Modelling of control options for an outbreak of Mycoplasma gallisepticum in egg production: a decision support tool.

Mycoplasma gallisepticum (MG) is a bacterium that causes respiratory disease in chickens, leading to reduced egg production. A dynamic simulation model was developed that can be used to assess the costs and benefits of control using antimicrobials or vaccination in caged or free range systems. The intended users are veterinarians and egg producers. A user interface is provided for input of flock specific parameters. The economic consequence of an MG outbreak is expressed as a reduction in expected egg output. The model predicts that either vaccination or microbial treatment can approximately halve potential losses from MG in some circumstances. Sensitivity analysis is used to test assumptions about infection rate and timing of an outbreak. Feedback from veterinarians points to the value of the model as a discussion tool with producers.

Physiological actions of the neuropeptide Antho-RNamide on antagonistic muscle systems in sea anemones.

Antagonistic contractions of longitudinal and circular muscles in the body wall cause shape changes in sea anemones. Single electrode voltage clamp recordings from Calliactis parasitica body wall preparations distinguish two cell types (Types 1 and 2) with different time courses of evoked inward current. The anthozoan neuropeptide Antho-RNamide (L-3-phenyllactyl-Leu-Arg-Asn-NH2) excites longitudinal body wall preparations and Type 1 cells, but inhibits circular body wall preparations and Type 2 cells. Type 1 and Type 2 cells may therefore belong to longitudinal and circular myoepithelium respectively. Antho-RNamide may open Ca2+ channels in longitudinal muscle cells and K+ channels in circular muscle cells. This opposite action of a neuropeptide is of significance in the control of antagonistic contractions.

The anthozoan neuropeptide Antho-RWamide I modulates Ca2+ current in sea anemone myoepithelial cells.

The anthozoan neuropeptide Antho-RWamide I (< Glu-Ser-Leu-Arg-Trp-NH2) excites contraction of endodermal muscles in sea anemones. Single electrode voltage clamp recordings from semi-intact preparations of endodermal myoepithelial cells reveal that Antho-RWamide I increases an inward Ca2+ current. Evidence for the involvement of a Ca2+ current in contraction was supported by the observation that Cd2+ abolished spontaneous contractions and reduced inward current. Contractions and inward currents induced by Antho-RWamide I were not, however, completely abolished in the presence of Cd2+. We conclude that Antho-RWamide, a putative neurotransmitter at sea anemone smooth muscle, acts by opening, either directly or indirectly, Ca2+ channels in the muscle membrane.

Synaptic potentials recorded from sea anemone muscle cells in situ.

Three pulse types, A,B, and C, can be recorded from column ring preparations of the sea anemone Calliactis parasitica (Couch). Lucifer Yellow injection confirmed that the recording sites are intracellular in endodermal myoepithelial cells. Type C pulses are spontaneously active depolarizing pulses, arising from a resting potential of around -60mV, and are usually < 5 mV in amplitude. When the cells were voltage-clamped these pulses reversed at around 0 mV membrane potential; therefore, we conclude that they are synaptic potentials. In some cells an increase in frequency of type C pulses accompanied contraction of the circular muscle field. Inward currents in these myoepithelial cells are carried by Ca2+, and although Na(+)-free solutions did not affect inward currents they did eliminate type C pulses, i.e., these synaptic potentials may result from activity of Na(+)-dependent nerve cells.

The expansion behaviour of sea anemones may be coordinated by two inhibitory neuropeptides, Antho-KAamide and Antho-RIamide.

Antho-KAamide (L-3-phenyllactyl-Phe-Lys-Ala-NH2) and Antho-RIamide (L-3-phenyllactyl-Tyr-Arg-Ile-NH2) are novel neuropeptides isolated from the sea anemone Anthopleura elegantissima. They both inhibited spontaneous contractions of isolated muscle preparations from a wide variety of anemone species (threshold around 10(-7) M). Their actions were universal in that they inhibited every muscle preparation tested, regardless of whether the muscle group was located in the ectoderm or endoderm, or was oriented in a circular or longitudinal direction. Injection of Antho-KAamide or Antho-RIamide into the coelenteron of intact sea anemones resulted in a marked expansion of the animals. Similar shape changes followed feeding or exposure to soluble food extracts. Therefore, we hypothesize that nerve cells that release Antho-KAamide and Antho-RIamide are involved in the expansion phase of feeding behaviour in sea anemones.

