Budding Mode Research Articles

Physiological and metabolic insights into the first cultured anaerobic representative of deep-sea Planctomycetes bacteria.

Planctomycetes bacteria are ubiquitously distributed across various biospheres and play key roles in global element cycles. However, few deep-sea Planctomycetes members have been cultivated, limiting our understanding of Planctomycetes in the deep biosphere. Here, we have successfully cultured a novel strain of Planctomycetes (strain ZRK32) from a deep-sea cold seep sediment. Our genomic, physiological, and phylogenetic analyses indicate that strain ZRK32 is a novel species, which we propose be named: Poriferisphaera heterotrophicis. We show that strain ZRK32 replicates using a budding mode of division. Based on the combined results from growth assays and transcriptomic analyses, we found that rich nutrients, or supplementation with NO3- or NH4+ promoted the growth of strain ZRK32 by facilitating energy production through the tricarboxylic acid cycle and the Embden-Meyerhof-Parnas glycolysis pathway. Moreover, supplementation with NO3- or NH4+ induced strain ZRK32 to release a bacteriophage in a chronic manner, without host cell lysis. This bacteriophage then enabled strain ZRK32, and another marine bacterium that we studied, to metabolize nitrogen through the function of auxiliary metabolic genes. Overall, these findings expand our understanding of deep-sea Planctomycetes bacteria, while highlighting their ability to metabolize nitrogen when reprogrammed by chronic viruses.

Physiological and metabolic insights into the first cultured anaerobic representative of deep-sea Planctomycetes bacteria

Planctomycetes bacteria are ubiquitously distributed across various biospheres and play key roles in global element cycles. However, few deep-sea Planctomycetes members have been cultivated, limiting our understanding of Planctomycetes in the deep biosphere. Here, we have successfully cultured a novel strain of Planctomycetes (strain ZRK32) from a deep-sea cold seep sediment. Our genomic, physiological, and phylogenetic analyses indicate that strain ZRK32 is a novel species, which we propose be named: Poriferisphaera heterotrophicis. We show that strain ZRK32 replicates using a budding mode of division. Based on the combined results from growth assays and transcriptomic analyses, we found that rich nutrients, or supplementation with NO3- or NH4+ promoted the growth of strain ZRK32 by facilitating energy production through the tricarboxylic acid cycle and the Embden-Meyerhof-Parnas glycolysis pathway. Moreover, supplementation with NO3- or NH4+ induced strain ZRK32 to release a bacteriophage in a chronic manner, without host cell lysis. This bacteriophage then enabled strain ZRK32, and another marine bacterium that we studied, to metabolize nitrogen through the function of auxiliary metabolic genes. Overall, these findings expand our understanding of deep-sea Planctomycetes bacteria, while highlighting their ability to metabolize nitrogen when reprogrammed by chronic viruses.

Cryptic growth strategies of the Cambrian coralCambroctoconus: flexible modes of budding and growth in immediate response to available space

AbstractThe Cambrian coralCambroctoconusoccurs selectively in the crypts of calcimicrobeEpiphytonreefs of the Zhangxia Formation (Miaolingian) in Shandong Province, North China. These cryptobionts preferentially grow laterally and/or downward, in some cases showing pendent growth from the ceilings of the reef framework. The upside‐down growth is clearly shown by the downward‐oriented aperture and the presence of fork‐like holdfasts at its base. The coral aggregations reveal two cryptic growth strategies: one involves the prevalence of far smaller individuals in vertically narrow spaces; the other involves the predominance of modular individuals in large cavernous spaces. Offset individuals frequently appear projecting downward and are connected to each other in a chain‐like fashion. Individuals often undergo rejuvenescence laterally and downward, when the growth directions may change. Therefore, the size and modularity of individuals and the direction of budding depend largely upon the cryptic microenvironments available at the time. The cryptobionts make the best use of vacant spaces by modifying the location of budding and altering their growth directions through rejuvenescence. The cryptic growth strategies found inCambroctoconusindicate a certain degree of morphological variability in the modules. More importantly, they indicate that the individuals flexibly and exquisitely utilized the microbial crypts that were still predominant even during the middle Cambrian. These modes of cryptic growth demonstrate the age‐specific exploitation of niches by sessile skeletal organisms that was facilitated by the development of a firm attachment structure.

