PrV-DeltaUL37, was impaired in normal cells; this defect could be complemented on cells stably expressing UL37. Ultrastructural analysis demonstrated that intranuclear capsid maturation and budding of capsids into and release from the perinuclear space were unimpaired. However, secondary envelopment was drastically reduced. Instead, apparently DNA-filled capsids accumulated in the cytoplasm in large aggregates similar to those observed in the absence of glycoproteins E/I and M but lacking the surrounding electron-dense tegument material. Although displaying an ordered structure, capsids did not contact each other directly. We postulate that the UL37 protein is necessary for correct addition of other tegument proteins, which are required for secondary envelopment. In the absence of the UL37 protein, capsids interact with each other through unknown components but do not acquire the electron-dense tegument which is normally found around wild-type capsids during and after secondary envelopment. Thus, apposition of the UL37 protein to cytoplasmic capsids may be crucial for the addition of other tegument proteins, which in turn are able to interact with viral glycoproteins to mediate secondary envelopment.

Animals↗

PubMed2001

Glycoprotein D-independent infectivity of pseudorabies virus results in an alteration of in vivo host range and correlates with mutations in glycoproteins B and H.

Infection of cells by herpesviruses is initiated by the interaction of viral envelope glycoproteins with cellular receptors. In the alphaherpesvirus pseudorabies virus (PrV), the causative agent of Aujeszky's disease in pigs, the essential glycoprotein D (gD) mediates secondary attachment of virions to target cells by binding to newly identified cellular receptors (R. J. Geraghty, C. Krummenacher, G. H. Cohen, R. J. Eisenberg, and P. G. Spear, Science 280:1618-1620, 1998). However, in the presence of compensatory mutations, infection can also occur in the absence of gD, as evidenced by the isolation in cell culture of an infectious gD-negative PrV mutant (PrV-gD(-) Pass) (J. Schmidt, B. G. Klupp, A. Karger, and T. C. Mettenleiter, J. Virol. 71:17-24, 1997). PrV-gD(-) Pass is replication competent with an only moderate reduction in specific infectivity but appears to bind to receptors different from those recognized by wild-type PrV (A. Karger, J. Schmidt, and T. C. Mettenleiter, J. Virol. 72:7341-7348, 1998). To analyze whether this alteration in receptor usage in vitro influences infection in vivo, the model host mouse and the natural host pig were intranasally infected with PrV-gD(-) Pass and were compared to animals infected by wild-type PrV. For mice, a comparable progress of disease was observed, and all animals infected with mutant virus died, although they exhibited a slight delay in the onset of symptoms and, correspondingly, a longer time to death. In contrast, whereas wild-type PrV-infected pigs showed clinical signs and histological and histopathological findings typical of PrV infection, no signs of disease were observed after infection with PrV-gD(-) Pass. Moreover, in these animals, virus-infected cells were not detectable by immunohistochemical staining of different organ samples and no virus could be isolated from nasal swabs. Mutations in glycoproteins B and H were found to correlate with, and probably contribute to, gD-independent infectivity. In conclusion, although PrV-gD(-) Pass is virulent in mice, it is apparently unable to infect the natural host, the pig. This altered host range in vivo correlates with a difference of receptor usage in vitro and demonstrates for the first time the importance of gD receptors in alphaherpesvirus infection of an animal host.

Animals↗

PubMed2001

The absence of glycoprotein gL, but not gC or gK, severely impairs pseudorabies virus neuroinvasiveness.

Penetration and propagation of herpesviruses in the nervous system require the action of several glycoproteins. To assay for a function of glycoproteins gC, gK, and gL in the neuroinvasiveness of pseudorabies virus (PrV), deletion mutants lacking one of these glycoproteins and corresponding rescuants were inoculated in the nasal cavity of adult mice. We demonstrate that the lack of gL almost prevented the virus from penetrating and propagating in trigeminal, sympathetic, and parasympathetic tracks innervating the nasal cavity, while the lack of gC and gK only slowed the invasion of the nervous system. The conclusion of this and previous studies is that only gB, gD, gH, and gL are indispensable for penetration into neurons, while gB, gH, and gL (and, in some categories of neurons, also gE and gI) are necessary for transneuronal transfer in the mouse model. The deletion of other glycoprotein genes has little effect on PrV neuroinvasiveness although it may affect the dissemination of the virus.

Administration, Intranasal↗

PubMed2001

Restoration of function of carboxy-terminally truncated pseudorabies virus glycoprotein B by point mutations in the ectodomain.

