Journal: Microbiology Spectrum

Article Title: Prolonged survival of venereal Tritrichomonas foetus parasite in the gastrointestinal tract, bovine fecal extract, and water

doi: 10.1128/spectrum.00429-23

Figure Lengend Snippet: Orally administered T. foetus survive through the bovine gastrointestinal tract and are discharged by feces. ( A ) Timeline, experimental design, and sample collection to evaluate T. foetus survival through the bovine gastrointestinal tract. ( B ) A representative optical microscopy image of spherical structures (white arrows) present in fresh feces samples after 4 days of T. foetus K strain oral administration. ( C ) A representative image analyzed by epifluorescence microscopy of pseudocyst-like structures stained with WGA in fresh feces samples after 4 days of T. foetus oral administration.

Article Snippet: The parasites were centrifuged, resuspended in a 40 ug/mL propidium iodide solution, incubated for 30 min at 37°C and counted using an epifluorescence microscope (Zeiss Axio Observer 7 inverted fluorescence microscope).

Techniques: Microscopy, Epifluorescence Microscopy, Staining

Journal: Microbiology Spectrum

Article Title: Prolonged survival of venereal Tritrichomonas foetus parasite in the gastrointestinal tract, bovine fecal extract, and water

doi: 10.1128/spectrum.00429-23

Figure Lengend Snippet: Parasite growth in an extract from bovine feces. ( A ) Analysis of the T. foetus kinetic growth curve. Replication curves of parasites' growth in extracts of bovine feces were compared. Parasite counts were collected at the indicated times on the X -axis. Each point represents the average parasite concentration in 10 different tubes containing different feces samples. ( B ) Growth rates of parasites grown in standard culture media (TYM) were compared to parasites grown in an extract of bovine feces (BFE). Error bars represent standard error, and asterisks denote statistically significant differences determined by t-Student ( P < 0.05). ( C ) An example of a pseudocyst stained with Calcofluor white (CFW) in a T. foetus grown in bovine feces extracts for 10 days. The samples were analyzed by epifluorescence microscopy. ( D ) Quantification of the number of nuclei per parasite. Fifty parasites from each population were counted in triplicate in three independent experiments. The percentages of parasites with one nucleus (1N), two nuclei (2N), or more than two nuclei (>2N) are shown. T. foetus grown under standard culture media (TYM) and parasites grown in extracts from bovine feces (BFE) were compared. The results represent the average of three independent experiments and the standard error (SE). ( E ) Comparative images of T. foetus parasites with one nucleus grown in standard culture media (TYM) (left panel) and parasites grown in extracts from bovine feces (BFE) (right panel). Nuclei were stained with DAPI (blue). PC, phase-contrast.

Article Snippet: The parasites were centrifuged, resuspended in a 40 ug/mL propidium iodide solution, incubated for 30 min at 37°C and counted using an epifluorescence microscope (Zeiss Axio Observer 7 inverted fluorescence microscope).

Techniques: Concentration Assay, Staining, Epifluorescence Microscopy

Journal: Microbiology Spectrum

Article Title: Prolonged survival of venereal Tritrichomonas foetus parasite in the gastrointestinal tract, bovine fecal extract, and water

doi: 10.1128/spectrum.00429-23

Figure Lengend Snippet: T. foetus water survival analysis. ( A ) A representative image of the viability of T. foetus maintained in water was determined by propidium iodide (PI) staining. Parasites were stained with PI and photographed under a fluorescent microscope. Alive parasites are not stained, whereas dead cells are stained red. ( B ) Quantification of the viability of T. foetus when they were incubated in water for 24, 48, 72, 96, and 120 h. One hundred parasites were counted in triplicate in three independent experiments. Bars represent the means of three independent assays. Samples were analyzed with ANOVA test. Mean differences were calculated using Tukey’s test with an alpha = 0. 05. ( C ) Estimation of the size of the nuclei of T. foetus grown for 24, 48, and 72 h in water. TYM: parasites T. foetus grown in TYM culture media (control). The images of 50 nuclei staining with DAPI were analyzed using Image J’s threshold tool and the “Analyze Particles” function. Experiments were performed in triplicate and repeated three times. Mean differences were calculated using the Kruskal-Wallis test with a P value of 0.05. ( D ) Indirect immunofluorescence using an anti-alpha tubulin antibody shows spherical parasites with internalized flagella (white arrow). DAPI (blue). PC, phase-contrast image ( E ) Representative images of a pseudocyst or cyst-like structure stained with Calcofluor white (CFW) in a T. foetus grown in vitro for 72 h, maintained in water. The samples were analyzed by epifluorescence microscopy. ( F ) A graph representing the DNA content profile of T. foetus grown in standard culture media conditions for 24 h (TYM) and T. foetus incubated in water for 24 h (Water). DNA content was measured by flow cytometry. ( G ) Image representative of an indirect immunofluorescence image showing PCNA-positive nuclei (green) of T. foetus parasites that were grown in water for 24 h. DAPI (blue). PC, phase-contrast image.

