The research was carried out in the Private Natural Heritage Reserve RPPN PROMATA, in São Francisco de Paula, in Rio Grande do Sul, Brazil’s southernmost state, at roughly 900 metres above sea level. The team sampled five randomly chosen points in each habitat, all within 150 metres of one another and no more than four kilometres apart at the furthest distance, on a single day in November 2022. After carefully removing leaf litter, they collected two grams of topsoil from the upper five centimetres at each point, kept the samples on ice, and transported them to the laboratory within six hours to minimise shifts in community composition before DNA extraction.

The molecular work relied on high-throughput sequencing on an Illumina MiSeq platform, with separate primer sets targeting the 16S rRNA gene for bacteria and archaea and the 18S rRNA gene for eukaryotes. Raw sequences were quality filtered and denoised into Amplicon Sequence Variants, or ASVs, using the DADA2 pipeline, and taxonomy was assigned against the SILVA and PR2 reference databases with a confidence threshold of at least 80 percent at the family level. In total, the study identified 247 distinct eukaryotic ASVs, 4,201 bacterial ASVs spanning 30 phyla, and 130 archaeal ASVs, providing one of the most detailed multi-domain snapshots of soil life in this highly endemic region.

The taxonomic portraits of the two habitats diverged most clearly among the eukaryotes. In the forest soil, the phylum Cercozoa dominated the unicellular eukaryotic community, with the group Filosa-Sarcomonadea reaching 55.1 percent relative abundance, whereas in the grassland the parasitic phylum Apicomplexa took over, with the class Gregarinomorphea accounting for 83.9 percent of sequences. Among soil animals, annelid worms dominated most forest sites at 57.1 percent mean relative abundance, while nematodes were far more prevalent in grassland, representing 46.7 percent of identified sequences there compared with just 7.55 percent in the forest. Tardigrades appeared exclusively at one grassland site, and ciliates were entirely absent from grassland samples.

In contrast, the prokaryotic communities looked remarkably similar across the habitat boundary. Bacterial communities in both ecosystems were dominated by the phyla Pseudomonadota, Acidobacteriota and Actinomycetota, which together made up 79.08 percent of relative abundance, with the class Alphaproteobacteria reaching 29.7 percent in forest and 37.5 percent in grassland. Archaeal communities were overwhelmingly dominated by the phylum Thermoproteota, whose class Nitrososphaeria reached 85.1 percent in forest and 87.3 percent in grassland. Notably, the ratio between Alphaproteobacteria and Acidobacteriae, often used as an indicator of soil health, was about 1.40 in forest and 1.93 in grassland, values above one that point to healthy soils in both habitats, likely a benefit of long-term protection from disturbance.

Diversity statistics reinforced the forest’s edge. Using incidence-based rarefaction and Hill numbers, the researchers found that richness and both the Shannon and Simpson diversity indices were significantly higher in the Native Forest across all four organism groups, with non-overlapping 95 percent confidence intervals. The gap was widest for bacteria, whose estimated richness in forest soil was almost three times that of the grassland, the greatest disparity observed among the studied groups. Non-metric multidimensional scaling based on Jaccard distances revealed clear spatial separation between the habitats for unicellular eukaryotes, bacteria and archaea, and a significant, though graphically less tractable, separation for metazoans, confirming high beta diversity between the two systems.

One of the study’s most technically interesting contributions is its use of zeta diversity, a multi-site framework that tracks how many species are shared across increasing numbers of sampling sites, avoiding the bias toward rare species that can distort single beta diversity indices. Unicellular eukaryotes started with roughly 35 shared species and then declined steeply, indicating communities dominated by rare, site-specific taxa. Metazoans began with about 15 shared species and followed a similar abrupt decline, with an exponential regression providing the best fit and suggesting a core of specialised species. Bacteria, by contrast, started with around 650 shared species and declined only moderately in a long-tailed pattern best described by a power law, consistent with their habitat-generalist nature. Archaea began with about 17 shared species and showed steep early declines, hinting at communities of rare or highly specialised, possibly endemic organisms.

