Schistosoma mansoni: male-biased sex ratios in snails and mice

Adult sex ratios of Schistosoma mansoni, in mice, were shown to be biased toward males (3:1) despite the finding that sex ratios of miracidia were 50:50. The adult male bias was caused by greater male infectivity of miracidia for snails and cercariae for mice. A significantly higher percentage of male miracidia developed to cercarial production in unimiracidial infections (57 male, 34 female), and a significantly higher percentage of male cercariae developed to adulthood in mice (143 male, 79 female worms resulted from 900 male and 900 female cercariae). No significant differences were found between male and female parasites for longevity of miracidia (both sexes, 10 hr) and cercariae (males 21.3 +/- 5.75 hr, females 25.0 +/- 7.02 hr), prepatent periods in snail hosts (male 34 +/- 2.92 days, females 33 +/- 2.36 days), longevity of snail infections (males 96.6 +/- 25.15 days, females 115.2 +/- 82.43 days), and the numbers of cercariae produced per snail lifetime (males 30,751.44 +/- 18,064.33, females 34,083.00 +/- 33,732.82). Present results provide a better understanding of the life cycle of S. mansoni, are of theoretical significance for theories of biased sex ratios (which at present cannot account for the male-biased ratio of S. mansoni), and also suggest that schistosomiasis transmission models assuming a 50:50 sex ratio at all stages of the life cycle should be reassessed.     

The influence of host sex on the L3 to L4 molt of Brugia malayi

Immunocompetent male mice are more susceptible to experimental infection with Brugia spp. than are females. Because permissive male SCID mice (severe combined immunodeficient mice), which lack T and B cells, also possess higher worm burdens, the mechanism is not solely immune mediated. Recovery of fewer adult worms from the female SCID mouse suggests that females do not provide sufficient nutrients for larval growth. This study assessed the potential of the female SCID mouse to support the L3 to L4 molt of Brugia malayi. Unexpectedly, worms grown in females molted at earlier time points of recovery than those harvested from males. This suggests that the early stage of development of B. malayi is delayed in the male murine host. To determine whether the effect of host sex on molting may be similar in humans, worms were cultured in media supplemented with serum from male or female donors. Worms grown in serum obtained from female donors exhibited a significantly higher percentage of complete molts over those cultured with serum from males. Host-derived molecules required for the L3 to L4 molt may be more abundant in the female, perhaps allowing the worms to survive a vulnerable developmental stage in a less permissive environment.     

Schistosoma japonicum: Infection characteristics in mice of various strains and a difference in the response to eggs

Female mice of 12 inbred strains were exposed to 20–25 cercariae of Schistosoma japonicum and infection status determined at day 40 by counting numbers of adult worms, eggs in faeces and eggs in a segment of liver. Most mouse strains appeared to be ‘permissive’ hosts although at least one strain (129/J) was shown to be relatively resistant in terms of day 40 adult worm numbers. In a radioisotopic lung assay for sensitivity to eggs, and developed as a rapid means of assessing granuloma formation, CBA/H mice were shown to differ from C57BL/6 mice in being non-responders. Histological examination of lungs of sensitized CBA/H and C57BL/6 mice injected intravenously with eggs established that granuloma formation was much more intense in C57BL/6 than CBA/H mice. Preliminary indications are that infected CBA/H mice are also low anti egg circumoval precipitin (COP) responders. Analysis of immune responses to isolated egg antigens in these two strains, and identification of the antigens of eggs to which such responses are directed in C57BL/6 mice, should provide insights into immunological disease processes (such as granulomatous inflammation) in this model system of japonicum schistosomiasis.     

