In the previous paper it was concluded that those aged [ges ] 15 years of both sexes could comprise the indicator group for rapid epidemiological assessment (REA) of onchocerciasis in the Amazonian focus. This paper explores relationships between community microfilarial (mf) prevalence, intensity, and nodule prevalence in 20 Yanomami communities, that would allow identification of REA methods in the region. The mean nodule ratio (prevalence of nodules/prevalence of mf) was 0·54 when onchocercomata in the indicator group were considered. The Spearman correlation coefficient between mf and nodule prevalence was 0·686 (P = 0·001). Palpation of nodules had 92% specificity and 32% sensitivity when compared to skin-snipping for the diagnosis of onchocerciasis. The predictive value positive increased from 75% to 81% when the indicator group was used. A microfilarial prevalence > 75% in this group would be indicative of hyperendemic status in the village, between 30 and 75% of mesoendemicity, and < 30% of hypoendemicity. For the assessment of infection intensity, biopsies may be taken from the iliac crest for all endemicity levels. Five of the hyperendemic villages surveyed in this work had a community microfilarial load (CMFL) greater than 10 mf/skin snip; the remaining 5 had a CMFL between 5 and 9. These levels of infection merit high priority ivermectin treatment. In Latin America, communities at both moderate and severe risk are included in mass chemotherapy programmes (i.e. when mf prevalence is over 20%). Roughly, a nodule prevalence in the indicator group > 10% would suggest a community mf prevalence > 20% with a sensitivity of 85% and a specificity of 71%. A multiple linear regression model of the arc-sine transformed mf prevalence in the village (all ages) on nodule prevalence in those aged [ges ] 15 years and altitude of the village explained 72% of the variance. The model combining nodule and altitudinal information had a sensitivity of 92% and a specificity of 71% in comparison to an estimated mf prevalence of 21% or more. It is suggested that the usefulness of the REA methods proposed be assessed in other areas of the Amazonian onchocerciasis focus.

The morphology of the post-hatch moulting and developmental sequence of an amphimictic population of Rotylenchulus reniformis has been examined in living and fixed material. Under suitable conditions of temperature and in a moist environment the 2nd-stage larva (L2) undergoes a series of 3 moults and developmental changes which result in the formation of males and immature females. After hatching and prior to the start of the first of these moults (the second moult), there is a pre-moult period, usually of 3–5 days duration. The morphology of the entire moulting and developmental sequences, from L2 to just after the final moult, has been followed in single living specimens of a parasitic nematode using differential interference contrast optics, and sections cut through different stages have been observed under the transmission electron microscope. These moulting sequences (at 24 °C) take place at similar times in developing males and females. The second moult takes place on the second day after commencement of moulting, the third on the third or fourth days and the fourth and final moult on the sixth or seventh days, followed by further development over several days to give rise to the adult male and the immature female. The ultrastructure associated with some of the more obvious of these developmental changes is described and includes the morphology of the head region and cuticles of L2, L4, adult males and immature females, the oesophageal glands of the immature female and the copulatory spicules and spermatozoa of the male.

The interesting Note by Dr Sant' Anna, which precedes this paper, leads me to record some observations of a somewhat similar nature to those he has reported. My information is derived from three letters received from Africa, and from Dr Hindle.

This bibliography—the first of the kind hitherto published—deals with zoological and medical publications relating to the two species of lice (Pediculits humanus Linn. 1758 and Phthirus pubis Leach, 1815) infesting man, the part they play in pathology, the prophylactic measures and means of destruction employed against lice, their anatomy and biology and other matters pertaining to them.

Our understanding of the neurobiology of the Platyhelminthes has come in large part from free-living turbellarians. In addition to providing considerable information about the capabilities of the rudimentary nervous system present in all members of the phylum, turbellarians have provided the most definitive information about the variety of ion channels present in the membranes of neurones and muscle cells, and about the physiology and pharmacology of those channels. Furthermore, preparations of single, viable muscle cells have provided some of the most conclusive evidence about the variety of transmitters present, and the types of response they evoke. Here, we review what is known about the physiology and pharmacology of the turbellarian neuromuscular system. Particular attention is given to the triclad flatworm Bdelloura Candida, the best studied species in this respect, but other species are included where relevant.

