Publisher Summary Chemo-, thermo-, and hygroreceptors are primary sensory cells, which bear one or several modified cilia whose membranes are specialized to take part in the transduction process. These membranes must be exposed to the stimuli but also be protected against desiccation, mechanical, and other injury. Hence, a variety of structural specializations is found that serves these functions. In insects, chemo-, thermo-, and hygroreceptive sensory cells are located exclusively in sensilla, which are organelles built up by a definite number of characteristic cells. During development, the innermost enveloping cell produces a cuticular sheath (scolopale), which envelops the dendritic processes for varying distances along their course through the receptor lymph cavity. This sheath producing cell is surrounded by the trichogen cell, which is itself surrounded by the trichogen cell. These cells build up shaft and socket, respectively, of the cuticular outer structures during development. The perikarya of all cells of the sensillum are enclosed within the epidermis.

Ultrastructure of invertebrate chemo-, thermo-, and hygroreceptors and its functional significance.

Granulomas are organized aggregates of macrophages, often with characteristic morphological changes, and other immune cells These evolutionarily ancient structures form in response to persistent particulate stimuli-infectious or noninfectious-that individual macrophages cannot eradicate Granulomas evolved as protective responses to destroy or sequester particles but are frequently pathological in the context of foreign bodies, infections, and inflammatory diseases We summarize recent findings that suggest that the granulomatous response unfolds in a stepwise program characterized by a series of macrophage activations and transformations that in turn recruit additional cells and produce structural changes We explore why different granulomas vary and the reasons that granulomas are protective and pathogenic Understanding the mechanisms and role of granuloma formation may uncover new therapies for the multitude of granulomatous diseases that constitute serious medical problems while enhancing the protective function of granulomas in infections

The Formation and Function of Granulomas.

In studies designed to elucidate the mechanisms that may effect or influence the susceptibility and/or nonsusceptibility of mollusks to invading microorganisms, it is essential that consideration be given toward understanding how the internal defense mechanisms of mollusks function when confronted with nonself substances. It should be apparent that such information is essential if rational approaches are to be taken for the development of potential biologic control agents for undesirable mollusks or, on the other hand, if we are to understand and possibly prevent the microbial diseases that weaken or destroy economically important mollusks. Since Tripp 2-.L demonstrated that microorganisms experimentally introduced into a pelecypod, the American oyster, Crassostrea virginica, and a pulmonate gastropod, Biomphularia glabrata, are phagocytized and degraded intracellularly, it has become evident that phagocytosis by molluscan hemolymph cells is the major line of defense in at least these species of mollusks against invading microorganisms. This information is extremely important for a third practical reason. Specifically, only when we have elucidated the biochemical bases for the recognition of self and nonself, the phagocytosis of nonself materials, and the intracellular degradation of foreign substances can we logically and rationally develop and design methods to depurate edible mollusks that have become biologically polluted. In other words, only when all of the parameters that influence or affect the phagocytosis and degradation of microorganisms that enter mollusks are understood can we devise methods to enhance this innate self-cleansing process and bring about the total and efficient clearing of real and potential pathogens from oysters, clams, and other edible shellfish. When one considers the various types of reactions of hosts exposed to microorganisms, the development of antibody-mediated immunity naturally comes to mind. This, however, does not appear to be the case in mollusks. The evidence for this is as follows. Cheng,5 as a result of a quantitative electrophoretic analysis of the serum of the pulmonate Helisomu duryi normale, has concluded that hyperglobulinemia does not occur when this snail is challenged with different species of bacteria and has also concluded that immunoglobulins are not synthesized in mollusks. It is noted, however, that Feng and Canzonier fi and Gress and Cheng have demonstrated that in C. virginica parasitized by larvae of the trematode Bu-

