Cannibalism and competition can increase parasite abundance for parasites with complex life history strategies.

Ecological interactions among hosts are critical to consider when predicting disease dynamics. Most theory predicts that intraguild predation (IGP) and cannibalism negatively impact parasite populations, but this is based primarily on assumptions of simple or single-host life cycles. Here we investigate the effects of cannibalism in a size-structured host population on two digenean trematodes that have complex, multihost life cycles. A high incidence of cannibalism among paratenic hosts produced higher parasite infection loads and abundance, whereas cannibalism among obligate hosts reduced parasite abundances. We attributed this difference to trophic transmission aggregating parasites in larger, potentially fitter hosts and also to transmission among paratenic hosts via cannibalism. Moreover, we found evidence of indirect competitive interactions between parasites that can also increase infections at small scales. Our results show there are multiple mechanisms through which high cannibalism environments can benefit parasites that use paratenic hosts and trophic transfer to complete their life cycles.

Histomorphological Evidence of Complex Cannibalism-An Uncharted Territory in Oral Squamous Cell Carcinoma.

Complex cannibalism (CxC) is an emerging new seeming entity in oral squamous cell carcinoma (OSCC) embody as a prime metabolic event in determining the aggressive potential. Owing to paucity in literature regarding it, the aim of the present study to deciphering the occurrence of CxC in OSCC. Further, the expression of cluster of differentiation 68 (CD68) was studied in tumor cells and correlated with CxC to ascertain the biological behaviour of OSCC. 30 Hematoxylin and Eosin stained sections of various grades of OSCC were scanned for CxC and correlated with clinicopathological parameters. Immunohistochemical analysis using CD68 was performed. While observing CxC in OSCC, statistically significant in age (p-0.048) and histological grades (p-0.004). CD68 expression in tumor cells was statistically significant in histopathological grades (p < 0.001) whereas on correlating with CxC (p - 0.171) was non-significant. The analogous rise in CxC and CD68 with increasing histopathological grades could aid in recognising CxC as a precise histopathological parameter to assess the aggressive biological potential in OSCC.

Cannibalism facilitated by parasite infection induces dispersal in a semi-aquatic insect.

Parasites are known to have direct effects on host dispersal ability and motivation. Yet, parasites have a variety of impacts on host populations, including shaping predation and cannibalism rates, and therefore may also have indirect effects on host dispersal; these indirect pathways have not been studied. We tested the hypothesis that parasites influence host dispersal through effects on cannibalism using backswimmers (Notonecta undulata) and Hydrachnidia freshwater mites. Mite parasitism impedes swimming in backswimmers, which we found increased their vulnerability to cannibalism. We imposed a manipulation that varied cannibalism rates across experimental populations consisting of a mix of backswimmers with and without simulated parasites. Using simulated parasites allowed us to examine the effects of cannibalism without introducing infection risk. We found that the odds of dispersal for infected backswimmers increased by 2.25× with every 10% increase in the risk of being cannibalized, and the odds of dispersal for healthy backswimmers increased by 2.34× for every additional infected backswimmer they consumed. Our results suggest that cannibalism was used as an energy source for dispersal for healthy individuals, while the risk of being eaten motivated dispersal in infected individuals. These results elucidate the complex ways that parasites impact host populations and strengthen our understanding of host-parasite interactions, including host and parasite population stability and spread. This article is part of the theme issue 'Diversity-dependence of dispersal: interspecific interactions determine spatial dynamics'.

Impact of pre-copulatory sexual cannibalism on genetic diversity and efficacy of selection.

Factors that increase reproductive variance among individuals act to reduce effective population size (Ne), which accelerates the loss of genetic diversity and decreases the efficacy of purifying selection. These factors include sexual cannibalism, offspring investment and mating system. Pre-copulatory sexual cannibalism, where the female consumes the male prior to mating, exacerbates this effect. We performed comparative transcriptomics in two spider species, the cannibalistic Trechaleoides biocellata and the non-cannibalistic T. keyserlingi, to generate genomic evidence to support these predictions. First, we estimated heterozygosity and found that genetic diversity is relatively lower in the cannibalistic species. Second, we calculated dN/dS ratios as a measure of purifying selection; a higher dN/dS ratio indicated relaxed purifying selection in the cannibalistic species. These results are consistent with the hypothesis that sexual cannibalism impacts operational sex ratio and demographic processes, which interact with evolutionary forces to shape the genetic structure of populations. However, other factors such as the mating system and life-history traits contribute to shaping Ne. Comparative analyses across multiple contrasting species pairs would be required to disentangle these effects. Our study highlights that extreme behaviours such as pre-copulatory cannibalism may have profound eco-evolutionary effects.