Novel budding mode in Polyandrocarpa zorritensis: a model for comparative studies on asexual development and whole body regeneration

BackgroundIn tunicates, the capacity to build an adult body via non-embryonic development (NED), i.e., asexual budding and whole body regeneration, has been gained or lost several times across the whole subphylum. A recent phylogeny of the family Styelidae revealed an independent acquisition of NED in the colonial species Polyandrocarpa zorritensis and highlighted a novel budding mode. In this paper, we provide the first detailed characterization of the asexual life cycle of P. zorritensis.ResultsBud formation occurs along a tubular protrusion of the adult epidermis, the stolon, in a vascularized area defined as budding nest. The bud arises through a folding of the epithelia of the stolon with the contribution of undifferentiated mesenchymal cells. This previously unreported mode of bud onset leads to the formation of a double vesicle, which starts to develop into a zooid through morphogenetic mechanisms common to other Styelidae. The budding nest can also continue to accumulate nutrients and develop into a round-shaped structure, designated as spherule, which represents a dormant form able to survive low temperatures.ConclusionsTo understand the mechanisms of NED and their evolution, it is fundamental to start from a robust phylogenetic framework in order to select relevant species to compare. The anatomical description of P. zorritensis NED provides the foundation for future comparative studies on plasticity of budding and regeneration in tunicates.

Ecological and demographic correlates of cooperation from individual to budding dispersal.

Identifying the ecological and demographic factors that promote the evolution of cooperation is a major challenge for evolutionary biologists. Explanations for the adaptive evolution of cooperation seek to determine which factors make reproduction in cooperative groups more favourable than independent breeding or other selfish strategies. A vast majority of the hypotheses posit that cooperative groups emerge in the context of philopatry, high costs of dispersal, high population density and environmental stability. This route to cooperation, however, fails to explain a growing body of empirical evidence in which cooperation is not associated with one or more of these predictors. We propose an alternative evolutionary path towards the emergence of cooperation that accounts for the disparities observed in the current literature. We find that when dispersal is mediated by a group mode of dispersal, commonly termed budding dispersal, our mathematical model reveals an association between cooperation and immigration, lower costs of dispersal, low population density and environmental variability. Furthermore, by studying the continuum from the individual to the partial and full budding mode of dispersal, we can explicitly explain why the correlates of cooperation change under budding. This enables us to outline a general model for the evolution of cooperation that accounts for a substantial amount of empirical evidence. Our results suggest that natural selection may have favoured two major contrasting pathways for the evolution of cooperation depending on a set of key ecological and demographic factors.

Evolution of Complex Asexual Reproductive Strategies in Jellyfish.

Many living organisms in terrestrial and aquatic ecosystems rely on multiple reproductive strategies to reduce the risk of extinction in variable environments. Examples are provided by the polyp stage of several bloom-forming jellyfish species, which can reproduce asexually using different budding strategies. These strategies broadly fall into three categories: (1) fast localized reproduction, (2) dormant cysts, or (3) motile and dispersing buds. Similar functional strategies are also present in other groups of species. However, mechanisms leading to the evolution of this rich reproductive diversity are yet to be clarified. Here we model how risk of local population extinction and differential fitness of alternative modes of asexual reproduction could drive the evolution of multiple reproductive modes as seen in jellyfish polyps. Depending on environmental parameters, we find that evolution leads to a unique evolutionarily stable strategy, wherein multiple reproductive strategies generally coexist. As the extinction risk increases, this strategy shifts from a pure budding mode to a dual strategy and finally to one characterized by allocation into all three modes. We identify relative fitness-dependent thresholds in extinction risk where these transitions can occur and discuss our predictions in light of observations on polyp reproduction in laboratory and natural systems.

Tumor budding is an independent prognostic marker in early stage oral squamous cell carcinoma: With special reference to the mode of invasion and worst pattern of invasion

