Comparative Susceptibility of Different Mouse Strains to Liver-Stage Infection With Plasmodium berghei Sporozoites Assessed Using In Vivo Imaging.

BACKGROUND: The liver stages of Plasmodium parasites are important targets for the discovery and development of prophylactic drugs. METHODS: A real-time in vivo imaging system was used to determine the level of luminescence measured from firefly luciferase expression by sporozoites developing in hepatocytes in different strains of mice. RESULTS: The luminescence values (photon counts/sec) measured from the anatomical liver location in the untreated mice infected with 10,000 Plasmodium berghei sporozoites were 8.15 × 105 for C57BL/6 Albino, 2.12 × 105 for C3H/HeNCrL, 0.91 × 105 for C57BL/6 WT, 0.28 × 105 for BALB/c, and 0.16 × 105 for ICR/CD-1 mice. This data suggests that the C57BL/6 Albino strain is most susceptible to luminescent photon, mainly because the less light scattering and absorption from deeper tissues and the skin in the strain of mouse. The photon count observed in black C57BL/6 wild type mice was shown to be 88.83% lower compared to C57BL/6 Albino mice. Although the highest growth rate of sporozoites in hepatocytes was found for C57BL/6 wild type mice in this study, the black skin of this mouse significantly reduced parasite-associated bioluminescence. CONCLUSIONS: The minimal light scattering and absorption and also enhanced susceptibility to liver infection of C57BL/6 Albino mice makes this strain preferable sensitivity for discovery and development of prophylactic antimalarial drugs.

Plasmid requirements for infection of ticks by Borrelia burgdorferi.

Borrelia burgdorferi strain B31 MI commonly loses one or more of its complement of 21 extrachromosomal plasmids during normal handling procedures and during genetic manipulations. Certain plasmid losses cause an inability or reduction in the ability of spirochetes to infect mice. In the current study, nine strains of spirochetes with varying plasmid profiles were used to identify plasmids necessary for nymphal tick infection. Nymphal ticks were artificially fed the nine spirochete strains as well as the parental strain containing a full complement of plasmids. The capillary fed nymphs were allowed to feed on mice for at least 63 h and then examined for the presence of spirochetes in their guts and salivary glands. All spirochete strains tested were able to infect ticks guts, but to different degrees. We determined that the plasmids lp5, lp28-1, and cp9 were not required for infecting tick guts, whereas loss of lp25 and lp28-4 was associated with reduced gut infectivity. A reduction in the ability of spirochetes to invade salivary glands was seen in bacteria that did not have lp28-1, whereas cp9 was not required for salivary gland infection. This study has pinpointed specific plasmids whose absence is deleterious to infecting nymphal tick guts and salivary glands.

Phenotypic plasticity in adult worms of Schistosoma mansoni (Trematoda:Schistosomatidae) evidenced by brightfield and confocal laser scanning microscopies.

A comparative morphometric study was performed to identify host-induced morphological alterations in Schistosoma mansoni adult worms. A wild parasite population was obtained from a naturally infected rodent (Nectomys squamipes) and then recovered from laboratory infected C3H/He mice. Furthermore, allopatric worm populations maintained for long-term under laboratory conditions in Swiss Webster mice were passed on to N. squamipes. Suckers and genital system (testicular lobes, uterine egg, and egg spine) were analyzed by a digital system for image analysis. Confocal laser scanning microscopy (CLSM) showed details of the genital system (testicular lobes, vitelline glands, and ovary) and the tegument just below the ventral sucker. Significant morphological changes (p < 0.05) were detected in male worms in all experimental conditions, with no significant variability as assessed by CLSM. Significant changes (p < 0.05) were evident in females from the wild population related to their ovaries and vitelline glands, whereas allopatric females presented differences only in this last character. We conclude that S. mansoni worms present the phenotypic plasticity induced by modifications in the parasite's microenvironment, mainly during the first passage under laboratory conditions.

A comparative parasitologic study on Biomphalaria glabrata snail and C3H/He mice infected with human and murine isolates of Schistosoma mansoni derived from Sumidouro, Rio de Janeiro, Brazil.