Glycoprotein B (gB) of pseudorabies virus (PrV) is essential for virus entry into target cells and direct viral cell-to-cell spread. Recently, we described a carboxy-terminally truncated derivative of PrV gB, gB-007, which was inefficiently incorporated into virions, was unable to complement infectivity, but was fully capable of restoring direct viral cell-to-cell spread of gB-negative PrV (R. Nixdorf, B. G. Klupp, and T. C. Mettenleiter, J. Virol. 74:7137-7145, 2000). Since recombinant PrV-007, which expresses gB-007 instead of wild-type gB, was able to spread directly from cell to cell, we attempted to obtain compensatory mutations leading to restoration of the entry defect by performing serial passages in cell culture. This procedure has previously been used to successfully restore entry defects in gD- or gL-deficient PrV mutants. From an initial titer of 100 PFU per ml in the supernatant, titers increased, reaching wild-type levels of up to 10(7) PFU after ca. 20 passages. One single-plaque isolate of the passaged mutant, designated PrV-007Pass, was further characterized. PrV-007Pass gB was efficiently incorporated into the viral envelope and restored infectivity to a gB-negative PrV mutant, PrV-gB(-). Interestingly, localization of PrV-007Pass gB in the plasma membrane was similar to that of PrV-007. In contrast, wild-type gB is mainly found in intracellular vesicles. Marker rescue experiments and trans-complementation assays demonstrated the presence of compensatory mutations within the gB gene of PrV-007Pass. DNA sequencing revealed two point mutations in the gB open reading frame of PrV-007Pass, resulting in amino acid substitutions at positions 305 and 744 of gB, both of which are required for compensation of the defect in PrV-007. Our data again demonstrate the power of reversion analysis of herpesviruses and suggest that cytosolic and ectodomains play a role in incorporation of gB into virions.

Amino Acid Sequence↗

PubMed2001

Egress of alphaherpesviruses: comparative ultrastructural study.

Egress of four important alphaherpesviruses, equine herpesvirus 1 (EHV-1), herpes simplex virus type 1 (HSV-1), infectious laryngotracheitis virus (ILTV), and pseudorabies virus (PrV), was investigated by electron microscopy of infected cell lines of different origins. In all virus-cell systems analyzed, similar observations were made concerning the different stages of virion morphogenesis. After intranuclear assembly, nucleocapsids bud at the inner leaflet of the nuclear membrane, resulting in enveloped particles in the perinuclear space that contain a sharply bordered rim of tegument and a smooth envelope surface. Egress from the perinuclear cisterna primarily occurs by fusion of the primary envelope with the outer leaflet of the nuclear membrane, which has been visualized for HSV-1 and EHV-1 for the first time. The resulting intracytoplasmic naked nucleocapsids are enveloped at membranes of the trans-Golgi network (TGN), as shown by immunogold labeling with a TGN-specific antiserum. Virions containing their final envelope differ in morphology from particles within the perinuclear cisterna by visible surface projections and a diffuse tegument. Particularly striking was the addition of a large amount of tegument material to ILTV capsids in the cytoplasm. Extracellular virions were morphologically identical to virions within Golgi-derived vesicles, but distinct from virions in the perinuclear space. Studies with gB- and gH-deleted PrV mutants indicated that these two glycoproteins, which are essential for virus entry and direct cell-to-cell spread, are dispensable for egress. Taken together, our studies indicate that the deenvelopment-reenvelopment process of herpesvirus maturation also occurs in EHV-1, HSV-1, and ILTV and that membrane fusion processes occurring during egress are substantially different from those during entry and direct viral cell-to-cell spread.

Alphaherpesvirinae↗

PubMed2001

Activity of cardiorespiratory networks revealed by transsynaptic virus expressing GFP.

A fluorescent transneuronal marker capable of labeling individual neurons in a central network while maintaining their normal physiology would permit functional studies of neurons within entire networks responsible for complex behaviors such as cardiorespiratory reflexes. The Bartha strain of pseudorabies virus (PRV), an attenuated swine alpha herpesvirus, can be used as a transsynaptic marker of neural circuits. Bartha PRV invades neuronal networks in the CNS through peripherally projecting axons, replicates in these parent neurons, and then travels transsynaptically to continue labeling the second- and higher-order neurons in a time-dependent manner. A Bartha PRV mutant that expresses green fluorescent protein (GFP) was used to visualize and record from neurons that determine the vagal motor outflow to the heart. Here we show that Bartha PRV-GFP-labeled neurons retain their normal electrophysiological properties and that the labeled baroreflex pathways that control heart rate are unaltered by the virus. This novel transynaptic virus permits in vitro studies of identified neurons within functionally defined neuronal systems including networks that mediate cardiovascular and respiratory function and interactions. We also demonstrate superior laryngeal motorneurons fire spontaneously and synapse on cardiac vagal neurons in the nucleus ambiguus. This cardiorespiratory pathway provides a neural basis of respiratory sinus arrhythmias.

