Development of Schizont Stages of <em>Toxoplasma gondii</em> in Primary Cell Culture of Feline Enterocytes

Intestinal epithelial cell cultures are a potentially applicable model for investigating enteropathogens such as the protozoan Toxoplasma gondii, the etiological agent of toxoplasmosis. Felids such as domestic cats are the only known definitive hosts where the parasite undergoes sexual reproduction, which occurs in the enterocytes. Primary feline intestinal epithelial cell (FIEC) cultures were obtained from the fetal small gut of felines, and the epithelial nature of these cells was confirmed by the revelation of cytokeratin and intestinal alkaline phosphatase content by fluorescence microscopy, besides alignment, microvilli, and adherent intercellular junctions by ultrastructural analysis. FIECs infected with T. gondii bradyzoite forms showed that the parasite:cell ratio was determinant for establishing the lytic cycle and cystogenesis and the induction of schizont-like forms. Type C and D schizonts were identified by light and electron microscopies, which showed morphological characteristics like those previously described based on the analysis of cat intestines experimentally infected with T. gondii. These data indicate that FIECs simulate the microenvironment of the felid intestine, allowing the development of schizogony and classic endopolygeny. This cellular framework opens new perspectives for the in vitro investigation of biological and molecular aspects involved in the T. gondii enteric cycle

Establishment of a murine model of congenital toxoplasmosis and validation of a qPCR assay to assess the parasite load in maternal and fetal tissues

Toxoplasma gondii is the causative agent of toxoplasmosis, a disease that affects warm-blooded animals and one third of the human population worldwide. Pregnant women who have never been exposed to the parasite constitute an important risk group, as infection during pregnancy often leads to congenital toxoplasmosis, the most severe form of the disease. Current therapy for toxoplasmosis is the same as it was 50 years ago and has little or no effect when vertical transmission occurs. Therefore, it is urgent to develop new strategies to prevent mother-to-fetus transmission. The implementation of experimental animal models of congenital toxoplasmosis that reproduces the transmission rates and clinical signs in humans opens an avenue of possibilities to interfere in the progression of the disease. In addition, knowing the parasite load in maternal and fetal tissues after infection, which may be related to organ abnormalities and disease outcome, is another important step in designing a promising intervention strategy. Therefore, we implemented here a murine model of congenital toxoplasmosis with outbred Swiss Webster mice infected intravenously with tachyzoites of the ME49 strain of T. gondii that mimics the frequency of transmission of the parasite, as well as important clinical signs of human congenital toxoplasmosis, such as macrocephaly, in addition to providing a highly sensitive quantitative real-time PCR assay to assess parasite load in mouse tissues. As the disease is not restricted to humans, also affecting several domestic animals, including companion animals and livestock, they can also benefit from the model presented in this study

Platelet Activating Factor Blocks Interkinetic Nuclear Migration in Retinal Progenitors through an Arrest of the Cell Cycle at the S/G2 Transition

Nuclear migration is regulated by the LIS1 protein, which is the regulatory subunit of platelet activating factor (PAF) acetyl-hydrolase, an enzyme complex that inactivates the lipid mediator PAF. Among other functions, PAF modulates cell proliferation, but its effects upon mechanisms of the cell cycle are unknown. Here we show that PAF inhibited interkinetic nuclear migration (IKNM) in retinal proliferating progenitors. The lipid did not, however, affect the velocity of nuclear migration in cells that escaped IKNM blockade. The effect depended on the PAF receptor, Erk and p38 pathways and Chk1. PAF induced no cell death, nor a reduction in nucleotide incorporation, which rules out an intra-S checkpoint. Notwithstanding, the expected increase in cyclin B1 content during G2-phase was prevented in the proliferating cells. We conclude that PAF blocks interkinetic nuclear migration in retinal progenitor cells through an unusual arrest of the cell cycle at the transition from S to G2 phases. These data suggest the operation, in the developing retina, of a checkpoint that monitors the transition from S to G2 phases of the cell cycle

First record of Thyene cf. pulchra (Araneae: Salticidae: Plexippina) in Brazil

Mariante, Rafael M., Hill, David E. (2018): First record of Thyene cf. pulchra (Araneae: Salticidae: Plexippina) in Brazil. Peckhamia 173 (1): 1-6, DOI: http://doi.org/10.5281/zenodo.716973

First report of the African jumping spider Thyene coccineovittata (Araneae: Salticidae: Plexippina) in Brazil

Mariante, Rafael M., Hill, David E. (2019): First report of the African jumping spider Thyene coccineovittata (Araneae: Salticidae: Plexippina) in Brazil. Peckhamia 173 (2): 1-23, DOI: http://doi.org/10.5281/zenodo.373271