Experiments were carried out to analyze the biological characteristics of two sympatric isolates of Schistosoma mansoni derived from humans and murines in a low endemic transmission area (Sumidouro county, state of Rio de Janeiro, Brazil). Sympatric reared-laboratory Biomphalaria glabrata and C3H/He mice were used as experimental hosts. Parameters assessed comprised: precercarial period, infectivity and mortality (snails), prepatent period, infectivity (percentage of cercariae maturation into adult worm) and intestinal egg count (mice). The murine isolate showed a shorter precercarial period and higher infectivity than human isolate (p<0.05). This biological heterogenicity did not correspond to the vertebrate data because any biological parameter presented significant difference (p>0.05). These data suggest that both isolates are local sub-populations, providing support for the hypotheses that in a same biotope mixed populations or sub-populations circulate among their main host (human beings) and/or rodent as an anfixenous infection.

Natural Schistosoma mansoni infection in Nectomys squamipes: histopathological and morphometric analysis in comparison to experimentally infected N. squamipes and C3H/He mice.

Histopathologic and morphometric (area, perimeter, major and minor diameters) analysis of hepatic granulomas isolated from twelve naturally infected Nectomys squamipes were compared to four experimentally infected ones and six C3H/He mice. Liver paraffin sections were stained for cells and extracellular matrix. Both groups of N. squamipes presented peculiar granulomas consisting predominantly of large macrophages, full of schistosome pigment, characterizing an exudative-macrophage granuloma type, smaller than the equivalent granuloma type in mouse. Naturally infected animals exhibited granulomas in different stages of development, including large number of involutional types. Morphometric analysis showed that all measurements were smaller in naturally infected animals than in other groups. The results demonstrated that both N. squamipes groups reproduced, with small variations, the hepatic granuloma aspects already described in cricetidium (Calomys callosus), showing a genetic tendency to set up strong macrophage responses and small granulomas. Unexpectedly, natural infection did not engender distinguished histopathological characteristics distinct from those derived from experimental single infection, showing changes predominantly secondary to the duration of infection. It appears that the variability of the inocula (and the number of infections?) interfere more with the quantity than with the quality of the pathological changes, denoting some morpho-functional determinism in the response to schistosomal infection dependent on the animal species.

Growth and development of Gymnophalloides seoi in immunocompetent and immunosuppressed C3H/HeN mice.

The growth and development of Gymnophalloides seoi were studied in C3H/HeN mice and effects of immunosuppression of the host on the worm development were observed. Two hundred metacercariae of G. seoi were orally administered to each mouse, and worms were recovered on days 1, 3, 5, 7, 14 and 21 post-infection (PI). The worm recovery rate was significantly higher in immunosuppressed (ImSP) mice than in immunocompetent (ImCT) mice except on days 1 and 3 PI. The worms attained sexual maturity by day 3 PI with eggs in the uterus, and worm dimensions and the number of uterine eggs continuously increased until day 14 PI in ImSP mice. Worms recovered from ImSP mice were significantly larger in size than those from ImCT mice on days 1 and 3 PI, and the number of uterine eggs was significantly larger in ImSP mice on days 5 and 7 PI. Genital organs such as the ovary, testes, and vitellaria, that were already developed in the metacercarial stage, grew a little in size until day 14 PI. The results show that the C3H/HeN mouse is, though not excellent, a suitable laboratory host for G. seoi.

Susceptibility of various species of animals and strains of mice to Gymnophalloides seoi infection and the effects of immunosuppression in C3H/HeN mice.

Susceptibility to Gymnophalloides seoi infection was studied in 8 species of animals, including 7 strains of mice; the effects of immunosuppression on susceptibility were examined in C3H/HeN mice. One hundred metacercariae of G. seoi isolated from naturally infected oysters were orally administered to each animal. Worm recovery rate (WRR), worm dimensions, and the number of uterine eggs were obtained at day 3 and day 7 postinfection (PI). Average WRR from gerbils, hamsters, and cats at day 7 PI was 28.0%, 14.2%, and 10.9%, respectively, the former 2 figures of which were significantly higher than the rate of 0.0-4.0% from Sprague-Dawley rats, dogs, ducks, guinea pigs, and chicks. In the case of mice, average WRR at day 7 PI was 12.4% (KK strain), 11.8% (C3H/HeN), 9.6% (ICR), 6.4% (BALB/c), and 6.3% (ddY), respectively; the first 3 figures were significantly higher than the rates from other strains, which were 1.8% (A) and 0% (C57BL/6). At day 3 PI, WRR was much higher in all strains except C57BL/6. Worm maturation was the highest in C3H/HeN mice. Immunosuppression of C3H/HeN mice by injecting prednisolone for 7, 14, or 21 days prior to infection increased WRR at day 7 PI to 27.8%, 33.8%, or 67.5%, respectively. The results show that gerbils, hamsters, cats, and KK, C3H/HeN, ICR. BALB/c, and ddY mice are laboratory hosts that are fairly susceptible to G. seoi infection. In C3H/HeN mice, susceptibility was markedly enhanced by immunosuppression.