Article Snippet: The parasites were centrifuged, resuspended in a 40 ug/mL propidium iodide solution, incubated for 30 min at 37°C and counted using an epifluorescence microscope (Zeiss Axio Observer 7 inverted fluorescence microscope).

Techniques: Staining, Microscopy, Incubation, Immunofluorescence, In Vitro, Epifluorescence Microscopy, Flow Cytometry

Journal: Microbiology Spectrum

Article Title: Prolonged survival of venereal Tritrichomonas foetus parasite in the gastrointestinal tract, bovine fecal extract, and water

doi: 10.1128/spectrum.00429-23

Figure Lengend Snippet: T. foetus pH survival analysis. ( A ) Simplified schematic diagram of the ruminant digestive tract and pH conditions in various regions. The pH of the digestive tract is variable in the different regions: pH 5.5 and 7 (esophagus-reticulum-rumen), pH 2–2.5 (abomasum), and pH 6–7 (large intestine). The digestive transit times are as follows: 24–48 h for the esophagus, reticulum, and rumen; 1–3 h for the abomasum; and 10–20 h for the large intestine. ( B ) The growth rates of parasites grown in culture medium at pH 5 and 7 were compared. Error bars represent standard error, and asterisks denote statistically significant differences determined by t -Student ( P < 0.05). ( C ) A representative image of a parasite stained with Calcofluor white (CFW) in a T. foetus grown in culture media at pH 2 for 3 h. The samples were analyzed by epifluorescence microscopy. ( D ) Quantification of viable T. foetus incubated for 3 h in TYM culture media at pH 2. One hundred parasites were counted in triplicate in three independent experiments. Bars represent the means of three independent assays. Samples show significant differences when analyzed with the Kruskal-Wallis test. LP (live parasites). DP (dead parasites). ( E ) A representative image of the viability of T. foetus maintained in water was determined by propidium iodide (PI) staining. Parasites were stained with PI and photographed under a fluorescent microscope. Alive parasites are not stained (white asterisks), whereas dead cells are stained red. ( F ) T. foetus kinetic growth curve depicting the percentage of motile trophozoites. T. foetus parasites were incubated in water for 24 h, then incubated for 3 h in culture media at pH 2, and then recovered in Diamond’s medium for 24, 48, and 72 h. Parasite counts were collected at the indicated times on the X -axis. Each data point on the graph represents the average percentage of motile parasites from three independent experiments. The error bars indicate the standard deviation.

Article Snippet: The parasites were centrifuged, resuspended in a 40 ug/mL propidium iodide solution, incubated for 30 min at 37°C and counted using an epifluorescence microscope (Zeiss Axio Observer 7 inverted fluorescence microscope).

Techniques: Staining, Epifluorescence Microscopy, Incubation, Microscopy, Standard Deviation

Journal: The Journal of Neuroscience

Article Title: Structure and Emergence of Specific Olfactory Glomeruli in the Mouse

doi: 10.1523/JNEUROSCI.21-24-09713.2001

Figure Lengend Snippet: Olfactory epithelium of OMP-GFP mice. A, Medial whole-mount view of a half-head of an adult mouse. Olfactory epithelium (left) and olfactory bulb (right) are intensely fluorescent. Images were photographed using an epifluorescence stereomicroscope (Leitz MZ12; Leitz, Stuttgart, Germany). Image width, 6 mm. B, Close-up whole-mount view of the olfactory epithelium. Green dots represent OSNs; dark areas correspond to the non-GFP-expressing supporting cells. Images were photographed using a Leitz MZ12 stereomicroscope. Image width, ∼1 mm. C, Low-power view of a histological section through the nose. Green fluorescent OSNs line the convoluted surface of the turbinates. Their axons are assembled in bundles underneath the epithelium. Images were photographed using an epifluorescence wide-field microscope (Zeiss Axioplan 2). Image width, ∼1.4 mm. D, Medium-power view of a histological section through the nose. The horizontal white lines indicate the approximate levels of the optical sections shown in E-H. Images were photographed using a confocal laser scanning microscope (Zeiss LSM 510). Image width, ∼256 μm. E, Optical section through OSN dendrites produced by TPLSM from a 4-d-old mouse. Individual dendritic knobs are visible. Image width, ∼256 μm. F, Optical section at the level of the cell bodies of OSNs produced by TPLSM. Image width, ∼256 μm. G, Optical section at the level of the axons of the cell bodies produced by TPLSM. Single axons are clearly visible, even when imaged through the brightly labeled cell body layer, which saturated the detector. Image width, ∼256 μm. H, Optical section at a level below the epithelium produced by TPLSM. OSN axons coalesce to form ribbon-like fascicles. Image width, ∼256 μm.

Article Snippet: B , Close-up whole-mount view of glomeruli in the olfactory bulb, photographed with an epifluorescence stereomicroscope (Zeiss Stemi SV11).

