Minimally Invasive Injectable Cosmetic Procedures Increase Feelings of Authenticity.

BACKGROUND: Minimally invasive cosmetic dermatology procedures continue to be increasingly popular; however, the extant literature has poorly documented the psychological antecedents of interest in cosmetic procedures and their psychological consequences. OBJECTIVE: To better inform dermatologists on their patients' motivations for cosmetic enhancement. MATERIALS AND METHODS: In a general population survey, an online representative sample of 984 Americans reported the extent to which they feel authentic using the validated authenticity scale and whether they were interested in undergoing a cosmetic procedure. In a prospective dermatology office survey, 102 participants reported their feelings of authenticity immediately before and 2 weeks after receiving a minimally invasive injectable cosmetic procedure. RESULTS: In the general population survey, participants interested in cosmetic procedures felt significantly less authentic than participants who were not interested ( p = .003). In the prospective dermatology office survey, participants felt significantly more authentic 2 weeks after their minimally invasive injectable cosmetic procedure than before ( p = .018). CONCLUSION: Lower feelings of authenticity are associated with interest in cosmetic procedures. Participants felt more authentic 2 weeks after receiving a minimally invasive injectable cosmetic procedure. Cosmetic procedures may present patients with an opportunity to feel more like their real, genuine selves.

An ex vivo model of Toxoplasma recrudescence reveals developmental plasticity of the bradyzoite stage.

IMPORTANCE: The classical depiction of the Toxoplasma lifecycle is bradyzoite excystation conversion to tachyzoites, cell lysis, and immune control, followed by the reestablishment of bradyzoites and cysts. In contrast, we show that tachyzoite growth slows independent of the host immune response at a predictable time point following excystation. Furthermore, we demonstrate a host cell-dependent pathway of continuous amplification of the cyst-forming bradyzoite population. The developmental plasticity of the excysted bradyzoites further underlines the critical role the cyst plays in the flexibility of the lifecycle of this ubiquitous parasite. This revised model of Toxoplasma recrudescence uncovers previously unknown complexity in the clinically important bradyzoite stage of the parasite, which opens the door to further study these novel developmental features of the Toxoplasma intermediate life cycle.

Motivations for Seeking Cosmetic Enhancing Procedures of the Face: A Systematic Review.

BACKGROUND: Cosmetic enhancing procedures continue to grow in demand. Physicians should understand the complex factors that drive patient motivation for seeking such procedures. OBJECTIVE: In contrast to a lens of psychopathology, this review reveals the driving power of everyday intrapersonal, social, and behavioral factors that motivate interest in elective facial cosmetic procedures. MATERIALS AND METHODS: The review was conducted according to PRISMA guidelines and included studies with at least 50 adult patients seeking facial cosmetic enhancements between January 1, 2000, and July 1, 2022. RESULTS: Among 1,239 identified publications, 21 studies with 9,005 participants were selected for inclusion. The review documents everyday factors as patient motivators for pursuing cosmetic enhancements of the face, with the majority of work focusing on intrapersonal factors (17 of 21 studies), such as preventing aging or negative appearance based self-appraisals. For studies reporting social factors (15 of 21 studies), the most common motivators were the patient's social network and a desire to promote social standing. Behavioral factors revealed that social media and media consumption impact patient motivation for cosmetic enhancements (5 of 21 studies). CONCLUSION: In summary, this review demonstrates that patient motivations for facial cosmetic enhancements may be best understood through everyday intrapersonal, social, and behavioral factors.

Restriction Checkpoint Controls Bradyzoite Development in Toxoplasma gondii.