Co-occurrence network analysis among eukaryotes revealed two well-structured clusters: one linking rotifers, flatworms and Sagenista, and another connecting nematodes, the class Elardia, Coccidiomorphea and Cercozoa. The significant association between Coccidiomorphea, a group within Apicomplexa, and Nematoda is particularly intriguing, since both may infect overlapping categories of host organisms, although the authors caution that such patterns should be interpreted as exploratory and may reflect shared responses to local micro-environmental conditions rather than direct ecological interactions.

The findings carry broader implications for conservation. Primary forests are increasingly recognised as biodiversity reservoirs whose long-term genetic and functional value cannot be quickly replicated by recovering or secondary forests, and this study adds soil microbes and soil animals to that argument. At the same time, the grassland, often described as an underground or dwarf rainforest because of its massive belowground biomass and evolutionary antiquity, made a substantial contribution to regional diversity despite its comparatively lower within-habitat heterogeneity. The clear separation of communities between the two habitats points to environmental filtering driven by vegetation structure, soil physicochemical properties and microclimatic conditions as the prevailing mechanism shaping these assemblages.

The authors are careful to note the limits of their design. With five replicates per habitat and a single sampling date, the observed patterns should be read as indicative rather than exhaustive, and additional sampling could uncover further rare taxa and refine diversity estimates in such heterogeneous soils. Still, the work delivers a foundational characterisation of two adjacent, well-preserved habitats and demonstrates the value of pairing multi-domain metabarcoding with zeta diversity analysis to capture fine-scale community differentiation. Future studies with larger sample sizes, broader sequencing coverage and longitudinal data will be needed to determine whether these community shifts are driven by localised environmental filters, stochastic drift, or a combination of both, but the message is already clear: protecting a landscape means protecting not just its trees and grasses, but the intricate, largely unseen soil worlds beneath them.

Subject of Research: Soil microbial and metazoan community composition in adjacent preserved Atlantic Forest and Highland Grassland habitats assessed by eDNA metabarcoding

Article Title: Soil microbial community composition in two distinct preserved habitats

Article References: Huber, C. S. R., Huber, L. K., & Utz, L. R. P. (2026). Soil microbial community composition in two distinct preserved habitats. Metabarcoding and Metagenomics, 10, Article e183786. https://doi.org/10.3897/mbmg.10.183786

Image Credits: AI Generated

DOI: 10.3897/mbmg.10.183786

Keywords: soil biodiversity, eDNA metabarcoding, Atlantic Forest, Highland Grassland, microbiome, zeta diversity, bacteria, archaea, Cercozoa, Apicomplexa, Nematoda, conservation

The yellow mealworm, Tenebrio molitor, occupies a peculiar dual role in human affairs. It is a notorious pest that infests stored cereal products, yet it is simultaneously one of the most valuable and nutritionally rich insects used in both animal and human food industries. Researchers describe it as a beneficial pest precisely because of this contradiction. Beyond its agricultural significance, the species can degrade polystyrene and plastic waste, making it an object of interest for waste management research, and it serves as a model organism in laboratories worldwide.

Gregarines are parasitic protozoans belonging to the phylum Apicomplexa, a group that includes some of the most notorious parasites of invertebrates. These single-celled organisms colonize the digestive tracts of many different insect species, and their effects on hosts have proven remarkably complex. Numerous experimental studies have documented both detrimental and beneficial outcomes. On the harmful side, gregarines can arrest host development, cause mortality, and inhibit reproduction. On the beneficial side, some studies have shown they can increase host growth, fitness, and longevity, complicating any simple assessment of their ecological role.

The research team dissected forty larval and forty adult mealworms from each location, examining their intestines under microscopes at magnifications ranging from 10x to 1000x. When parasitic infections were observed, the scientists prepared Giemsa-stained smears to visualize the parasites in greater detail. The slides were air-dried, fixed with methanol, and stained with a freshly prepared 5 percent Giemsa solution before re-examination. The team then measured key anatomical structures of the parasites, including the length of the epimerite, deutomerite, and protomerite, along with total body length and various width ratios that help distinguish between gregarine species.