Pathologically proven regional lymph node metastasis affects the prognosis in early stage oral cancer. Therefore we investigated invasive tumor patterns predicting nodal involvement and survival in patients with clinically node-negative T1 and T2 oral squamous cell carcinoma (cT1,2N0M0 OSCC). Ninety-one cases of cT1,2N0M0 OSCC treated with transoral resection of the primary tumor were assessed based on 3 types of invasive tumor patterns on histopathologic and pancytokeratin-stained immunohistological sections: the mode of invasion, worst pattern of invasion (WPOI), and tumor budding. The correlations among invasive tumor patterns, regional metastasis, and disease-free survival were analyzed. Of the 91 cases, 22 (24%) had pathologically proven regional metastasis. The mode of invasion (p<0.01) and tumor budding (p<0.01) were associated with regional metastasis as well as lymphovascular invasion (p = 0.04) in univariate analysis. In logistic regression analysis, however, tumor budding was the only independent predictor of regional metastasis (hazard ratio (HR) = 3.05, 95% confidence interval (CI) = 0.29–5.30, p<0.01). All three invasive patterns, the mode of invasion, WPOI, and tumor budding, were found to be significant predictors of 5-year disease-free survival (p<0.01, p = 0.03, and p<0.01, respectively) as well as lymphovascular invasion (p = 0.02) and perineural invasion (p = 0.02). A final model for Cox multivariate analysis identified the prognostic advantage of the intensity of tumor budding (HR = 2.19, 95% CI = 1.51–3.18, p<0.01) compared with the mode of invasion and WPOI in disease-free survival. Our results indicate that the intensity of tumor budding may be a novel diagnostic biomarker, as well as a therapeutic tool, for regional metastasis in patients with cT1,2N0M0 OSCC. If the pancytokeratin-based immunohistochemical features of more than five buds, and a grade 4C or 4D mode of invasion are identified, careful wait-and-see follow-up in a short period with the use of imaging modalities is desirable. If there are more than ten buds, a grade 4D mode of invasion, or WPOI-5 in the same section, wide resection of the primary tumor with elective neck dissection should be recommended.

Environmental control of asexual reproduction and somatic growth of Aurelia spp. (Cnidaria, Scyphozoa) polyps from the Adriatic Sea.

Polyps of two moon jellyfish species, Aurelia coerulea and A. relicta, from two Adriatic Sea coastal habitats were incubated under multiple combinations of temperature (14, 21°C), salinity (24, 37 ppt) and food regime (9.3, 18.6, 27.9 μg C ind−1 week−1) to comparatively assess how these factors may influence major asexual reproduction processes in the two species. Both species exhibited a shared pattern of budding mode (Directly Budded Polyps: DBP; Stolonal Budded Polyps: SBP), with DBP favoured under low food supply (9.3 μg C ind −1 week−1) and low temperature (14°C), and SBP dominant under high temperature (21°C). However, A. coerulea showed an overall higher productivity than A. relicta, in terms of budding and podocyst production rates. Further, A. coerulea exhibited a wide physiological plasticity across different temperatures and salinities as typical adaptation to ecological features of transitional coastal habitats. This may support the hypothesis that the invasion of A. coerulea across coastal habitats worldwide has been driven by shellfish aquaculture, with scyphistoma polyps and resting stages commonly found on bivalve shells. On the contrary, A. relicta appears to be strongly stenovalent, with cold, marine environmental optimal preferences (salinity 37 ppt, T ranging 14–19°C), corroborating the hypothesis of endemicity within the highly peculiar habitat of the Mljet lake. By exposing A. relicta polyps to slightly higher temperature (21°C), a previously unknown developmental mode was observed, by the sessile polyp regressing into a dispersive, temporarily unattached and tentacle-less, non-feeding stage. This may allow A. relicta polyps to escape climatic anomalies associated to warming of surface layers and deepening of isotherms, by moving into deeper, colder layers. Overall, investigations on species-specific eco-physiological and ontogenetic potentials of polyp stages may contribute to clarify the biogeographic distribution of jellyfish and the phylogenetic relationships among evolutionary related sister clades.

Research on the Method of Selecting Construction Participating Companies for the Owner in a Project under Bidding Mode in China

Construction participating companies play very important roles in the project for they undertake the constructing part of construction projects. Therefore, selecting a good company is the prerequisite to accomplish project goals including project process, quality and investment, etc. Based on analyzing the basic rules of how an owner can select construction companies under budding mode, the paper presents the specific methods for the owner to select the companies for a construction project including management service company, exploration design company, construction company, and subcontractors in construction industry in China.

Research on the Method of Selecting Construction Participating Companies for the Owner in a Project under Bidding Mode in China

Construction participating companies play very important roles in the project for they undertake the constructing part of construction projects. Therefore, selecting a good company is the prerequisite to accomplish project goals including project process, quality and investment, etc. Based on analyzing the basic rules of how an owner can select construction companies under budding mode, the paper presents the specific methods for the owner to select the companies for a construction project including management service company, exploration design company, construction company, and subcontractors in construction industry in China.