Human toxoplasmosis is a life-threatening disease caused by the apicomplexan parasite Toxoplasma gondii. Rapid replication of the tachyzoite is associated with symptomatic disease, while suppressed division of the bradyzoite is responsible for chronic disease. Here, we identified the T. gondii cell cycle mechanism, the G1 restriction checkpoint (R-point), that operates the switch between parasite growth and differentiation. Apicomplexans lack conventional R-point regulators, suggesting adaptation of alternative factors. We showed that Cdk-related G1 kinase TgCrk2 forms alternative complexes with atypical cyclins (TgCycP1, TgCycP2, and TgCyc5) in the rapidly dividing developmentally incompetent RH and slower dividing developmentally competent ME49 tachyzoites and bradyzoites. Examination of cyclins verified the correlation of cyclin expression with growth dependence and development capacity of RH and ME49 strains. We demonstrated that rapidly dividing RH tachyzoites were dependent on TgCycP1 expression, which interfered with bradyzoite differentiation. Using the conditional knockdown model, we established that TgCycP2 regulated G1 duration in the developmentally competent ME49 tachyzoites but not in the developmentally incompetent RH tachyzoites. We tested the functions of TgCycP2 and TgCyc5 in alkaline induced and spontaneous bradyzoite differentiation (rat embryonic brain cells) models. Based on functional and global gene expression analyses, we determined that TgCycP2 also regulated bradyzoite replication, while signal-induced TgCyc5 was critical for efficient tissue cyst maturation. In conclusion, we identified the central machinery of the T. gondii restriction checkpoint comprised of TgCrk2 kinase and three atypical T. gondii cyclins and demonstrated the independent roles of TgCycP1, TgCycP2, and TgCyc5 in parasite growth and development. IMPORTANCE Toxoplasma gondii is a virulent and abundant human pathogen that puts millions of silently infected people at risk of reactivation of the chronic disease. Encysted bradyzoites formed during the chronic stage are resistant to current therapies. Therefore, insights into the mechanism of tissue cyst formation and reactivation are major areas of investigation. The fact that rapidly dividing parasites differentiate poorly strongly suggests that there is a threshold of replication rate that must be crossed to be considered for differentiation. We discovered a cell cycle mechanism that controls the T. gondii growth-rest switch involved in the conversion of dividing tachyzoites into largely quiescent bradyzoites. This switch operates the T. gondii restriction checkpoint using a set of atypical and parasite-specific regulators. Importantly, the novel T. gondii R-point network was not present in the parasite's human and animal hosts, offering a wealth of new and parasite-specific drug targets to explore in the future.

Toxoplasma gondii AP2XII-2 Contributes to Proper Progression through S-Phase of the Cell Cycle.

Toxoplasma gondii is a protozoan parasite that causes lifelong chronic infection that can reactivate in immunocompromised individuals. Upon infection, the replicative stage (tachyzoite) converts into a latent tissue cyst stage (bradyzoite). Like other apicomplexans, T. gondii possesses an extensive lineage of proteins called ApiAP2s that contain DNA-binding domains first characterized in plants. The function of most ApiAP2s is unknown. We previously found that AP2IX-4 is a cell cycle-regulated ApiAP2 expressed only in dividing parasites as a putative transcriptional repressor. In this study, we purified proteins interacting with AP2IX-4, finding it to be a component of the recently characterized microrchidia (MORC) transcriptional repressor complex. We further analyzed AP2XII-2, another cell cycle-regulated factor that associates with AP2IX-4. We monitored parallel expression of AP2IX-4 and AP2XII-2 proteins in tachyzoites, detecting peak expression during S/M phase. Unlike AP2IX-4, which is dispensable in tachyzoites, loss of AP2XII-2 resulted in a slowed tachyzoite growth due to a delay in S-phase progression. We also found that AP2XII-2 depletion increased the frequency of bradyzoite differentiation in vitro These results suggest that multiple AP2 factors collaborate to ensure proper cell cycle progression and tissue cyst formation in T. gondiiIMPORTANCEToxoplasma gondii is a single-celled parasite that persists in its host by converting into a latent cyst stage. This work describes a new transcriptional factor called AP2XII-2 that plays a role in properly maintaining the growth rate of replicating parasites, which contributes to signals required for development into its dormant stage. Without AP2XII-2, Toxoplasma parasites experience a delay in their cell cycle that increases the frequency of latent cyst formation. In addition, we found that AP2XII-2 operates in a multisubunit complex with other AP2 factors and chromatin remodeling machinery that represses gene expression. These findings add to our understanding of how Toxoplasma parasites balance replication and dormancy, revealing novel points of potential therapeutic intervention to disrupt this clinically relevant process.

Aberrant DNMT3B7 expression correlates to tissue type, stage, and survival across cancers.

