Changes in microbiome composition during ontogeny and dispersal of the coral boring sponge Thoosa mismalolli

Bautista-Guerrero, Eric; Carballo, José Luis; Rodríguez Zaragoza, Fabián Alejandro; Goméz-Gil, Bruno; García-Gasca, Alejandra; Rodríguez-Troncoso, Alma P.; González-Castillo, Adrián

doi:10.1038/s41598-025-85622-x

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Published: 18 January 2025

Changes in microbiome composition during ontogeny and dispersal of the coral boring sponge Thoosa mismalolli

Eric Bautista-Guerrero¹,
José Luis Carballo^2,3,
Fabián Alejandro Rodríguez Zaragoza⁴,
Bruno Goméz-Gil⁵,
Alejandra García-Gasca⁶,
Alma P. Rodríguez-Troncoso¹ &
…
Adrián González-Castillo⁷

Scientific Reports volume 15, Article number: 2355 (2025) Cite this article

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Abstract

Dispersal is an important life history trait that plays a key role in the demography and evolution of species. We employed a combined approach of DNA sequencing and transmission electron microscopy to examine the changes in the microbiome during the ontogeny and dispersal of the coral-excavating sponge Thoosa mismalolli. The results show that sponge can acquired their associated bacteria via both vertical (VT) and horizontal transmission (HT). Adult sponges, brooding larvae, and early free-swimming sponge larvae harbor a similar high-diversity microbial assemblage, dominated by Proteobacteria and Chloroflexi, which change throughout the larval dispersal phase. Larvae collected offshore showed a reorganization of their microbiome with a significant reduction of the dominance of inherited bacteria (Proteobacteria and Chloroflexi), and an enrichment of environmentally derived bacteria taxa (Bacteroidetes, Tenericutes, and Firmicutes). TEM confirmed a substantial change in cell structure and microbial composition, attributed to symbionts’ massive phagocytosis. This research provides information on microbiome dynamics through the sponge ontogeny and sheds on their possible role in the dispersal capacity of their larvae.

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Introduction

Marine sponges (phylum Porifera) are one of the most primitive and early diverging metazoans¹ with 9,686 valid species to date² distributed in marine and non-marine benthic environments³. Particularly in tropical ecosystems (i.e., coral reefs), sponges contribute not only to their biological biomass but also become a component of the energy requirements via a pathway defined as the “sponge loop”^4,5.

Sponges are sessile filter feeders which take up microorganisms from seawater and digest them by phagocytosis. They often harbor a dense and diverse community of symbiotic microorganisms such as archaea, heterotrophic and autotrophic bacteria, unicellular algae, fungi, and viruses, representing up to 35–50% of the sponge’s total biomass^{6,7,8,9,10,11,12}. For this reason, sponges are often defined as “metaorganisms” or “holobionts”, which refer to the complex relationships with hosts and the multitude of symbiotic microorganisms that inhabit them. These microorganisms, with their genes and metabolites, are collectively known as the sponge’s “microbiome”¹³, whose communities perform multiple vital functions in the host, such as nutrition¹⁴, immunity¹⁵, chemical defense¹⁶, secondary metabolite production¹⁷, and enhance competitive capacity¹⁸. Meanwhile, symbionts also benefit from the host, who provides shelter, a nutrient-rich niche, and the ability to respond to environmental stressors¹³. The host-associated microbial communities mostly depend on the symbionts’ vertical transmission (VT); the heritage obtained from parent to offspring via gametes^19,20 and larvae²¹ or during asexual reproduction (fragmentation, budding, or production of gemmules)²². In addition, the sponges can acquire symbionts from the surrounding environment, defined as horizontal transmission (HT)^23,24. A transmission route is not exclusive since the holobiont results from a mixture of VT and HT strategies^14,25.