The results revealed a dramatic contrast between the two populations. In the Tbilisi samples, gregarine infection was present in both life stages, with several life stages of the parasite found in the midgut lumens of larvae and adults alike. Each trophozoite was elongate with a bud-type epimerite measuring 5.5 micrometers, and the largest trophozoite observed reached 67.9 micrometers in length. The sporadins found in the midgut lumen were solitary, with a clearly distinct septum separating the protomerite from the deutomerite, and the deutomerite was ovoid to ellipsoid with a spherical nucleus averaging 10.3 micrometers in diameter.

Infection rates differed sharply between life stages. Twenty-nine of the forty examined larvae were infected, representing a 72.5 percent infection rate, while only seven of the forty examined adults carried the parasite, corresponding to just 17.5 percent. This substantial difference suggests that mealworm larvae are considerably more susceptible to gregarine infection than adults, a finding that carries practical weight for insect farmers who rear larvae as the primary product for food and feed applications.

In stark contrast, the Antalya population from Türkiye showed no gregarine infection whatsoever, neither in larvae nor in adults. This complete absence of the parasite in one geographic population while another shows high infection rates raises intriguing questions about how gregarines spread within and between insect colonies, and what environmental or management factors might explain such divergent outcomes between facilities located in different countries.

To identify the parasites, the researchers compared their morphological measurements and ratios against a well-documented comparative study by Clopton and colleagues from 1991, which described the morphology and general appearance of gregarine species infecting T. molitor. According to a recent review of harmful and beneficial symbionts of the mealworm, four eugregarine species are known to infect this insect: Gregarina cuneata, G. polymorpha, G. niphandrodes, and G. steini, with G. cuneata considered the most impressive and complex among them. The parasites found in the Georgian population resembled both G. cuneata and G. niphandrodes in body morphology and general appearance, yet they differed from published descriptions in their morphological measurements and ratios.

This discrepancy becomes particularly interesting when considering host stage specificity. Clopton and colleagues reported that G. polymorpha and G. steini are found only in mealworm larvae, whereas G. niphandrodes was reported only from adults. Gregarina cuneata, by contrast, infects both larvae and adults. Because the Georgian parasites were detected in both life stages, the researchers suggest that more than one gregarine species might be infecting the Georgian population, or that multiple species could coexist within the same individual host. The possibility even remains that a previously unknown gregarine species could be identified from Georgian mealworm populations.

The findings arrive at a critical moment for the edible insect industry. High-density rearing conditions associated with mass production may increase the risk of disease and infections by parasitic symbionts, promoting their spread within insect colonies. These pathogens could potentially be passed to humans and animals consuming mealworms as food, making parasite surveillance a food safety concern as well as a production issue. The researchers call for further characterization of the gregarines in Georgian mealworm populations, including molecular phylogeny studies, and recommend documenting the presence and distribution of these parasites across other Georgian populations. Such work will be essential for understanding whether the parasites represent a threat to commercial production or simply another facet of the complex biology of an insect that humanity increasingly relies upon for sustainable protein.

Subject of Research: Gregarine parasite infection in yellow mealworm populations in Georgia and Türkiye

Article Title: Occurrence of gregarine parasites of Tenebrio molitor ( Coleoptera : Tenebrionidae ) in Georgia and Türkiye

Article References: Yaman, M., Kereselidze, M., & Güvendik, T. S. (2025). Occurrence of gregarine parasites of Tenebrio molitor (Coleoptera: Tenebrionidae) in Georgia and Türkiye. Caucasiana, 4, 119-124. https://doi.org/10.3897/caucasiana.4.e164996

Image Credits: AI Generated

DOI: 10.3897/caucasiana.4.e164996

Keywords: gregarine parasites, Tenebrio molitor, yellow mealworm, Apicomplexa, insect farming, edible insects, Georgia, Türkiye, parasitology, food safety, entomology, sustainable protein

The life cycle begins when a host ingests an oocyst, a resilient, wall-encased structure containing four sporozoites. In the gastrointestinal tract the oocyst excysts, releasing sporozoites that invade the epithelial cells lining the gut. Cryptosporidia are obligate intracellular parasites: they attach to the surface of a gut cell and pull the host membrane around themselves to form a parasitophorous vacuole, inside which they replicate. The sporozoite develops into a meront, duplicating its nucleus and organelles, and after roughly twelve hours divides into eight motile merozoites. These egress and invade neighbouring epithelial cells, repeating the asexual cycle. Remarkably, live-cell imaging of Cryptosporidium parvum infecting HCT-8 cells, together with single-cell RNA sequencing data from in vitro and in vivo infections, indicates that merozoites from the third round of asexual replication are committed directly to sexual development, apparently without a morphologically distinct type II meront of the kind described in other species such as C. muris and C. mortiferum.