Growth of Trypanosoma cruzi as colonies on solid medium has not been widely used as an experimental procedure. We therefore sought to establish a reliable and routine plating method. The optimal results were achieved with a matrix of 0·65% low melting point agarose onto which epimasigotes from the mid-to-late logarithmic phase of growth were spread. Colonies could be isolated after incubation for 21 days in a humidified 5% CO2 environment at 28°C. Plating efficiencies in the range of 40% were obtained by this method and clones could be recovered into liquid medium or onto blood-agar slopes with a high success rate. The procedure has also been adapted for the isolation of genetically transformed clones after electroporation of epimastigotes with either plasmid or cosmid vectors. This was best achieved by inclusion of the electroporated cell inoculum in a 0·6% agarose overlay containing G418 as the selective drug, on top of a 0·8% agar base. Transformation efficiencies were as high as 10−5 cells per μg of DNA. A reliable plating method for T. cruzi will have many applications and is a significant step towards the use of ‘shotgun transformation’ to generate libraries of T. cruzi recombinants.

The abomasum of living sheep infected with 50000 larvae of Haemonchus contortus was examined before and during infection, by means of fibre optic endoscopy. Biopsy specimens were removed from the abomasum of the living sheep and were examined by means of scanning electron microscopy. Changes were noted in the surface structure of the abomasum 2 days after infection and larvae were seen on, and burrowing into, the mucosa. These changes became more pronounced as the infection proceeded, especially after day 10 of the infection when the adult worms had appeared. The behaviour of adult nematodes was observed within the abomasum and on occasions they were seen to move from surrounding areas into an area of haemorrhage, caused by removal of a biopsy specimen. Individual nematodes were removed from the abomasum by means of the biopsy forceps. Fibre optic endoscopy was shown to be a useful tool in the study of parasitic nematodes in vivo.

FhCaBP2 is a Fasciola hepatica protein which belongs to a family of helminth calcium-binding proteins which combine an N-terminal domain containing two EF-hand motifs and a C-terminal dynein light chain-like (DLC-like) domain. Its predicted structure showed two globular domains joined by a flexible linker. Recombinant FhCaBP2 interacted reversibly with calcium and manganese ions, but not with magnesium, barium, strontium, copper (II), colbalt (II), iron (II), nickel, lead or potassium ions. Cadmium (II) ions appeared to bind non-site-specifically and destabilize the protein. Interaction with either calcium or magnesium ions results in a conformational change in which the protein's surface becomes more hydrophobic. The EF-hand domain alone was able to interact with calcium and manganese ions; the DLC-like domain was not. Alteration of a residue (Asp-58 to Ala) in the second EF-hand motif in this domain abolished ion-binding activity. This suggests that the second EF-hand is the one responsible for ion-binding. FhCaBP2 homodimerizes and the extent of dimerization was not affected by calcium ions or by the aspartate to alanine substitution in the second EF-hand. The isolated EF-hand and DLC-like domains are both capable of homodimerization. FhCaBP2 interacted with the calmodulin antagonists trifluoperazine, chlorpromazine, thiamylal and W7. Interestingly, while chlorpromazine and thiamylal interacted with the EF-hand domain (as expected), trifluoperazine and W7 bound to the DLC-like domain. Overall, FhCaBP2 has distinct biochemical properties compared with other members of this protein family from Fasciola hepatica, a fact which supports the hypothesis that these proteins have different physiological roles.

1. The term Schizogregarinae Léger (1900) is the name now given to a sub-order of the Gregarinida, the remaining members of which are known as the Eugregarinae. The Schizogregarines were formerly known as Amoebosporidia Aimé Schneider (1884), a name given in misapprehension of the character of the cytoplasmic processes, fixative in function, present in the genus Ophryocystis. Two species of Ophryocystis (0. buetschlii and 0. francisci) were the only members of this sub-order known before 1900.

2. At present the sub-order Schizogregarinae contains five families: Ophryocystidae, Schizocystidae, Selenidiidae, Merogregarinidae, and Aggregatidae.

3. All these organisms show well-marked schizogonic stages in their life-history, and—with the possible exception of the Aggregatidae—follow after the Eugregarinae in their methods of sporogony.

4. In Ophryocystis and Schizocystis the schizogony is extracellular, that is, these forms are ectoschizous. The life-cycle of the former is shown in Fig. 1. In these parasites the number of the nuclei in the schizont increases simultaneously with its volume.

