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MBBGNR trichrome-stained smear of Colpodella sp. and B. caudatus cysts. aColpodella sp. cysts (black arrows) show irregularly shaped, blue-purple and white demilune pattern. Bodo caudatus cysts (black arrowheads) were detected as red-pink spherical cysts generally in clusters. Some B. caudatus cysts had a ring (halo) surrounding the cyst. Encysting Colpodella sp. after feeding is indicated by the open arrow. b MBBGNR trichrome-stained smear of Colpodella sp. and B. caudatus cysts. Mature Colpodella sp. cysts in four-way division were detected (black arrows). Cysts of B. caudatus were stained red-pink (black arrowheads). × 1000
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Colpodella species are free-living close relatives of apicomplexans that were recently reported to cause red blood cell infection in an immunocompromised human host and in a tick-borne human infection resulting in neurological symptoms. Unambiguous identification of the life cycle stages of Colpodella sp. using routine stains for light microscopy w...
Citations
... We developed Sam-Yellowe's trichrome staining protocols to identify life cycle stages of Colpodella sp. (ATCC 50594) [34]. The staining protocol was used to identify previously undocumented life cycle stages of Colpodella sp. ...
Colpodella species are free-living opportunistic pathogens that cause human and animal infections and use ticks and flies as vectors. Trophozoite and cyst stages of Colpodella species can be distinguished from stages of the prey Parabodo caudatus using Sam-Yellowe’s trichrome staining. Colpodella species obtain nutrients by attaching to their prey, aspirating the prey’s cytoplasmic contents into a posterior food vacuole and encysting. It is unclear whether both trophozoite and cyst stages are present during infection. Molecular techniques identified Colpodella species in all reported infections. However, no morphological information was reported to aid life cycle stage identification in hosts. This review discusses the increased incidence of Colpodella infection in animals and in vectors and the need to identify stages used for transmission and pathogenesis. The potential for zoonotic infections through tick and fly bites is a public health concern. We previously used Sam-Yellowe’s trichrome staining to identify life cycle stages of Colpodella sp. In order to gain a better understanding of transmission and pathogenesis, identifying Colpodella life cycle stages in infected tissue and in vectors will provide important insights regarding nutrient uptake in hosts by determining whether attachment of trophozoites to host cells occurs, and identifying trophozoites and cysts in infected hosts.
... Accordingly, transmission studies in H. dromedarii and epidemiological research in camels are required to close a gap in scientific knowledge of Colpodella spp. The identification of Colpodella spp. in such research may be aided by molecular biology techniques involving the use of the recently established trichrome staining method (Sam-Yellowe et al., 2020). ...
In Egypt, tick-borne diseases pose a significant threat to human and animal health, and the threat to dromedaries (Camelus dromedarius), the country's dominant camelid species, is of particular concern. These animals are frequently infested with ticks, and may thus develop tick-borne diseases or become reservoirs of tick-borne pathogens. However, there is a paucity of data on tick infestation in Egyptian camels, especially in the south of the country. Accordingly, we aimed to determine the prevalence of tick infestation in southern Egyptian camel populations (in Luxor and Aswan governorates), and identify the hemoprotozoan parasites carried by camel-infesting ticks. Camels were checked for ticks during veterinary examination at quarantine and household checks, and ticks were collected from infested camels for species identification using morphological examination and PCR analyses. Tick and hemoprotozoan species were identified using Basic Local Alignment Search Tool analysis with subsequent confirmation in phylogenetic analyses. All camel-infesting ticks belonged to the species Hyalomma dromedarii, and were clustered with ticks of this species previously found in Egypt in a phylogenetic tree based on the 16S rRNA gene. Molecular analysis targeting the 18S rRNA gene revealed the presence of hitherto undetected hemoprotozoan parasites, Colpodella spp., in 30/297 (10.1 %) camel-infesting ticks. In phylogenetic analysis, these Colpodella spp. were highly homologous (94-98.6 %) with Colpodella spp. previously deposited in GenBank with accession numbers OQ540590Q, MH208621, and GQ411073, which relate to Col-podella spp. previously detected from Haemaphysalis longicornis, Rhipicephalus haemaphysaloides, and humans in China. PCR analyses with spherical body protein-4 (SBP-4) gene-specific primers revealed Babesia bovis in 16/ 297 (5 %) of camel-infesting ticks, however, Babesia bigemina and Theileria annulata were not detected. Here, we report the first detection of Colpodella spp. in H. dromedarii in Egypt. Further epidemiological studies are needed to assess the risk to camels and humans, and the transmission dynamics. Based on the high tick infestation rates in Egyptian camels and the identification of previously unreported protozoan hemoparasites in ticks, we consider that the dromedary should be subject to surveillance as a sentinel species for tick-borne diseases in Egypt. Our findings underline the need for surveillance and collecting data on lesser known pathogens circulating in camel-infesting ticks, as part of a public health strategy for dealing with tick-borne diseases in Egypt.