Functional morphology and biochemistry of molluscan phagocytes

The transmission success of free-living larval stages of endohelminths is generally modulated by a variety of abiotic and biotic environmental factors. Whereas the role of abiotic factors (including anthropogenic pollutants) has been in focus in numerous studies and summarized in reviews, the role of biotic factors has received much less attention. Here, we review the existing body of literature from the fields of parasitology and ecology and recognize 6 different types of biotic factors with the potential to alter larval transmission processes. We found that experimental studies generally indicate strong effects of biotic factors, and the latter emerge as potentially important, underestimated determinants in the transmission ecology of free-living endohelminth stages. This implies that biodiversity, in general, should have significant effects on parasite transmission and population dynamics. These effects are likely to interact with natural abiotic factors and anthropogenic pollutants. Investigating the interplay of abiotic and biotic factors will not only be crucial for a thorough understanding of parasite transmission processes, but will also be a prerequisite to anticipate the effects of climate and other global changes on helminth parasites and their host communities.

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The role of biotic factors in the transmission of free-living endohelminth stages.

Publisher Summary The study of intramolluscan single and double trematode infections has proved to be a useful approach in better understanding host–parasite relationship Modification and disruption of snail infection with trematodes is reviewed and the parameters and characteristics of various infection combinations are described Biologically, however, the inter-trematode reactions and the responses of the snail host offer remarkable opportunities to examine and isolate various aspects of this host–parasite relationship The single species infection patterns differ widely between the pure sporocyst type, exemplified by S mansoni, and the redial type, typified by P segregatum Characteristics of each are reviewed Their interaction is then reviewed, based on original data and comparison with other studies Redial predation (“direct antagonism”) is discussed in terms of relative activity, efficiency, and predatory response to triggering stimuli in the snail Possible mechanisms of indirect antagonism are reviewed, with a fuller consideration of snail immunity as induced by these infections and measured experimentally The parameters or characteristics of single-species infection are described and related to one another in a preliminary fashion as an “adaptation index” (AI), which includes such parameters as infection rate, period for 50% snail death, and period to cercarial shed The trematode method is locally applicable, using indigenous strains supplied and sustained at a high level by continual reseeding with eggs of the controlling trematode (or processed faeces from the appropriate final hosts) Effective biocontrol as a self-sustaining life-cycle appears highly improbable, as is the expectation that this method can be adopted on a wide scale with the same controlling agent

Intramolluscan inter-trematode antagonism: a review of factors influencing the host-parasite system and its possible role in biological control.