Cancer cells are known for aberrant methylation patterns leading to altered gene expression and tumor progression. DNA methyltransferases (DNMTs) are responsible for regulating DNA methylation in normal cells. However, many aberrant versions of DNMTs have been identified to date and their role in cancer continues to be elucidated. It has been previously shown that an aberrant version of a de novo methylase, DNMT3B7, is expressed in many cancer cell lines and has a functional role in the progression of breast cancer, neuroblastoma, and lymphoma. It is clear that DNMT3B7 is important to tumor development in vitro and in vivo, but it is unknown if expression of the transcript in all of these cell lines translates to relevant clinical results. In this study, a bioinformatics approach was utilized to test the hypothesis that DNMT3B7 expression corresponds to tumor progression in patient samples across cancer types. Gene expression and clinical data were obtained from the Genomic Data Commons for the 33 cancer types available and analyzed for DNMT3B7 expression with relation to tissue type in matched and unmatched samples, staging of tumors, and patient survival. Here we present the results of this analysis indicating a role for DNMT3B7 in tumor progression of many additional cancer types. Based on these data, future in vitro and in vivo studies can be prioritized to examine DNMT3B7 in cancer and, hopefully, develop novel therapeutics to target this aberrant transcript across multiple tumor types.

Transcriptional repression by ApiAP2 factors is central to chronic toxoplasmosis.

Tachyzoite to bradyzoite development in Toxoplasma is marked by major changes in gene expression resulting in a parasite that expresses a new repertoire of surface antigens hidden inside a modified parasitophorous vacuole called the tissue cyst. The factors that control this important life cycle transition are not well understood. Here we describe an important transcriptional repressor mechanism controlling bradyzoite differentiation that operates in the tachyzoite stage. The ApiAP2 factor, AP2IV-4, is a nuclear factor dynamically expressed in late S phase through mitosis/cytokinesis of the tachyzoite cell cycle. Remarkably, deletion of the AP2IV-4 locus resulted in the expression of a subset of bradyzoite-specific proteins in replicating tachyzoites that included tissue cyst wall components BPK1, MCP4, CST1 and the surface antigen SRS9. In the murine animal model, the mis-timing of bradyzoite antigens in tachyzoites lacking AP2IV-4 caused a potent inflammatory monocyte immune response that effectively eliminated this parasite and prevented tissue cyst formation in mouse brain tissue. Altogether, these results indicate that suppression of bradyzoite antigens by AP2IV-4 during acute infection is required for Toxoplasma to successfully establish a chronic infection in the immune-competent host.

Toxoplasma gondii AP2IX-4 Regulates Gene Expression during Bradyzoite Development.

Toxoplasma gondii is a protozoan parasite of great importance to human and animal health. In the host, this obligate intracellular parasite persists as a tissue cyst that is imperceptible to the immune response and unaffected by current therapies. The tissue cysts facilitate transmission through predation and give rise to chronic cycles of toxoplasmosis in immunocompromised patients. Transcriptional changes accompany conversion of the rapidly replicating tachyzoites into the encysted bradyzoites, and yet the mechanisms underlying these alterations in gene expression are not well defined. Here we show that AP2IX-4 is a nuclear protein exclusively expressed in tachyzoites and bradyzoites undergoing division. Knockout of AP2IX-4 had no discernible effect on tachyzoite replication but resulted in a reduced frequency of tissue cyst formation following alkaline stress induction-a defect that is reversible by complementation. AP2IX-4 has a complex role in regulating bradyzoite gene expression, as the levels of many bradyzoite mRNAs dramatically increased beyond those seen under conditions of normal stress induction in AP2IX-4 knockout parasites exposed to alkaline media. The loss of AP2IX-4 also resulted in a modest virulence defect and reduced cyst burden in chronically infected mice, which was reversed by complementation. These findings illustrate that the transcriptional mechanisms responsible for tissue cyst development operate across the intermediate life cycle from the dividing tachyzoite to the dormant bradyzoite. IMPORTANCEToxoplasma gondii is a single-celled parasite that persists in its host as a transmissible tissue cyst. How the parasite converts from its replicative form to the bradyzoites housed in tissue cysts is not well understood, but the process clearly involves changes in gene expression. Here we report that parasites lacking a cell cycle-regulated transcription factor called AP2IX-4 display reduced frequencies of tissue cyst formation in culture and in a mouse model of infection. Parasites missing AP2IX-4 lose the ability to regulate bradyzoite genes during tissue cyst development. Expressed in developing bradyzoites still undergoing division, AP2IX-4 may serve as a useful marker in the study of transitional forms of the parasite.

Opposing Transcriptional Mechanisms Regulate Toxoplasma Development.