Onset of resistance to light Hymenolepis nana infection in mice of different strains.

Rapidity in onset of resistance against Hymenolepis nana egg infection after a light primary infection was studied in low and high responder mice challenged at different time intervals. A very rapid acquisition of protection was observed in C57 and a delayed response in C3H mice. In both cases the effect of resistance on weight or worm number was related to the time of challenge infection, suggesting a "race against time" involving host response and parasite development, the outcome varying according to host genetic background.

Immune stimulation and Hymenolepis nana cysticercoid expulsion in C3H/He and BALB/c mice.

Adoptive transfer of immunity with heterologous and homologous immune serum, and drug-abbreviated immunizations were used in C3H and BALB/c mice to determine the strain-characteristic time of expulsion of H. nana cysts. Transfer of immune serum did not accelerate worm expulsion in C3H, while elimination of worms was virtually complete by day 8 in BALB/c mice. Loss of worms was also obtained when BALB/c mice were stimulated with abbreviated infections using 20 or 1000 H. nana eggs. The immunizing infection terminated immediately after the tissue phase. After similar immunizations C3H mice again appeared slow responders but were able to affect the intestinal worms population after the higher immunizing infection. The data obtained suggest that the time of worm expulsion was related to the genetically-determined ability of the mice to respond and was independent of the stimulations used for immunization. A quantitative difference in response is proposed to explain the slow responder status of C3H.

Expulsion of Echinostoma trivolvis (Cort, 1914) Kanev, 1985 and retention of E. caproni Richard, 1964 (Trematoda: Echinostomatidae) in C3H mice: pathological, ultrastructural, and cytochemical effects on the host intestine.

C3H mice were infected with 30 metacercarial cysts of either echinostome to study the pathological, ultrastructural, and cytochemical effects of the infection on the mouse small intestine. In mice infected with Echinostoma caproni, the intestine showed villous atrophy with fused or eroded villi. The microvilli of the enterocytes were sparse and distorted and showed reduced alkaline phosphatase activity. The crypts of Lieberkuhn were hyperplastic and showed a marked reduction in goblet and Paneth cells. As compared with uninfected controls, there was a marked reduction in glucose-6-phosphatase activity in the enterocytes of the infected gut. Collagen fibers and the number of fibroblasts were increased under the epithelium. In mice infected with E. trivolvis, the tips of the intestinal villi were bent and blunted. The microvilli of the enterocytes were less tightly packed than those of uninfected controls. The mitochondria in the enterocytes were irregularly shaped, contained intracristal bodies, and showed increased cytochrome oxidase activity as compared with those of uninfected controls. The crypts were hyperplastic but showed an increase in the numbers of goblet and Paneth cells. The fibroblasts and collagen fibers showed abnormal development. The ultrastructural and cytochemical differences seen in this study reflect the uniqueness of the host-parasite relationship of each of these echinostome species in the gut of the C3H mouse.

A study of peritoneal cells from healthy and Schistosoma mansoni-infected mice with special reference to myofibroblasts arising in culture.

Adherent, trypsin-resistant, peritoneal cells from mice with chronic schistosomiasis mansoni, and from control mice, were cultivated in vitro up to 20 days. Fibroblasts regularly appeared, about 6 days after seeding, in cultures of the manyfold more numerous cells from infected mice, concomitantly with a dramatic increase, detected by autoradiography, in the percentage of DNA-replicating cells of the monocyte-macrophage lineage. Peritoneal cells from healthy and from infected mice were fractionated on discontinuous Percoll gradients. Eight cell subsets were harvested in both cases, quantitated, and studied by electron microscopy. Two fractions (2 and 3: 1.041 < densities < 1.060 g/ml) from infected mice were greatly enriched in monoblasts and promonocytes. The cells of the different subsets were seeded separately, trypsin-treated and cultivated in vitro. Cultures of cell fractions 2 and 3 from infected mice contained the majority of the DNA-synthesizing cells and gave regularly rise to fibroblasts. Cultures of the different fractions were used for sequential morphological observations (2-11 days) at the electron microscope level. Early cultures were also used for the ultrastructural detection of the Mac-1 (CD 18/CD 11b) surface antigen by gold immunocytochemistry. A few fibroblasts were rarely observed in cultures of fractions 2 and 3 from control mice, while cells with ultrastructural features of myofibroblasts were regularly observed in cultures of the same fractions harvested from mice with chronic schistosomiasis. Fractions 2 and 3 from infected mice contained a large number of Mac-1 positive monoblasts. The correlations between the presence of monoblasts, DNA replication in cells of the monocyte-macrophage lineage and the appearance of myofibroblasts in cultures of the same fractions derived from infected mice are discussed.

