Allogromia laticollaris: A Foraminiferan with an Unusual Apogamic Metagenic Life Cycle*

SYNOPSIS. The life cycles of 3 strains of Allogromia laticollaris, a monothalamous foraminiferan, have been studied. Each of the strains had a different, nonclassical, and basically apogamic, life cycle. The Cold Spring Harbor (CSH) strain regularly alternated between 2 agamontic forms: agamont I (uninucleate and diploid) and agamont II (multinucleate and diploid). The complete life cycle took 26 days. Sexual reproduction was rare (0.01%) and autogamous. Small numbers of organisms also underwent budding, binary fission, and cytotomy. The life cycles of the Towd Point (TPA) and Sippewissett (SIP) strains were comparatively abbreviated. Agamont II dominated their typical life cycles, which were completed in 16-18 days. The life cycle of SIP was basically a continuous cycling of the agamont II phase. Approximately 75% of the schizozoites of the TPA strain developed into agamont II. The other 25% alternated between agamont II and agamont I phase. In the CSH strain schizozoites with ～ 8 (range 5-15) nuclei characterized newly formed agamonts II. More nuclei (～ 25) were found in the other 2 strains. The nuclei in young agamonts II underwent rapid morphologic changes leading to a “mushroom-like” chromosome appearance and extensive RNA synthesis. Nucleolar material accumulated at the nuclear periphery and eventually was discharged to the cytoplasm. Karyokinesis took place without the breakdown of the nuclear membrane. The single nucleus of young agamont I forms was proportionally quite large. The S₁ phase occurred quite early (2-5 days) in this part of the life cycle. RNA in the CSH strain formed a compact, subcortical, coarsely granular ring, while in the TPA it was cortical and differentiated into finely granular matrix with randomly distributed coarse granules. During the G₂ phase the nucleus became further enlarged and eventually amoeba-form. Intermediate stages in nuclear breakdown were not found.     

Compositional factors influencing cell surface hydrophobicity ofPasteurella multocida variants

The influence of macromolecules other than lipopolysaccharide on the hydrophobic properties ofPasteurella multocida was investigated by assessing cell surface hydrophobicity (CSH) after experimentally modifying surfaces of various strains. CSH of hydrophobic variants was enhanced by growth on blood-supplemented medium and mechanical shearing, whereas chloramphenicol, oxytetracycline, trypsin, and pronase E treatments decreased CSH. No such modifications were observed for hydrophilic strains. Microscopic observations revealed hydrophilic strains to be heavily encapsulated in contrast to hydrophobic strains. Repeated subculturing reduced encapsulation with a concomitant increase in CSH for one hydrophilic strain while exerting no changes in the other hydrophilic strain examined. Hyaluronidase removal of capsular material from a serotype A strain resulted in increased CSH; subsequent exposure to pronase E resulted in partial restoration of hydrophilicity. These data suggest the encapsulation of hydrophilicP. multocida strains masks a relatively hydrophobic surface that is conferred, at least in part, by the presence of one or more surface-exposed proteins common to both hydrophilic and hydrophobic variants.     

Flocculation in ale brewing strains of <Emphasis Type="Italic">Saccharomyces cerevisiae</Emphasis>: re-evaluation of the role of cell surface charge and hydrophobicity

Flocculation is an eco-friendly process of cell separation, which has been traditionally exploited by the brewing industry. Cell surface charge (CSC), cell surface hydrophobicity (CSH) and the presence of active flocculins, during the growth of two (NCYC 1195 and NCYC 1214) ale brewing flocculent strains, belonging to the NewFlo phenotype, were examined. Ale strains, in exponential phase of growth, were not flocculent and did not present active flocculent lectins on the cell surface; in contrast, the same strains, in stationary phase of growth, were highly flocculent (>98%) and presented a hydrophobicity of approximately three to seven times higher than in exponential phase. No relationship between growth phase, flocculation and CSC was observed. For comparative purposes, a constitutively flocculent strain (S646-1B) and its isogenic non-flocculent strain (S646-8D) were also used. The treatment of ale brewing and S646-1B strains with pronase E originated a loss of flocculation and a strong reduction of CSH; S646-1B pronase E-treated cells displayed a similar CSH as the non-treated S646-8D cells. The treatment of the S646-8D strain with protease did not reduce CSH. In conclusion, the increase of CSH observed at the onset of flocculation of ale strains is a consequence of the presence of flocculins on the yeast cell surface and not the cause of yeast flocculation. CSH and CSC play a minor role in the auto-aggregation of the ale strains since the degree of flocculation is defined, primarily, by the presence of active flocculins on the yeast cell wall.     