Species of the genus Thoosa Hancock, 1849, play a significant role in bioerosion within reef ecosystems across the Eastern Tropical Pacific^26,27. The excavating sponges (i.e. Thoosa mismalolli and T. calpulli Carballo, Cruz-Barraza & Gómez, 2004)²⁸ brood and release an unciliated hoplitomella larva, characterized by an atypical siliceous skeleton, unique among species of the suborder Thoosina. They are hermaphroditic and viviparous, and their larvae are released through the channels into the water column, reaching a full planktonic stage^29,30. Sponge larvae are typically classified as lecithotrophic larvae, which cannot feed on the water column and have limited swimming abilities³¹. However, Hoplitomella larvae can remain in the water column for extended periods in full oceanic conditions, thereby increasing their capacity to disperse^29,32.

We investigated the microbiome composition of the coral excavating sponge T. mismalolli, focusing on the patterns of microbiome diversity and variation over its ontogeny and during the planktonic larval phase through a combined approach based on DNA barcoding, 16S rRNA gene amplicon sequencing (NGS), and transmission electron microscopy (TEM).

Results

Taxonomic and molecular identification of sponge adults and larvae.

Adult sponges and larvae were morphologically identified as Thoosa mismalolli Carballo, Cruz-Barraza, and Gómez, 2004 (Class: Demospongiae, order: Tetractinellida, suborder: Thoosina, family: Thoosidae, genus: Thoosa). Adult sponges were found on the coral substrate only in the alpha stage. Their spiculation is typical of the genus. (i) Amphiasters with a verrucose surface. (ii) Oxyasters with smooth and microspined rays that have an elongated or irregular center at the junction, these are biradiate, triradiate, tetraradiate forms. (iii) Centrotylote oxea are smooth and/or spined, and a low proportion of slender tylostyles were found.

Hoplitomella larvae were globular, pale yellow, and 320–399 μm in diameter. The outer cover of the larvae was composed of a monolayer of monoaxonic flat discs combined with 9–11 long radial protrusions supported by internal slender styles (Figs. 1A, 2A,B). Molecular markers of the COI mtDNA gene confirmed a 100% nucleotide similarity between adults identified as T. mismalolli and all larvae collected inshore and offshore (1–3 km). Additionally, the topologies obtained with maximum likelihood (ML) analysis (suppl. Fig. S1) supported all hoplitomella larvae and adult sponges clustered together with sequences from two adult specimens of T. mismalolli deposited in the GenBank database (Accession number: MN587873, KU559628). Molecular and morphological data confirmed the identity of all specimens and supported the conclusion that adults and larvae belonged to the species T. mismalolli.

Transmission electron microscopy

Sponge adult and brooded larvae

TEM observations revealed a high abundance and multiple morphotypes of symbiotic bacteria within the mesohyl of parental sponges and inside hoplitomella larvae. Specifically, the mesohyl of the brooder sponge was homogeneously filled by dispersed collagen fibrils with freely and randomly distributed bacterial symbionts.

The larvae were found within the brood chambers (Fig. 1A), which exhibited well-developed follicular epithelia and nurse cells (Fig. 1B). Each larva possessed 6–9 underdeveloped radial protrusions, which consisted of small and globular protrusions supported by internal slender styles (Fig. 1A). The mesohyl of the progenitor was connected between monodiscs with the larvae, where we suspected the transfer of bacterial symbionts.

Anatomical and ultra-structural characteristics showed that hoplitomella larvae exhibit three distinct regions: the outer zone (OZ), middle zone (MZ), and inner zone (IZ) (Fig. 1B). The OZ (5–7 µm thick) is composed of a single layer of imbricate disk-shaped siliceous spicules (monoaxonic flat disc) oriented horizontally to the larva surface (Fig. 1B). Each spicule is surrounded by a complex reticulate meshwork of collagen fibrils (Fig. 1A,B) below which, are two fusiform microsclerocytes with long cytoplasmatic processes (like filopodia) that surround each monoaxonic flat disc (Fig. 1C). The cytoplasm of each microsclerocyte contains numerous smooth-surfaced vesicles, no phagosomes, endosomes with electro-transparent granules and a prominent large oval nucleus with double-membraned and some aggregations of heterochromatin (Fig. 1C). Symbiotic bacteria are not commonly found in this zone.