The skunk Conepatus chinga as new host of Trypanosoma cruzi in Argentina.

We report the first systematic epidemiological research carried out in Argentina on the skunk Conepatus chinga. Forty-nine animals were captured in the settlements of Amamá, Trinidad, and nearby forested areas located in the Department of Moreno, Province of Santiago del Estero, between April 1985 and May 1989. Isolation of parasites was done through xenodiagnosis, and their identification as Trypanosoma cruzi was achieved by biological and biochemical criteria. The isolate was highly virulent and pathogenic in inoculated C3H mice. Prevalence was 4.1% (2 of 49). Two facts account for a possible domestic source of infection: both infected skunks were captured near Trinidad, in an area that had never been treated with insecticides, and electrophoretic isoenzyme patterns of the parasites isolated from the skunks were identical to those found in humans. Because extensive deforestation probably would increase the distribution area of C. chinga, further investigation should be performed to evaluate the epidemiological role of this wild mammal.

Early expression in mice of genetic resistance to intestinal amebiasis.

Some strains of mice are known to be relatively resistant to hepatic or intestinal amebic infections. In order to know if the intestinal resistance is expressed few hours after infection, we inoculated axenic amebae in three inbred strains of mice either by direct intracecal injection or by infection of a washed-closed cecal loop. We found that amebae do not survive in conventional animals but they colonize longer in animals with the cecal loop. However, the survival was low after 24 hours postinfection. Balb/c mice were more susceptible and CBA mice more resistant. Our results suggest that genetic resistance to intestinal amebiasis is expressed in mice in the early phases of infection.

Changes in the isoenzyme and kinetoplast DNA patterns of Trypanosoma cruzi strains induced by maintenance in mice.

Culture forms of thirteen Trypanosoma cruzi strains from 4 zymodemes and 9 schizodemes were inoculated and kept by successive passages in C3H mice. The strains were initially from the following zymodemes: 3 from A, 3 from B, 4 from C and 2 from D and 1 from AB mixed zymodemes. After approximately 18 months maintenance the parasites were isolated by hemoculture and again typed according to their isoenzyme and kinetoplast DNA patterns. The zymodeme A strains kept their initial patterns; from the 3 zymodeme B strains, two kept the initial patterns and one changed to zymodeme A; from the 4 zymodeme C, two kept the initial pattern and two changed to zymodeme B; from the 2 zymodeme D strains, one kept the initial pattern and one changed to zymodeme A. The strain from AB mixed zymodeme was reduced to zymodeme. A. The zymodeme changes were accompanied by schizodeme changes. Although not simultaneously, in one T. cruzi strain the parasitemia change was followed by zymodeme and schizodeme changes. The results showed that prolonged maintenance of T. cruzi in mice by successive passages alters the isoenzyme and k-DNA patterns of some strains and that these alterations tend to move towards zymodeme A, suggesting a selective effect of mice over these T. cruzi populations.

Strain-dependent differences in susceptibility of mice to experimental Angiostrongylus costaricensis infection.

Experimental Angiostrongylus costaricensis infection was carried out in inbred strains of mice (C57BL/6 BALB/c, DBA/2 and C3H/He). All strains became infected with this parasite. Marked differences in mortality and in worm burden were found among inbred strains of mice tested. A significant reduction was shown in worm length from mice compared to that from cotton rats.

Relationship of route of infection to susceptibility and immune response of inbred mice to Y strain Trypanosoma cruzi.

