Dickson D. Despommier

Toxocariasis: Clinical Aspects, Epidemiology, Medical Ecology, and Molecular Aspects

SUMMARY Toxocariasis is caused by a series of related nematode species (ascarids) that routinely infect dogs and cats throughout the world. The eggs from these ascarids are common environmental contaminants of human habitation, due largely to the fact that many kinds of dogs and cats serve as pets, while countless others run wild throughout the streets of most urban centers. The eggs, present in dog and cat feces, become infectious within weeks after they are deposited in the local environment (e.g., sandboxes, city parks, and public beaches, etc.). Humans, particularly children, frequently ingest these eggs by accident and become infected. Infection in humans, in contrast to their definitive hosts, remains occult, often resulting in disease caused by the migrating larval stages. Visceral larva migrans (VLM) and ocular larva migrans (OLM) are two clinical manifestations that result in definable syndromes and present as serious health problems wherever they occur. Diagnosis and treatment of VLM and OLM are difficult. These issues are summarized in this review, with emphasis on the ecology of transmission and control of spread to both humans and animals through public health initiatives employing treatment of pets and environmental intervention strategies that limit the areas that dogs and cats are allowed within the confines of urban centers.

How Does Trichinella spiralis Make Itself at Home

The nurse cell-parasite complex of Trichinella spiralis is unlike anything else in Nature. It is derived from a normal portion of striated skeletal muscle cell and develops in a matter of 15 to 20 days after the larva invades that cell type. What are the molecular mechanisms at work that result in this unique relationship? Here, Dickson Despommier presents a hypothesis to account for its formation, in which secreted tyvelosylated proteins of the larva play a central role. These proteins are always present in the intracellular niche of the larva from Day 7 after infection and may be responsible for redirecting host genomic expression, leading to nurse cell formation.

Infectivity of the newborn larva of Trichinella spiralis in the rat.

A method is described for the isolation of large numbers of newborn larvae from adult Trichinella spiralis. These isolated larvae were used in experiments which tested their ability to infect mice via the intraduodenal, intravenous, and intraperitoneal routes, and to infect rats via the intraperitoneal and intravenous routes. Encysted muscle larvae were seen in diaphragm preparations from mice injected intravenously and intraperitoneally 14 days previously with newborn larvae. No muscle larvae were seen in diaphragm muscle preparations from mice which had received newborn larvae intraduodenally. Quantitative studies on larval infectivity were carried out in rats. Larval counts showed that 66% of the larvae were recoverable as mature muscle larvae from rats injected iv, while only 2% of the original dose was recovered from ip injected rats. In another experiment using larger numbers of rats in each group, newborn larvae were 73% infective in iv injected rats, and only 9% infective in ip injected rats. Nembutal, administered ip to rats prior to infection with newborn larvae, had no major effect on larval infectivity. Limited numbers of newborn larvae have been isolated from host body cavities (Matoff, 1940; Berntzen, 1965; Shanta and Meerovitch, 1967) and blood (Matoff, 1943b; Gould et al., 1955; Phillipson and Kershaw, 1961) during early phases of intestinal infection and have been described in morphological detail (Richels, 1955; Ali Khan, 1966). Denham (1967) collected parenterally infective newborn larvae after incubation of gravid females for 24 to 48 hr. However, isolation of large numbers of newborn larvae of a determinable age has not previously been described, although Larsh (1963) and others have suggested that this would be necessary in order to better correlate the stage of the parasite with host immunity. Inoculation of gravid female worms and newborn larvae of Trichinella spiralis demonstrated that newborn larvae could infect dogs and rats by the parenteral route (Matoff, 1943a). However, quantitative data about infectivity, pathReceived for publication 3 February 1970. t The Rockefeller University, New York, New York 10021. +t Please address reprint requests to author Dickson D. Despommier. * This work was supported, in part, by USPHS Training Grant F02-A-142057, USPHS Grant AI-04842, and NIH Post-Doctoral Fellowship 1F2-AI--31, 188. ogenicity, and immunogenicity of the newborn larva of T. spiralis are still lacking. We describe here a reproducible system for obtaining newborn larvae of a known age in large numbers. Isolation techniques are described, and a quantitative comparison is made of the infectivity of pure preparations of larvae inoculated into rats via the intraperitoneal and intravenous routes. We show that both mice and rats can be infected with this stage via the ip and iv routes and that the newborn larvae are more infective for the rat when they are inoculated iv as opposed to ip. MATERIALS AND METHODS Mature muscle larvae were obtained from stockinfected CFW male mice by the method of Larsh and Kent (1949). Male Wistar rats (120 g) were inoculated orally with approximately 10,000 muscle larvae suspended in a mixture of 0.6% nutrient broth and 2% gelatin with the aid of a syringe fitted with a blunt 18-gauge needle. We removed the food from the rats on the 6th day postinfection and killed them on the 7th. The entire small intestine was then removed from each animal, slit longitudinally, cut into 2-cm sections, and placed in a modified Baermann apparatus containing 0.85% saline solution at 37 C. Adult worms were collected over a period of 4 hr, and resedimented in flasks, then washed 4 times at intervals of 0.5 hr with saline at 37 C. Worms thus treated appeared free of debris and were viable. The worms were then transferred to a large, glass moisture chamber containing 300 cc of the following me-