5. In Selenidium and Merogregarina the schizogony is intracellular, in other words these forms are endoschizous. The life-cycle of the former is illustrated in Fig. 3. In these forms the schizont is uninucleate during its growth, only becoming multinucleate at the end of the growing period.

6. Ophryocystis forms only one sporocyst, a fact which has been emphasised by Léger and Duboscq (1908), by the placing of the Ophryocystidae in a special section, the Monospora. However, this apparent peculiarity is easily explained by a process of reduction and degeneration having taken place, affecting with one exception all the gametes formed from each gametocyte. There is good morphological evidence in support of this explanation (see p. 382, and Fig. 4, D).

7. Figures of the interesting form Schizocystis gregarinoides (Léger, 1900) are not yet published, but a paper dealing with this organism is promised by Prof. Léger at an early date.

8. Aggregata differs from other Schizogregarines in that its schizogony takes place in one host (crab), while its sporogony occurs in another (Cephalopod mollusc). In this respect Aggregata resembles the Haemosporidia. The schizogonic phases in Crabs were formerly regarded as belonging to a gymnosporous Gregarine, Aggregata Frenzel, while the sporogonic phases were considered to belong to a Coccidian, Eucoccidium (Benedenia) in cuttlefishes and Octopus. Regarding this, Léger and Duboscq (1908, p. 102) write “…Aggregata, avec un changement de cycle coïncidant avec un changement d'hôte, c'est à dire qui soient à la fois digénétiques et hétéroïques.”

9. It is evident therefore that the Aggregatidae stand apart. On this account, I suggest a division of the Schizogregarinae into two sections, termed respectively, the Homoïica (to include the first four families discussed in this paper, wherein schizogony and sporogony take place in the same host) and the Heteroïca (for the Aggregatidae).

10. Among the Homoïca we have extracellular schizogony (ectoschizous forms) in the Ophryocystidae and Schizocystidae, and intracellular schizogony (endoschizous forms) in the Selenidiidae and Merogregarinidae. This difference is not merely superficial, it requires to be emphasised, and for this reason I would divide the section Homoïca into two sub-sections, termed respectively Ectoschiza and Endoschiza.

11. The classification of the Schizogregarines, which I would propose, is as follows:

12. Much further research is needed on the life-cycles of the Endoschiza, especially among the Selenidiidae, which occur so frequently in the Annelida. Sporogonic stages are at present unknown in Eleutheroschizon and Siedleckia.

13. In connection with the Aggregatidae, and to a less extent with the Selenidiidae, stress is laid upon the necessity of carefully distinguishing between “coelomic” and “gut” parasites. (See pp. 397 and 387.)

14. The Schizogregarinae form a most interesting link between the Eugregarinae and the Coccidiidea.

The microsporidian Encephalitozoon cuniculi is an obligate intracellular parasite that develops asynchronously inside parasitophorous vacuoles. Spore differentiation involves the construction of a cell wall commonly divided into an outer layer (exospore) and a thicker, chitin-rich inner layer (endospore). The developmental patterns of protein deposition and mRNA expression for 2 different spore wall proteins were studied using immunocytochemical and in situ hybridization procedures with ultrathin frozen sections. The onset of deposition of an exospore-destined protein (SWP1) correlated with the formation of lamellar protuberances during meront-to-sporont conversion. No evidence for a release of SWP1 towards the parasitophorous vacuole lumen was obtained. An endospore-destined protein (EnP1) was detected early on the plasma membrane of meronts prior to extensive accumulation within the chitin-rich layer of sporoblasts. swp1 mRNA was preferentially synthesized in early sporogony while enp1 mRNA was transcribed during merogony and a large part of sporogony. The level of both mRNAs was reduced in mature spores. Considering the availability of the E. cuniculi genome sequence, the application of nucleic and/or protein probes to cryosections should facilitate the screening of various genes for stage-specific expression during microsporidian development.