The Toxoplasma biology that underlies human chronic infection is developmental conversion of the acute tachyzoite stage into the latent bradyzoite stage. We investigated the roles of two alkaline-stress-induced ApiAP2 transcription factors, AP2IV-3 and AP2IX-9, in bradyzoite development. These factors were expressed in two overlapping waves during bradyzoite development, with AP2IX-9 increasing expression earlier than AP2IV-3, which peaked as AP2IX-9 expression was declining. Disruption of the AP2IX-9 gene enhanced, while deletion of AP2IV-3 gene decreased, tissue cyst formation, demonstrating that these factors have opposite functions in bradyzoite development. Conversely, conditional overexpression of FKBP-modified AP2IX-9 or AP2IV-3 with the small molecule Shield 1 had a reciprocal effect on tissue cyst formation, confirming the conclusions of the knockout experiments. The AP2IX-9 repressor and AP2IV-3 activator tissue cyst phenotypes were borne out in gene expression studies that determined that many of the same bradyzoite genes were regulated in an opposite manner by these transcription factors. A common gene target was the canonical bradyzoite marker BAG1, and mechanistic experiments determined that, like AP2IX-9, AP2IV-3 regulates a BAG1 promoter-luciferase reporter and specifically binds the BAG1 promoter in parasite chromatin. Altogether, these results suggest that the AP2IX-9 transcriptional repressor and the AP2IV-3 transcriptional activator likely compete to control bradyzoite gene expression, which may permit Toxoplasma to better adapt to different tissue environments and select a suitable host cell for long-term survival of the dormant tissue cyst. IMPORTANCEToxoplasma infections are lifelong because of the development of the bradyzoite tissue cyst, which is effectively invisible to the immune system. Despite the important clinical consequences of this developmental pathway, the molecular basis of the switch mechanisms that control tissue cyst formation is still poorly understood. Significant changes in gene expression are associated with tissue cyst development, and ApiAP2 transcription factors are an important mechanism regulating this developmental transcriptome. However, the molecular composition of these ApiAP2 complexes and the operating principles of ApiAP2 mechanisms are not well defined. Here we establish that competing ApiAP2 transcriptional mechanisms operate to regulate this clinically important developmental pathway.

Toxoplasma gondii Cyclic AMP-Dependent Protein Kinase Subunit 3 Is Involved in the Switch from Tachyzoite to Bradyzoite Development.

UNLABELLED: Toxoplasma gondii is an obligate intracellular apicomplexan parasite that infects warm-blooded vertebrates, including humans. Asexual reproduction in T. gondii allows it to switch between the rapidly replicating tachyzoite and quiescent bradyzoite life cycle stages. A transient cyclic AMP (cAMP) pulse promotes bradyzoite differentiation, whereas a prolonged elevation of cAMP inhibits this process. We investigated the mechanism(s) by which differential modulation of cAMP exerts a bidirectional effect on parasite differentiation. There are three protein kinase A (PKA) catalytic subunits (TgPKAc1 to -3) expressed in T. gondii Unlike TgPKAc1 and TgPKAc2, which are conserved in the phylum Apicomplexa, TgPKAc3 appears evolutionarily divergent and specific to coccidian parasites. TgPKAc1 and TgPKAc2 are distributed in the cytomembranes, whereas TgPKAc3 resides in the cytosol. TgPKAc3 was genetically ablated in a type II cyst-forming strain of T. gondii (PruΔku80Δhxgprt) and in a type I strain (RHΔku80Δhxgprt), which typically does not form cysts. The Δpkac3 mutant exhibited slower growth than the parental and complemented strains, which correlated with a higher basal rate of tachyzoite-to-bradyzoite differentiation. 3-Isobutyl-1-methylxanthine (IBMX) treatment, which elevates cAMP levels, maintained wild-type parasites as tachyzoites under bradyzoite induction culture conditions (pH 8.2/low CO2), whereas the Δpkac3 mutant failed to respond to the treatment. This suggests that TgPKAc3 is the factor responsible for the cAMP-dependent tachyzoite maintenance. In addition, the Δpkac3 mutant had a defect in the production of brain cysts in vivo, suggesting that a substrate of TgPKAc3 is probably involved in the persistence of this parasite in the intermediate host animals. IMPORTANCE: Toxoplasma gondii is one of the most prevalent eukaryotic parasites in mammals, including humans. Parasites can switch from rapidly replicating tachyzoites responsible for acute infection to slowly replicating bradyzoites that persist as a latent infection. Previous studies have demonstrated that T. gondii cAMP signaling can induce or suppress bradyzoite differentiation, depending on the strength and duration of cAMP signal. Here, we report that TgPKAc3 is responsible for cAMP-dependent tachyzoite maintenance while suppressing differentiation into bradyzoites, revealing one mechanism underlying how this parasite transduces cAMP signals during differentiation.