Why and how the parasite commits to sexual stages after exactly three asexual rounds remains unknown. What is clear is that the committed merozoite invades a new epithelial cell and differentiates into either a macrogamont, which develops a single female gamete, or a microgamont, which produces sixteen male gametes. Mature males egress and seek out females, which never leave their host cell and instead mature intracellularly until fertilised. Fertilisation produces a zygote that undergoes meiosis, constructs the oocyst wall and completes sporogony to yield four haploid sporozoites within the protective shell. Because standard co-culture of C. parvum with HCT-8 cells does not support observation of fertilisation or subsequent zygote and oocyst development, sexual commitment, fertilisation, meiosis, sporogony and oocyst construction remain among the most mysterious phases of the life cycle.

Sexual commitment itself is mandatory for the third generation of merozoites and appears to be governed by an intrinsic, non-chromosomal mechanism. Live imaging has shown that merozoites from a single meront give rise to both sexes: of the eight merozoites in a meront, approximately two to three become male and the remaining five to six become female. This produces an excess of small, motile microgametes, presumably improving the odds that a female will be found and fertilised. One known player is the transcription factor Myb-M, expressed early in male development; experimental overexpression of Myb-M in C. parvum is sufficient to drive differentiation into males. No equivalent trigger for female commitment has been identified, leading the authors to suggest that female fate may be the default programme of sexual differentiation.

Female parasites, once committed, undertake one of the most demanding developmental programmes in the parasite’s repertoire. They carry a large nucleus, reflecting their status as the most transcriptionally and translationally active life cycle stage. Early in development, females express putative transporters, membrane proteins and secreted proteins hypothesised to mediate fertilisation, possibly facilitating male attachment and entry. Later, they mount two distinct waves of gene expression devoted to building the oocyst wall. The wall itself is a multilayered structure: an external glycocalyx, an outer layer of acid-fast lipids, and an inner layer of glycoproteins. These components are synthesised and stockpiled in female-exclusive organelles called Wall Forming Bodies, or WFBs, of which there are two subtypes distinguished by electron density. The prevailing hypothesis is that WFB Type I contents are secreted first to build the outer layer and WFB Type II contents follow to form the inner layer.

Only a handful of WFB components are confirmed. COWP1 is currently the only validated marker for WFB Type I, while COWP6 and COWP8 show punctate localisation in developing females, suggesting they also reside in these organelles. Transcriptomic and oocyst proteomic datasets have generated a comprehensive candidate list of WFB proteins, but how the organelles are synthesised, what exactly they contain, and how their secretion is regulated remain open questions. The single-cell atlas of C. parvum reveals that the first expression wave contains the Cryptosporidium Oocyst Wall Protein family and accessory enzymes such as proteases and protein modifiers for assembling the proteinaceous shell, while the second wave expresses glycosyltransferases and other secreted proteins including mucins and C-type lectins. These components are secreted into the space between the parasite plasma membrane and the host cell membrane, allowing the wall to be built while the parasite is still inside the gut cell. Having wall components ready just in time presumably keeps pace with the rapid turnover of epithelial host cells.

Females also provision their offspring. Genetic crosses using isolates tagged with different fluorescent proteins have shown that maternal cytoplasmic proteins persist in the sporozoites of first-generation progeny, including proteins involved in assembling dense granule organelles, which remodel the host cell after invasion. Females additionally express the proteins of the crystalloid body, a structure whose function is unknown but is likely involved in sporogony. Post-transcriptional control may be equally important: the maternal RNA-binding protein F-RBP, expressed in early females, binds female RNA transcripts, and conditional knockouts in vivo produce sterile sporozoites. F-RBP may therefore be essential for regulating maternally inherited RNA required to generate infectious sporozoites in the next generation.