Acephalic cysticercus (Ac), a rarely developed multilobulated and nonencysted form of larval Taenia, causes hydrocephalus or adhesive arachnoiditis in the ventricles and subarachnoidal space that often lead to fatal outcome in affected patients. Ac has been proposed to originate from T. solium on the basis of morphological features, while no molecular data supporting the presumption have been available. In the present study, we investigated the immunological properties as well as molecular characteristics of Ac that was obtained surgically from 6 patients. Immunoblotting of the cyst fluid from Ac samples demonstrated the constitutive expression of a T. solium metacestode (TsM) 10 kDa protein. Specific antibodies against the truncated 10 kDa protein, which appears to be species specific for TsM cysticercosis, were detected in both serum and cerebrospinal fluid samples of Ac patients. Nucleotide sequences of mitochondrial cytochrome c oxidase subunit I (COI) and NADH dehydrogenase subunit 1 (ND1) genes of Ac were almost identical to those of T. solium but differed substantially from those of the other Taenia species. In phylogenetic analysis, Ac clustered with T. solium in a well-supported clade. Our results strongly suggest that Ac may have originated from T. solium.

A total of 28 components were detected in the free amino acid (FAA) pool of hydatid fluid from primary and secondary equine cysts, secondary ovine cysts and host plasma. Examination of data from equine cysts revealed that the majority of FAAs were present in significantly greater concentrations in secondary cysts, glycine being over 30 times more concentrated. Values for total carbohydrates and glucose did not, however, differ significantly and total protein content was greater in primary cysts. Comparison of the (FAA) pool of secondary equine and ovine cysts revealed strain variation. It was demonstrated that most FAAs were more concentrated in hydatid fluid than in the corresponding host plasma, many concentration ratios exceeding 10. The possible contribution that mediated amino acid transport across the cyst wall and parasite amino acid metabolism makes to the composition of the FAA pool was discussed. No significant plasma aminoacidaemia was associated with infection.

Previous work has shown that the surface of infective larvae of parasitic nematodes will not bind the fluorescent lipid analogue 5-N(octadecanoyl)aminofluorescein (AF18) until after exposure of the parasite to mammalian tissue-culture conditions. In this study, culture media which are permissive or non-permissive for the acquisition of lipophilicity for AF18 were altered in order to examine possible stimuli involved. This showed that external alkaline pH and high sodium ion concentration were highly stimulatory. The internal signalling pathways which may be involved in the surface alteration were then examined using agents which are known to affect intracellular signalling in mammalian cells. The results indicated that elevation of cGMP levels was stimulatory whereas inhibition of a putative Na+/H+ antiporter or calcium mobilization was inhibitory, and it is argued that high intracellular levels of cAMP may be inhibitory. Whilst the precise effects of the agents used on nematode cells remain to be established, these results provide a framework for the examination of the processes involved in the modification of the nematode surface which takes place immediately after the infection event.

The series of egg envelopes around the acanthor in the mature egg of the palaeacanthocephalan, Polymorphus minutus, has been investigated with light and electron microscopy. An attempt has been made to characterize the macromolecules from which the envelopes are constructed using histochemistry, an analysis of stabilizing bond types and comparisons of envelope fine structure with that of other structural protein systems.The outermost envelope (I) of the mature Polymorphus egg is a fertilization membrane. It is produced by membrane-bounded inclusions in the cytoplasm of the unfertilized oocyte which discharge their contents into the extracellular space around the egg. Envelope II is composed largely of keratin, and its subzones consist of different packing arrangements of 8 nm diameter electron-lucid filaments in an electron-dense matrix. The electron microscopical appearances which this filamentous keratin can present due to section orientation artefacts are discussed. The innermost envelope (III) is 20–30 nm thick and highly corrugated. The spaces between envelopes I and II and between envelope III and the inner surface of envelope II contain material with a granular or fibrous ultrastructure. These spaces show histochemical evidence for the presence of acid mucopolysaccharides.

