Comparison of Trypanosoma cruzi infection in dogs inoculated with blood or metacyclic trypomastigotes of Berenice-62 and Berenice-78 strains via intraperitoneal and conjunctival routes

This paper aimed to verify the influence of the inoculum source (blood or metacyclic trypomastigote) and the route of inoculation (intraperitoneal or conjunctival) on the course of T. cruzi infection in dogs, using comparatively the T. cruzi strains Berenice-62 and Berenice-78. All dogs inoculated intraperitoneally became infected independently of the T. cruzi strain and source of trypomastigotes used. High level of infectivity was also observed when metacyclic trypomastigotes of both strains were inoculated by conjunctival route. However, when blood trypomastigotes were inoculated by conjunctival route the percentages of infectivity were significantly lower in dogs inoculated with both strains. Parasitaemia was significantly higher in animals infected with metacyclic trypomastigotes via the conjunctival route independently of the T. cruzi strain used. All animals infected with Berenice-78 strain showed severe acute myocarditis. On the other hand, animals infected with Berenice-62 showed severe acute myocarditis only when infected with metacyclic trypomastigote, via the intraperitoneal route. The results suggest that the source of the inoculum and the route of inoculation remarkably influence the evolution of the infection for the T. cruzi in the vertebrate host even when the same strain of the parasite is used.

Metastatic melanoma positively influences pregnancy outcome in a mouse model: could a deadly tumor support embryo life?

The incidence of melanoma is increasing worldwide. It is one of the leading cancers in pregnancy and the most common malignancy to metastasize to placenta and fetus. There are no publications about experimental models of melanoma and pregnancy. We propose a new experimental murine model to study the effects of melanoma on pregnancy and its metastatic process. We tested several doses of melanoma cells until we arrived at the optimal dose, which produced tumor growth and allowed animal survival to the end of pregnancy. Two control groups were used: control (C) and stress control (SC) and three different routes of inoculation: intravenous (IV), intraperitoneal (IP) and subcutaneous (SC). All the fetuses and placentas were examined macroscopically and microscopically. The results suggest that melanoma is a risk factor for intrauterine growth restriction but does not affect placental weight. When inoculated by the SC route, the tumor grew only in the site of implantation. The IP route produced peritoneal tumoral growth and also ovarian and uterine metastases in 60% of the cases. The IV route produced pulmonary tumors. No placental or fetal metastases were obtained, regardless of the inoculation route. The injection of melanoma cells by any route did not increase the rate of fetal resorptions. Surprisingly, animals in the IV groups had no resorptions and a significantly higher number of fetuses. This finding may indicate that tumoral factors released in the host organism to favor tumor survival may also have a pro-gestational action and consequently improve the reproductive performance of these animals.

Rapid transplacental infection with bovine pestivirus following intranasal inoculation of ewes in early pregnancy

Despite the importance of congenital viral infections in both veterinary and human medicine, only limited experimental work has been carried out to elucidate the mechanisms involved in transplacental virus infections. To further an understanding of fetal infection with pestiviruses, the distribution of bovine pestivirus in the uterine and fetal tissues of ewes in early pregnancy, following a natural route of infection, was investigated. On the 18th day of pregnancy, nine ewes were inoculated by the intranasal route with 1 X 10(5) 50% tissue culture infective doses of an Australian isolate of noncytopathic bovine pestivirus (bovine viral diarrhea virus genotype 1). All ewes were ovariohysterectomized at approximately 100 hours postinfection. Samples from the reproductive tract and conceptus were examined histologically and tested for bovine pestivirus by nested reverse transcription-polymerase chain reaction (RT-PCR) and immunohistochemistry and for interferon-tau mRNA expression by nonnested RT-PCR. Although no histopathologic changes were observed in the maternal or fetal tissues, virus was detected in the reproductive tract of all nine ewes and in all of the conceptuses examined. Al; the time of surgery, only two of the nine ewes were demonstrably viremic. This study demonstrates that bovine pestivirus can spread from a natural site of infection to the ovine fetus within 4 days in the absence of maternal immunity and despite the presence of interferon expression in the reproductive tract.