First report of the Asian jumping spider Menemerus nigli (Araneae: Salticidae: Chrysillini) in Brazil

Mariante, Rafael M., Hill, David E. (2020): First report of the Asian jumping spider Menemerus nigli (Araneae: Salticidae: Chrysillini) in Brazil. Peckhamia 205 (1): 1-21, DOI: http://doi.org/10.5281/zenodo.387520

Review of specimens corresponding to three species of Thyene (Araneae: Salticidae: Plexippini) in the Peckham Collection

Hill, David E., Mariante, Rafael M. (2019): Review of specimens corresponding to three species of Thyene (Araneae: Salticidae: Plexippini) in the Peckham Collection. Peckhamia 185 (1): 1-12, DOI: 10.5281/zenodo.509336

Thyene coccineovittata

&lt;i&gt;Thyene coccineovittata&lt;/i&gt; (Simon 1886) &lt;p&gt; &lt;i&gt;Hyllus coccineovittatus&lt;/i&gt; Simon 1886 &male; &female;&lt;/p&gt; &lt;p&gt; &lt;i&gt;Thyene crudelis&lt;/i&gt; Peckham &amp; Peckham 1903 &male;, synonomy by Berland &amp; Millot 1941&lt;/p&gt; &lt;p&gt; &lt;i&gt;Thyene pulchra&lt;/i&gt; Peckham &amp; Peckham 1903 &female;, &lt;b&gt; &lt;i&gt;new synonymy&lt;/i&gt;:&lt;/b&gt; Weso&lstrok;owska &amp; Haddad 2009 &female;: Hill &amp; Mariante 2019 &female;&lt;/p&gt; &lt;p&gt; &lt;i&gt;Thyene coccineovittata&lt;/i&gt; Berland &amp; Millot 1941 &male; &female; (but not &female; &lt;i&gt;T. ogdeni&lt;/i&gt;): Weso&lstrok;owska &amp; Haddad 2009 &male; (only): Dawidowicz &amp; Weso&lstrok;owska 2016 &male;: Oger &amp; Van Keer 2017 &male;: Hill &amp; Mariante 2019 &male; (&lt;i&gt;T. crudelis&lt;/i&gt;)&lt;/p&gt; &lt;p&gt; &lt;i&gt;Thyene&lt;/i&gt; cf. &lt;i&gt;pulchra&lt;/i&gt; Mariante &amp; Hill 2018 &female;&lt;/p&gt; &lt;p&gt; &lt;i&gt;Diagnosis.&lt;/i&gt; Each pedipalp of the male &lt;i&gt;Thyene coccineovittata&lt;/i&gt; has a very short retrolateral tibial apophysis (RTA), often difficult to observe behind the many coils of the embolus. The epigynum of the female is very lightly sclerotized, generally with only a small anteromedial &lt;i&gt;parens&lt;/i&gt; visible through the cuticle. The male is dark brown to black, the female light yellow-brown and translucent. Both sexes have a middorsal opisthosomal band comprised of broad, flat white-yellow scales, and many of these scales are present on the carapace and/or the chelicerae. On either side this band is flanked by dark stripes, interrupted with paired spots toward the rear, and highlighted with bright, scarlet scales. The species group name (&lt;i&gt;bearing scarlet stripes&lt;/i&gt;) is almost certainly based on the appearance of these flanking stripes, but their coloration varies and in some males they are black without any scarlet scales. In females the flanking stripes are usually broken into large, black spots. Other characters, such as the femoral stripes of females, are shared with other species of &lt;i&gt;Thyene&lt;/i&gt; and are thus of limited use for purposes of identification. Here (Figures 3- 22) we present photographs of male, female and immature &lt;i&gt;T. coccineovittata&lt;/i&gt; with reference to the sites (Figure 2) where they have been observed in the state of Rio de Janeiro.&lt;/p&gt; &lt;p&gt; &lt;i&gt;Habitat&lt;/i&gt; (Figure 23). Our two study sites (Figure 2: sites 1-2) are both in Rio de Janeiro, although other observers have found &lt;i&gt;Thyene coccineovittata&lt;/i&gt; near Cabo Frio on the eastern side of the state of Rio de Janeiro (Figure 22). At both of these sites &lt;i&gt;T. coccineovittata&lt;/i&gt; was found on vegetation in inhabited areas. One of the authors (RMM, 9 OCT 2018; Mariante &amp; Hill 2018) first found this species standing on a leaf of &lt;i&gt;Triplaris&lt;/i&gt; sp. (Polygonaceae, &lt;i&gt;Ant Tree&lt;/i&gt;), a common native tree in Brazil, at the Oswaldo Cruz Institute (site 2), a campus of Fiocruz, a scientific institution for research and development in the biological sciences. This campus has many native trees separated by the stairs, sidewalks and paths that give access to the buildings. Many Neotropical salticids (e.g., &lt;i&gt;Bryantella&lt;/i&gt;, &lt;i&gt;Chira&lt;/i&gt;, &lt;i&gt;Colonus&lt;/i&gt;, &lt;i&gt;Corythalia&lt;/i&gt;, &lt;i&gt;Frigga&lt;/i&gt;, &lt;i&gt;Lyssomanes&lt;/i&gt;, &lt;i&gt;Maeota&lt;/i&gt;, &lt;i&gt;Phiale&lt;/i&gt;, &lt;i&gt;Sarinda&lt;/i&gt;, &lt;i&gt;Sumampattus&lt;/i&gt; and &lt;i&gt;Metaphidippus&lt;/i&gt;) have previously been found at the same location. We do not presently consider this species to be a synanthrope in this area as it has not been found living on man-made structures, but its survival in Brazil may nonetheless be dependent on cultivated plants in a semi-urban environment. We do not know if this spider has invaded unaltered natural habitats in Brazil, but the finding of this species in a semi-natural habitat in the state of Rio de Janeiro should not be ignored, since some introduced spiders can become invasive and displace native species. Although the best-known introduced spiders are synanthropic and cosmopolitan (Cutler 1990; Taucare-R&iacute;os &amp; Edwards 2012; Taucare-R&iacute;os 2013), some recent salticid introductions that are neither synanthropic nor cosmopolitain have also been very successful (Kaldari et al. 2011; Gall &amp; Edwards 2016).&lt;/p&gt;Published as part of &lt;i&gt;Mariante, Rafael M. &amp; Hill, David E., 2019, First report of the African jumping spider Thyene coccineovittata (Araneae: Salticidae: Plexippina) in Brazil, pp. 1-23 in Peckhamia 173 (2)&lt;/i&gt; on pages 1-20, DOI: &lt;a href="http://zenodo.org/record/3732711"&gt;10.5281/zenodo.3732711&lt;/a&gt