A high cellular density and an intercellular matrix rich in collagen fibrils characterize the MZ (13–15 µm thick). Additionally, this zone contains a distinctive peripheral stratified layer of gray cells with a triangular shape whose long axis is oriented horizontally to the larval surface. The cytoplasm of these cells contains numerous spherical to ellipsoidal electrodense yolk inclusions, spherical lipid droplets, dispersed small granules of glycogen, oval mitochondria, a well-developed Golgi complex, and a central and prominent nucleus with islands of tightly packed form of DNA heterochromatin (Fig. 1D). Beneath this peripheral layer are various types of cells that are overcrowded and contain large electro-transparent inclusions. These include abundant spherulous cells, large ameboid archaeocytes, and bacteriocytes (Fig. 1B,E). The spherulous cells were the most abundant in this region and arranged as a wide alveolar-like layer due to their numerous cytoplasmic electro-transparent inclusions that are empty. The cytoplasm of these cells contains abundant and dispersed granular glycogen, spherical lipid droplets, and some phagosomes with bacteria (Fig. 1E). Another type of cell present in MZ are archeocytes (Fig. 1E). Their cytoplasm contains osmophilic inclusions with variable contents, dispersed granular glycogen, some intravacuolar bacteria, small vesicles, and a spherical nucleus with double membrane surrounded by 5–6 well-developed Golgi complexes (Fig. 1F). Additionally, bacteriocytes are present deeper in the MZ (Fig. 1E), where a high abundance of intercellular symbiotic bacteria are also present (Fig. 1G). The cytoplasm of bacteriocytes contains a distinctive and subspherical nucleolate nucleus with a double membrane surrounded by multiples (3–5) well-developed Golgi complexes, as well as many distinctive vacuoles with electro-dense and electro-transparent zones, surrounded by abundant small heterogeneous and granular inclusions of glycogen (Fig. 1E). Their cytoplasm also includes numerous phagosomes with 1–3 bacteria of different morphotypes most of which, are well-preserved, while a smaller proportion shows evidence of degradation. Externally, the phagosomes are surrounded by numerous but dispersed cytoplasmatic glycogen granules (Fig. 1G). This is the earliest larval stage in which there is evidence of sponge cell phagocytosis of symbiotic bacteria.

The inner zone (IZ) contains a disrupted extracellular matrix and a loose network of collagen bundles. A distinctive and prominent cavity is lined by flattened pinacocytes, resembling a precociously formed choanocyte chamber (Fig. 1H). Numerous extracellular bacteria surround this cavity, some undergoing evident bipartition (Fig. 1H).

Pelagic swimming larvae

During this larval stage, the regional differentiation (OZ, MZ, and IZ) persists within the larval body. However, these larvae undergo significant modifications and cell migration, leading to the remodeling of all internal structure.

The OZ is characterized by a single layer of imbricate monoaxonic flat disc spicules and well-developed 9–11 long radial protrusions supported by internal slender styles. Each protrusion is entirely covered by a thin and translucent cellular layer of an organic matrix that widens at the distal end to form a paddle-like structure with a ribbed surface (Fig. 2A). Below each monoaxonic flat disc spicule, microsclerocytes are also present but in few numbers and withdrawn (Fig. 2B).