Animals↗

PubMed2001

[DNA vaccines for veterinary medicine].

DNA vaccination represents one of the most recent novel approaches to vaccine development. Experimentally, DNA vaccines induce a broad range of long lasting immune responses including humoral and cell-mediated immunity against infectious diseases in humans and animals. Furthermore, DNA vaccines are potentially useful for the treatment of autoimmune diseases or cancer. However, most information on the efficacy of DNA vaccines has been generated in mice and studies in larger animals are limited. In this review, the potential application of DNA vaccines in livestock and pet animals are discussed. The principle of this new technology, its potency and future perspectives for use in veterinary medicine will be outlined.

Animal Diseases↗

PubMed2000

A study of the ability of a TK-negative and gI/gE-negative pseudorabies virus (PRV) mutant inoculated by different routes to protect pigs against PRV infection.

The capacity of a TK-negative (TK-) and gI/gE-negative (gI/gE-) pseudorabies virus (PRV) mutant to protect pigs against Aujeszky's disease carried out by experimental infection with a virulent PRV strain, was tested. There were three groups, each of four susceptible pigs which were inoculated twice by two different schedules. Group 1 received the modified virus by the intradermal (first inoculation)-intramuscular (second inoculation) routes; group 2 was treated by the intranasal (first inoculation)-intramuscular (second inoculation) routes. The third group was left untreated as the control. All of the pigs were challenged intranasally with a virulent PRV strain and they were subsequently injected with dexamethasone. Two pigs in each group were necropsied on days 5 and 15 after dexamethasone inoculation. The challenge exposure resulted in mild clinical signs, increase in growth and a shorter period of virus shedding in vaccinated pigs, whereas the control group showed severe signs of Aujeszky's disease. No difference in the titre of the virulent virus which was excreted by pigs of all three groups, was observed and all animals seroconverted. Both the mutant strain and the wild-type virus established a latent infection although only the latter was reactivated and shed. Slight lesions were observed in target tissues of the vaccinated animals and no significant differences were detected between the two inoculation schedules.

Administration, Intranasal↗

PubMed2000

The non-essential UL50 gene of avian infectious laryngotracheitis virus encodes a functional dUTPase which is not a virulence factor.

The DNA sequence of the infectious laryngotracheitis virus (ILTV) UL50, UL51 and UL52 gene homologues was determined. Although the deduced UL50 protein lacks the first of five conserved domains of the corresponding proteins of mammalian alphaherpesviruses, the ILTV gene product was also shown to possess dUTPase activity. The generation of UL50-negative ILTV mutants was facilitated by recombination plasmids encoding green fluorescent protein (GFP), and expression constructs of predicted transactivator proteins of ILTV (alphaTIF, ICP4) were successfully used to increase the infectivity of viral genomic DNA. A GFP-expressing UL50-deletion mutant of ILTV showed reduced cell-to-cell spread in vitro, and was attenuated in vivo. A similar deletion mutant without the foreign gene, however, propagated like wild-type ILTV in cell culture and was pathogenic in chickens. We conclude that the viral dUTPase is not required for efficient replication of ILTV in the respiratory tract of infected animals. The replication defect of the GFP-expressing ILTV recombinant is most likely caused by toxic effects of the reporter gene product, since spontaneously occurring inactivation mutants exhibited wild-type-like growth.

Amino Acid Sequence↗

PubMed2000

Characterization of the replication origin (Ori(S)) and adjoining parts of the inverted repeat sequences of the pseudorabies virus genome.

The DNA sequence of a 2.4 kbp fragment located in the internal and terminal inverted repeat sequences of the pseudorabies virus genome determined in this study closes a gap between the previously described genes for the ICP4 and ICP22 homologues. The novel sequence contains no conserved herpesvirus open reading frames. Northern blot and cDNA analyses revealed a viral immediate-early transcript of 1.8 kb, which is spliced by the removal of two small introns close to its 5' end and which presumably represents the mRNA of the downstream open reading frame encoding the ICP22 homologue. Upstream of the transcribed region, an imperfect set of three directly repeated sequences was identified. Each of them contains a complementary pair of the alphaherpesvirus origin-binding protein recognition motif GTTCGCAC, spaced by AT-rich sequences. In vitro studies confirmed that the DNA fragment analysed includes a functional origin of viral DNA replication.

Animals↗