New insights from the oyster Crassostrea rhizophorae on bivalve circulating hemocytes.

Hemocytes are the first line of defense of the immune system in invertebrates, but despite their important role and enormous potential for the study of gene-environment relationships, research has been impeded by a lack of consensus on their classification. Here we used flow cytometry combined with histological procedures, histochemical reactions and transmission electron microscopy to characterize the hemocytes from the oyster Crassostrea rhizophorae. Transmission electron microscopy revealed remarkable morphological characteristics, such as the presence of membranous cisternae in all mature cells, regardless of size and granulation. Some granular cells contained many cytoplasmic granules that communicated with each other through a network of channels, a feature never previously described for hemocytes. The positive reactions for esterase and acid phosphatase also indicated the presence of mature cells of all sizes and granule contents. Flow cytometry revealed a clear separation in complexity between agranular and granular populations, which could not be differentiated by size, with cells ranging from 2.5 to 25 µm. Based on this evidence we suggest that, at least in C. rhizophorae, the different subpopulations of hemocytes may in reality be different stages of one type of cell, which accumulates granules and loses complexity (with no reduction in size) as it degranulates in the event of an environmental challenge

Interferon-Gamma Promotes Infection of Astrocytes by <i>Trypanosoma cruzi</i>

<div><p>The inflammatory cytokine interferon-gamma (IFNγ) is crucial for immunity against intracellular pathogens such as the protozoan parasite <i>Trypanosoma cruzi</i>, the causative agent of Chagas disease (CD). IFNγ is a pleiotropic cytokine which regulates activation of immune and non-immune cells; however, the effect of IFNγ in the central nervous system (CNS) and astrocytes during CD is unknown. Here we show that parasite persists in the CNS of C3H/He mice chronically infected with the Colombian <i>T. cruzi</i> strain despite the increased expression of IFNγ mRNA. Furthermore, most of the <i>T. cruzi</i>-bearing cells were astrocytes located near IFNγ⁺ cells. Surprisingly, <i>in vitro</i> experiments revealed that pretreatment with IFNγ promoted the infection of astrocytes by <i>T. cruzi</i> increasing uptake and proliferation of intracellular forms, despite inducing increased production of nitric oxide (NO). Importantly, the effect of IFNγ on <i>T. cruzi</i> uptake and growth is completely blocked by the anti-tumor necrosis factor (TNF) antibody Infliximab and partially blocked by the inhibitor of nitric oxide synthesis L-NAME. These data support that IFNγ fuels astrocyte infection by <i>T. cruzi</i> and critically implicate IFNγ-stimulated <i>T. cruzi</i>-infected astrocytes as sources of TNF and NO, which may contribute to parasite persistence and CNS pathology in CD.</p></div