... In previous studies we described the life cycle of Colpodella sp. ATCC 50594, investigated the process of myzocytosis and identified previously undocumented life cycle stages using Sam-Yellowe's trichrome staining protocols [23][24][25][26]. Sam-Yellowe's trichrome staining protocol was developed to identify and distinguish cyst stages of Colpodella sp. ...
... ATCC 50594 and its prey Parabodo caudatus. Both protists are biflagellated and form cysts of similar size, indistinguishable by Giemsa staining [25]. We identified the trophozoites of V. pontica and P. cosmopolitus in earlier studies using Giemsa staining, but were unable to differentiate the cyst stages [27]. ...
... ATCC 50594 since these stages can be obscured by bacteria or other debris in samples and in cultures. In previous staining studies, we obtained the dyes neutral red, brilliant green and methylene blue from the same vendors [23][24][25]. The vendor supplying neutral red closed operations resulting in the purchase of neutral red from a different vendor. ...
The colpodellids consist of free-living predatory protists phylogenetically related to pathogenic Apicomplexan parasites such as Plasmodium species, Cryptosporidium sp. and Toxoplasma gondii. Among the colpodellids, Colpodella species prey on bodonids, algae, and ciliates. Voromonas pontica preys on Percolomonas species. In the past decade, Colpodella species have been reported to cause opportunistic infections in humans and animals, and having the potential for zoonotic infections in humans due to the presence of Colpodella species in ticks. In the reported cases, transmission and pathogenic stages of Col-podella were not identified by light microscopy. In the colpodellid V. pontica, cyst stages of both the pred-ator and its prey Percolomonas cosmopolitus have not been identified by light microscopy. In this study we evaluated the dye components used for Sam-Yellowe’s trichrome staining, from different major vendors to determine the consistency and reproducibility of the staining protocol to aid the use of the staining proto-col for diagnosis. We show that although slight variations in the color of the stained cells are obtained, life cycle stages of Colpodella ATCC 50594 could be clearly identified, regardless of the vendor used. For the first time, Sam-Yellowe’s trichrome staining series, could identify cysts of V. pontica and its prey, demonstrating that the staining protocol can identify life cycle stages of other colpodellids. Sam-Yellowe's trichrome staining will aid molecular diagnosis of infections caused by colpodellids in human and ani-mals, identify colpodellids in ticks and flies and from environmental samples, when used for staining.
... The size of aspirated nutrients has not been reported for Colpodella species, and in our previous light microscopy and ultrastructural studies, particulate nutrients were not observed in the pre-cysts and cysts of Colpodella sp. (ATCC 50594) [23][24][25]. ...
... (ATCC 50594) and the prey's membrane in [24] and the current study. The sequential progression of myzocytosis involving attachment to the prey, the uptake of the prey's membrane into the cytoplasm of the predator and dissolution of the membrane to permit aspiration of the prey's cytoplasmic contents into the posterior food vacuole argue against phagocytosis and trogocytosis [24,25]. Moreover, the presence of a "myzocytotic apparatus" posterior to the tip of the pseudoconoid demonstrates that attachment is not followed by the piercing of the prey's plasma membrane by the pseudoconoid. ...
Colpodella sp. (ATCC 50594) is a free-living biflagellate predator closely related to pathogenic Apicomplexa such as Plasmodium, Cryptosporidium and Toxoplasma gondii. Colpodella sp. (ATCC 50594) obtain nutrients by preying on Parabodo caudatus using myzocytosis. The organization of the myzocytic apparatus and the mechanism of nutrient uptake into the posterior food vacuole of Colpodella species is unknown. In this study, we investigated myzocytosis using light and transmission electron microscopy. We investigated the uptake of 40 nm and 100 nm fluorescent nanoparticles and E. coli BioParticles by Colpodella sp. (ATCC 50594) in a diprotist culture. Transmission electron microscopy was used to investigate the morphology of the tubular tether formed during myzocytosis. E. coli BioParticles were taken up by P. caudatus but not by Colpodella sp. (ATCC 50594). Both protists took up the 100 nm and 40 nm beads, which were observed distributed in the cytoplasm of free unattached Colpodella sp. (ATCC 50594) trophozoites, and also in feeding Colpodella sp. (ATCC 50594) trophozoites and in the pre-cysts. Fragments of the nucleus and kinetoplast of P. caudatus and the nanoparticles were identified in the tubular tether being aspirated into the posterior food vacuole of Colpodella sp. (ATCC 50594). Unattached Colpodella sp. (ATCC 50594) endocytose nutrients from the culture medium independently from myzocytosis. The mechanisms of myzocytosis and endocytosis among Colpodella species may provide important insights into nutrient uptake among the pathogenic apicomplexans.