Anthropogenic noise disrupts early-life development in a fish with paternal care.

Anthropogenic noise is a global pollutant but its potential impacts on early life-stages in fishes are largely unknown. Here, using controlled laboratory experiments, we tested for impacts of continuous or intermittent exposure to low-frequency broadband noise on early life-stages of the common goby (Pomatoschistus microps), a marine fish with exclusive paternal care. Neither continuous nor intermittent noise exposure had an effect on filial cannibalism, showing that males were capable and willing to care for their broods. However, broods reared in continuous noise covered a smaller area and contained fewer eggs than control broods. Moreover, although developmental rate was the same in all treatments, larvae reared by males in continuous noise had, on average, a smaller yolk sac at hatching than those reared in the intermittent noise and control treatments, while larvae body length did not differ. Thus, it appears that the increased consumption of the yolk sac reserve was not utilised for increased growth. This suggests that exposure to noise in early life-stages affects fitness-related traits of surviving offspring, given the crucial importance of the yolk sac reserve during the early life of pelagic larvae. More broadly, our findings highlight the wide-ranging impacts of anthropogenic noise on aquatic wildlife living in an increasingly noisy world.

Control Potential of Multiple Nucleopolyhedrovirus (SfMNPV) Isolated from Fall Armyworm in Nigeria (West Africa).

The fall armyworm (FAW) Spodoptera frugiperda (Lepidoptera, Noctuidae) has now become an invasive pest of global concern. The pest was first detected in Central and Western Africa in early 2016. Sustainable management options explored by stakeholders during early FAW invasion in Africa included the use of biopesticides and biological control. The current study aimed to compare the susceptibility of FAW larvae to SfMNPV with the assumption that the virus isolated from FAW populations in Africa has higher virulence compared with an isolate from Argentina (SfMNPV-ARG). We also hypothesized that host plant plays a role in SfMNPV efficacy and that cannibalism mediates horizontal and vertical transmission of the virus. This work provides pioneering data on the virulence of the new SfMNPV isolate from Nigeria (SfMNPV-KA1), which proved more effective than its exotic counterpart from Argentina (SfMNPV-ARG). The host plant effect made a significant difference between maize and onion with more FAW death in the larvae fed with contaminated onion 5 days post treatment. The study demonstrates and discusses the effect of cannibalism on virus transmission.

Molecular, behavioral, and growth responses of juvenile yellow catfish (Tachysurus fulvidraco) exposed to carbamazepine.

Carbamazepine (CBZ) is an anticonvulsant medication used to treat epilepsy and bipolar disorder. Due to its persistence and low removal rate in wastewater treatment plants, it is frequently detected in the environment, raising concerns regarding its potential adverse effects on aquatic organisms and ecosystems. In this study, we aimed to assess the impact of CBZ on the behavior and growth of juvenile yellow catfish Tachysurus fulvidraco, a native and economically important species in China. Fish were exposed to CBZ at three concentrations of 1, 10, or 100 µg/L for 14 days. The fish exposed to 10 and 100 µg/L of CBZ exhibited decreased feeding, and a significant increase in cannibalistic tendencies was observed in fish exposed to 100 µg/L CBZ. Acetylcholinesterase activity was increased in the brain of fish exposed to 100 µg/L CBZ. CBZ also inhibited the growth of yellow catfish. To better elucidate mechanisms of toxicity, transcriptomics was conducted in both the brain and liver. In the brain, gene networks associated with neurotransmitter dysfunction were altered by CBZ, as well as networks associated with mitochondrial dysfunction and metabolism. In the liver, gene networks associated with the immune system were altered by CBZ. The current study improves comprehension of the sub-lethal effects of CBZ and reveals novel insight into molecular and biochemical pathways disrupted by CBZ, identifying putative key events associated with reduced growth and altered behavior. This study emphasizes the necessity for improved comprehension of the effects of pharmaceutical contaminants on fish at environmentally relevant levels.

Filial cannibalism of Nabis pseudoferus is not evolutionarily optimal foraging strategy.