Experiments demonstrated that infestation with the oligochaete worm, Chaetogaster limnaei, afforded a degree of protection to the snail Australorbis glabratus when the snail was exposed to Schistosoma mansoni miracidia. To a lesser extent, the oligochaetes protected snails exposed to echinostome cercariae. The presence of C. limnaei within the kidney of the snail Physa heterostropha is reported. An intimate association between the oligochaete worm Chaetogaster limnaei von Baer and various species of aquatic snails has long been recognized (von Baer, 1827), yet the nature of this relationship has not been adequately defined. Early naturalists considered C. limnaei to be a true parasite which was thought to feed upon the "slime" produced by the host. However, Wagin (1931) and subsequent investigators demonstrated that the oligochaete fed principally on microorganisms, and thus the worm has been considered a commensal. Wagin also observed that the oligochaete ingested cercariae and suggested that C. limnaei might be of value in controlling trematode transmission. While Krasnodebski (1936) confirmed Wagin's field observations and experimentally demonstrated that the oligochaete would ingest various types of cercariae, he did not believe that Chaetogaster could play a significant role in the control of trematodes. Backlund (1949) observed that cercariae of Fasciola hepatica were ingested by Chaetogaster, and Ruiz (1951) later reported a similar observation with respect to Schistosoma mansoni. Coelho (1957) noted that Chaetogaster frequently ingested S. mansoni miracidia, but commented that the worms did not protect snails from infection. HowReceived for publication 3 March 1964. * These studies were supported in part by Grants AI-00513 and 5 TI AI 46 from the National Institute of Allergy and Infectious Diseases, National Institutes of Health, Public Health Service, Bethesda, Maryland. 441 ever, Khalil (1961) presented circumstantial evidence suggesting that snails infested with C. limnaei were refractory to infection with Fasciola hepatica under laboratory conditions. The present study was initiated to determine quantitatively whether Australorbis glabratus infested with C. limnaei were protected against invasion by miracidia of S. mansoni and by cercariae of an echinostome. MATERIALS AND METHODS Specimens of C. limnaei were obtained from Physa heterostropha collected in Boston. The oligochaetes were freed by immersing the snails in 1% Urethan (ethyl carbamate) for 3 to 5 min. Anesthetized worms were collected by pipette, allowed to recover in charcoal-filtered tap water, and used immediately or colonized for later use (Brandwein, 1937). A Puerto Rican strain (PR-1) of Australorbis glabratus was used in all experiments. This snail strain has been maintained in our laboratory for several years and has been free from infestation with C. limnaei. Miracidia were hatched from eggs obtained from the livers of mice infected with a Puerto Rican strain of S. mansoni and were used within 30 min of emergence (Michelson, 1964). Cercariae of an unidentified echinostome were obtained from Physa heterostropha collected from local ponds. Five C. limnaei were added to individually isolated A. glabratus in 5-ml beakers containing 3 ml of charcoal-filtered tap water. The snail was confined with the oligochaetes for 1 hr or until all worms became attached. At this time and depending upon the experiment, one to five miracidia or ten cercariae were added to each beaker. After an exposure of approximately 16 hr, the snails from any one experiment were maintained together in a battery jar containing 2.5 liters of water. The battery jars were continually aerated, maintained at 25 ?+ 1 C, and the snails fed Romaine lettuce. ConThis content downloaded from 157.55.39.157 on Tue, 17 May 2016 04:47:34 UTC All use subject to http://about.jstor.org/terms 442 THE JOURNAL OF PARASITOLOGY, VOL. 50, NO. 3, JUNE 1964 TABLE I. Influence of Chaetogaster limnaei infestation on the rate of infection of snails exposed to S. mansoni miracidia and to echinostome cercariae. Snails with C. limnaeil Control snails Group no. No. positive/ No. positive/ no. exposed no. exposed 5 miracidia of S. mansoni per snail 12 2/7 4/8 22 7/18 14/20 32 1/5 5/6 4 5/10 8/10 5 2/10 10/10 Total 17/50 (34%) 41/54 (76%) 1 miracidium of S. mansoni per snail 6 1/10 5/10 7 2/15 7/15 Total 3/25 (12%) 12/25 (48%) 10 cercariae of echinostome sp. per snail 8 7/10 10/10 9 7/10 10/10 Total 14/20 (70%) 20/20 (100%) 1 All snails infested with 5 worms each. 2 Test groups 1, 2, and 3 had one, two, and one snails die before they could be examined for the presence of infection; these have not been included with the data. trol snails, free of C. limnaei, were exposed to miracidia or cercariae in the same manner. Two weeks postexposure, control and Chaetogaster-infested snails were