Techniques: Expressing, Microscopy, Laser-Scanning Microscopy, Produced, Labeling

Journal: The Journal of Neuroscience

Article Title: Structure and Emergence of Specific Olfactory Glomeruli in the Mouse

doi: 10.1523/JNEUROSCI.21-24-09713.2001

Figure Lengend Snippet: Olfactory bulb of OMP-GFP mice. A, Whole-mount view of the dorsal surface of both olfactory bulbs. Top is posterior; bottom is anterior. The outer nerve layer and the glomerular layer produce intense fluorescence that is easily detectable with an epifluorescence stereomicroscope (Zeiss Stemi SV11). Image width, 5 mm. B, Close-up whole-mount view of glomeruli in the olfactory bulb, photographed with an epifluorescence stereomicroscope (Zeiss Stemi SV11). Image width, ∼1.2 mm. C, Optical section at the level of the glomerular layer (right) and the outer nerve layer (left) produced by TPLSM. Glomeruli are discrete globose structures covered with a thick mat of fibers running across the surface of the bulb without obvious direction or stereotyped organization. Image width, ∼256 μm. D, Optical section at the level of the glomeruli, below the outer nerve layer, produced by TPLSM. The nonfluorescent areas inside the glomeruli presumably correspond to the dendrites of interneurons and second-order neurons and to glia and blood vessels, which do not express OMP and thus do not express GFP in these mice. Image width, ∼256 μm.

Article Snippet: B , Close-up whole-mount view of glomeruli in the olfactory bulb, photographed with an epifluorescence stereomicroscope (Zeiss Stemi SV11).

Techniques: Fluorescence, Produced

Journal: The Journal of Neuroscience

Article Title: Structure and Emergence of Specific Olfactory Glomeruli in the Mouse

doi: 10.1523/JNEUROSCI.21-24-09713.2001

Figure Lengend Snippet: M72 glomeruli in M72-IRES-tauGFP mice. A, Whole-mount view of the dorsal surface of the olfactory bulbs of a mature M72-IRES-tauGFP mouse. Orientation is identical to that in Figure 3A. A pair of medial and lateral green fluorescent glomeruli can be discerned within each bulb. This study concentrated on the lateral M72 glomeruli because they reside in a flattened region of the dorsal surface of the olfactory bulb and are readily accessible for imaging. The dark lines are blood vessels within the meningi. Images were photographed using an epifluorescence stereomicroscope (Zeiss Stemi SV11). Image width, 5 mm. B, Schematic representation of image shown in A. P, Posterior; M, medial; A, anterior; L, lateral, R, right; L, left. The interbulbar symmetry of the positions of the M72 glomeruli is apparent. The intrabulbar symmetry of the positions of the M72 glomeruli is along a plane that intersects with the midline at an ∼30° angle, such that the lateral glomeruli are more anterior and more dorsal than the medial glomeruli. C, Stereo pair of three-dimensional TPLSM reconstruction of the right lateral glomerulus of a PD18 M72-IRES-tauGFP mouse (strain T15/loxP). A few major fascicles terminate in the glomerulus. Image width, ∼256 μm. D, Stereo pair of the left lateral glomerulus of the mouse shown in C. There is no bilateral symmetry; the pattern of fascicles is different from that in C. Image width, ∼256 μm. E, Stereo pair of a lateral glomerulus from a PD18 M72-IRES-tauGFP mouse, a littermate of the mouse shown in C and D. Multiple small fascicles converge onto the glomerulus from widely varying angles. Image width, ∼256 μm.

Article Snippet: B , Close-up whole-mount view of glomeruli in the olfactory bulb, photographed with an epifluorescence stereomicroscope (Zeiss Stemi SV11).

Techniques: Imaging

Journal: The Journal of Biological Chemistry

Article Title: Inhibition of Nucleotide Sugar Transport in Trypanosoma brucei Alters Surface Glycosylation *

doi: 10.1074/jbc.M113.453597

Figure Lengend Snippet: TbNST4 is localized at the Golgi apparatus. A, genomic PCR analysis of in situ C-terminal HA-tagged tbnst4 in T. brucei BSF. Genomic DNA was isolated from untagged (lane 1) and tagged (lane 2) TbNST4 cells. In situ tagging locus was PCR amplified with a forward primer complementary to TbNST4 coding sequence and a reverse primer complementary to the 3′ UTR (outside the recombination sequence). The expected genomic PCR products are indicated. B, Western blot analysis of the in situ tagged TbNST4HA protein expression. Cell lysates from untagged (lane 1) and tagged TbNST4HA (lane 2) cells were subjected to SDS-PAGE and Western blotting with anti-HA antibody detection. The position of the expressed TbNST4 is indicated. C, immunofluorescence analysis of TbNST4HA localization. BSF cells carrying the tagged TbNST4 were stained with anti-HA for TbNST4 (green), anti-Ty for TbGT15 (red, upper panel), or anti-p67 (red, lower panel) antibodies. Cell images were captured with Nikon wide-field epifluorescence microscope. Blue, DAPI staining for nuclei and kinetoplasts.

Article Snippet: Cell images were captured with a Nikon wide-field epifluorescence microscope.

Techniques: In Situ, Isolation, Amplification, Sequencing, Western Blot, Expressing, SDS Page, Immunofluorescence, Staining, Microscopy