The distinct role of catalase and DNA repair systems in protection against hydrogen peroxide in Escherichia coli

The katEkatG mutant of E. coli, UM1, had no assayable catalase activities in the extract and showed increased (about 20 fold) sensitivity to killing by H2O2 when compared with its parental strain CSH7. The mutant strain was able to reactivate H2O2-damaged lambda phage. On the other hand, recA and polA mutants were also highly sensitive to H2O2, but they had normal level of catalase activities. RecA derivatives of UM1 were much more sensitive to H2O2 than UM1 and recA strains. The induction of umu operon occurred in UM1 at lower (1/10-1/20) doses of H2O2 than in CSH7. From the results it is concluded that the lethal effect of H2O2 is due to DNA damage induced by it and that catalase and DNA repair systems have a distinct role in protection against H2O2 in E. coli.     

Effect of endocrine disruptor para-nonylphenol on the cell growth and oxygen radical generation in Escherichia coli mutant cells deficient in catalase and superoxide dismutase

para-Nonylphenol (NP) had previously been found to have strong suppressive effects of growth of bacterial and yeast cells, and these effects were associated with NP-induced generation of radical oxygen species (ROS). In the present study, we determined that wild-type strains of Escherichia coli (CSH 7, SY-11, and IFO-3545) were resistant to NP compared with other sensitive microorganisms reported previously. To elucidate the relationship between NP-induced ROS generation and cell growth inhibition in more detail, we analyzed the effect of NP on cell growth and survival of wild-type and mutant E. coli strains deficient in ROS-scavenging enzymes such as catalase and superoxide dismutase (SOD). The SOD-deficient strain QC 774 (sod A- and sod B-) was much more sensitive to NP than wild-type (CSH 7) and catalase-deficient (UM 1 kat E- and kat G-) strains. As a comparative experiment, when hydrogen peroxide was applied to the same growth and survival assays, UM 1 cells were more sensitive to hydrogen peroxide than QC 774 and CSH 7. A chemiluminescence (CHL) experiment using MCLA (2-methyl-6-Lf-methylphenyl]-3,7-dihydroimidazc [1,2-alpha] pyrazin-3-one) reflecting predominantly superoxide generation showed that NP caused marked CHL generation in QC 774 cells, but not in CSH 7 and UM 1 cells. However, the CHL experiment using L-012 reflecting predominantly hydroxyl radical and hypochlorite did not exhibit significant CHL generation in QC 774 cells at the same concentrations of NP. Furthermore, supplementation with SOD prevented NP-induced ROS generation and cell survival inhibition of QC 774 cells, but the catalase and metal-chelating agent deferoxamine did not have significant effects. These results suggest that one of the primary actions of NP in cells is the generation of superoxide which may be responsible for NP-induced cell growth inhibition.     

Whole Genome Sequence of Treponema pallidum ssp. pallidum,Strain Mexico A,Suggests Recombination between Yaws and Syphilis Strains

Background

Treponema pallidum ssp. pallidum (TPA), the causative agent of syphilis, and Treponema pallidum ssp. pertenue (TPE), the causative agent of yaws, are closely related spirochetes causing diseases with distinct clinical manifestations. The TPA Mexico A strain was isolated in 1953 from male, with primary syphilis, living in Mexico. Attempts to cultivate TPA Mexico A strain under in vitro conditions have revealed lower growth potential compared to other tested TPA strains.

Methodology/Principal Findings

The complete genome sequence of the TPA Mexico A strain was determined using the Illumina sequencing technique. The genome sequence assembly was verified using the whole genome fingerprinting technique and the final sequence was annotated. The genome size of the Mexico A strain was determined to be 1,140,038 bp with 1,035 predicted ORFs. The Mexico A genome sequence was compared to the whole genome sequences of three TPA (Nichols, SS14 and Chicago) and three TPE (CDC-2, Samoa D and Gauthier) strains. No large rearrangements in the Mexico A genome were found and the identified nucleotide changes occurred most frequently in genes encoding putative virulence factors. Nevertheless, the genome of the Mexico A strain, revealed two genes (TPAMA_0326 (tp92) and TPAMA_0488 (mcp2-1)) which combine TPA- and TPE- specific nucleotide sequences. Both genes were found to be under positive selection within TPA strains and also between TPA and TPE strains.