Toxoplasma development - turn the switch on or off?

Toxoplasma gondii exhibits a complex, multi-stage life cycle in which the need for parasite expansion is balanced with the production of transmissible forms. For human disease the key developmental switch is from the tachyzoite to the mature bradyzoite, which is not well understood at the molecular level. This review highlights the role of the tachyzoite in regulating the initiation of bradyzoite differentiation through newly discovered transcription factors of the ApiAP2 family that must be turned off for development to unfold. Exit from the tachyzoite cell cycle is also tightly co-ordinated with the induction of bradyzoite gene expression, which is strongly influenced by the host cell environment. New evidence suggests a parasite casein kinase II and host anti-growth factor CDA1 can influence specific pathways that are responsible for sensing the host cell environment and informing the parasites decision to continue replication or to develop into bradyzoites. These results indicate tachyzoite gene expression mechanisms and signal transduction pathways likely hold the keys to tissue cyst formation in Toxoplasma.

A nucleolar AAA-NTPase is required for parasite division.

Apicomplexa division involves several distinct phases shared with other eukaryote cell cycles including a gap period (G1) prior to chromosome synthesis, although how progression through the parasite cell cycle is controlled is not understood. Here we describe a cell cycle mutant that reversibly arrests in the G1 phase. The defect in this mutant was mapped by genetic complementation to a gene encoding a novel AAA-ATPase/CDC48 family member called TgNoAP1. TgNoAP1 is tightly regulated and expressed in the nucleolus during the G1/S phases. A tyrosine to a cysteine change upstream of the second AAA+ domain in the temperature sensitive TgNoAP1 allele leads to conditional protein instability, which is responsible for rapid cell cycle arrest and a primary defect in 28S rRNA processing as confirmed by knock-in of the mutation back into the parent genome. The interaction of TgNoAP1 with factors of the snoRNP and R2TP complexes indicates this protein has a role in pre-rRNA processing. This is a novel role for a cdc48-related chaperone protein and indicates that TgNoAP1 may be part of a dynamic mechanism that senses the health of the parasite protein machinery at the initial steps of ribosome biogenesis and conveys that information to the parasite cell cycle checkpoint controls.

ApiAP2 transcription factor restricts development of the Toxoplasma tissue cyst.

Cellular differentiation leading to formation of the bradyzoite tissue cyst stage is the underlying cause of chronic toxoplasmosis. Consequently, mechanisms responsible for controlling development in the Toxoplasma intermediate life cycle have long been sought. Here, we identified 15 Toxoplasma mRNAs induced in early bradyzoite development that encode proteins with apicomplexan AP2 (ApiAP2) DNA binding domains. Of these 15 mRNAs, the AP2IX-9 mRNA demonstrated the largest expression increase during alkaline-induced differentiation. At the protein level, we found that AP2IX-9 was restricted to the early bradyzoite nucleus and is repressed in tachyzoites and in mature bradyzoites from 30-d infected animals. Conditional overexpression of AP2IX-9 significantly reduced tissue cyst formation and conferred alkaline pH-resistant growth, whereas disruption of the AP2IX-9 gene increased tissue cyst formation, indicating AP2IX-9 operates as a repressor of bradyzoite development. Consistent with a role as a repressor, AP2IX-9 specifically inhibited the expression of bradyzoite mRNAs, including the canonical bradyzoite marker, bradyzoite antigen 1 (BAG1). Using protein binding microarrays, we established the AP2 domain of AP2IX-9 binds a CAGTGT DNA sequence motif and is capable of binding cis-regulatory elements controlling the BAG1 and bradyzoite-specific nucleoside triphosphatase (B-NTPase) promoters. The effect of AP2IX-9 on BAG1 expression was direct because this factor inhibits expression of a firefly luciferase reporter under the control of the BAG1 promoter in vivo, and epitope-tagged AP2IX-9 can be immunoprecipitated with the BAG1 promoter in parasite chromatin. Altogether, these results indicate AP2IX-9 restricts Toxoplasma commitment to develop the mature bradyzoite tissue cyst.

The transcription of bradyzoite genes in Toxoplasma gondii is controlled by autonomous promoter elements.