Trichinella spiralis: role of non-H-2 genes in resistance to primary infection in mice

Two strains of mice which share identical H-2 genes but differ in their genetic backgrounds were compared for their ability to resist infection with Trichinella spiralis. The two strains of mice, C3HeB/FeJ and AKR/J, share the H-2k haplotype which is associated with susceptibility to primary infection with T. spiralis in H-2 congenic strains of mice. AKR/J mice, infected with 150 infective muscle larvae, harbored significantly fewer muscle larvae 30 days postinfection than did mice of the strain C3HeB/FeJ. Approximately equal numbers of worms establish in the small intestine of AKR and C3H mice, but the AKR mice expelled adult worms from the gut more rapidly than did mice of the C3H strain. By Day 9 postinfection, 50% of the worms had been expelled by the AKR mice whereas expulsion of worms from C3H mice was delayed beyond Day 9 and occurred primarily between Days 10 and 12. Over this same experimental period (Days 6-12), fecundity of female worms from AKR mice, measured as the mean newborn larvae/female/hour, was approximately one-half that of worms taken from C3H mice. These results support the conclusion that genes outside of the mouse H-2 complex regulate expulsion of adult worms from the gut. These background genes also markedly influence the fecundity of female worms.     

Defective resistance to Plasmodium yoelii in CBA/N mice.

The role of B lymphocytes in resistance to malaria was studied in defective and normal F1 mice derived from CBA/N mice, a strain with an X-linked B cell defect. When infected with normally nonlethal Plasmodium yoelii, immune defective F1 male mice had higher parasitemias and more prolonged infections than normal F1 mice, as well as a 50% mortality rate. Before infection the plasma levels of IgM and IgG were lower in defective F1 males than normal F1 mice. The polyclonal IgM and IgG responses of infected abnormal F1 mice were delayed and lower in absolute magnitude than those of normal F1 mice. Furthermore, specific IgM and IgG anti-plasmodial antibody titers, as determined by radioimmunoassay, were depressed on day 12 in the defective F1 males. Although IgG titers approached those of the normal F1 mice on day 19, defective F1 male IgM titers remained depressed. These data demonstrate that an X-linked gene that affects B cell function influences malarial resistance in mice, presumably via a decreased specific IgM response, and the slow development of a specific IgG response to P. yoelii infection.     

Infections of forest rat filaria, Breinlia booliati, in neonate and juvenile laboratory white rats

The kinetics of Breinlia booliati infection in 3 inbred rat strains (Lewis, Wistar and Sprague Dawley) were investigated. One group of rats was infected as neonates (less than 24 hours of age) with third-stage larvae of B. booliati and the other group was infected as juveniles (4 weeks of age). The results showed that infection in the neonates were significantly different from the infection in the juveniles. The 60 rats infected as neonates, when necropsied between 8 to 10 months postinfection, yielded adult worms. The 2 neonatal infection groups of Lewis and Wistar strains showed highest susceptibility to the infections. The mean prepatent period was 85 days. Ninety to 95% of the infected rats were patent with microfilaraemia and a large percentage (33 to 47%) of them had high microfilaraemia counts exceeding 3000 mff/20 mm3 of blood and larger sizes (mean 157.11 mm for female adult worms and 61.88 mm for male adult worms. The adult worms were distributed equally in both the pleural (57%) and peritoneal cavity (43%). In most aspects, the neonatal infection group of the Sprague-Dawley strain was intermediate in susceptibility between the 2 neonatal infection groups of the Lewis and Wistar strains and the 3 juvenile infection groups. In contrast to neonatal infection groups, the 3 juvenile infection groups exhibited low infection rates (37%, 58% and 47% for the Lewis, Wistar and Sprague Dawley strains respectively), longer prepatent periods (mean 101 days), lower recovery rates (2 to 4%), lower adult worm loads (mean 0.4 to 0.8 female worms, and 0.2 to 0.8 male worms per rat), and smaller sizes (mean 141.24 mm for female adult worms and 53.75 mm for male adult worms). Forty-four to 57% of these infected rats harboured either single male or single female adult worms in the body cavity. Most (92%) of the adult worms recovered from the juvenile infection groups resided in the pleural cavity and the remaining 8% were recovered from the peritoneal cavity. Microfilaraemia could be detected in only 3/20 Lewis rats, 5/20 Wistar rats and 5/20 Sprague Dawley rats. The mean peak microfilaraemia of the 3 pooled juvenile infection groups was 632 mff/20 mm3 of blood, ranging from 7 mff/20 mm3 to 1856 mmf/20 mm3. Our results indicate that the susceptibility to B. booliati infection in white rats is both genetic and age-associated. The responses of the 2 distinct infection groups to B. booliati infections are discussed.     