The MZ is disorganized and discontinuous, containing bundles of collagen fibrils and a low cell density. The distinctive peripheral stratified layer of gray cells is now discontinuous and less evident (Fig. 2B). Fusiform gray cells are present, but their density is lower than the MZ of brooded larvae. Some intercellular bacteria were directly engulfed by the remanent grey cells through the formation of well-developed pseudopodia and uptake into the cytoplasm via endocytosis (Fig. 2B). Their cytoplasm contains numerous spherical to ellipsoidal yolk inclusions, spherical lipid droplets, small, dispersed granules, oval mitochondria, a well-developed Golgi complex, and in some cases, a prominent nucleus filled with diffuse euchromatin (Fig. 2C). This characteristic suggests high synthetic activity. The remaining cells underwent a massive migration away from the surface or experienced apparent degeneration (Fig. 2D). The layer composed of spherulous cells resembling the wide alveolar structure is absent. These cells lose their cytoplasmic electro-transparent inclusions, leading to evident degeneration (Fig. 2D). The cells become overcrowded below the residual peripheral layer. Bacteriocytes densely populated the MZ’s deeper region, and archeocytes with variable cytoplasmic contents. Additionally, there was evidence that a highly abundant and diverse morphotype of symbiotic bacteria that can move freely into the extracellular space (Fig. 2E). Although at present, archaeocytes were less abundant than in brooded larvae. The cytoplasm contained osmophilic inclusions with variable contents, granular glycogen, intravacuolar bacteria undergoing digestion, small vesicles, and a spherical nucleus with a double membrane surrounded by 2–3 well-developed Golgi complexes. Bacteriocytes were also present in the MZ, where a high abundance of intercellular symbiotic bacteria was detected. Internally, their cytoplasm includes a distinctive and subspherical nucleus with diffuse euchromatin and a double membrane. It is surrounded by multiple (5–6) well-developed Golgi complexes (Fig. 2F), distinctive vacuoles with electro-dense and electro-transparent zones, as well as numerous and massive phagosomes with bacteria (8–10) in advanced digestion and dense inclusion with heterogeneous material. This may be the ending stage of digestive vacuoles, probably used to elaborate the yolk.

In addition to the inner zone (IZ), many bacteria were observed in the intercellular medium (Fig. 2G). Bacteriocytes and archaeocytes internalized these bacteria through a process of reorganization of the cytoplasmic membrane and the formation of multiple phagosomes. The cells exhibited high efficiency in engulfing several bacteria (Fig. 2H).

Microbial composition across life stages of T. mismalolli

We obtained 16S rRNA metagenome amplicon libraries from the different life stages of T. mismalolli, and the surrounding seawater to compare their bacterial community composition (libraries of ≈1.5 million sequence reads with lengths ≥ 280 bp). A total of 214,272 (218 to 253 bp) clean reads (sample depth ranging from 12,713 to 18,704) were analyzed from fifteen samples per sponge/larvae (149,151 reads) and six SW samples (71,581 reads). A total of 3,778 OTUs were recovered from parental adult sponges, larvae, and SW samples, which were assigned to the Bacteria domain at a 97% identity.

The count of unique OTUs varied from 221–223 OTUs associated to adult sponges, 178–491 OTUs for brooding larvae, 196–352 OTUs in early free-swimming larvae collected inshore, 403–512 OTUs in free-swimming pelagic larvae collected offshore to 1 km, 64–127 for free-swimming pelagic larvae collected offshore to 3 km, 647–703 OTUs in samples of SW inshore and 276–669 in SW offshore (Suppl. Table S1).

Taxonomic composition and relative abundance

The bacterial assemblage was composed of 29 non-redundant bacteria phyla, comprising 73 classes, 490 families, and 1,181 genera. In general, 96% of the bacterial diversity (i.e., taxonomic composition and relative abundance) was represented by seven phyla dominated mainly by Proteobacteria (49%), Cyanobacteria (18.6%), Chloroflexi (12.8%), Bacteroidetes (8.2%), Actinobacteria (4.3%), Firmicutes (1.9%) and Acidobacteria (1.3%), while other bacterial phyla accounted < 1% (Fig. 3A). At the class level, Alphaproteobacteria (23.8%) dominated the microbiome, followed by Gammaproteobacteria (22.2%), Oxyphotobacteria (16.3%), Dehalococcoidia (15.3%), Bacteroidia (8.8%), and Actinobacteria (2.7%). The other bacterial class accounted for < 2% (Fig. 3B). Sample-based rarefactions showed the bacterial Phyla observed richness had 84.5% average representativeness regarding the richness expected by chance, suggesting an adequate bacterial record (Suppl. Fig. S2).