Isolation of

Using a radioimmunoassay against the C-terminal sequence Arg-Pro-NH2 (RPamide) we have isolated the neuropeptide

Isolation of Leu-Pro-Pro-Gly-Pro-Leu-Pro-Arg-Pro-NH2 (Antho-RPamide), an N-terminally protected, biologically active neuropeptide from sea anemones.

Using a radioimmunoassay against the C-terminal sequence Arg-Pro-NH2 (RPamide), we have isolated the peptide Leu-Pro-Pro-Gly-Pro-Leu-Pro-Arg-Pro-NH2 (Antho-RPamide) from an extract of the sea anemone Anthopleura elegantissima. Antho-RPamide is located in neurons of sea anemones. Application of low concentrations of Antho-RPamide to tentacle preparations of sea anemones strongly increased the frequency and duration of spontaneous contractions, suggesting that this peptide is involved in neurotransmission. Antho-RPamide has a free N-terminus, yet its X-Pro-Pro sequence makes it relatively resistant to degradation by nonspecific aminopeptidases. Thus, we have discovered another strategy by which sea anemones protect the N-termini of their bioactive neuropeptides.

Isolation of two novel neuropeptides from sea anemones: the unusual, biologically active L-3-phenyllactyl-Tyr-Arg-Ile-NH2 and its des-phenyllactyl fragment Tyr-Arg-Ile-NH2.

Using a radioimmunoassay for the carboxyl-terminal sequence Arg-Val-NH2, two novel peptides were purified from extracts of the sea anemone Anthopleura elegantissima. These peptides were L-3-phenyllactyl-Tyr-Arg-Ile-NH2 (name: Antho-RIamide I) and its des-phenyllactyl fragment Tyr-Arg-Ile-NH2 (Antho-RIamide II). Immunocytochemical staining showed that these peptides were localized in neurons of sea anemones. Application of low concentrations (10(-8) M) of Antho-RIamide I inhibited spontaneous contractions in several muscle groups of sea anemones, whereas Antho-RIamide II was inactive. Antho-RIamide I is the second neuropeptide from sea anemones that bears the unusual, amino-terminal L-3-phenyllactyl blocking group. We suggest that this group renders the peptide resistant agaist degradation by nonspecific aminopeptidases. In addition, the L-3-phenyllactyl residue might also play a role in receptor binding.

Effects of three anthozoan neuropeptides, Antho-RWamide I, Antho-RWamide II and Antho-RFamide, on slow muscles from sea anemones.

Antho-RWamide I (less than Glu-Ser-Leu-Arg-Trp-NH2) and Antho-RWamide II (less than Glu-Gly-Leu-Arg-Trp-NH2), the second and third anthozoan neuropeptides to be identified, both induced slow contractions of several endodermal muscles in four species of sea anemone. In a fifth species, Protanthea simplex, Antho-RWamide II, but not Antho-RWamide I, evoked contractions of body wall muscles. Isolated, trimmed sphincter muscle preparations of Calliactis parasitica contracted at a threshold concentration of 10(-9) mol l-1 Antho-RWamide II. Antho-RWamide II was more potent than Antho-RWamide I. Unlike the responses to Antho-RFamide (the first anthozoan neuropeptide described), these were simple contractions with no change in spontaneous activity. The Antho-RWamides did not excite electrical activity in any of the three known conducting systems (the through-conducting nerve net and the slow systems 1 and 2), indicating that they may be acting directly on endodermal muscles. Application of peptides to smooth muscle cells, isolated from the sphincter of C. parasitica, confirmed that Antho-RWamide I and II act directly on the muscle. We conclude that the Antho-RWamides may be neurotransmitters at some neuromuscular synapses in sea anemones.

Neurones and neuropeptides in coelenterates.