Effects of low-protein diet on Schistosoma mansoni morphology visualized by morphometry and confocal laser scanning microscopy

Previous studies have shown that protein deficiencies can hamper both the course of experimental schistosomiasis and normal development of adult worms. To further investigate this relationship, we compared adult male and female Schistosoma mansoni from malnourished and well-fed mice through morphometric and confocal laser scanning microscopy analysis. Swiss mice were fed protein-deficient diets (8%) and infected subcutaneously with approximately 80 S. mansoni cercariae (BH strain, Brazil). Control mice were fed a standard rodent diet (23% protein). The nutritional status was evaluated by body weight gain and albumin values. Mice were sacrificed 63 days post-infection. Recovered worms were stained with hydrochloric carmine and preserved as whole-mounts for bright-field examination and confocal microscopy. The body weight gain and serum albumin concentrations were significantly lower (P < 0.05) in malnourished mice than in controls. In general, all morphometric values of specimens grown in malnourished mice were lower than those of control mice. Schistosome worms grown in malnourished mice had statistically significant differences (P < 0.05) in the reproductive system and tegument than those grown in mice fed standard diets. In female worms, vitelline glands showed few remaining follicles and ovaries lacked mature oocytes. In male parasites, tubercles were fewer in number on the dorsal surface and testicular lobes presented fewer differentiated germinal cells. In summary, we describe novel data supporting the view that low-protein diets may influence the development of adult worms.     

Reappraisal of experimental infections with cercariae and schistosomula of a Brazilian strain of Schistosoma mansoni in mice.

The present investigation involves a reevaluation of previous results obtained after experimental infection of Swiss Webster mice with cercariae and schistosomula of the Schistosoma mansoni LE strain maintained under laboratory conditions. Three experimental groups of mice were considered: the animals of the first group were percutaneously (ring method) infected with cercariae, those of the second were subcutaneously inoculated with cercariae and the mice of the third were inoculated by the same route with schistosomula transformed in vitro. The data obtained so far indicated that the most effective method of infection is the subcutaneous injection with schistosomula, with a mean adult worm burden recovery of 54.1% when compared to the abdominal percutaneous and subcutaneous routes of infection with cercariae, in which the values were 36.7% and 32.4%, respectively. This suggests that, in experimental infections of SW mice with a LE S. mansoni strain, the skin is to be considered an effective attrition site in the percutaneous route, whereas in the case of inoculation with cercariae, a small amount of larvae fails to be transformed into viable schistosomula, possibly due to skin phase avoidance. A brief discussion about attrition sites and elimination of larval S. mansoni worms in mice is presented.     

Tegumental changes in adult Schistosoma mansoni harbored in mice treated with artemether

A detailed temporal examination was made of alterations induced by artemether in the tegument of adult Schistosoma mansoni worms using scanning electron microscopy (SEM). Mice infected with S. mansoni cercariae 42 days previously were treated intragastrically with artemether at a single dose of 400 mg/kg. Groups of 3 mice were killed at 24 hr, 72 hr, and 7 days after treatment; the worms were collected by perfusion and examined by SEM. Twenty-four hours after artemether treatment, focal damage to the tubercles on the tegumental surface of male worms was seen. In both male and female worms, there was focal swelling and fusion of tegumental ridges, and sometimes peeling. After 72 hr, the damage to the tegument had increased, especially in female worms, with extensive swelling, fusion, and peeling of the tegumental ridges. In the most severely damaged worms, host leukocytes were seen to be adhered to the damaged tegument. Damage to the oral sucker was also occasionally seen in both male and female worms. Seven days after treatment, the appearance of the tegument had returned to normal in some male and female worms, whereas others still showed apparent damage. The results demonstrate that artemether damages the tegument of adult S. mansoni, and the intensity of damage is more severe in female worms than in males.     