Transcriptional regulation of female development is only beginning to be mapped. Two female-specific Apetala-2-type DNA-binding proteins have been identified: ApiAP2-1 is required for female differentiation, and ApiAP2-F is required for oocyst shedding. Insulinase-like protease 1 also contributes to female development and oocyst shedding. Intriguingly, females show reduced expression of ribosomal proteins, indicative of translational repression, and the authors hypothesise that epigenetic mechanisms such as chromatin remodelling may operate, consistent with the expansion of the female nucleus. Whatever the precise combination of mechanisms, they are fundamental to completing the life cycle and represent attractive targets for intervention.

What happens after fertilisation is even less understood. Fertilisation is presumed to trigger oocyst wall formation, sporogony and meiosis, but no culture system permits direct observation, so the signals regulating post-fertilisation development remain unknown across the coccidia. In Cryptosporidium, meiotic divisions appear to precede and partially overlap with deposition of the oocyst wall. Genes related to meiosis and DNA repair are expressed by females, including the DNA meiotic recombinase DMC1, detected in female nuclei. Under experimental conditions, C. parvum chromosomes show Mendelian inheritance with high frequencies of meiotic recombination, and frequent sex between genetically distinct parasites could accelerate the evolution and spread of traits such as virulence or drug resistance, as suggested by clofazimine resistance observed between C. parvum subtypes. Unlike other apicomplexans, Cryptosporidium oocysts are shed fully sporulated and immediately infectious, ready either to be excreted into the environment or to excyst in situ and reinfect the current host, a process that underpins the chronic disease seen in vulnerable patients.

A long-standing model proposes two oocyst types: thin-walled oocysts that excyst within the host to drive autoinfection, and thick-walled oocysts that are excreted and transmitted. The evidence for thin-walled oocysts rests on a few electron micrographs, and single-cell data show no transcriptional signature suggesting that some females are programmed to produce thin-walled oocysts. The authors instead hypothesise that all oocysts are thick-walled and environmentally robust, but that a small number excyst in the gut before excretion; given the exponential asexual cycle, even rare reinfections can sustain heavy parasite burdens. Improved culture systems, including organoid co-culture, are now making post-fertilisation biology experimentally accessible for the first time. Because oocyst biogenesis is exclusive to females and conserved across coccidia, dissecting this female-driven process could expose factors essential for transmission and open new avenues for drugs or vaccines. As the authors argue, it is time to stop overlooking Cryptosporidium females, the true engines of the parasite’s success.

Subject of Research: The biology of female sexual stages and oocyst formation in Cryptosporidium parasites

Article Title: Mysterious girls: The biology of female Cryptosporidium parasites

Article References: Stevens, S., Goddard, P., Walzer, K. A., & Pawlowic, M. C. (2026). Mysterious girls: The biology of female Cryptosporidium parasites. PLOS Pathogens, 22(10), e1014678. https://doi.org/10.1371/journal.ppat.1014678

Image Credits: AI Generated

DOI: 10.1371/journal.ppat.1014678

Keywords: Cryptosporidium, parasitology, oocyst, sexual development, gametogenesis, apicomplexa, Cryptosporidium parvum, transmission, diarrhoeal disease, Wall Forming Bodies, meiosis, global health

The research team, led by Huan Li and Lei Ma of Hebei Normal University together with Jing Huang of Hebei Children’s Hospital, assembled an unusually large sample set for this kind of survey: 2,448 human fecal specimens and 215 ticks of the species Haemaphysalis longicornis, an aggressive, wide-ranging biting tick common across East Asia. Using polymerase chain reaction assays designed to amplify the 18S rRNA gene, a standard molecular marker for identifying eukaryotic microorganisms, the researchers screened every sample for Colpodella DNA. The 18S rRNA gene is favored in studies of protist diversity because it is present in all eukaryotes, contains both conserved and variable regions, and allows newly recovered sequences to be compared against a deep archive of reference sequences from known organisms.