Experimental evidence suggests that apicomplexan parasites possess bipartite promoters with basal and regulated cis-elements similar to other eukaryotes. Using a dual luciferase model adapted for recombinational cloning and use in Toxoplasma gondii, we show that genomic regions flanking 16 parasite genes, which encompass examples of constitutive and tachyzoite- and bradyzoite-specific genes, are able to reproduce the appropriate developmental stage expression in a transient luciferase assay. Mapping of cis-acting elements in several bradyzoite promoters led to the identification of short sequence spans that are involved in control of bradyzoite gene expression in multiple strains and under different bradyzoite induction conditions. Promoters that regulate the heat shock protein BAG1 and a novel bradyzoite-specific NTPase during bradyzoite development were fine mapped to a 6-8 bp resolution and these minimal cis-elements were capable of converting a constitutive promoter to one that is induced by bradyzoite conditions. Gel-shift experiments show that mapped cis-elements are bound by parasite protein factors with the appropriate functional sequence specificity. These studies are the first to identify the minimal sequence elements that are required and sufficient for bradyzoite gene expression and to show that bradyzoite promoters are maintained in a 'poised' chromatin state throughout the intermediate host life cycle in low passage strains. Together, these data demonstrate that conventional eukaryotic promoter mechanisms work with epigenetic processes to regulate developmental gene expression during tissue cyst formation.

The transcriptome of Toxoplasma gondii.

BACKGROUND: Toxoplasma gondii gives rise to toxoplasmosis, among the most prevalent parasitic diseases of animals and man. Transformation of the tachzyoite stage into the latent bradyzoite-cyst form underlies chronic disease and leads to a lifetime risk of recrudescence in individuals whose immune system becomes compromised. Given the importance of tissue cyst formation, there has been intensive focus on the development of methods to study bradyzoite differentiation, although the molecular basis for the developmental switch is still largely unknown. RESULTS: We have used serial analysis of gene expression (SAGE) to define the Toxoplasma gondii transcriptome of the intermediate-host life cycle that leads to the formation of the bradyzoite/tissue cyst. A broad view of gene expression is provided by >4-fold coverage from nine distinct libraries (approximately 300,000 SAGE tags) representing key developmental transitions in primary parasite populations and in laboratory strains representing the three canonical genotypes. SAGE tags, and their corresponding mRNAs, were analyzed with respect to abundance, uniqueness, and antisense/sense polarity and chromosome distribution and developmental specificity. CONCLUSION: This study demonstrates that phenotypic transitions during parasite development were marked by unique stage-specific mRNAs that accounted for 18% of the total SAGE tags and varied from 1-5% of the tags in each developmental stage. We have also found that Toxoplasma mRNA pools have a unique parasite-specific composition with 1 in 5 transcripts encoding Apicomplexa-specific genes functioning in parasite invasion and transmission. Developmentally co-regulated genes were dispersed across all Toxoplasma chromosomes, as were tags representing each abundance class, and a variety of biochemical pathways indicating that trans-acting mechanisms likely control gene expression in this parasite. We observed distinct similarities in the specificity and expression levels of mRNAs in primary populations (Day-6 post-sporozoite infection) that occur prior to the onset of bradyzoite development that were uniquely shared with the virulent Type I-RH laboratory strain suggesting that development of RH may be arrested. By contrast, strains from Type II-Me49B7 and Type III-VEGmsj contain SAGE tags corresponding to bradyzoite genes, which suggests that priming of developmental expression likely plays a role in the greater capacity of these strains to complete bradyzoite development.

A change in the premitotic period of the cell cycle is associated with bradyzoite differentiation in Toxoplasma gondii.

We have demonstrated that bradyzoites return to the tissue-cyst stage by a developmental pathway that is indistinguishable from that initiated by sporozoites. Mature bradyzoites, like sporozoites from oocysts, were non-proliferative as they contained uniform 1N DNA contents, and replication occurred only in parasites that de-differentiated back into tachyzoites. Moreover, tachyzoites emergent from the bradyzoite-initiated pathway underwent a spontaneous growth shift prior to the onset of tissue cyst formation in a timeframe that was identical to cultures infected with sporozoites. In sporozoite-infected cultures, a novel premitotic, near-diploid subpopulation was detected during bradyzoite differentiation that co-expressed tachyzoite and bradyzoite markers. These observations suggest that activation of a G2-related cell cycle mechanism is required during bradyzoite development, and indicates that equivalent cell cycle mechanisms may govern development in the intermediate life cycle regardless of the origin of infection.