Influence of primary infection on the population dynamics of Nematospiroides dubius after challenge infections in mice

Dobson C., Sitepu P. and Brindley P. J. 1985. Influence of primary infection on the population dynamics of Nematospiroides dubius after challenge infections in mice. International Journal for Parasitology15: 353–359. Similar proportions of the inoculum of Nematospiroides dubius larvae reached sexual maturity by 14 days after administration of 50–400 larvae but more adult worms had been expelled by day 63 after infection from those mice infected with 50 vs 400 larvae. There was a significant correlation between time and worm expulsion for all inoculum size groups except for mice given 400 larvae.In mice reinfected with 100 larvae, after termination of primary infections derived from 10 through 400 larvae, more worms from the challenging dose were recovered from mice given greater compared with those given smaller numbers of larvae at primary infection. The N. dubius population size after challenge infection was correlated positively both with number of larvae administered as the primary infection and with the resultant population size during that infection. The serum anti-N. dubius antibody titres after reinfection were higher in mice given 400 compared with those given fewer larvae at primary infection, and the fecundity and female to male sex ratio of the N. dubius populations decreased in proportion to these antibody titres.Protective immunity against challenge N. dubius infection, in mice which had been drenched free of adult worms established from 400 larvae for 5 down to 1 weeks before reinfection, increased from 45% (1 week) to 80% (5 weeks). There was a negative correlation between the population size of N. dubius during challenge infection and the duration between anthelmintic treatment and challenge infection.     

Dipetalonema viteae: Response of spleen cells in experimental mouse filariasis to mitogens and antigens

Balb/c mice were infected by transplanting 3, 5, 10, or 20 female adult Dipetalonema viteae under the dorsal skin. The microfilaremias resulting from infections with 3 or 5 adult worms were of lesser magnitude and of shorter duration than those produced in infections with 10 or 20 worms. Spleen cells taken from these mice at various intervals after infection were assayed in vitro for their ability to respond to phytohemagglutinin (PHA) or lipopolysaccharide (LPS). There was no depression in the response to LPS or to PHA in mice given infections of 3 or 5 D. viteae adult worms. In contrast, the response to PHA was significantly depressed in groups receiving 10 or 20 adult female worms 12 days after infection and by Day 25, the depression was severe. Thereafter the PHA responsiveness recovered gradually to reach control values on Day 60. In mice transplanted with 10 or 20 D. viteae adult worms there was no significant depression in the response to LPS at any time during the infection, but the response was increased slightly sporadically during infection. These results indicate that in mice, this infection causes an initial suppression in the function of PHA-sensitive T cells but has little effect on the B cells which respond to LPS. A factor present in serum taken on Day 25 from mice infected with 10 or 20 adult worms inhibited the proliferative response to PHA by spleen cells from normal mice. The recovery of PHA responsiveness in mice given the heavier infections coincided with death of the adult worms, but mitogen reactivity and microfilaremia were unrelated. Antigens from male or female worms induced cell division in spleen cells taken from infected mice after microfilaremia had ceased whether they were implanted with 3 or 10 adult worms.     

Influence of immunizing dose and presence or absence of adult worms on the development of resistance to Nematospiroides dubius challenge infections of mice

H-2 congenic strains expressing resistant (H-2q, H-2f) or susceptible (H-2k) haplotypes were compared for their ability to resist challenge infection with N. dubius following a 6- or 14-day ivermectin-abbreviated immunizing infection. B10.BR mice (H-2k) were considerably more resistant to infection when the priming interval was shortened from 14 to 6 days. B10.Q (H-2q) and B10.M (H-2f) mice resisted challenge regardless of which immunization regimen was used. The influence of parasite numbers on the response to challenge was studied by comparing infections in resistant DBA/1 (H-2q) and susceptible CBA/J (H-2k) mice that differ at both H-2 and non-H-2 genes. DBA/1 mice, immunized with 50 or 150 L3 of N. dubius for 14 days, resisted challenge, whereas mice receiving 300 worms did not. In contrast, CBA/J mice failed to resist challenge at all priming doses tested. When the immunizing infection was shortened from 14 to 6 days, DBA/1 mice resisted challenge regardless of priming dose and CBA/J mice resisted challenge only when the highest dose of 300 worms was used for priming. The data suggest that susceptible strains of mice may be preferentially immunosuppressed, particularly at low infective doses, and that suppression is associated with adult worms present in the lumen of the small intestine.     