C3HeB/FeJ (C3H) mice infected ip with 10(6), 5 x 10(5), and 10(5) blood-form trypomastigotes (BFTs) of the Y strain of Trypanosoma cruzi were more resistant than C57B1/6 (B6) mice infected in the same manner. This pattern of susceptibility is opposite that reported for other stocks of this parasite. In a second experiment, C3H and B6 mice were infected ip or sc with 2 x 10(6), 10(6), 5 x 10(5), 10(5), or 10(3) Y strain BFTs. C3H mice infected ip with the 3 highest doses were again more resistant than the B6 mice, while mice infected ip with the 2 lowest doses were essentially equivalent in resistance. Thus, the difference in susceptibility was detectable, in terms of parasitemia levels and survival, primarily at the higher infection doses. For the groups infected sc, the pattern of susceptibility reversed. B6 mice infected with the 3 highest doses had lower parasitemia levels than the corresponding C3H mice, while C3H and B6 mice infected with 10(5) or 10(3) BFTs were similar in resistance. Blastogenic responses of lymphoid cells to phytohemagglutinin (PHA) and a soluble trypanosome extract (STE) were compared for C3H mice infected ip or sc to determine if the susceptibility to infection obtained with the 2 routes would be associated with differences in immune responses. Mesenteric lymph node cells (MLNCs) of mice infected ip were responsive to the STE early in infection, while superficial lymph node cells (SLNCs) of these mice were not. C3H mice infected sc had SLNCs which yielded strong responses to STE, while their MLNCs were relatively unresponsive. PHA stimulated responses by lymphoid cells from mice infected ip or sc were similar.(ABSTRACT TRUNCATED AT 250 WORDS)

Caracterizaçäo biológica de clones das cepas Y, CL e MR de Trypanosoma cruzi em camundongos C3H isogênicos / Biological characterization of Trypanosoma cruzi clones from Y, CL and MR strains in C3H inbred mice

Dez clones isolados das cepas Y, CL e MR foram caracterizados segundo infectividade das culturas, curvas de parasitemia, polimorfismo e mortalidade em camundongos C3H isogênicos. Entra os clones das cepas Y e CL foram encontradas diferenças intragrupos bastante significativas. Os clones da cepa MR apresentaram maior homogeneidade. Estes resultados indicam que as cepas do T. cruzi podem apresentar diferentes graus de heterogeneidade. Também sugerem que as condiçöes utilizadas para a manutençäo de cepas de T. cruzi podem resultar em vantagens seletivas para algumas subpopulaçöes, podendo uma cepa ser o resultado da interaçäo destas subpopulaçöes (clones) selecionadas após alguns anos de manutençäo em laboratório

[Biological characterization of clones of the Y, CL and MR strains of Trypanosoma cruzi in inbred C3H mice]. / Caracterização biológica de clones das cepas Y, CL e MR de Trypanosoma cruzi em camundongos C3 H isogênicos.

Ten clones of Trypanosoma cruzi isolated from Y, CL and MR strains were studied. The infectivity of culture forms, parasitemia pattern, polymorphism and mortality were studied in C3H inbred mice. Significant intra-group differences among Y and CL clones were found. MR clones showed higher homogeneity. These data indicate that T. cruzi strains can show different degrees of heterogeneity. It is suggested that conditions used to maintain T. cruzi strains may result in a selective advantage for some subpopulations (clones) after many years of laboratory maintenance.

Trypanosoma cruzi: constancy of clone pathogenicity for inbred mice during long-term in vitro maintenance.

The reproducibility of infection of C3H/He mice with T. cruzi clones Sylvio-X10/4 and Sylvio-X10/7 maintained in the laboratory for 946 and 496 days respectively was assayed. Clone X10/7 from 15 different in vitro passages consistently induced an acute lethal infection (94.3% mortality) and constant survival time (Mean = 24.6 d.p.i.). Female mice survived significantly longer than males and mice older than 30 days at the time of infection survived significantly longer than younger mice. The mortality of mice infected with clone X10/4 from 23 different in vitro passages was lower (5.1%) and their survival longer (Mean = 42.7 d.p.i.) than mice infected with clone X10/7. High anti-T. cruzi IgG titres were detected in the plasma of all mice killed after 30 d.p.i. Histologically, the heart, skeletal muscles and/or large intestine contained intracellular parasites in 59% of the mice; parasites were found in mice of all groups tested. The hearts of all mice were comparably inflammed regardless of parasite passage number or duration of infection. These data demonstrate that the presentation and course of infection of inbred mice with two T. cruzi clones is not changed by either the duration or protocol used for in vitro maintenance; the reported loss or change in virulence of T. cruzi strains during long-term in vitro maintenance did not occur. Consequently, T. cruzi clones and inbred mice provide a dependable and reproducible means to study host-parasite interactions.