... Colpodella sp. (ATCC 50594) in diprotist cultures was fixed using 5% formalin as described [33]. Briefly, a viable diprotist culture was mixed with equal volumes of 10% formalin directly in the culture flask and incubated for ten minutes at room temperature [31]. ...
Free-living relatives of the Apicomplexa such as Colpodella species, Alphamonas species, and Voromonas pontica are predators that prey on ciliate, bodonid, and algal prey using the process of myzocytosis. During myzocytosis, the pseudoconoid is used to attach to the prey leading to aspiration of cytoplasmic contents of the prey into a posterior food vacuole formed in the predator, aided by secretions from the apical complex organelles. The conoid and associated proteins are conserved among the apicomplexa. However, the organization and function of the pseudoconoid during myzocytosis are not well understood. In this study, we investigated the morphology and ultrastructure of Colpodella sp. (ATCC 50594) during the stages of myzocytosis and cyst formation in the life cycle using light microscopy and transmission electron microscopy (TEM) in order to identify the organization of the tubular tether involved in nutrient aspiration by Colpodella sp. Tubular tethers of varying lengths were identified by light microscopy. We report that initial contact by Colpodella sp. trophozoites with Parabodo caudatus prey is by an area posterior to the apical tip of the rostrum that engulfs the membrane of the prey pulling it into the cytoplasm of the predator. The tubular tether that forms contains membranes of both predator and prey and is facilitated by microtubule organization and the cytoskeleton at the point of contact. Cytochalasin D treatment of diprotist cultures resulted in morphological distortions of trophozoites and the tubular tether suggesting a role of actin in the formation of the tubular tether. This mechanism of predation may provide insight into the mode of invasion observed in pathogenic apicomplexan zoites during host cell entry.
... However, it remains unclear whether the ticks are mechanical or biological transmitters, which requires further study. On the basis of molecular biology, recent modified light microscopy using a new trichrome staining technique by Sam-Yellowe et al. (2020) will aid in diagnosis of infections of Colpodella sp. and provide another available avenue for future surveillance and research. ...
Colpodella species, which mainly feed on protists and algae, are free-living close relatives of apicomplexans. Recent reports have identified Colpodella sp. infections in an immunocompromised individual and a suspected case of tick-transmitted infection resulting in neurological symptoms. Our molecular examination of piroplasmosis-infected horses in China identified nearly whole 18S rRNA gene sequences that are closely related to Colpodella sp. ATCC 50594 isolated from brown woodland soil at Gambrill State Park, located in Frederick, MD, shedding light on an underreported emerging zoonotic pathogen.
... (ATCC 50594) in resting cultures. Cells were collected every 24 h for eight days and formalin-fixed for staining with Sam-Yellowe's trichrome staining [18]. Days five, seven and eight cultures were fixed, stained and counted. ...
... Three time course experiments with four replicates of experiment one were performed. Cells were formalin-fixed and stained with Sam-Yellowe's trichrome [5,18] stains to view the cells. We used the first four time course replicates to demonstrate duration and reproducibility of the life cycle stages and timing of stage transitions in Colpodella sp. ...
... The appearance of the early cyst stage of Colpodella sp. (ATCC 50594) shows an irregular dual-colored, dark blue-purple and white colored cyst which we designated a demilune cyst [5,18]. The stage before the demilune cyst when the food vacuole and nucleus are still visible and the anterior end of the trophozoite is disintegrated, we have designated as the pre-cyst stage. ...
Colpodella species are free living bi-flagellated protists that prey on algae and bodonids in a process known as myzocytosis. Colpodella species are phylogenetically related to Apicomplexa. We investigated the life cycle of Colpodella sp. (ATCC 50594) to understand the timing, duration and the transition stages of Colpodella sp. (ATCC 50594). Sam-Yellowe’s trichrome stains for light microscopy, confocal and differential interference contrast (DIC) microscopy was performed to identify cell morphology and determine cross reactivity of Plasmodium species and Toxoplasma gondii specific antibodies against Colpodella sp. (ATCC 50594) proteins. The ultrastructure of Colpodella sp. (ATCC 50594) was investigated by transmission electron microscopy (TEM). The duration of Colpodella sp. (ATCC 50594) life cycle is thirty-six hours. Colpodella sp. (ATCC 50594) were most active between 20–28 h. Myzocytosis is initiated by attachment of the Colpodella sp. (ATCC 50594) pseudo-conoid to the cell surface of Parabodo caudatus, followed by an expansion of microtubules at the attachment site and aspiration of the prey’s cytoplasmic contents. A pre-cyst formed at the conclusion of feeding differentiates into a transient or resting cyst. Both DIC and TEM microscopy identified asynchronous and asymmetric mitosis in Colpodella sp. (ATCC 50594) cysts. Knowledge of the life cycle and stages of Colpodella sp. (ATCC 50594) will provide insights into the development of intracellular parasitism among the apicomplexa.