Schistosoma mansoni: stage-specific expression of muscle-specific genes

It was previously shown that an antigen preparation termed 9B obtained from Schistosoma mansoni cercarial extracts partially (34%) protects mice from challenge infection with cercariae (R. Tarrab-Hazdai et al., J. Immunol. 135, 2772, 1985). To characterize some of the proteins which comprise this preparation, rabbit antibodies to the 9B antigen preparation were used to screen cDNA libraries of cercariae and adult worms. We isolated and sequenced cDNA clones encoding three proteins: calcium-binding protein, paramyosin, and myosin. The calcium-binding protein was previously shown to be expressed in cercariae but not in sporocysts or adult worms (D. Ram et al., Mol. Biochem. Parasitol. 34, 167, 1989). Northern blots showed the presence of paramyosin and myosin mRNAs in sporocysts and adult worms but not in cercariae. Antibodies to paramyosin detected the protein in sporocysts and adult worms as well as in cercariae. These findings explain, in part, the protective activity of the 9B antigen preparation against challenge infection.     

In vivo and in vitro egg production by Nematospiroides dubius during primary and challenge infections in resistant and susceptible strains of mice

Experiments were conducted to examine adult worm burdens, fecal egg output, and in vitro fecundity of Nematospiroides dubius in resistant LAF1 and susceptible CBA mice 12, 15, 18, and 21 days following primary and challenge infections. A strong correlation was obtained on the number of eggs produced by worms cultured in vitro and the egg production as assessed by fecal egg count. Worm counts, fecal egg counts, and in vitro fecundity were similar on all days studied following a primary infection in both mouse strains. However, after challenge infection, LAF1 mice showed lower worm burdens, fecal egg output, and in vitro egg production when compared to CBA mice. Although the egg production of surviving female worms from immune LAF1 mice was decreased, it never fell below a threshold of 100 eggs/day. The reduced fecundity may be a manifestation of a general anti-worm response rather than responses directed specifically at worm reproduction.     

Relationships of deer mouse movement, vegetative structure, and prevalence of infection with Sin Nombre virus

The effects of vegetative structure on movement of deer mice (Peromyscus maniculatus) were examined in two distinct vegetation associations, one near Hesperus and the other near Molina in western Colorado (USA) from June-October 1994 to October 1998. We monitored movement by live-trapping small mammals within Gambel's oak/mixed-grass (Hesperus) and sage brush/juniper (Molina) vegetation types. Vegetative structure differed between the sites with Molina having more cover provided by shrubs and Hesperus having more cover provided by forbs. Adult male deer mice moved greater distances at Hesperus than at Molina. Sub-adult males tended to move greater distances than did adult females. Relative abundances of deer mice tended to differ by season, but the average relative abundance of deer mice was greater at Molina. Long-term prevalence of infection with SNV was greater at Hesperus and was greatest in adult males at Hesperus (36.1%). Adult males at Molina exhibited a prevalence of infection with SNV of 25.0%. Infection with SNV was highly associated with scars or wounds for adult male, adult female, and juvenile male deer mice at Hesperus, but only for adult female deer mice at Molina. The presence of scars or wounds tended to be associated with greater age, but male deer mice at Hesperus were more likely to have wounds than female deer mice of the same age class. A similar pattern, excluding juveniles, was observed at Molina. Intraspecific interactions and environmentally elicited long-distance movements of deer mice may play a role in prevalence of infection with SNV in these animals.     

Nicarbazin in schistosome infections: I. Antibody formation in mice and hamsters infected with Schistosoma mansoni.

The immunoprecipitin response of mice infected with Schistosoma mansoni and treated with nicarbazin, an egg suppressive agent, is significantly lower than in untreated-infected mice. The precipitin response to cercarial extract is virtually abolished in treated mice indicating common antigenic determinants with eggs of the same species. Haemagglutinins to adult worms are also significantly diminished in treated mice. Finally, circumoval precipitins are absent in treated mice when the drug is given continuously to infected mice in order to prevent egg laying by the female adult parasite. The results suggest that a significant portion of the precipitating antibody produced in schistosome infections reactive with cercariae and adult worms, as well as eggs, is probably a secondary antibody response due to common antigenic determinants found in eggs.     