The results were striking in their consistency across the two very different sample types. Colpodella DNA was detected in 136 of the human fecal samples, a detection rate of 5.56 percent, and in 11 of the tick samples, a rate of 5.12 percent. The near-identical prevalence in feces and ticks is one of the more intriguing aspects of the study, hinting at some shared route of exposure or a common environmental reservoir linking the human and tick samples, though the design of the study cannot distinguish among the many possible explanations, including ingestion of contaminated food or water, environmental pass-through, or a genuine biological association with the tick vector.

Every PCR-positive sample yielded a high-quality 18S rRNA sequence when the amplified DNA was read by Sanger sequencing, a validation step that strengthens confidence in the specificity of the detection. Because some of the tick-derived sequences were identical to one another, the researchers collapsed the data into representative sequence types, ultimately selecting one tick-derived sequence and 21 distinct human-derived sequence types for deeper analysis. These 22 representative sequences were deposited in GenBank under accession numbers QB017305 through QB017326, making them available to other researchers for comparison and future surveillance work.

Phylogenetic analysis, the technique biologists use to place newly recovered gene sequences on the tree of life, showed that all 22 representative sequences from the study clustered within previously reported Colpodella lineages. In other words, the DNA found in human stool and ticks belongs to the same evolutionary neighborhood as Colpodella species that have been described from aquatic and terrestrial habitats around the world. The sequences did not branch off into entirely novel territory, which suggests that the organisms detected in Hebei are related to known forms rather than representing a radically divergent new lineage, although species-level identification in this group remains difficult because morphology and molecular data do not always align neatly.

To probe the genetic variation within their dataset, the team performed a haplotype analysis, a method that groups sequences by their shared sets of variable sites. Among the analyzed sequences, they identified three haplotypes, indicating a modest but real degree of genetic diversity among the Colpodella organisms whose DNA was recovered from the samples. Haplotype networks and diversity statistics are standard tools in population genetics, and applying them to environmental protist DNA can reveal whether a detection reflects a single widespread genotype or a mixture of related strains circulating in the environment. In this case, the limited number of haplotypes suggests relatively constrained diversity in the sampled region, though the authors caution that broader sampling would be needed to draw firmer conclusions.

Understanding why this matters requires a short detour into evolutionary biology. Colpodella species are predatory flagellates that feed by a process called myzocytosis, in which they attach to another microbe, puncture its membrane, and suck out its cytoplasmic contents. This dramatic feeding strategy has fascinated cell biologists in part because it may resemble the evolutionary steps that gave rise to intracellular parasitism. Colpodellids are considered the free-living sister group to the Apicomplexa, the phylum that includes Plasmodium, the malaria parasite, and Toxoplasma gondii. Studying Colpodella therefore offers a window into how some of humanity’s most burdensome parasites may have evolved from free-living ancestors that once hunted their prey from the outside.

The new study also fits into a rapidly expanding literature on where Colpodella DNA shows up outside of ponds and soil. Recent surveys have reported the organism’s genetic traces in goats, foxes, and birds; in Hyalomma ticks infesting camels in Egypt; in ticks feeding on ruminants in Italy; in horses; in zoo animals; and even in the urine of a human patient. Clinically, isolated case reports have described Colpodella-like parasites in a woman in China and in a patient with neurological symptoms whose infection was linked to a potential novel tick-borne Colpodella species. A 2025 scoping review of the genus’s biological characteristics and zoonotic potential underscored how much remains unknown about whether these detections represent transient environmental exposure, colonization, or true infection.

That uncertainty is central to how the Hebei findings should be interpreted, and the authors are explicit about the limits of their method. PCR-based detection confirms only that Colpodella DNA was present in the sample; it cannot determine whether live organisms were present, whether they were actively replicating, whether they colonized the human gut or the tick, or whether they caused any disease. DNA can persist after an organism has died, and environmental contamination or passive passage through the body can both produce positive signals. Distinguishing among these possibilities will require complementary approaches, including attempts to culture the organisms, sequencing of additional genetic markers beyond the 18S rRNA gene, and biological studies of how Colpodella interacts with animal hosts.