Conclusions/Significance

The observed mosaic character of the TPAMA_0326 and TPAMA_0488 loci is likely a result of inter-strain recombination between TPA and TPE strains during simultaneous infection of a single host suggesting horizontal gene transfer between treponemal subspecies.     

La hidrofobicidad de la superficie celular como indicador de otros factores de virulencia en Candida albicans

ObjectiveTo evaluate virulence factors involved in the adhesion process, such as cell surface hydrophobicity (CSH), adherence to plastic capacity, adherence capacity to buccal epithelial cells (BEC), and biofilm formation, in 17 strains of C. albicans isolated from bronchial aspirates of critically ill patients.MethodThe CSH of the strains of C. albicans was determined using the MATH method, a microbial adhesion to hydrocarbons test. The study of adherence to plastic was performed in microtitre plates in accordance with Christensen's technique. Biofilm formation was studied in polystyrene microtitre plates, according to the method of Ramage. Adherence to BEC was evaluated by quantifying the percentage of adhered yeasts to cells.ResultsAll the strains studied showed factors directly involved in adhesion, with variability in the degree of expression among them. Medium-high levels of CSH were found in 52.9% of the strains. The percentage of strains with high values in adherence to plastic was 35.3%. The most hydrophobic strains were the most adherent to plastic, with a correlation coefficient of 0.76. Of the 12 biofilm-producing strains, 6 were high producers. These strains had also high levels of CSH and adherence to plastic, with significant results. All the strains studied adhered to BEC, with results ranging widely from 45 to 157 yeasts/100 BEC, with no significant correlation with the rest of the parameters studied, although CSH was seen to be an indispensable prior requisite for adherence to cells.ConclusionCSH is a variable characteristic in C. albicans and is directly related to adherence to plastic and biofilm formation. Ease in evaluating CSH permits its quantification, and could be used as an indicator of the presence of other determinants of pathogenicity.     

Life cycle as a stable trait in the evaluation of diversity of Nostoc from biofilms in rivers

The diversity within the genus Nostoc is still controversial and more studies are needed to clarify its heterogeneity. Macroscopic species have been extensively studied and discussed; however, the microscopic forms of the genus, especially those from running waters, are poorly known and likely represented by many more species than currently described. Nostoc isolates from biofilms of two Spanish calcareous rivers were characterized comparing the morphology and life cycle in two culture media with different levels of nutrients and also comparing the 16S rRNA gene sequences. The results showed that trichome shape and cellular dimensions varied considerably depending on the culture media used, whereas the characteristics expressed in the course of the life cycle remained stable for each strain independent of the culture conditions. Molecular phylogenetic analysis confirmed the distinction between the studied strains established on morphological grounds. A balanced approach to the evaluation of diversity of Nostoc in the service of autecological studies requires both genotypic information and the evaluation of stable traits. The results of this study show that 16S rRNA gene sequence similarity serves as an important criterion for characterizing Nostoc strains and is consistent with stable attributes, such as the life cycle.     

Assessment of lipopolysaccharide-binding activity of Bifidobacterium and its relationship with cell surface hydrophobicity, autoaggregation, and inhibition of interleukin-8 production

This study was performed to screen probiotic bifidobacteria for their ability to bind and neutralize lipopolysaccharides (LPS) from Escherichia coli and to verify the relationship between LPS-binding ability, cell surface hydrophobicity (CSH), and inhibition of LPS-induced interleukin-8 (IL-8) secretion by HT-29 cells of the various bifidobacterial strains. Ninety bifidobacteria isolates from human feces were assessed for their ability to bind fluorescein isothiocyanate (FITC)-labeled LPS from E. coli. Isolates showing 30-60% binding were designated LPS-high binding (LPS-H) and those with less than 15% binding were designated LPS-low binding (LPS-L). The CSH, autoaggregation (AA), and inhibition of LPS-induced IL-8 release from HT-29 cells of the LPS-H and LPS-L groups were evaluated. Five bifidobacteria strains showed high levels of LPS binding, CSH, AA, and inhibition of IL-8 release. However, statistically significant correlations between LPS binding, CSH, AA, and reduction of IL-8 release were not found. Although we could isolate bifidobacteria with high LPS-binding ability, CSH, AA, and inhibition of IL-8 release, each characteristic should be considered as strain dependent. Bifidobacteria with high LPS binding and inhibition of IL-8 release may be good agents for preventing inflammation by neutralizing Gram-negative endotoxins and improving intestinal health.     