examined for the presence of S. mansoni sporocysts by the technique of Chernin and Dunavan (1962). Snails exposed to echinostome cercariae were examined 7 days postexposure for the presence of metacercariae. For this purpose the snail was removed from its shell, the mantle cut along its left margin and reflected to the right, and the pallial cavity examined under a 13 X dissecting microscope. Experimental results In five experiments a total of 54 snails, each infested with five Chaetogaster limnaei, were exposed individually to five S. mansoni miracidia and 17 (34%) of 50 snails which survived were found infected (Table I). By comparison, in the control groups of snails 41 of 54 snails (76%) were infected. Thus, a mean reduction in the rate of infection of 42% was associated with the presence of C. limnaei. In two additional experiments (Table I), snails harboring five C. limnaei were exposed individually to a single miracidium of S. mansoni. Of a total of 25 snails so exposed, three (12%) became infected. An infection rate of 48% (12/25 snails) occurred, however, in the two groups of control snails. A mean reduction of 36% in the infection rate was again observed to be associated with the presence of C. limnaei. Snails were exposed to echinostome cercariae in two experiments (Table I), and all the control snails were later found infected. However, only 70% (14/20) of the snails infested with C. limnaei had metacercarial cysts. In the first experiment 38 and 58 metacercarial cysts were recovered from the Chaetogaster-infested and control snails respectively, and in the second experiment 22 and 69 cysts respectively. Infected snails were found to have from three to eight cysts each. Miscellaneous observations The intensity of Chaetogaster infestation in snails under field conditions would appear to vary considerably. Backlund (1949) observed that no more than ten Chaetogaster were found on specimens of Lymnaea stagnalis and L. ovata; however, Krasnodebski (1936) noted as many as 300 worms on a single specimen of L. stagnalis, up to 60 worms on Physa fontinalis, and a mean of 1.3 worms on specimens of Ancylus lacustris. In our own experience, as many as 25 C. limnaei have been recovered from a single Physa heterostropha. In one collection of 105 P. heterostropha there was a mean of nine worms per snail. The heaviest infestations were found on older and larger snails, particularly those which had overwintered. Quantitative data concerning natural infestations of Chaetogaster on the snail Auistralorbis glabratus are not available, although infestations of field populations have been reported (Coelho, 1957). Natural infestations of South African populations of Bulinus tropicus, Bulinus africanus, and Biomphalaria pfeifferi have also been noted by Bayer (1955). Likewise, laboratory colonies of A. glabratus, as well as Bulinus truncatus and B. tropicus, have been observed to be infested with Chaetogaster (Khalil, 1961). Although it has generally been thought that C. limnaei acts as a commensal of snails, feeding principally on aquatic microorganisms, there is some evidence to suggest that this oligochaete may also be a parasite. The observation of Lankester (1870) of C. limnaei in the kidney of a single L. stagnalis has been overlooked. Our examination of both field and laboratory populations of P. heterostropha This content downloaded from 157.55.39.157 on Tue, 17 May 2016 04:47:34 UTC All use subject to http://about.jstor.org/terms MICHELSON-PROTECTIVE ACTION OF CHAETOGASTER ON SNAILS EXPOSED TO SCHISTOSOMA MANSONI TABLE II. Distribution of Chaetogaster limnaei in a collection of Physa heterostropha. Number C. limnaei present Snail no.1 On body surface and Inside in mantle cavity kidney 1 1 0 2 0 7 3 8 4 0 5 5 2 1 6 6 5 7 1 3 8 0 7 9 5 10 1 1 2 5 12 0 2 3 3 1 14 1 5 5 5 18 1 Five snails in this collection had no Chaetogaster and are not included in the table. revealed that under some circumstances large numbers of C. limnaei may be found within the kidney of the snail. In one group of P. heterostropha, examined 3 days after collection, 15 of 20 snails were infested. Fourteen of the 15 infested snails harbored 1 to 18 worms within the kidney; whereas only nine of these 15 snails had external infestations (Table II). Of further interest is the fact that snails could be free of external infestation while harboring numerous worms in their kidney. Histologic studies demonstrated that most of the oligochaetes were in the anterior portion of the kidney, but any part of the organ might harbor the worms (Fig. 1). No tissue reactions were observed in sections of snails so infected, although some oligochaetes appeared attached to the renal epithelium and renal concretions were observed in the intestinal tract of a few worms. Oligochaetes were not found in the kidney of experimentally infested A. glabratus.