Using a recursion model with real parameters of Nabis pseudoferus, we show that its filial cannibalism is an optimal foraging strategy for life reproductive success, but it is not an evolutionarily optimal foraging strategy, since it cannot maximize the descendant's number at the end of the reproductive season. Cannibalism is evolutionarily rational, when the number of newborn offspring produced from the cannibalized offspring can compensate the following two effects: (a) The cannibalistic lineage wastes time, since the individuals hatched from eggs produced by cannibalism start to reproduce later. (b) Cannibalism eliminates not only one offspring, but also all potential descendants from the cannibalized offspring during the rest of reproductive season. In our laboratory trials, from conspecific prey Nabis pseudoferus did not produce newborn nymphs enough to compensate the above two effects.

Extra Nestlings That Are Condemned to Die Increase Reproductive Success in Hoopoes.

AbstractThe adaptive value of routinely laying more eggs than can be successfully fledged has intrigued evolutionary biologists for decades. Extra eggs could, for instance, be adaptive as insurance against hatching failures. Moreover, because recent literature demonstrates that sibling cannibalism is frequent in the Eurasian hoopoe (Upupa epops), producing extra offspring that may be cannibalized by older siblings might also be adaptive in birds. Here, directed to explore this possibility in hoopoes, we performed a food supplementation experiment during the laying period and a clutch size manipulation during the hatching stage. We found that females with the food supplement laid on average one more egg than control females and that the addition of a close-to-hatch egg at the end of the hatching period increased the intensity of sibling cannibalism and enhanced fledging success in hoopoe nests. Because none of the extra nestlings from the experimental extra eggs survived until fledging, these results strongly suggest that hoopoes obtain fitness advantages by using temporarily abundant resources to produce additional nestlings that will be cannibalized. These results therefore suppose the first experimental demonstration of the nutritive adaptive function of laying extra eggs in vertebrates with parental care.

Robert Hertz, Anthropophagic Practices and Traditional South Fore Mortuary Rites in Papua New Guinea.

In this paper we examine the traditional mortuary rites of the South Fore people of Papua New Guinea using Robert Hertz's theory of secondary burial and the three mechanisms of mourning identified by Daniel Lagache. The ethnographic data that we obtained on South Fore interpretations of their own mortuary rites showed that all forms of corpse handling achieved the same end results through the process of secondary burial. Furthermore, the three mechanisms of mourning applied equally to all forms of corpse disposal and we found no evidence to support psychosexual interpretations of mortuary anthropophagy which emphasise aggression. South Fore interpretations of mortuary anthropophagy show that the dead were eaten out of love, and to protect the mourners from the painful emotion of witnessing the decomposition of the corpse. These findings affirm the relevance of the concepts of Hertz and Lagache to the universal human experiences of death and mourning.

Brobdingnagians and Goliaths: two forms of gigantism in fish.

Two forms of gigantism are differentiated in fish, Brobdingnagian and Goliathan gigantism, the former applying to populations whose individuals are all larger than is typical for the taxon, the latter to single individuals within a population. While Brobdingnagian gigantism is largely explained by various ecological and evolutionary rules, Goliathan gigantism is not. A mechanistic hypothesis is proposed which explains Goliathan gigantism in terms of the reduction of oxygen requirements of individual fish via moving to cooler temperatures and/or acquiring larger, more energy-dense prey, which enable them to get bigger, and, in the process, sometimes generate bimodal size distributions that may qualify as gradual forms between Goliathan and Brobdingnagian gigantism. This mechanism, which relies on the manner in which their gill surface area grows, is more likely to operate in fish that can get big in the first place than in fish that remain small.

Examining the relationship between different naturally-occurring maxillary beak shapes and their ability to cause damage in commercial laying hens.

1. Using chicken models to avoid unnecessary harm, this study examined the relationship between naturally-occurring maxillary (top) beak shapes and their ability to cause pecking damage.2. A selection of 24 Lohmann Brown laying hens from a total population of 100 were sorted into two groups based on their maxillary beak shape, where 12 were classified as having sharp beaks (SB) and 12 as having blunt beaks (BB).3. All hens were recorded six times in a test pen which contained a chicken model (foam block covered with feathered chicken skin) and a video camera. During each test session, the number of feathers removed from the model, the change in skin and block weight (proxies for tissue damage) and the percentage of successful pecks (resulting in feather and/or tissue removal) were recorded.4. SB hens removed more feathers from the model and had a greater change in skin weight than BB hens. The mean number of pecks made at the model did not differ between the beak shape groups; however, SB hens had a greater percentage of successful pecks, resulting in feather and/or tissue removal, compared to BB hens.5. In conclusion, SB hens were more capable of removing feathers and causing damage. Birds performed more successful pecks resulting in feather and/or tissue removal as they gained experience pecking at the model.