The protein nature of the substance inducing female monogamy in Aedes aegypti

Abstract Virgin females of Aedes aegypti are inseminated only once. Subsequent insemination is inhibited by a substance from the male accessory gland that is transferred to the female in seminal fluid at the initial mating. This substance, called ‘matrone’, can be extracted from the whole bodies of male mosquitoes. Injection of matrone into females renders them sterile for life. Gel filtration chromatography with Sephadex G-10, G-25, G-50, and G-100 indicates matrone activity can be separated into at least two components, both macro-molecules. The active material is temperature and pH sensitive and is completely inactivated by treatment with 5% trichloroacetic acid or an amyl-alcohol-chloroform mixture. In addition treatment with the proteolytic enzyme, protease, will result in a complete loss of activity. On the basis of these observations it was concluded at least one of the molecular components which make up active matrone is a protein. Partial purification was achieved by extraction methods using cold acetone and butanol, ammonium sulphate, manganous chloride, gel filtration chromatography with Sephadex, and ion-exchange chromatography with diethylaminoethanol-cellulose.

Trichinella spiralis: Mediation of the intestinal component of protective immunity in the rat by multiple, phase-specific, antiparasitic responses

Abstract Rats infected orally with Trichinella spiralis developed an immunity that was induced by and expressed against separate phases of the parasites enteral life cycle. Infectious muscle larvae generated an immune response (rapid expulsion) that was directed against the very early intestinal infection and resulted in the expulsion of worms within 24 hr. This response eliminated more than 95% of worms in an oral challenge inoculum. Developing larvae (preadults) also induced an immune response that was expressed against adult worms. The effect on adults was dependent upon continuous exposure of worms to the immune environment throughout their enteral larval development. Immunity induced by preadult T. spiralis was not expressed against adult worms transferred from nonimmune rats. While adult worms were resistant to the immunity engendered by preadults they induced an efficient immunity that was autospecific. Both “preadult” and “adult” immunities were expressed in depression of worm fecundity as well as in the expulsion of adults from the gut. However, the two reactions differed in respect to their kinetics and their efficiency against various worm burdens. Preadult immunity was directed mainly against fecundity whereas adult immunity favored worm expulsion. All responses (rapid expulsion, preadult and adult immunity, and antifecundity) acted synergistically to produce sterile immunity against challenge infections of up to 5000 muscle larvae. These findings indicate that the host protective response to T. spiralis is a complex, multifactorial process that operates sequentially and synergistically to protect the host against reinfection.

The in vivo and in vitro analysis of immunity to Trichinella spiralis in mice and rats.

Wistar-Furth strain male rats and CFW strain male mice were immunized against Trichinella spiralis using an antigenic fraction derived from a cell-free homogenate of mature muscle larvae. In rats, animals immunized with 250 mug of antigen harboured significantly fewer (135000) muscle larvae 30 days after oral challenge than controls (231000). Further 7-day-old adult worms derived from immunized rats shed 48% fewer (P less than 0.001) newborn larvae over a 24h period in vitro than adult worms from non-immunized animals. Mice were injected with either 10 or 100 mug of antigen. In comparison with non-immunized controls, mice immunized with 100 mug of antigen harboured significatnly fewer adult worms at days 7 and 9 after oral challenge infection, while female worms recovered from immune mice on days 6-10 after challenge shed fewer newborn larvae in vitro. Finally, mice immunized with 100 mjg of antigen harboured significantly fewer (10391) muscle larvae at 30 days after challenge than did controls (47750). Immunization of mice with 10 mug of antigen did not induce a statistically significant reduction in adult worms at either day 7 or 9 after challenge (P less than 0.5). However, adult female worms from mice receiving 10 mug of antigen still shed fewer larvae than did adults from control mice (P less than 0.05). Mice immunized with 10 mug of antigen harboured significantly fewer (13700) recoverable muscle larvae than did controls at 30 days after challenge (39000).

The stichosome and its secretion granules in the mature muscle larva of Trichinella spiralis.

