Parental state and offspring recognition in the biparental cichlid fish Pelvicachromis pulcher

Alloparental care was investigated in the biparental West African cichlid, Pelvicachromis pulcher. Non-breeding adults typically consumed young conspecifics but this trait was inhibited in both sexes during reproductive attempts. Alien conspecific young were accepted into the brood if they were of a similar age/developmental stage to the parents' own young but not if they were much older or much younger. If not accepted they were consumed by whichever adult located them. Parents separated from their brood for up to 4 days accepted their young on reunion but separation for more than 4 days resulted in the young being consumed. This latter response occurred if chemical stimuli from the young were available during the separation but not if visual stimuli were available. In this latter case parental responsiveness was maintained. Both sexes of this externally fertilizing species appeared to have the same information about their young and showed the same changes in responsiveness and the same discriminatory abilities. (C) 1997 The Association for the Study of Animal Behaviour.

IMMUNOCYTOCHEMICAL DEMONSTRATION OF A SALMFAMIDE-LIKE NEUROPEPTIDE IN THE NERVOUS-SYSTEM OF ADULT AND LARVAL STAGES OF THE HUMAN BLOOD FLUKE, SCHISTOSOMA-MANSONI

The localization and distribution of SALMFamide immunoreactivity (IR), SI(GFNSALMFamide), in the nervous system of both the adult and larval stages of the trematode Schistosoma mansoni has been determined by an indirect immunofluorescent technique in conjunction with confocal scanning laser microscopy (CSLM). Immunostaining was widespread in the nervous system of adult male and female S. mansoni. In the central nervous system (CNS), IR was evident in nerve cells and fibres in the anterior ganglia, cerebral commissure and dorsal and ventral nerve cords. In the peripheral nervous system (PNS), IR was apparent in nerve plexuses associated with the subtegmental musculature, oral and ventral suckers, the lining of the gynaecophoric canal, and in fine nerve fibres innervating the dorsal tubercles of the male worm. In the reproductive system of male and female worms, S1-IR was only observed around the ootype/Mehlis' gland complex in the female. Immunostaining was also evident in the nervous system of both miracidium and cercarial larval stages. A post-embedding, IgG-conjugated colloidal gold immunostaining technique was employed to examine the subcellular distribution of SALMFamide-IR in the CNS of S. mansoni. Gold labelling of peptide was localized over dense-cored vesicles within nerve cell bodies and fibres constituting the neuropile of the anterior ganglia, cerebral commissure and nerve cords of the CNS. Antigen pre-absorption studies indicated that the results obtained do suggest S1-like immunostaining and not cross-reactivity with other peptides, in particular FMRFamide.

FASCIOLA-HEPATICA - TEGUMENTAL SURFACE CHANGES IN ADULT AND JUVENILE FLUKES FOLLOWING TREATMENT IN-VITRO WITH THE SULFOXIDE METABOLITE OF TRICLABENDAZOLE (FASINEX)

The effects of the novel benzimidazole, triclabendazole (Fasinex, Ciba-Geigy), in its active sulphoxide metabolite form (TCBZ-SX), on the tegumental surface of Fasciola hepatica has been examined in vitro. The tegument of adult flukes incubated in TCBZ-SX (50 mug/ml) appeared swollen and blebbed after only 6 h. In addition, progressive spine loss at the oral cone was evident following 12 h treatment. After 24 h, the tegumental syncytium and spines had completely sloughed away, leaving an exposed basal lamina and empty spine sockets. Juvenile flukes (3 weeks old) also demonstrated tegumental alterations after treatment with TCBZ-SX (20 mug/ml). The syncytium became extremely roughened and corrugated on both dorsal and ventral surfaces after only 3 h. Following 6- and 9-h incubations, there were many deep furrows, which were especially pronounced on the ventral surface, and by 18 h, the juvenile tegument was severely disrupted, especially on the ventral surface. In all cases, the effects were more marked than in the previous incubation periods. The results confirm the potent activity of triclabendazole against F. hepatica and suggest that the tegument of adult and juvenile flukes may be a target organ for this important fasciolicide.