Rickettsial Infection Caused by Accidental Conjunctival Inoculation

The most common transmission route of tick-borne Rickettsia is through tick bite; nevertheless, other transmission routes should also be considered. We report a case of rickettsial infection in a 15-year-old boy caused by accidental contamination of the conjunctiva through the infected fluid of a crushed engorged tick removed from a dog. Right eye pain, conjunctival hyperaemia with mucopurulent exudate, chemosis and eyelid oedema were the first signs and symptoms. Two days later, the boy developed fever, myalgia, headache, abdominal pain and was vomiting; physical examination showed multiple cervical adenopathies but no rash. He was treated with doxycycline (200 mg/day) for 7 days with progressive resolution of clinical signs. Rickettsial infection was confirmed by immunofluorescence assay with serological seroconversion in two consecutive samples. Rickettsia conorii or Rickettsia massiliae were the possible causal agents since they are the Rickettsia spp found in the Rhipicephalus sanguineus dog tick in Portugal.

Trypanosoma (herpetosoma) Rangeli tejera, 1920: influence of host weight, size of inoculum, and route of infection upon experimental parasitemia

To study the influence of host age, inoculum size, and route of infection on Trypanosoma (Herpetosoma) rangeli, 12 lots of 6.0 g albino mice (NMRI strain) were infected up. with from 25x10¹ to 25x10(6) trypomastigotes/gram body weight harvested from LIT medium. The lower inocula produced low but persistent parasitemias, while the higher inocula produced high levels of parasitemia that fell quickly, suggesting the mobilization of resistance mechanisms. In other experiments, i.p. inoculation produced higher parasitemias than s.c. inoculation, and 6.0 g mice had higher parasitemias than 16.0 or 26.0 g mice. Thus, a standard methodology would seem to be necessary in the study of the various strains and/or species that may make up the T. rangeli complex.

Severe pneumonia due to Parachlamydia acanthamoebae following intranasal inoculation: a mice model.

Parachlamydia acanthamoebae is an obligate intracellular bacterium naturally infecting free-living amoebae. The role of this bacterium as an agent of pneumonia is suggested by sero-epidemiological studies and molecular surveys. Furthermore, P. acanthamoebae may escape macrophages microbicidal effectors. Recently, we demonstrated that intratracheal inoculation of P. acanthamoebae induced pneumonia in 100% of infected mice. However, the intratracheal route of infection is not the natural way of infection and we therefore developed an intranasal murine model. Mice inoculated with P. acanthamoebae by intranasal inoculation lost 18% of their weight up to 8 days post-inoculation. All mice presented histological signs of pneumonia at day 2, 4, 7, and 10 post-inoculation, whereas no control mice harboured signs of pneumonia. A 5-fold increase in bacterial load was observed from day 0 to day 4 post-inoculation. Lungs of inoculated mice were positive by Parachlamydia-specific immunohistochemistry 4 days post-inoculation, and P. acanthamoebae were localized within macrophages. Thus, we demonstrated that P. acanthamoebae induce a severe pneumonia in mice. This animal model (i) further supports the role of P. acanthamoebae as an agent of pneumonia, confirming the third Koch postulate, and (ii) identified alveolar macrophages as one of the initial cells where P. acanthamoebae is localized following infection.

Toxoplasma gondii: Infection natural congenital in cattle and an experimental inoculation of gestating cows with oocysts

Two studies, of a natural infection and an experimental infection, were performed in order to study congenital transmission of Toxoplasma gondii in cattle. In the first study, 50 fetuses were harvested from gestating cows that were eutanasied at a municipal slaughterhouse in Jaboticabal, São Paulo state, Brazil. In the second study, 11 gestating cows were divided into four groups for inoculation with T. gondii: GI consisted of three cows inoculated with 1.0 x 10(5) oocysts during their first trimester of gestation; GII consisted of three cows inoculated with 1.0 x 10(5) oocysts during their second trimester of gestation; GIII consisted of three cows inoculated with 1.0 x 10(5) oocysts during their last trimester of gestation; and GIV consisted of two control cows, one during its first and the other during its second trimester of gestation. In both studies, the presence of T. gondii was confirmed both indirectly by immunofluorescence assay (IFAT). In the natural infection experiment, 18% (9/50) of the gestating cows were confirmed to have specific antibodies (IFAT - 1:64) against T. gondii. The bioassay was able to diagnose the presence of T. gondii in the tissue samples from three calves. In the second experiment, the nine cows from groups I, II and III presented with specific antibodies (IFAT) against T. gondii. In contrast, T. gondii could not be detected by IFAT, histopathological examination or the bioassay in any of the nine calves born to cows experimentally infected with T. gondii oocysts. Based on the results from both studies, we conclude that congenital infection of T. gondii in cattle, while infrequent, does occur naturally. The pathogenicity of the strain of T. gondii may influence the likelihood of this route of transmission. (C) 2010 Elsevier B.V. All rights reserved.