Nevertheless, the study marks a meaningful addition to surveillance data on an understudied group of protists. The large number of human samples screened, the near-matching detection rates in feces and ticks, the successful sequencing of every positive sample, and the public deposition of 22 representative sequences collectively provide a foundation for future work. As molecular surveys continue to sweep ever more environments and host species, Colpodella keeps appearing in unexpected places, and each new detection sharpens the questions that make this genus scientifically compelling: how these predatory relatives of apicomplexan parasites move between environmental reservoirs, animals, and occasionally people, and whether any of them pose a genuine health risk. For now, the answer in Hebei Province is a well-documented molecular presence and a clear agenda for the research that must follow.

Subject of Research: Molecular detection and genetic diversity of Colpodella protist DNA in human fecal and tick samples in China

Article Title: Molecular Detection and Genetic Characterization of Colpodella spp. DNA in Human Fecal and Tick Samples from Hebei Province, China

Article References: Li, H., Huang, J., Zhang, M., Zhang, Y., Chen, Y., & Ma, L. (2026). Molecular Detection and Genetic Characterization of Colpodella spp. DNA in Human Fecal and Tick Samples from Hebei Province, China. Acta Parasitologica, 71(5), Article 234. https://doi.org/10.1007/s11686-026-01419-6

Image Credits: AI Generated

DOI: 10.1007/s11686-026-01419-6

Keywords: Colpodella, protists, Apicomplexa, Haemaphysalis longicornis, ticks, 18S rRNA, PCR detection, genetic diversity, haplotypes, Hebei Province, zoonotic potential, myzocytosis

Caryospora species are single-celled parasites belonging to the phylum Apicomplexa, the same broad group that includes the malaria parasite and Toxoplasma. These organisms belong to the family Eimeriidae and are unusual among coccidia for their distinctive oocysts, the tough environmental transmission stages shed in host feces. Caryospora oocysts are monosporocystic and octozoic, meaning each contains a single sporocyst housing eight infectious sporozoites, and each sporocyst bears a Stieda body, a specialized plug used during excystation inside a new host. The genus is also remarkable for its life cycle: transmission to snakes occurs either directly through the fecal-oral route or indirectly when a snake preys upon a rodent harboring caryocysts, dormant parasite stages embedded in the prey’s tissues.

That second route involves one of the more striking examples of host manipulation in the parasite world. Caryospora species induce cyst formation in the oral musculature of rodents, impairing the animals’ feeding and agility and thereby making them easier prey for snakes. This contrasts sharply with their closest relatives, the raptor parasites now assigned to the genus Eumonospora, which form cysts in the skin and connective tissue of their rodent hosts, preserving mobility while keeping the prey visible to hunting birds of prey. The two strategies reflect deep evolutionary commitment to two entirely different predator-prey pathways, and they underscore why Caryospora species that parasitize snakes are considered to have a particularly robust taxonomic foundation.

Until recently, the genus had been poorly explored at the molecular level. Since the last comprehensive review, the number of Caryospora species recorded in snakes has grown from 18 to 56, yet only seven nominal species have been documented in vipers, the family that includes rattlesnakes, lanceheads, and true vipers. The new study focused on two of Brazil’s most important venomous snakes: the Jararaca, Bothrops jararaca, and the Cascabel Rattlesnake, Crotalus durissus. Researchers collected fecal samples from five individuals of each species that had been rescued from urban environments in the municipality of Miguel Pereira and temporarily housed at a municipal zoonosis control facility.

The results were striking. Oocysts were detected in 60 percent of the Jararacas and 80 percent of the rattlesnakes, an overall prevalence of 70 percent. Under the microscope, the oocysts were subspheroidal, measuring roughly 14 by 14 micrometres, with a bi-layered wall, no micropyle, and a polar granule present in only about half of the specimens. Each broadly ellipsoidal sporocyst carried a flattened, knob-like Stieda body and a rounded to trapezoidal sub-Stieda body. These features placed the parasites squarely within the morphotype of the genus’s type species, Caryospora simplex, first described from the Asp Viper in Europe more than a century ago. But morphology alone could not settle the identification, because the quantitative differences among related species proved unreliable.