Chemosignals from isolated females have antimutagenic effect in dividing the cells of bone marrow from male mice of the CBA strain

A level of X-ray induced mitotic disturbances in the cells of the bone marrow of male mice was studied under the modifying influence of chemosignals from isolated adult female mice of the CBA strain. It has been shown that the frequency of chromosomal aberrations in irradiated (4 Gr) males after exposing them for 24 hours on bedding soiled with female chemosignals is lower than in irradiated males in cages with clean bedding. The mechanisms and importance of the antimutagenic effect of female house mouse chemosignals are discussed.     

X-Y chromosome dissociation in mouse strains difering in efficiency of spermatogenesis: Elevated frequency of univalents in pubertal males

Dissociation of the X-Y chromosome bivalent in diakinesis-metaphase I spermatocytes of adult mice was significantly more frequent in the CBA strain (29%) than in C57, KP, or KE strains (7–11%). Autosome dissociatio (1–5%) involved only the smallest chromosome pairs. Eleyatedfrequency of X-Y dissociation in the CBA strain correlates with significantly lower testes weight and lower yield of spermatogenesis, which suggests that sex bivalent dissociation man be responsible for some loss of spermatogenic cells. However, sperm quality is not affected, the percentage of normal spermatozoa and their fertlizing capacity being higher in CBA thatn in the remaining strains. Two congenic strains, KE and KE. CBA (the latter with the Y chromosome introduced from CBA), had the same level of X-Y dissociatios, suggesting that the Y chromosome plays no rle in the determination of this character. In comparison with adult males pubertal (27–29 day-old) males had twice as hig a frequency of X-Y dissociation in KE an KP strains, and combined frequeicies of dissociated sex and autosome bivalents were significantly higher in pubertal males of all tested strains. Although te level of chromosome dissociation is not sufficient to explain increased mortality of germ cells observed in pubertal males, it could be one of the contributing factors.     

Penetration and maturation of Schistosoma mansoni in suckling and adult Swiss Webster and DBA/2 mice

Murine schistosomosis is a widely used experimental model of the human disease. Different methods have been employed to infect mice with Schistosoma mansoni cercariae, such as subcutaneous or intraperitoneal injections, the tail immersion technique, and the use of a metal ring placed on the abdominal skin of anaesthetized animals. An alternative method of infection that requires no anaesthesia and no restraint is to place suckling mice (10 days old) on a Petri dish with a small volume of water containing cercariae. In this study, we compared the penetration and maturation of S. mansoni in mice infected by this alternative method with those noted in mice infected at an older age (45 days) by the tail immersion technique. Besides evaluating the effects of age and method of infection, we also compared the susceptibility of two strains of mice (Swiss Webster (SW) and DBA/2). Mice were exposed to 100 cercariae and worms (by portal-hepatic perfusion) as well as eggs were recovered in the liver and intestines on postinfection (PI) days 35, 55 and 90. Skin penetration was very efficient (about 100%) irrespective of the mouse strain, sex, age and method of infection. Worm and egg recoveries were higher in SW mice at any PI interval, but strain differences tended to be less pronounced on PI day 90. In both strains, recoveries of worms and eggs were clearly higher in mice infected at a younger age (10 days old). This study thus suggests that infection of free-moving suckling mice is a suitable alternative to other methods of infection with S. mansoni.     

Surface antigens on cercariae,schistosomula and adult worms of Schistosoma mansoni

Shah J. and Ramasamy R. 1982. Surface antigens on cercariae, schistosomula and adult worms of Schistosoma mansoni. International Journal for Parasitology12: 451–461. The surface protein antigens of Schistosoma mansoni were radiolabelled by lactoperoxidase catalysed I¹²⁵-iodination and analysed by immune-precipitation and polyacrylamide gel electrophoresis. The results showed that regularly labelled surface antigens of mol. wts >150,000, 78,000, 45,000 and 22,000 were present on adult worms. Common surface antigens were observed on the cercariae, schistosomula and adult worms. It is suggested that surface antigens released from living adult worms can sensitise a host to react against the invading schistosomula of a secondary infection. However, the failure to vaccinate mice using material containing adult worm surface antigens suggests that the induction of protective immunity is a complex phenomenon.