Many liver-invasive parasites cause extensive liver damage which may result in an impaired ability to catabolize endotoxin. The influence of endogenous endotoxin on the progress of liver-invasive parasitic diseases has been investigated in murine Mesocestoides corti infection. Invasion of liver tissue by tetrathyridia resulted in extensive parenchymal destruction with fibrosis. In association with this, undetoxified endotoxin, in potentially biologically active concentration, was found on peritoneal macrophages, 5 months post-M. corti infection. Host susceptibility was influenced by the Lps gene for responsiveness to lipopolysaccharide (LPS). The parasite burden of LPS-responsive (C3H/HeN) mice was significantly increased in the livers of these mice when compared to LPS-resistant (C3H/HeJ) mice. LPS reduced the ability of normal peritoneal macrophages to kill tetrathyridia, when co-cultured in vitro. LPS also abrogated the ability of recombinant interferon-γ (r.IFN-γ) to enhance macrophage larvicidal activity. These in vitro findings were confirmed in vivo. Daily intraperitoneal administration of LPS, at low concentration, caused a 4-fold increase in parasite burden in the liver, while r.IFN-γ at optimal concentration reduced parasite burden by 57%. Post-infection macrophages have previously been shown to be refractory to cytokine-activation for larval killing. In this report, we conclude that (1) this refractoriness may be due to the presence of undetoxified endotoxin on post-infection macrophages and (2) endotoxin may reduce host resistance by abrogating effector macrophage response to IFN-γ.

Reversal of chloroquine (CQ) resistance by verapamil, a Ca2+ antagonist, has been shown in CQ-resistant human and rodent malaria parasites. Here, we report ultrastructural changes associated with this phenomenon in CQ-resistant Plasmodium chabaudi (AS strain) after infected mice were administered CQ and verapamil. At parasitaemias of 5–7%, CQ at 6 mg/kg caused little morphological effect on CQ-resistant parasites. In contrast, co-administration of CQ and verapamil at 50 mg/kg induced swelling of food vacuoles with clumped pigment at 2.5 h. Morphological changes other than food vacuole enlargement occurred at 21 and 45 h: disappearance of endoplasmic reticulum, formation of myelin structures, focal cytoplasmic vacuolization and coarse clumping of electron-dense material in nuclei. These structural changes appeared to be very similar to those observed in CQ-sensitive P. chabaudi in mice injected with CQ alone or CQ plus verapamil. On the other hand, verapamil at 50 mg/kg alone did not induce any effect on both CQ-sensitive and CQ-resistant P. chabaudi. These results suggest that swelling of the food vacuoles is an initial event associated with reversal of CQ-resistance by verapamil.

A significant polyclonal activation of B lymphocytes was observed during experimental infection of C57BL/10J mice with Schistosoma mansoni. The isotypic pattern of this expansion, assessed by the Protein-A plaque-forming cell method, was compared with and found to differ from those occurring after infection by Trypanosoma cruzi or injection of bacterial LPS. In the infection of S. mansoni an early expansion of most immunoglobulin isotypes occurs together with a late, sustained expansion of IgG1-secreting cells. High levels of polyclonal B cell activation were observed after adoptive transfer of spleen cells from infected mice to isogenic recipients pre-treated with hydroxyurea.

Codon usage of Crytosporidium parvum was compared with those of other Eimeriorina Toxoplasma gondii and Eimeria tenella and revealed a biased use of synonymous codons with a preference for NNU (40·0%) and NNA (33·4%). There was no close resemblance of the codon usage of C. parvum to T. gondii (correlation coefficient, r = 0·14) or E. tenella (r = 0·14) but it was similar to Entamoeba histolytica (r = 0·75) and Plasmodium falciparum (r = 0·5). Analysis of the codon usage in homologous gene sequences (actin, β-tubulin) also failed to reveal a close relationship between C. parvum and T. gondii or E. tenella. The low usage codons in C. parvum were most frequently used codons in T. gondii and E. tenella. These observations are consistent with 18S rRNA sequence analysis which shows no close relationship of Cryptosporidium with other Eimeriorina (Sarcocystis, Toxoplasma and Eimeria) and questions the validity of the current classification of C. parvum.

The hatching organ of Lipeurus columbae Linn. is described. It is very similar to that already described in sucking lice, and unlike that of any other group of insects. This resemblance is further evidence for the affinity of the Mallophaga and Siphunculata.

A description is given of the life history and pathogenicity of a species of Eimeria, derived from a single oocyst infection in a coccidia-free bird, and subsequently identified as E. meleagridis.

There are two asexual generations followed by the sexual generation. The first schizogony cycle occurs in the small intestine and the second and sexual generations are restricted to the caeca.

The organism is not pathogenic but the sexual stages produce marked caseous lesions in the caeca. It is immunologically distinct from a pure strain of E. adenoeides and cannot be transmitted to chickens.