Non-mitotic proliferation of malignant cancer cells revealed through live-cell imaging of primary and cell-line cultures.

INTRODUCTION: Anti-mitosis has been a key strategy of anti-cancer therapies, targeting at a fundamental property of cancer cells, their non-controllable proliferation due to overactive mitotic divisions. For improved anti-cancer therapies, it is important to find out whether cancer cells can proliferate independent of mitosis and become resistant to anti-mitotic agents. RESULTS: In this study, live-cell imaging was applied to both primary-cultures of tumor cells, and immortalized cancer cell lines, to detect aberrant proliferations. Cells isolated from various malignant tumors, such as Grade-III hemangiopericytoma, atypical meningioma, and metastatic brain tumor exhibit distinct cellular behaviors, including amoeboid sequestration, tailing, tunneling, nucleic DNA leakage, as well as prokaryote-like division such as binary fission and budding-shedding, which are collectively referred to and reported as 'non-mitotic proliferation' in this study. In contrast, benign tumors including Grade-I hemangiopericytoma and meningioma were not obvious in such behaviors. Moreover, when cultured in medium free of any anti-cancer drugs, cells from a recurrent Grade-III hemangiopericytoma that had been subjected to pre-operation adjuvant chemotherapy gradually shifted from non-mitotic proliferation to abnormal mitosis in the form of daughter number variation (DNV) and endomitosis, and eventually regular mitosis. Similarly, when treated with the anti-cancer drugs Epirubicin or Cisplatin, the cancer cell lines HeLa and A549 showed a shift from regular mitosis to abnormal mitosis, and further to non-mitosis as the dominant mode of proliferation with increasing drug concentrations. Upon removal of the drugs, the cells reversed back to regular mitosis with only minor occurrences of abnormal mitosis, accompanied by increased expression of the stem cell markers ALDH1, Sox, Oct4 and Nanog. CONCLUSIONS: The present study revealed that various types of malignant, but not benign, cancer cells exhibited cellular behaviors indicative of non-mitotic proliferation such as binary fission, which was typical of prokaryotic cell division, suggesting cell level atavism. Moreover, reversible transitions through the three modes of proliferation, i.e., mitosis, abnormal mitosis and non-mitosis, were observed when anticancer drug concentrations were grossly increased inducing non-mitosis or decreased favoring mitosis. Potential clinical significance of non-mitotic proliferation in cancer drug resistance and recurrence, and its relationship with cancer stem cells are worthy of further studies.

Bibliometric analysis on cannibalism/infanticide and maternal aggression towards pups in laboratory rodents.

Animal welfare has evolved during the past decades to improve not only the quality of life of laboratory rodents but also the quality and reproducibility of scientific investigations. Bibliometric analysis has become an important tool to complete the current knowledge with academic databases. Our objective was to investigate whether scientific research on cannibalism/infanticide is connected with maternal aggression towards the offspring in laboratory rodents. To carry out our research, we performed a specific search for published articles on each concept. Results were analyzed in the open-source environment RStudio with the package Bibliometrix. We obtained 253 and 134 articles for the first search (cannibalism/infanticide) and the second search (maternal aggression towards the pups) respectively. We observed that the interest in infanticide/cannibalism started in the 1950s, while researchers started showing interest in maternal aggression towards the pups 30 years later. Our analyses indicated that maternal aggression had better citations in scientific literature. In addition, although our results showed some common features (e.g. oxytocin or medial preoptic area in the brain), we observed a gap between cannibalism/infanticide and maternal aggression towards the pups with only 14 published articles in common for both the searches. Therefore, we recommend researchers to combine both concepts in further investigations in the context of cannibalism for better dissemination and higher impact in laboratory rodents' welfare research.

m-Aminophenylacetylene induces maternal care in a predatory spider.