The stichosome of the mature muscle larva of Trichinella spiralis consists of a single row of 45 to 55 stichocytes. Each stichocyte is about 25 mum in diameter and possesses a single nucleus. A duct leads from each stichocyte to the lumen of the esophagus. The stichocyte cytoplasm contains mitochondria, structures resembling Golgi-complexes, rough endoplasmic reticulum, and usually 1 of 2 types of secretory granules. The alpha-granule measures about 800 mn in diameter, contains a prominent inclusion, and has a granular matrix. The beta-granule is about 600 mn in diameter and is homogeneous in appearance. Both granule types are surrounded by a single membrane. Ten to thirteen stichocytes containing alpha-granules are confined to the posterior portion of the stichosome. After isopycnic centrifugation in sucrose gradient of large granule fractions obtained from cell-free homogenates, the alpha- and beta-granules show characteristic distribution patterns as revealed by the morphology of the fractions. The median equilibrium density of the alpha-granules is 1.245, while that of the beta-granules is 1.230. There is a correlation between the distribution of the granules and of antigens reacting with hyperimmune antitrichinella seruma. At least 4 unique antigens can be attributed to each of the granule types. Fractions enriched in mitochondria do not contain specific antigens. Antigens from both types of secretory granules cross react totally with those present in the excretion-secretion products of living muscle larvae. Cytoimmunochemical data show that antigens are distributed in a patchy fashion throughout the stichocyte cytoplasm. This finding is consistent with the distribution of the secretory granules in the intact stichocyte.

Trichinella spiralis: Secreted antigen of the infective L1 larva localizes to the cytoplasm and nucleoplasm of infected host cells

Antibodies were elicited against a purified antigen with an apparent molecular weight of 43K. This antibody preparation also detected a second antigen consisting of a group of closely related components of 45-50K. These antigens are stage specific for the infective first stage larva of Trichinella spiralis and are among the repertoire of secreted antigens originating from the stichosome. Antibody raised against the 43K antigen reacted with the stichosome and cuticle of the mature larva and the cytoplasm and nucleoplasm, but not nucleolus, of all nuclei of infected host cells (Nurse cells) in sections of infected tissues. Studies on sections of synchronously infected muscle tissue revealed that antigen was present only within the worm on Day 7 of the infection. On Day 9 after infection, the stichosome and cuticular surface of the larva and the cytoplasm and nucleoplasm of each nucleus of the Nurse cell reacted with antibody. Nurse cell cytoplasmic and nuclear reactivity increased in intensity until Day 18 after infection. These results suggest that stichocyte-specific antigens are synthesized during the early phase of infection in the muscle, and that as the Nurse-parasite complex develops, some of the antigen is secreted into the milieu of the Nurse cell. The presence of antigen in the cytoplasm and nucleoplasm of the infected host cell is discussed in relation to Nurse cell formation and maintenance.

Trichinella spiralis and the concept of niche.

Trichinella spiralis is an intracellular parasite as both a larva and an adult. The first-stage larva lives in a modified portion of a skeletal muscle cell, the nurse cell, and can reside there for the life span of the host. Adult worms occupy a nonmembrane-bound portion of columnar epithelium, living there as intramulticellular parasites. The newborn larva is the only nonintracellular stage, living free in the circulation. Trichinella spiralis induces modifications in each of its intracellular niches. Parasite signals secreted into the milieu of the developing nurse cell results in the reprogramming of host genomic expression, reflected in loss of muscle-specific proteins, over-expression of collagen, and the development of a circulatory rete. Formation of the nurse cell is complex, presumably involving many steps; yet there is not a large series of related intermediate forms in nature. Trichinella pseudospiralis induces an incomplete nurse cell. Adult parasites cause the death of the infected epithelium. The precise nature of most of the signals from parasite to host and from host to parasite has not been determined. As a direct consequence of exposure to some of them, the host develops long-lasting immunity to reinfection. This may confer advantages both for the parasite, as well as the host, because strong immune responses should reduce intraspecific competition.

Farming up the city: the rise of urban vertical farms

The invention of agriculture dates back some 10 000 years and arose spontaneously at multiple sites throughout the world (Mexico, China, The Middle East, and Borneo). It rapidly spread to almost every culture, offering a better life to those who embraced it. The human population was estimated then to be around 1 million. Farming in all its forms increased over the next millennia by leaps and bounds, eventually changing forever the face of the Earth. Yet, for thousands of years, right up to modern times, agriculture was essentially practiced in the same way as the original farmers derived it: dig a hole, plant a seed, fertilize it, irrigate it, pick out the weeds, harvest the crop, ship/store/sell it.