The first nervous system probably evolved in coelenterates. Many neurons in coelenterates have morphological characteristics of both sensory and motor neurones, and appear to be multifunctional. Using immunocytochemistry with antisera to the sequence Arg-Phe-NH2 (RFamide), RFamide-like peptides were demonstrated in the nervous systems of all classes of coelenterates. Using a radioimmunoassay for RFamide, three such peptides were isolated from the sea anemone Anthopleura elegantissisma and sequenced: less than Glu-Gly-Arg-Phe-NH2 (Antho-RFamide), less than Glu-Ser-Leu-Arg-Trp-NH2 (Antho-RWamide I) and less than Glu-Gly-Leu-Arg-Trp-NH2 (Antho-RWamide II). The general structure of these peptides can be described as less than Glu...Arg-X-NH2, where X is an aromatic amino acid. From the hydromedusa Polyorchis penicillatus, the peptide less than Glu-Leu-Leu-Gly-Gly-Arg-Phe-NH2 (Pol-RFamide I) was isolated, which also belongs to the less than Glu...Arg-X-NH2 family. Using specific antisera it was shown that all four peptides were located in neurones. Application of low doses of Antho-RFamide, or Antho-RWamide I or II induced contractions of endodermal muscles of sea anemones. This indicates that these neuropeptides play a role in neurotransmission.

Delayed initiation of SS1 pulses in the sea anemone Calliactis parasitica: evidence for a fourth conducting system.

1. Single electrical shocks to the column sometimes elicit a series of 1-6 pulses in the SS1 (ectodermal slow system) but the first pulse does not appear until 5-28 s after stimulation. These pulses occur in addition to the early SS1 pulse which follows every shock and which has a conduction delay of less than 1 s. 2. The threshold of the delayed SS1 response is different from the thresholds of the three known conducting systems (through-conducting nerve net, SS1, and SS2). 3. In the case of stimulation of the column, the delayed SS1 pulses do not arise at the point of stimulation but probably originate in the tentacles or upper column. The pulse origin can shift during a single burst. 4. The pathway from the point of stimulation to the site of origin of delayed SS1 pulses is endodermal. We propose that this pathway represents a fourth conducting system (Delayed Initiation System--DIS). The DIS must connect, across the mesogloea, with the ectodermal SS1. The long pulse delay and repetitive firing may derive from pacemaker activity in the DIS. The DIS pacemakers closely resemble the pacemakers connected to the through-conducting nerve net. The DIS may be neuronal. 5. Delayed SS1 pulse bursts from unattached anemones showed an earlier onset, and more pulses/burst, than those from attached anemones. 6. Delayed SS1 pulses can also be evoked by electrical, and in some cases mechanical, stimulation of the pedal disc, tentacles, and pharynx, but there are regional differences in the number of pulses evoked, in their delay, and in their site of origin.

Two slow conduction systems co-ordinate shell-climbing behaviour in the sea anemone Calliactis parasitica.

1. Pulses in two slow conducting systems, the ectodermal SS 1 and the endodermal SS 2, were recorded during shell-climbing behaviour. The mean pulse interval of SS 1 pulses was 7-4 s and that of SS 2 pulses was 6-4 s. Activity in both systems may arise as a sensory response of tentacles to shell contact, but the SS 1 and SS 2 may not share the same receptors. 2. Electrical stimulation of the SS 1 and SS 2 together, at a frequency of 1 shock every 5 s, elicits shell-climbing behaviour in the absence of a shell. 3. Low-frequency nerve-net activity (about 1 pulse every 15 s) accompanies column bending during both normal and electrically elicited responses. This activity probably arises as a result of column bending and is not due to a sensory response to the shell.

Control of mouth opening and pharynx protrusion during feeding in the sea anemone Calliactis parasitica.

1. Activity in all three known conducting systems (the nerve net, SS1, and SS2) may accompany feeding in Calliactis. The most marked response is an increase in pulse frequency in the SS2 (the endodermal slow conducting system) during mouth opening and pharynx protrusion. 2. Electrical stimulation of the SS2 at a frequency of one shock every 5 s elicits mouth opening and pharynx protrusion in the absence of food. 3. A rise in SS2 pulse frequency is also evoked by food extracts, some amino acids, and in particular by the tripeptide reduced glutathione, which produces a response at a concentration of 10(-5) M. 4. Although the SS2 is an endodermal system, the receptors involved in the response to food appear to be ectodermal. 5. The epithelium that lines the pharynx conducts SS1 pulses, but there is some evidence for polarization of conduction.