Maternal care is critically important for the survival of offspring in various animals. Spiders in the family Lycosidae are known for their hunting ability and maternal care behaviors. Predation on newly hatched spiderlings (pulli) by mother spiders decreases when they come into contact, and they carry the pulli on their dorsal surface. However, the factors inducing maternal care in lycosid spiders have not been elucidated. In this study, we investigated maternal care in Pardosa pseudoannulata (Araneae, Lycosidae) females. We proposed that the physical interaction between pulli and mother spiders induces maternal care via m-aminophenylacetylene (m-A), a novel regulator of maternal care. The presence of pulli on the dorsal abdomen of non-mother spiders suppressed pulli predation and increased the pulli-carrying rate, and the absence of pulli on the mother spiders increased pulli predation and decreased the pulli-carrying rate. The compound m-A was abundant in mother spiders, and it could be induced in non-mother spiders when they carried pulli. The topical application of m-A to non-mother spiders and m-A injection decreased pulli predation and increased the pulli-carrying rate, respectively; these findings indicate that m-A in both internal tissues and the integument is required for the induction of maternal care behavior, and the interaction between pulli and females induces the production of m-A. In-depth study of the regulatory mechanism of maternal care will enhance our understanding of spider biology and behavior.

Concurrent threats and extinction risk in a long-lived, highly fecund vertebrate with parental care.

Detecting declines and quantifying extinction risk of long-lived, highly fecund vertebrates, including fishes, reptiles, and amphibians, can be challenging. In addition to the false notion that large clutches always buffer against population declines, the imperiled status of long-lived species can often be masked by extinction debt, wherein adults persist on the landscape for several years after populations cease to be viable. Here we develop a demographic model for the eastern hellbender (Cryptobranchus alleganiensis), an imperiled aquatic salamander with paternal care. We examined the individual and interactive effects of three of the leading threats hypothesized to contribute to the species' demise: habitat loss due to siltation, high rates of nest failure, and excess adult mortality caused by fishing and harvest. We parameterized the model using data on their life history and reproductive ecology to model the fates of individual nests and address multiple sources of density-dependent mortality under both deterministic and stochastic environmental conditions. Our model suggests that high rates of nest failure observed in the field are sufficient to drive hellbender populations toward a geriatric age distribution and eventually to localized extinction but that this process takes decades. Moreover, the combination of limited nest site availability due to siltation, nest failure, and stochastic adult mortality can interact to increase the likelihood and pace of extinction, which was particularly evident under stochastic scenarios. Density dependence in larval survival and recruitment can severely hamper a population's ability to recover from declines. Our model helps to identify tipping points beyond which extinction becomes certain and management interventions become necessary. Our approach can be generalized to understand the interactive effects of various threats to the extinction risk of other long-lived vertebrates. As we face unprecedented rates of environmental change, holistic approaches incorporating multiple concurrent threats and their impacts on different aspects of life history will be necessary to proactively conserve long-lived species.

Sexual cannibalism as a female resistance trait: a new hypothesis.

Female spiders and praying mantises are renowned for their cannibalism of male partners before, during, or after mating. While several hypotheses have been proposed to explain species-specific examples of sexual cannibalism, much variation remains unexplained, including why the timing of cannibalism varies across taxa. Here, I outline how sexually cannibalistic behavior could evolve via sexually antagonistic selection as a type of behavioral resistance to male-imposed mating costs, and how such a generalizable interpretation provides a framework for understanding the evolution of both sexual cannibalism in females and anti-cannibalistic traits in males. I discuss how differences between mating systems that physiologically constrain males to mate only once (monogyny) or twice (bigyny) and systems where the sexes can potentially mate multiply (polygyny and polyandry) are likely to influence how sexual conflict shapes cannibalistic behavior. I review key examples from the literature that suggest how sexually cannibalistic behavior might function as a female resistance trait and provide comprehensive predictions for testing this hypothesis empirically.

El banquete humano. A propósito de Hannibal Lecter y el canibalismo. El silencio de los corderos (1991) de Jonathan Demme / The human banquet. About Hannibal Lecter and cannibalism. The Silence of the Lambs (1991) by Jonathan Demme