FASCIOLA-HEPATICA - THE EFFECT OF THE MICROTUBULE INHIBITORS COLCHICINE AND TUBULOZOLE-C ON THE ULTRASTRUCTURE OF THE ADULT FLUKE

The effect of the microtubule inhibitors colchicine (1 x 10(-3) M) and tubulozole-C(1 x 10(-6) M) on the ultrastructure of adult Fasciola hepatica has been determined in vitro by transmission electron microscopy (TEM), using both intact flukes and tissue-slice material. With colchicine treatment, the apical membrane of the tegument became increasingly convoluted and blebbed, while accumulations of T1 secretory bodies occurred in the basal region of the syncytium, leading to progressively fewer secretory bodies in the syncytium. In the tegumental cells there were distinct accumulations of Tl secretory bodies around the Golgi complexes, which remained active for up to 12 h incubation. Tubulozole-treated flukes showed more severe effects, with initial accumulations of secretory bodies, both at the tegumental apex and base. This was followed in the later time-periods by the sloughing of the tegumental syncytium. In the underlying tegumental cells, the granular endoplasmic reticulum (GER) cisternae were swollen and disrupted, becoming concentrated around the nucleus. The Golgi complexes were dispersed to the periphery of the cells and gradually disappeared from the cytoplasm. After treatment with both drugs, the cell population in the vitelline follicles was altered, with an abnormally large proportion of stem cells and relatively few intermediate type 1 cells. The nurse cell cytoplasm became fragmented and was no longer in contact with the vitelline cells, while the shell globule clusters within the intermediate type 2 and mature cells were loosely packed. In the mature vitelline cells, 'yolk' globules and glycogen deposits became fewer than normal and lipid droplets were observed. The results are discussed in relation to the different modes of action of the two drugs and potential significance of this to anthelmintic (benzimidazole) therapy.

FASCIOLA-HEPATICA - THE EFFECT OF THE MICROFILAMENT INHIBITOR CYTOCHALASIN-B ON THE ULTRASTRUCTURE OF THE ADULT FLUKE

The effect of the microfilament inhibitor cytochalasin B (10 and 100-mu-g/ml) on the ultrastructure of adult Fasciola hepatica was determined in vitro by scanning and transmission electron microscopy (SEM, TEM) using both intact flukes and tissue-slice material. SEM revealed that initial swelling of the tegument led to surface blebbing and limited areas of sloughing after 24 h treatment at 100-mu-g/ml. In the tegumental syncytium, basal accumulations of secretory bodies (especially T2s) were evident in the earlier time periods but declined with longer incubations, until few secretory bodies remained in the syncytium overall. Blebbing of the apical plasma membrane and occasional areas of breakdown and sloughing of the tegument were observed over longer periods of treatment at 100-mu-g/ml. In the tegumental cell bodies, the Golgi complexes gradually decreased in size and activity, and few secretory bodies were produced. In the later time periods, the cells assumed abnormal shapes, the cytoplasm shrinking in towards the nucleus. In the vitelline follicles, a random dispersion of shell protein globules was evident within the intermediate-type cells, rather than their being organized into distinct shell globule clusters. Disruption of this process was more severe at the higher concentration of 100-mu-g/ml and again was more evident in tissue-slice material. In the latter, after prolonged (12 h) exposure to cytochalasin B, the intermediate and mature vitelline cells were filled with loosely packed and expanded shell globule clusters, containing few shell protein globules. The mature vitelline cells continued to lay down "yolk" globules and glycogen deposits. Disruption of the network of processes from the nurse cells was evident at the higher concentration of cytochalasin. Spaces began to appear between the vitelline cells and grew larger with progressively longer incubation periods, and the cells themselves assumed abnormal shapes. A number of binucleate stem cells were observed in tissue-slice material at the longest incubation period (12 h).