Migratory route of Strongyloides venezuelensis in Lewis rats: Comparison of histological analyses and PCR

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Effect of immersion and inoculation in ovo of Lactobacillus spp. in embryonated chicken eggs in the prevention of Salmonella enteritidis after hatch

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Leishmania spp. parasite isolation through inoculation of patient biopsy macerates in interferon gamma knockout mice

Isolation of Leishmania parasite and species identification are important for confirmation and to help define the epidemiology of the leishmaniasis. Mice are often used to isolate pathogens, but the most common mouse strains are resistant to infection with parasites from the Leishmania (Viannia) subgenus. In this study we tested the inoculation of interferon gamma knockout (IFNγ KO) mice with biopsy macerates from Leishmania-infected patients to increase the possibility of isolating parasites. Biopsies from twenty five patients with clinical signs of leishmaniasis were taken and tested for the presence of parasites. Immunohistochemical assay (IHC) and conventional histopathology detected the parasite in 88% and 83% of the patients, respectively. Leishmania sp. were isolated in biopsy macerates from 52% of the patients by culture in Grace's insect medium, but 13% of isolates were lost due to contamination. Inoculation of macerates in IFNγ KO mice provides isolation of parasites in 31.8% of the biopsies. Most isolates belong to L. (Viannia) subgenus, as confirmed by PCR, except one that belongs to L. (Leishmania) subgenus. Our preliminary results support the use of IFNγ KO mice to improve the possibility to isolate New World Leishmania species.

Histopathological, immunohistochemical and ultraestructural evaluation of inflammatory response in Arius genus fish under experimental inoculation of BCG

The aim of this study was to evaluate the inflammatory response kinetics after experimental inoculation with BCG in the primitive Arius sp. fish. The BCG was applied through the intramuscular injection in the caudal peduncular region, and the samples were collected for the analyses at days 1, 3, 7, 14, 21, and 33 post-injection. Acute phase inflammatory infiltrate was characterized by the predominant mononuclear cells, intersticial edema, and muscular tissue necrosis. As the inflammatory response evolved, a large number of multinuclear giant cells were formed containing the BCG. These giant cells were positive for the S100 protein at the histochemical analysis, which demonstrate the macrofage activity, confirmed by the ultra-structural analysis showing the lack of the cytoplasmic membrane enveloping the many nuclei within the giant cell. These results led to the conclusion that Arius sp. fish injected with the BCG showed a difuse inflammatory response characterized by a large number of mononuclear cells, absence of granuloma formation, and predominant giant cells.

A Simple and Fast Method for the Production and Characterization of Methylic and Ethylic Biodiesels from Tucum Oil via an Alkaline Route

A simple, fast, and complete route for the production of methylic and ethylic biodiesel from tucum oil is described. Aliquots of the oil obtained directly from pressed tucum (pulp and almonds) were treated with potassium methoxide or ethoxide at 40 degrees C for 40 min. The biodiesel form was removed from the reactor and washed with 0.1 M HCl aqueous solution. A simple distillation at 100 degrees C was carried out in order to remove water and alcohol species from the biodiesel. The oxidative stability index was obtained for the tucum oil as well as the methylic and ethylic biodiesel at 6.13, 2.90, and 2.80 h, for storage times higher than 8 days. Quality control of the original oil and of the methylic and ethylic biodiesels, such as the amount of glycerin produced during the transesterification process, was accomplished by the TLC, GC-MS, and FT-IR techniques. The results obtained in this study indicate a potential biofuel production by simple treatment of tucum, an important Amazonian fruit.

Mycoplasma pneumoniae and/or Chlamydophila pneumoniae inoculation causing different aggravations in cholesterol-induced atherosclerosis in apoE KO male mice