To break the deadlock, the team turned to DNA. Individual oocysts, each photographed and documented before isolation, were subjected to repeated freeze-thaw cycles to rupture their walls, followed by proteinase K digestion and column-based DNA extraction. The researchers then amplified four non-overlapping genetic loci: three mitochondrial regions, designated MAVCOXI, MARI, and MACOIII, targeting cytochrome c oxidase subunits 1 and 3 and rDNA fragments, plus a nuclear locus spanning the 18S ribosomal RNA gene. Sequences from both host species were essentially identical to one another, differing at a single locus by just 0.4 percent, or three nucleotides out of 748. The concatenated four-locus dataset spanned 2,438 base pairs and was analyzed under a General Time Reversible model with gamma-distributed rate variation, using both Bayesian inference and maximum likelihood phylogenetics.

The molecular comparisons delivered a clear verdict. The Brazilian parasites matched most closely with Caryospora bigenetica, the only other Caryospora species from a viper to have been sequenced previously, isolated from the Massasauga rattlesnake in North America. Identity scores reached 100 percent at the nuclear 18S locus and between 97.7 and 99.3 percent at the mitochondrial loci, yielding 99 percent identity across the full concatenated dataset. In the phylogenetic trees, the two species formed a highly supported monophyletic clade, with posterior probability of 0.99 and bootstrap support of 100 percent. On this combined morphological and molecular evidence, the team confirmed the parasites as Caryospora jararacae, originally described from Jararacas in 1939, and reidentified earlier Brazilian records attributed to C. bigenetica as belonging to jararacae instead. This makes C. jararacae the sole Caryospora species known from Neotropical vipers and only the second genotypically characterized species from the entire viper family worldwide.

The broader taxonomic review that accompanied the sequencing led to the formal definition of the Caryospora simplex species complex, encompassing five nominal species that are morphologically indistinguishable: C. simplex from Eurasia, C. bigenetica from North America, C. jararacae from South America, C. maculatus from Africa, and C. bothriechis from Central America. Each occupies a different continent, effectively representing five geographic variants, or geotypes, of what appears to be a single young lineage. The researchers argue that these geotypes likely share a relatively recent common ancestor, and that their near-identical morphology and genetics reflect insufficient evolutionary time for structural divergence rather than ongoing genetic exchange across oceans. As a secondary hypothesis, they note that broad-ranging or migratory hosts, and even the international wildlife trade, could theoretically bridge geographic barriers, a possibility they flag for future population-level study.

The team deliberately retained all five names rather than collapsing them into a single species, because an integrative framework combining biogeography, host records, and molecular data allows each geotype to be delimited meaningfully. They also rejected a subspecies-based scheme, noting that such nomenclature is exceedingly rare within the Eimeriidae and would destabilize established names. Two additional viper parasites, C. matatu and C. saudiarabiensis, remain morphologically distinguishable from the complex and await molecular characterization. The researchers emphasize that sequencing C. simplex itself, ideally from its type host, will be essential to consolidate the emerging picture. For now, the study stands as a compelling demonstration that even among microscopic parasites, continents tell stories, and that some of evolution’s newest chapters are written in the feces of venomous snakes.

Subject of Research: Integrative taxonomy and first molecular characterization of the coccidian parasite Caryospora jararacae and the Caryospora simplex species complex in viperid snakes

Article Title: Integrative Taxonomy of the Caryospora simplex Species Complex (Apicomplexa: Eimeriidae) From Vipers (Squamata: Viperidae), With Redescription and First Molecular Characterization of Caryospora jararacae

Article References: da Silva, A. M. M., Ortúzar-Ferreira, C. N., Oliveira, M. D. S., Dorna-Santos, L., Ferreira, V. D. S., de Lima, V. M., & Berto, B. P. (2026). Integrative Taxonomy of the Caryospora simplex Species Complex (Apicomplexa: Eimeriidae) From Vipers (Squamata: Viperidae), With Redescription and First Molecular Characterization of Caryospora jararacae. Acta Parasitologica, 71(5), Article 229. https://doi.org/10.1007/s11686-026-01411-0

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DOI: 10.1007/s11686-026-01411-0

Keywords: Caryospora, Apicomplexa, Eimeriidae, vipers, Bothrops jararaca, Crotalus durissus, integrative taxonomy, molecular phylogeny, coccidia, species complex, host specificity, Neotropical region