Simple mathematical models do not accurately predict early SIV dynamics.

Upon infection of a new host, human immunodeficiency virus (HIV) replicates in the mucosal tissues and is generally undetectable in circulation for 1–2 weeks post-infection. Several interventions against HIV including vaccines and antiretroviral prophylaxis target virus replication at this earliest stage of infection. Mathematical models have been used to understand how HIV spreads from mucosal tissues systemically and what impact vaccination and/or antiretroviral prophylaxis has on viral eradication. Because predictions of such models have been rarely compared to experimental data, it remains unclear which processes included in these models are critical for predicting early HIV dynamics. Here we modified the “standard” mathematical model of HIV infection to include two populations of infected cells: cells that are actively producing the virus and cells that are transitioning into virus production mode. We evaluated the effects of several poorly known parameters on infection outcomes in this model and compared model predictions to experimental data on infection of non-human primates with variable doses of simian immunodifficiency virus (SIV). First, we found that the mode of virus production by infected cells (budding vs. bursting) has a minimal impact on the early virus dynamics for a wide range of model parameters, as long as the parameters are constrained to provide the observed rate of SIV load increase in the blood of infected animals. Interestingly and in contrast with previous results, we found that the bursting mode of virus production generally results in a higher probability of viral extinction than the budding mode of virus production. Second, this mathematical model was not able to accurately describe the change in experimentally determined probability of host infection with increasing viral doses. Third and finally, the model was also unable to accurately explain the decline in the time to virus detection with increasing viral dose. These results suggest that, in order to appropriately model early HIV/SIV dynamics, additional factors must be considered in the model development. These may include variability in monkey susceptibility to infection, within-host competition between different viruses for target cells at the initial site of virus replication in the mucosa, innate immune response, and possibly the inclusion of several different tissue compartments. The sobering news is that while an increase in model complexity is needed to explain the available experimental data, testing and rejection of more complex models may require more quantitative data than is currently available.

Mycological Characteristics of Nine Unrecorded Yeasts from Flowers in the Orchard of Yesan-gun, Chungcheongnam-do and Hanbat Arboretum in Daejeon City, Korea

Six unrecorded yeasts, Cryptococcus festucosus 41-3, Cryptococcus heveanensis 56-4, Debaryomyces nepalensis 95- 4, Issatchenkia occidentalis 142-1, Dioszegia zsoltii 39-1, and Kwoniella europala 47-2 were screened from 108 yeasts isolated from flowers and fruits in orchards of Yesan-gun, Chungcheongnam-do, Korea. The morphological and cultural characteristics of these unrecorded yeasts were investigated. They had various shapes, including ellipsoidal, globose, and oval, and also had budding mode in vegetable reproduction, except I. occidentalis 142-1 (fission mode). K. europaea 47-2 only formed pseudomy- celium. D. zsoliti 39-1 did not grow in yeast extract-malt extract medium, potato dextrose medium, and vitamin-free medium. C. festucosus 41-3 grew well in 5% NaCl-containing yeast extract-peptone-dextrose medium and had a growth pH range of 7.0~ 10.0. Three unrecorded yeasts Ogataea polymorpha HB45-1, Rhodotonula hinnulla HB62-2, and Cryptococcus rajasthanensis HB80-4 were screened from 51 yeasts isolated from flowers in Hanbat arboretum in Daejeon city, Korea. They were globose in shape and did not form pseudomycelium. In addition, O. polymorpha HB45-1 and C. rajasthanensis HB80-4 had budding mode in vegetable reproduction. All of them grew well in vitamin-free medium and C. rajasthanesis HB80-4 also grew in 50% glucose and 5% NaCl-containing YPD medium.