IMMUNOCYTOCHEMICAL AND RADIOIMMUNOMETRICAL DEMONSTRATION OF SEROTONIN-IMMUNOREACTIVITIES AND NEUROPEPTIDE-IMMUNOREACTIVITIES IN THE ADULT-RAT TAPEWORM, HYMENOLEPIS-DIMINUTA (CESTODA, CYCLOPHYLLIDEA)

Standard indirect immunocytochemical techniques have been interfaced with confocal scanning laser microscopy (for whole-mount preparations) and epifluorescence microscopy (for cryosections) to investigate the occurrence and distribution of serotoninergic and peptidergic nerve elements in adult H. diminuta. Serotonin (5-HT)-immunoreactivity (IR) was widespread throughout the worm, occurring in the paired cerebral ganglia, transverse commissure, the 10 longitudinal nerve cords and in a plethora of small nerve fibres of the peripheral nervous system. An abundance of serotoninergic nerve cell bodies was found in association with the lateral nerve cords. The genital atrium and accessory reproductive ducts were richly innervated with serotoninergic nerve fibres. Thirty-five antisera to 20 vertebrate regulatory peptides and 1 invertebrate peptide (FMRFamide) were used to screen the worm for neuropeptide IR. Immunostaining was obtained with antisera raised to pancreatic polypeptide (PP), peptide YY (PYY), neuropeptide Y (NPY), substance P (SP), peptide histidine isoleucine (PHI), xenopsin (XP) and FMRFamide. The most extensive pattern of IR occurred with antisera to PP and PYY, IR being evident in the cerebral ganglia, transverse commissure, longitudinal nerve cords and in small nerve fibres that ramified throughout the parenchyma. A series of bipolar nerve cell bodies between the median nerve cords displayed PP/PYY-IR. The distribution of FMRFamide-IR was reminiscent of the PP/PYY pattern but was less extensive. Comparison of the serotoninergic and peptidergic nervous systems has revealed general similarities and some distinct differences, especially with regard to the distribution of immunoreactive nerve cell bodies. Quantitative data are presented on the levels of PP-, SP-, PHI-, and gastrin-releasing peptide (GRP)-immunoreactivities demonstrable in acid-alcohol extracts of whole worms. The highest level of peptide IR determined was recorded for PP.

Survival of pathogenic bacteria during mesophilic anaerobic-digestion of animal waste

The survival of pathogenic bacteria was investigated during the operation of a full-scale anaerobic digester which was fed daily and operated at 28-degrees-C. The digester had a mean hydraulic retention time of 24 d. The viable numbers of Escherichia coli, Salmonella typhimurium, Yersinia enterocolitica, Listeria monocytogenes and Campylobacter jejuni were reduced during mesophilic anaerobic digestion. Escherichia coli had the smallest mean viable numbers at each stage of the digestion process. Its mean T90 value was 76-9 d. Yersinia enterocolitica was the least resistant to the anaerobic digester environment; its mean T90 value was 18.2 d. Campylobacter jejuni was the most resistant bacterium; its mean T90 value was 438.6 d. Regression analysis showed that there were no direct relationships between the slurry input and performance of the digester and the decline of pathogen numbers during the 140 d experimental period.

Comparative study of the spatial relationship between nicotinamide adenine dinucleotide phosphate-diaphorase activity, serotonin immunoreactivity, and GYIRFamide immunoreactivity and the musculature of the adult liver fluke, Fasciola hepatica (Digenea, Fasciolidae)

This is the first detailed description of the nitrergic nervous system in a fluke. In this study, the authors analysed the distribution of the nicotinamide adenine dinucleotide phosphate-diaphorase (NADPH-d) reactivity in neuronal and nonneuronal tissues of the adult fluke Fasciola hepatica and compared this with the distribution of the musculature using tetramethylrhodamine isothiocyanate-phalloidin. To assess the correlation between the number of muscle cells in different parts of the fluke and the NADPH-d-stained cells, the nuclei were stained with Hoechst 333 42, which is specific for chromatin. The spatial relation between the NADPH-d-positive nerves and the 5-hydroxytryptamine (serotonin; 5-HT)-immunoreactive (-IR) and GYIRFamide-IR nervous elements was also examined. The methods complement each other. NADPH-d-positive staining occurs in both in neuronal tissue and nonneuronal tissue. Large, NADPH-d-stained neurones were localised in the nervous system. The oral and ventral suckers are innervated with many large NADPH-d-stained neurones. Ln addition, the NADPH-d staining reaction follows closely the muscle fibres in both the suckers, in the body, and in the ducts of the reproductive organs. The presence of NADPH-d activity along muscle fibres in F. hepatica and in other flatworms supports a possible myoinhibitory role for nitric oxide. Neuronal nitric oxide synthase in flatworms may form a novel drug target, which would facilitate the development of a novel anthelminthic. (C) 2001 Wiley-Liss, Inc.