Influence of incubation conditions on cell surface hydrophobicity of Candida species fungi

Cell surface hydrophobicity (CSH) is considered to be one of several virulence factors of Candida yeast-like fungi. The aim of the study was a measurment of hydrophobic properties of Candida sp. depending on growth conditions. A total of 139 strains of Candida (80 - C. albicans and 59 - C. non-albicans) were examined. The method of salt aggregation test (SAT) was used. The strains were cultured on three different media, in two variants of incubation temperature and time. The incubation temperature and microbiological medium affected CSH of just C. albicans strains. The influence of incubation time on CSH of examined species of Candida was not occurred. There was a strong correlation between CSH and species of Candida demonstrated in the study Hydrophobic properties were more frequent and stronger among strains of C. non-albicans than C. albicans species. The results of the study indicates that CSH of Candida spp. is a dynamic feature. The ability to change surface properties may play a role in pathogenesis of candidosis.     

Growth Conditions Influence Expression of Cell Surface Hydrophobicity of Staphylococci and Other Wound Infection Pathogens

The initial adhesion of microbes to tissue and solid surfaces can be mediated by hydrophobic interaction. Expression of microbial cell surface hydrophobicity (CSH) is influenced by growth conditions, and often best expressed after growth under nutrient-poor conditions, or “starvation.” In the present study, the CSH of 133 strains of Enterobacteriaceae, Staphylococcus aureus, coagulase-negative staphylococci, Enterococcus faecalis, group A streptococcus, Pseudomonas aeruginosa, Clostridium perfringens, Bacteroides fragilis, Peptococcus magnus, and of 8 Candida albicans strains was measured by the salt aggregation test after growth on hematin agar in a 5% CO₂ atmosphere, or under anaerobiosis. Cells of all but 8 strains expressed pronounced or moderate CSH, i.e., they aggregated in 0.01-2 M ammonium sulfate. When the agar surface was covered by human serum (diluted 1:5) to mimic growth conditions in a wound, 94 strains expressed higher CSH, and 44 strains the same CSH as after growth without serum. The CSH of 12 strains of different species was measured after growth on blood, hematin and PDM agar, with or without serum, and in an aerobic or a 5% CO₂ atmosphere. The highest CSH was expressed after growth in 5% CO₂ with serum, and the lowest growth after on blood agar in aerobic atmosphere. Identical results were obtained with native and heat-inactivated (56 C, 20 min) serum. The reduced surface tension obtained in 5% CO₂, as well as yet unidentified serum factors, promotes expression of CSH.     

The effect of culture conditions on hydrophobic properties of Pseudomonas aeruginosa

The cell surface hydrophobicity (CSH) plays an important role in a adhesion of bacteria on solid surfaces. CSH of 62 Pseudomonas aeruginosa strains isolated from humans and different animals was assessed using the ammonium sulfate salt aggregation test. Bacteria were grown for 24 h and 48 h at a room temperature (22 degrees C) and 37 degrees C on enrichment broth and agar (Biomed) and tryptic soy agar (Difco). The hydrophobic properties of the Pseudomonas aeruginosa strains were depended on the temperature, time of the culture of bacteria and the kind of media. CSH properties were most frequently expressed when the analyzed strains were cultured in enrichment broth. In a such conditions Pseudomonas aeruginosa strains were more hydrophobic when grown at 22 degrees C (94% after 24 h and 87% after 48%) than those at 37 degrees C (72% after 24 h and 71% after 48 h). Among strains cultured in tryptic soy agar at 37 degrees C, 48% after 24 h and 75% after 48 h were autoaggregating, representing very strong hydrophobic properties.     

Aggregation of lactobacilli and bifidobacteria with<Emphasis Type="Italic">Escherichia coli</Emphasis> O157

A total of 5 Bifidobacterium spp. isolated from pig and children' feces and 6 Lactobacillus spp. from chicken feces were examined for expression of aggregation, cell surface hydrophobicity (CSH) and adherence to intestinal mucin. Co-aggregation activity was seen in 3 strains of auto-aggregative bifidobacteria and 4 auto-aggregative strains of Lactobacillus spp. with 2 enterohemorrhagic E. coli (O157). CSH correlated with Lactobacillus auto-aggregating activity and adherence to mucin but the correlation between Bifidobacterium adherence to mucin and CSH was not confirmed.     