Type I Interferon Imposes a TSG101/ISG15 Checkpoint at the Golgi for Glycoprotein Trafficking during Influenza Virus Infection

Several enveloped viruses exploit host pathways, such as the cellular endosomal sorting complex required for transport (ESCRT) machinery, for their assembly and release. The influenza A virus (IAV) matrix protein binds to the ESCRT-I complex, although the involvement of early ESCRT proteins such as Tsg101 in IAV trafficking remain to be established. We find that Tsg101 can facilitate IAV trafficking, but this is effectively restricted by the interferon (IFN)-stimulated protein ISG15. Cytosol from type I IFN-treated cells abolished IAV hemagglutinin (HA) transport to the cell surface in infected semi-intact cells. This inhibition required Tsg101 and could be relieved with deISGylases. Tsg101 is itself ISGylated in IFN-treated cells. Upon infection, intact Tsg101-deficient cells obtained by CRISPR-Cas9 genome editing were defective in the surface display of HA and for infectious virion release. These data support the IFN-induced generation of a Tsg101- and ISG15-dependent checkpoint in the secretory pathway that compromises influenza virus release.

Probable advanced hydroid from the Early Ordovician of China

A fossil of a colonial organism with pyritized soft tissues of elongated fusiform zooids, found in the middle part of the early Floian Fenxiang Formation in Hubei Province of China, probably represents the oldest reliable record of a hydroid cnidarian. The preservation of the fossil is sufficiently different from that of associated carbonized skeletons of graptolites to exclude affinities with this group. The fossil is unlikely to be a bryozoan because of the mode of budding from proximal, not distal, parts of parent zooids, which is typical rather of hydroids. Although no thecae are preserved, the fossil, named Sino- bryon elongatum gen. et sp. nov., is suggested to be a thecate hydroid, possibly related to the Haleciidae. The apparent presence of an advanced member of the thec- aphoran Macrocolonia clade in strata 470 Ma old means that much of the hydroid (and cnidarian) diversification preceded the Middle Ordovician.

Plastids division in shoot apical meristem during the tuberization of taro (Colocasia esculenta)

This paper reports the first unambiguous observation of the differentiation of proplastids to amyloplasts in shoot apical meristem during the tuberization of taro. The differentiation mechanism from proplastids to amyloplasts was examined during tuberization of taro. Shoot tips during in vitro tuberization of taro were sampled and observed every 2 days using a transmission electron microscope. The enlargement of proplastids, accumulation of thylakoids and grana were observed in the cells of shoot apical meristem. After 8 days of the culture, starch grains deposition was distinctly observed in the amyloplasts. Binary fission and budding mode of proplastid division were found at the beginning of tuberization in taro. However, binary fission was the only division type shown by the amyloplasts.

Regularity and polarity in budding of the colonial scleractinian Dendrophyllia ehrenbergiana : consequences of radio-bilateral symmetry of the scleractinian body plan

Sentoku, A. & Ezaki, Y. 2012: Regularity and polarity in budding of the colonial scleractinian Dendrophyllia ehrenbergiana: consequences of radio-bilateral symmetry of the scleractinian body plan. Lethaia, Vol. 45, pp. 586–593. Regularities and polarity in budding of the azooxanthellate scleractinian Dendrophyllia ehrenbergiana were examined with the aim of understanding the developmental constraints on the formation of colonies. Its mode of budding, in light of the orientations of directive septa of offsets and the inclination angle of budding, is consistent with that of other dendrophyllids; however, the offsets of D. ehrenbergiana only occur near the two primary septa on the convex side of individual corallites, showing a plane of bilateral symmetry with a distinct polarity. These regularities and polarity are seen in the axial and its derived corallites throughout growth. Of note, the polarity at individual corallites is clearly reflected in subsequent colony growth by the branching pattern and corallite number. These characteristics imply the presence of radio-bilateral symmetrical constraints on the asexual reproduction of the Scleractinia and give us invaluable clues to the understanding of shape-making mechanisms of marine modular organisms. □Asexual reproduction, azooxanthellate coral, budding, colony, Dendrophyllia ehrenbergiana, polarity.