Background: Chamydophila pneumoniae (CP) and/or Mycoplasma pneumoniae ( MP) are two bacteria detected in vulnerable atheromas. In this study we aimed to analyze whether CP and/or MP aggravates atherosclerosis induced by cholesterol-enriched diet in C57BL/6 apoE KO male mice. Thirty male apoE KO mice aged eight weeks fed by a diet containing 1% cholesterol until 32 weeks of age were divided into four groups: the first was inoculated with CP (n = 7), the second with MP (n = 12), the third with both CP + MP ( n = 5), and the fourth with saline (sham n = 6). The animals were re-inoculated at 36 weeks of age, and sacrificed at 40 weeks of age. Two ascending aorta and one aortic arch segments were sampled. In the most severely obstructed segment, vessel diameter, plaque height, percentage of luminal obstruction and the degree of adventitial inflammation were analyzed. The plaque area/intimal surface ratio was obtained by measuring all three segments. The adventitial inflammation was semiquantified (0 absent, 1 mild, 2 moderate, and 3 diffuse). Results: The mean and standard deviation of plaque height, % luminal obstruction, external diameter, the plaque area/intimal surface ratio and the adventitial inflammation values are the following for each group: MP (0.20 +/- 12 mm, 69 +/- 26%, 0.38 +/- 0.11 mm, 0.04 +/- 0.04 and 0.22 +/- 0.67), CP (0.23 +/- 0.08 mm, 90 +/- 26%, 0.37 +/- 0.08 mm, 0.04 +/- 0.03, and 0.44 +/- 0.53), MP + CP ( 18 +/- 0.08 mm, 84 +/- 4.0%, 0.35 +/- 0.25 mm, 0.03 +/- 0.03 and 1.33 +/- 0.82) and sham (0.08 +/- 0.09 mm, 42 +/- 46%, 0.30 +/- 0.10 mm, 0.02 +/- 0.03 and 0.71 +/- 0.76). A wider area of plaque/intimal surface was observed in MP + CP inoculated groups (p = 0.07 and 0.06) as well as an increased plaque height in CP (p = 0.01) in comparison with sham group. There was also an increased luminal obstruction (p = 0.047) in CP inoculated group in comparison to sham group. Adventitial inflammation in MP + CP inoculated group was higher than MP, CP and the sham groups (p = 0.02). Conclusion: Inoculation of CP, MP or both agents in C57BL/6 apoE KO male mice caused aggravation of experimental atherosclerosis induced by cholesterol-enriched diet, with distinct characteristics. CP inoculation increased the plaque height with positive vessel remodeling and co-inoculation of MP + CP caused the highest adventitial inflammation measures.

Tamoxifen Is Effective in the Treatment of Leishmania amazonensis Infections in Mice

Background: Chemotherapy is still a critical issue in the management of leishmaniasis. Until recently, pentavalent antimonials, amphotericin B or pentamidine compounded the classical arsenal of treatment. All these drugs are toxic and have to be administered by the parenteral route. Tamoxifen has been used as an antiestrogen in the treatment and prevention of breast cancer for many years. Its safety and pharmacological profiles are well established in humans. We have shown that tamoxifen is active as an antileishmanial compound in vitro, and in this paper we analyzed the efficacy of tamoxifen for the treatment of mice infected with Leishmania amazonensis, an etiological agent of localized cutaneous leishmaniasis and the main cause of diffuse cutaneous leishmaniasis in South America. Methodology/Principal Findings: BALB/c mice were infected with L. amazonensis promastigotes. Five weeks post-infection, treatment with 15 daily intraperitoneal injections of 20 mg/kg tamoxifen was administered. Lesion and ulcer sizes were recorded and parasite burden quantified by limiting dilution. A significant decrease in lesion size and ulcer development was noted in mice treated with tamoxifen as compared to control untreated animals. Parasite burden in the inoculation site at the end of treatment was reduced from 10(8.5 +/- 0.7) in control untreated animals to 10(5.0 +/- 0.0) in tamoxifen-treated mice. Parasite load was also reduced in the draining lymph nodes. The reduction in parasite number was sustained: 6 weeks after the end of treatment, 10(15.5 +/- 0.5) parasites were quantified from untreated animals, as opposed to 10(5.1 +/- 0.1) parasites detected in treated mice. Conclusions/Significance: Treatment of BALB/c mice infected with L. amazonensis for 15 days with tamoxifen resulted in significant decrease in lesion size and parasite burden. BALB/c mice infected with L. amazonensis represents a model of extreme susceptibility, and the striking and sustained reduction in the number of parasites in treated animals supports the proposal of further testing of this drug in other models of leishmaniasis.

Iridium(III)-surfactant complex immobilized in mesoporous silica via templated synthesis: a new route to optical materials

In this work we report the preparation of a new blue-emitting material based on the templated synthesis of mesoporous silica (MCM-41) using micellar solutions of the newly synthesized monocationic metallosurfactant complex bis[1-benzyl-4-(2,4-difluorophenyl)-1H-1,2,3-triazole](4,4'-diheptadecyl-2,2'- bipyridine)-iridium(III) chloride in hexadecyl-trimethyl-ammonium bromide (CTAB). Under ambient conditions, significant increases in excited state lifetime and quantum yield values (up to 45%), were obtained for the solid materials in comparison to the corresponding micellar solutions. Solid state (1)H and (19)F NMR spectroscopies were successfully employed for quantifying the luminophore content in terms of Ir-surfactant to CTAB and Ir-surfactant to silica ratios.