The Protective Action of Chaetogaster limnaei on Snails exposed to Schistosoma mansoni.

Summary
A pathogenic acid-fast bacillus was isolated from the fresh-water snail Helisoma anceps and has been successfully maintained in pure culture. Techniques for the isolation and maintenance of this organism, as well as observations on its pathogenicity, are described. The results of experimental transmission of the organism to 13 strains of fresh-water snails, representing 6 species in 4 genera, are also presented.

An acid-fast pathogen of fresh-water snails.

Chemoreception in the snail Australorbis glabratus.

Summary
The growth of an Australorbis glabratus population was effectively controlled by the leech, Helobdella fusca, under laboratory conditions. Small snails were found to be exceptionally vulnerable although snails of all sizes were killed by this leech. Furthermore, while the mature leeches were primarily responsible for killing the snails, it was found that young leeches may also play a contributory role. The manner in which the leech kills its prey is described and the possible practical aspects of the study are discussed.

Studies on the biological control of schistosome-bearing snails. II. The control of Australorbis glabratus populations by the leech, Helobdella fusca, under laboratory conditions.

This study was initiated to determine the immunologic specificity of snail hemolymph antigens and to evaluate their serotaxonomic usefulness. Hemolymph antigens were obtained from snails of the families Planorbidae and Lymnaeidae, representing 9 genera, 17 species, and 26 strains. The interfacial ring test (IRT), and a gel-diffusion system, were employed to compare the precipitating activity of the various antigens in the presence of specific snail antisera. Antigenic differences between genera were demonstrable in the IRT. Generic specific precipitin reactions were obtained following absorption, from the antisera, of familial cross-reacting antibodies. Congeneric species or strains of a species could not be differentiated by the IRT. Gel-diffusion studies confirmed the results of the IRT with respect to generic differences; in addition, the gel-diffusion technique proved sensitive enough to demonstrate, with some antisera, antigenic differences between congeneric species. However, strains of a species were not differentiated. The results of this study indicate that, in spite of certain limitations, serologic techniques may serve as valuable adjuncts to the methods now employed by the molluscan taxonomist. For example, present serologic evidence suggests that species assigned to the nominal genera Australorbis, Tropicorbis, and Biomphalaria are congeneric-a concept recently set forth by several malacologists on the basis of anatomical studies. Furthermore, serologic evidence indicates that snails now assigned to the genus Lymnaea may, in fact, belong to more than one genus. Requisite to control and to all biological studies on the snail hosts of schistosomes is a foundation of sound taxonomy. The present taxonomic status of many of the snails of medical importance has not changed appreciably since 1954 when it was noted that "Confusion was found to exist not only in the nomenclature of the species, but even in the delimitation of the genera and the family to which the snails belong" (Anonymous, 1954). Malacologists first used shell characters to establish species of snails. They failed, however, to appreciate individual variation or the plasticity of these animals in response to environmental challenges, and consequently the literature became flooded with a mass of specific names. For example, although 200 or more specific names have been ascribed to the South American Planorbidae (Harry, 1962), it is doubtful whether more than 25 valid species exist, of which a half dozen or so may serve as snail-hosts for Schistosoma mansoni. A similar profusion of synonyms exists for the African Planorbidae. Received for publication 18 September 1965. * These studies were supported, in part, by Grants AI-00513 and 5TI A-I 46 from the NIAID, NIH, U. S. Public Health Service, Bethesda, Maryland. t Portions of this paper were presented before the 2nd European Malacological Congress, Copenhagen, Denmark, on 11 August 1965. Recent applications of morphologic criteria, principally those associated with snail genitalia, have improved immeasurably our taxonomic concepts; however, even these criteria have failed to establish universally recognized genera and species. A system of classification can be no better than the criteria upon which it is based. Having reached an impasse with available criteria, the need for newer approaches for differentiating snail taxa is apparent. Some progress has been made in this direction by the application to molluscan taxonomy of chromatography (Kirk et al., 1954; Wright, 1959; Wright and Claugher, 1959), electrophoresis (Targett, 1962, 1963; Wright and Ross, 1963), and by the enumeration of snail karyotypes (Burch, 1960, 1961, 1964). Taxonomic problems associated with various plant and animal groups have been resolved, with considerable success, by means of serologic and immunologic techniques (Leone, 1964). However, except for the early studies of Erhardt (1931) and Wilhelmi (1944), who attempted to elucidate the phylogenetic relationships between Mollusca and other phyla by immunologic methods, and the immunoelectrophoretic study of Tran van Ky et al. (1962), in which the total proteins of four snail species were compared, malacologists have rarely employed these techniques. In the present study an attempt has been made to determine the serologic specificity of

Edward H. Michelson

Papers

The Protective Action of Chaetogaster limnaei on Snails exposed to Schistosoma mansoni.

An acid-fast pathogen of fresh-water snails.

Chemoreception in the snail Australorbis glabratus.

Studies on the biological control of schistosome-bearing snails. II. The control of Australorbis glabratus populations by the leech, Helobdella fusca, under laboratory conditions.

Specificity of hemolymph antigens in taxonomic discrimination of medically important snails.