Regularity in budding mode and resultant growth morphology of the azooxanthellate colonial scleractinian Tubastraea coccinea

Scleractinia exhibit a variety of growth forms, whether zooxanthellate or azooxanthellate, according to factors that control asexual reproduction and ensuing coral growth. The azooxanthellate branching scleractinian Dendrophyllia arbuscula shows regular modes of budding in terms of the locations of budding sites, the orientations of directive septa, and the inclination angle of budding throughout colonial growth. This study reports that such regularities are also found in the apparently different growth form of the massive dendrophylliid Tubastraea coccinea, which shows the following growth features: (1) the offsets (lateral corallites) always occur near four primary septa, except the two directive primary septa, meaning that the lateral corallites do not appear in the sectors of the two directive septa; (2) the two directive septa in lateral corallites tend to be oriented subperpendicular to the growth direction of the parental corallites; (3) the lateral corallites grow approximately diagonally upwards; and (4) these regularities are seen in the axial and derived lateral corallites among all generations during colony growth. Large differences in growth form are found between the branching D. arbuscula and massive T. coccinea, irrespective of the presence of specific regularities. It is likely that subtle modifications of certain parameters (e.g., budding interval, branch length, corallite size, and inclination angle of lateral corallites) have a strong effect on the overall growth morphology. A precise understanding of such regularities, which occur regardless of generation or taxonomic position, would contribute to understanding the “shape-controlling mechanism” of corals, which are an archetypal modular organism.

Roseovarius halotolerans sp. nov., isolated from deep seawater

A Gram-reaction-negative, non-motile, aerobic bacterium, designated HJ50(T), was isolated from deep seawater of the East Sea, South Korea. Cells were ovoid to rod-shaped (0.5-0.8x1.3-3.0 mum), often with unequal ends, suggesting a budding mode of reproduction. The strain had an absolute requirement for sea salts and tolerated up to 20 % (w/v) sea salts. Propionate, dl-lactate, 2-ketogluconate, 3-hydroxybutyrate and rhamnose were used as growth substrates, but not mannitol, salicin, 4-hydroxybenzoate or acetate. The major fatty acid was summed feature 7 (C(18 : 1)omega7c/omega9t/omega12t) and the DNA G+C content was 59.0+/-0.1 mol%. Phylogenetic analyses based on 16S rRNA gene sequences showed that this strain was affiliated with the genus Roseovarius. Similarities between the 16S rRNA gene sequence of strain HJ50(T) (1430 nt) and those of type strains of members of the genus Roseovarius were 94.1-96.3 %. DNA-DNA relatedness values between strain HJ50(T) and the type strains of members of the genus Roseovarius were low (1.3-24.6 %). Physiological and biochemical differences support assignment of strain HJ50(T) to the genus Roseovarius as a representative of a novel species. The name Roseovarius halotolerans sp. nov. is proposed, with HJ50(T) (=KCTC 22224(T) =LMG 24468(T)) as the type strain.

Nutrition induced pleomorphism and budding mode of reproduction in Deinococcus radiodurans

BackgroundMorphological adaptation is an important biological function of a microorganism to cope with its environment. Pleomorphism (to exist in a number of morphological forms) took centre stage in many discussions wherein a bacterium exhibits morphological transition and altered mode of reproduction in response to an environmental condition.FindingsTo strengthen the concept on pleomorphism in bacteria, we report here different cell morphologies of Deinococcus radiodurans in response to variation in nutrient concentration. From our studies we attempt primary evidence towards the presence of significant population of monomer cells of D. radiodurans in specific culture condition. In this report we also illustrate with scanning electron micrographs an unusual budding mode of reproduction in D. radiodurans which was not reported till date for this group of bacteria.ConclusionIn a holistic view the study reflects on bacterial shape (morphotypes) and the physiological adaptation to a particular nutrient environment. The discovery of budding mode of reproduction in Deinococcus will be of interest to microbiologists. It can serve as a model system to understand the mechanism of budding process at molecular level.

Larger foraminifera as a substratum for encrusting bryozoans (Late Oligocene, Tethyan Seaway, Iran)

Considering the diversity and abundance of larger foraminifera examined from a wide range of Late Oligocene to Early Miocene palaeoenvironments in the Tethyan Seaway, encrusting bryozoans make extremely little use of their tests as substratum. Significant encrustations by bryozoans were exclusively found on large (o c. 6 cm), undulating tests of Lepidocyclina spp., on which, however, a remarkable 34 taxa of encrusting bryozoans were recorded. This shallow-water fauna of Chattian age was analyzed in respect of the bryozoan taxa present, colony growth type, and mode of budding, colony size, as well as onset of reproduction. Taxic and morphological similarities between the fossil assemblage and modern faunas encrusting mobile substrata indicate a long history of bryozoans as part of the interstitial habitat, while the tests of certain larger foraminifera may have played a significant role in the evolution of shallow-water bryozoans by providing substrata for encrusting species in otherwise unfavorable environments.