Age- and strain-related differences in murine spleen cell responses to different activation signals.

The effect of age on the response of splenocytes to activation with anti-CD3 mAb and a combination of anti-CD3 mAb and TPA, as evidenced by interleukin-2 (IL-2) and interleukin-4 (IL-4) production and cell proliferation, was examined in the C57BL/6 and DBA/2 murine strains. Depending on the mode of activation, there were age and strain differences in IL-2 and IL-4 production. With all modes of activation, cells from the old C57BL/6 mice produced less IL-2 than their young counterparts. In DBA/2 mice there was no age-related difference in IL-2 production with anti-CD3 mAb activation alone, whereas when the same cell population was activated with anti-CD3 mAb and TPA an age-associated decrease in IL-2 production occurred. In both strains, there was an age-related increase in IL-4 production with anti-CD3 mAb activation. After addition of TPA, however, there was an age-related decrease in IL-4 production. An age-related decline in the proliferation occurred with all modes of activation in both mouse strains. There were also strain-related differences in proliferation after the addition of forskolin, an inhibitor of Th1-cell function. While forskolin inhibited the proliferation of cells from the young C57BL/6 mice only, in the DBA/2 mice proliferation of cells was inhibited in both age groups. There were no strain-related differences in inhibition by anti-transferrin receptor (TrfR) mAb, although cells from the old mice were slightly more sensitive to this inhibition.     

Phage t: a group T plasmid-dependent bacteriophage

Phage t was isolated from sewage from Pretoria. It formed plaques only on Escherichia coli and Salmonella typhimurium strains that carried plasmids belonging to incompatibility group T. Five of six group T plasmids permitted visible lysis of R+ host strains. There was no visible lysis of E. coli J53-2 or S. typhimurium LT2trpA8 carrying the T plasmid Rts1 although the strains supported phage growth as indicated by at least a 10-fold increase in phage titre. The latter strains transferred the plasmid at high frequency to E. coli strain CSH2 and the resulting transconjugants plated the phage. Proteus mirabilis strain PM5006(R402) failed to support phage growth although it transferred the plasmid and concomitant phage sensitivity to E. coli J53-2. The phage was hexagonal in outline, RNA-containing, resistant to chloroform and adsorbed to the shafts of pili determined by T plasmids.     

Influence of culture conditions on cell surface hydrophobicity of rods of genus Serratia

The cell surface hydrophobicity (CSH) is a non-specific adhesion factor that is important in the proliferation of microorganisms on solid surfaces. Serratia spp. is a bacterium that has been increasingly implicated as a primary pathogen in numerous human infections, particularly in urinary tract infections. CSH of 60 Serratia spp. strains isolated from clinical materials was evaluated using the ammonium sulfate salt aggregation test. Bacteria were grown for 24 h and 48 h at room temperature (22 degrees C) and 37 degrees C on enrichment broth and agar (Biomed), enrichment agar with 5% human blood and medium composed of agar granulated (Becton Dickinson), neopeptone (Difco) and 1% (v/v) glycerol. CSH was estimated most frequently when the analyzed strains in enrichment broth were cultured. When grown in enrichment broth cells of Serratia spp. at room temperature were more hydrophobic (43% after 24 h and 47% after 48 h) than those at 37 degrees C (30% after 24 h and 33% after 48 h). CSH of the examined Serratia spp. strains were depended on the temperature, time of the culture of bacteria and the kind of media. The influence of the culture conditions on the changes in CSH of the analyzed bacteria may suggest significance of these properties in the pathogenesis of Serratia spp.     

Contribution of cell surface hydrophobicity protein 1 (Csh1p) to virulence of hydrophobic Candida albicans serotype A cells

The CSH1 gene product is the first protein implicated to affect the phenotype of cell surface hydrophobicity in Candida albicans. Ablation of expression of CSH1 resulted in a 75% loss of the cell surface hydrophobicity (CSH) phenotype. When the C. albicans csh1 knockout derivative was cultured from frozen stocks, it had reacquired CSH levels similar to the parent strain and isogenic reintegrant in the absence of Csh1p re-expression through an unknown mechanism. Prior to reacquisition of CSH, the knockout was less adherent to fibronectin than the parent. Comparison of the csh1 knockout and CSH1 reintegrant in a hematogenous dissemination model allows analysis of Csh1p contribution to virulence using matched strains with similar levels of CSH. No statistical significance between the knockout and reintegrant was found in virulence based on median day of survival, although a reproducible delay in onset of lethal infection for the knockout was observed. A modest difference in mucosal colonization in a vaginal infection model was also observed between the knockout and reintegrant. The present study demonstrates that Csh1p contributes to virulence of C. albicans in mice, but other gene products also contribute to the CSH phenotype and virulence.     