... Similarly, the process of excystation and egress of trophozoites is not understood. In a previous study, we developed new trichrome staining protocols that differentiated cysts and trophozoites of Colpodella sp. and its prey Bodo caudatus (Sam-Yellowe et al. 2019b). ...
... Supernatant was discarded and the pellets washed in 1× Dulbecco's phosphate buffered saline (dPBS) and used for preparing smears on glass slides as described (Yadavalli and Sam-Yellowe 2017;Sam-Yellowe and Yadavalli 2018). Formalin-fixed cells were stained with recently described trichrome stains (Sam-Yellowe et al. 2019b) (Table 1), Kinyoun's carbol fuchsin (KCF), and Giemsa stain followed by observations using light microscopy. All stained smears were examined under oil immersion at ×1000 magnification, and images were captured using an Olympus BX43 compound microscope attached to an Infinity HD Lumenera digital camera and Olympus U-TV0.35xc-2 ...
Colpodella spp. are free-living flagellates closely related to the apicomplexans. Human infections by Colpodella sp. have been reported. A biflagellated trophozoite and cyst stage comprise the known life cycle stages of Colpodella sp. However, the process of encystation and excystation within the life cycle is unclear. Life cycle stages initiating human infections are unknown. We performed a detailed investigation of the life cycle of Colpodella sp. (ATCC 50594) in culture using Sam-Yellowe’s trichrome stains and differential interference contrast (DIC) for light microscopy and fluorescence microscopy of Congo red-stained cells and investigated ultrastructure using transmission electron microscopy (TEM). We report previously undocumented stages of Colpodella sp. Asymmetric and asynchronous division was detected inside cysts by trichrome staining and by TEM. Odd-numbered juveniles and cysts containing more than four juvenile trophozoites were identified. Live imaging of active cultures captured the excystation and egress of juvenile trophozoites and confirmed the presence of multinucleate cysts. The ultrastructure of the multinucleate cyst is reminiscent of apicomplexan schizonts. Insights gained from the life cycle stages observed in culture allowed the construction of the life cycle of Colpodella sp. Knowledge of the life cycle will aid biochemical and molecular characterization of Colpodella sp. and help identify stages in human infections.
Buku ini merupakan bagian dari program bidang Mikrobiologi dan Parasitologi yang ditulis beberapa tenaga pengajar dan dikemas dalam satu topik. Penulis terdiri dari tenaga pengajar aktif di seluruh Indonesia, dan ahli pada bidang Mikrobiologi dan Parasitologi. Secara garis besar, cakupan materinya Buku ini meliputi: 1) Bentuk, Sifat dan Karakteristik Mikroorganisme. 2) Bakteriologi. 3) Virologi. 4) Konsep Pencegahan dan Pengendalian Infeksi (Sterilisasi dan Disinfeksi). 5) Nutrisi, Pertumbuhan, dan Keragaman Mikroorganisme. 6) Mikologi. 7) Pemeriksaan Mikrobiologi. 8) Konsep Pewarnaan dan Mikrobiologi. 9) Konsep Infeksi Nosokomial. 10) Entomologi. Secara umum, semua materi tersebut meliputi kajian dasar dari tema Mikrobiologi dan Parasitologi.
Buku ini merupakan bagian dari program bidang Mikrobiologi
dan Parasitologi yang ditulis beberapa tenaga pengajar dan dikemas
dalam satu topik. Penulis terdiri dari tenaga pengajar aktif di seluruh
Indonesia, dan ahli pada bidang Mikrobiologi dan Parasitologi.
Secara garis besar, cakupan materinya Buku ini meliputi: 1)
Bentuk, Sifat dan Karakteristik Mikroorganisme. 2) Bakteriologi. 3)
Virologi. 4) Konsep Pencegahan dan Pengendalian Infeksi (Sterilisasi
dan Disinfeksi). 5) Nutrisi, Pertumbuhan, dan Keragaman
Mikroorganisme. 6) Mikologi. 7) Pemeriksaan Mikrobiologi. 8) Konsep
Pewarnaan dan Mikrobiologi. 9) Konsep Infeksi Nosokomial. 10)
Entomologi. Secara umum, semua materi tersebut meliputi kajian
dasar dari tema Mikrobiologi dan Parasitologi.