El dragón rojo (1981) de Thomas Harris es la carta de presentación del perverso psiquiatra Hannibal Lecter, un psicópata asesino que brilla singularmente en el firmamento de los homicidas más abyectos de la historia del cine. Esta colección se completó con la publicación de El silencio de los corderos (1988), Hannibal (1999) y Hannibal: El origen del mal (2006), hasta ahora su última secuela. Fuente de inspiración cinematográfica, la saga se inició con la galardonada El silencio de los corderos (1991) de Jonathan Demme, continuada por Hannibal (2001) de Ridley Scott, El dragón rojo (2002) de Brett Ratner y Hannibal: El origen del mal (2007) de Peter Webber, si bien además existe una primera adaptación de la novela El dragón rojo titulada Hunter/Cazador de hombres (1986) de Michael Mann, donde los agentes del FBI se enfrentan con el temible asesino en serie Dr. Hannibal Lecktor (Brian Cox). Asimismo, el escritor y productor estadounidense Bryan Fuller creó la serie televisiva Hannibal (2013-2015), 39 episodios en los que el actor danés Mads Mikkelsen encarna al perverso antropófago. Independiente de sus connotaciones antropológicas, morales y culturales, la mayor parte de las conductas de canibalismo están relacionadas con trastornos psicóticos, como por ejemplo la esquizofrenia paranoica, cuando estos enfermos asesinan y se comen parte de los cuerpos de sus víctimas dentro de sus cuadros delirantes patológicos.(AU)

The Red Dragon (1981) by Thomas Harris is the introduction of the perverse psychiatrist Hannibal Lecter, a murdeous psychopath who shines singularly in the firmament of the most heinous murderers in the history of cinema. This collection was completed with the publication of The Silence of the Lambs (1988), Hannibal (1999) and Hannibal: The Origin of Evil (2006), until now its last sequel. A source of cinematographic inspirations, the saga began with the Oscar-winning The Silence of the Lambs (1991) by Jonathan Demme, followed by Hannibal (2001) by Ridley Scott, The Red Dragon (2002) by Brett Rainer, and Hannibal: The Origin of Evil (2007) by Peter Webber, although there is also a first adaptation of the novel The Red Dragon titled Manhunter (1986) by Michael Mann, where FBI agents confront the fearsome serial killer Dr. Hannibal Lecktor (Bryan Cox). Likewise, the American writer and producer Bryan Fuller created the television series Hannibal (2013-2015), 39 episodes in which the danish actor Mads Mikkelsen embodies the perverse cannibal. Regardless of its anthropological, moral and cultural connotations, most cannibalism behaviors are related to psychotic disorders, such as paranoid schyzophrenia, when this patients kill an d eat part of the bodies of their victims in their delusional pictures.(AU)

Overcompensation of transient and permanent death rate increases in age-structured models with cannibalistic interactions.

There has been renewed interest in understanding the mathematical structure of ecological population models that lead to overcompensation, the process by which a population recovers to a higher level after suffering a permanent increase in predation or harvesting. Here, we apply a recently formulated kinetic population theory to formally construct an age-structured single-species population model that includes a cannibalistic interaction in which older individuals prey on younger ones. Depending on the age-dependent structure of this interaction, our model can exhibit transient or steady-state overcompensation of an increased death rate as well as oscillations of the total population, both phenomena that have been observed in ecological systems. Analytic and numerical analysis of our model reveals sufficient conditions for overcompensation and oscillations. We also show how our structured population partial integrodifferential equation (PIDE) model can be reduced to coupled ODE models representing piecewise constant parameter domains, providing additional mathematical insight into the emergence of overcompensation.

Mapping Cell-in-Cell Structures in Oral Squamous Cell Carcinoma.

Cell-in-cell (CIC) structures contribute to tumor aggressiveness and poor prognosis in oral squamous cell carcinoma (OSCC). In vitro 3D models may contribute to the understanding of the underlying molecular mechanisms of these events. We employed a spheroid model to study the CIC structures in OSCC. Spheroids were obtained from OSCC (HSC3) and cancer-associated fibroblast (CAF) lines using the Nanoshuttle-PLTM bioprinting system (Greiner Bio-One). Spheroid form, size, and reproducibility were evaluated over time (EvosTM XL; ImageJ version 1.8). Slides were assembled, stained (hematoxylin and eosin), and scanned (Axio Imager Z2/VSLIDE) using the OlyVIA System (Olympus Life Science) and ImageJ software (NIH) for cellular morphology and tumor zone formation (hypoxia and/or proliferative zones) analysis. CIC occurrence, complexity, and morphology were assessed considering the spheroid regions. Well-formed spheroids were observed within 6 h of incubation, showing the morphological aspects of the tumor microenvironment, such as hypoxic (core) and proliferative zone (periphery) formation. CIC structures were found in both homotypic and heterotypic groups, predominantly in the proliferative zone of the mixed HSC3/CAF spheroids. "Complex cannibalism" events were also noted. These results showcase the potential of this model in further studies on CIC morphology, formation, and relationship with tumor prognosis.