Long-term influence of the presence of a non-aqueous phase on the cell surface hydrophobicity of Pseudomonas in two-phase partitioning bioreactors

The long-term influence of silicone oil 200 cSt (SO200) and 2, 2, 4, 4, 6, 8, 8-heptamethylnonane (HMN) on the cell surface hydrophobicity (CSH) of a hexane-degrading Pseudomonas aeruginosa strain and a toluene-degrading Pseudomonas putida strain was assessed in two-phase partitioning bioreactors under batch and continuous operation. CSH was evaluated using a modified BATH method based on optical density (CSH_OD) and colony-forming unit (CSH_CFU) measurements. In the presence of HMN, P. aeruginosa turned hydrophobic over the time course as shown by the gradual increase in CSH_OD (61 ± 1%) and CSH_CFU (53 ± 3%) under batch degradation and in CSH_OD (49 ± 0%) under continuous operation. However, P. putida turned hydrophobic only under continuous operation (CSH_OD = 28 ±2% {\hbox{CS}}{{\hbox{H}}_{\rm{OD}}} = 28 \pm 2\% ). On the other hand, no significant CSH enhancement was observed in both Pseudomonas strains in the presence of SO200. These results suggested that CSH is species, non-aqueous phase, and cultivation mode dependant, and an inducible property of bacteria. Maximum hexane elimination capacities increased by 2 and 3 in the presence of SO200 and HMN, respectively. Based on the absence of CSH in P. aeruginosa in the presence of SO200, the higher elimination capacities recorded were likely due to an improved hexane mass transfer (physical effect). However, in the presence of HMN, a direct hexane uptake from the non-aqueous phase (biological effect) might have also contributed to this enhancement.     

氯氰菊酯降解菌株L12的分离鉴定及降解特性

[目的]通过分离纯化氯氰菊酯降解菌,研究降解特性为实际应用提供理论依据.[方法]利用富集驯化培养技术分离菌株,通过形态、生理生化特征及16S rRNA基因序列分析鉴定菌株.测定培养液中氯氰菊酯的浓度和代谢产物以及菌体细胞的密度和细胞表面疏水性,研究菌株的降解特性.[结果]从生产氯氰菊酯的农药厂污水曝气池中,分离到1株能...     

Alteration of cell cycle progression in human leukemia cell line (KOPM-28) induced by 12-o-tetradecanoylphorbol-13-acetate

Terminal cell differentiation usually results in an irreversible arrest in the G1 phase of the cell cycle and loss of cell renewal ability. Human promyelocytic leukemia HL-60 cells induced with 12-o-tetradecanoylphorbol-13-acetate (TPA) differentiate into monocytes/macrophages and accumulate in G1. We determined the effect of TPA on the growth kinetics of a human leukemia cell line (KOPM-28), which developed several of the characteristics of megakaryocytes in response to TPA, such as the surface antigen complex IIb/IIIa, platelet peroxidase and polyploidy. Cell growth was immediately and completely inhibited by TPA. Flow cytometric analysis of cellular DNA content revealed a gradual decrease in cells in G1 and an accumulation of cells in G2. These data suggest that TPA prolonged G1 and rapidly arrested the cells in G2. Synchronized cells were utilized to further analyze the rapid G2 arrest. Cells arrested with aphidicolin at the G1/S interphase were released, and the effects of TPA (added at different intervals) on cell cycle progression were examined 14 h after release. The results showed that TPA added at the end of the S phase, as well as at the G1/S interphase incompletely but distinctly arrested cells in G2. Moreover, G2 arrest was observed when TPA was added to cells released from a colcemid-induced G2/M block, suggesting that cells already in G2 were inhibited by TPA from moving through M to G1. Since some cells became multi-nucleated in the course of incubation with TPA, this G2 accumulation may have resulted at least in part from a prolongation of the phase or a transient G2 block. These changes in cell cycle progression induced by TPA may be characteristic of and/or related to megakaryocytic differentiation of hemopoietic precursor cells.