The bacterial assemblage composition among adult sponges, larvae, and the SW samples differed notably at the phyla level (Fig. 3A). In adult sponges and brooding larvae, Chloroflexi was the dominant phyla, followed by Proteobacteria, Acidobacteria, and Actinobacteria (Suppl. Fig. S3A, B). Instead, in early free-swimming larvae collected inshore, Proteobacteria was the dominant phyla, followed by Chloroflexi (Suppl. Fig. S3C). Interestingly, the microbiome of free-swimming pelagic larvae collected offshore to 1 km (Suppl. Fig. S3D) and those found in SW samples collected inshore and offshore (Suppl. Fig. S3F-G) showed a high similarity in the dominant bacterial phyla composition. In these last larvae category and SW samples (inshore and offshore), Proteobacteria and Cyanobacteria were the dominant phyla. In contrast, in the microbiome of free-swimming pelagic larvae collected offshore to 3 km, there was a significant reduction in the dominance of Proteobacteria and Cyanobacteria, and evident depletion of Chloroflexi, Acidobacteria, Poribacteria, and other nine phyla were registered. Unexpectedly, these larvae found a remarkable species-specific enrichment in Bacteroidetes, Tenericutes, and Firmicutes (Suppl. Fig. S3E).

At the class level, we found that in adult sponges and brooding larvae, members of Dehalococcoidia (60%, 46.2%) were the main class, followed by members of the class Gammaproteobacteria (10.7%, 15.3%) and Alphaproteobacteria (9.5%, 16.4%), respectively (Suppl. Fig. S3A-B). Instead, Alphaproteobacteria were mainly found in the microbiome of early free-swimming larvae collected inshore (32.1%) and offshore to 1 km (39.4%). The same pattern was observed for the Gammaproteobacteria, highly represented in the microbiome of early free-swimming larvae collected inshore (19.9%) and low in larvae collected at 1 km (14%) (Suppl. Fig. S3C-D). Comparatively, Bacteroidia (54%) and Gammaproteobacteria were the dominant class in free-swimming pelagic larvae collected at 3 km (Suppl. Fig. S3E). Finally, Gammaproteobacteria and Oxyphotobacteria made up 63–45% of the microbial assemblage composition in the SW samples inshore (38.8%, 25.1%) and offshore (16.4%, 29.2%), respectively (Suppl. Fig. S3F-G).

The shade plot exhibited a clear aggrupation of bacterial phyla − considering their number of OTUs − found in all sponge life stages and SW samples (Fig. 4). The bacteria found in adult sponges clustered closer to the assemblage found in brooding larvae and early free-swimming larvae collected inshore. Members of the phyla Chloroflexi, Dadabacteria, and Nitrospirota integrated into the first subgroup, and phylum Proteobacteria and Actinobacteria conformed to the second subgroup. The bacterial assemblage structure was highly similar and clustered in two principal subgroups in free-swimming pelagic larvae collected offshore to 1 km and all SW samples (WIS and WOS). The first one was composed of members of the phyla Proteobacteria and Actinobacteria, while bacteria of the phylum Cyanobacteria mainly integrated the second one, SAR406_Clade and Eremiobacteraeota (Fig. 4). In contrast, the bacterial assemblages of free-swimming pelagic larvae collected offshore to 3 km exhibited clear and consistent shifts in composition, resulting in an independent cluster. The microbiome of free-swimming pelagic larvae collected offshore to 3 km was integrated by bacterial OTUs corresponding to the phyla Bacteroidetes, Tenericutes, and Synergistetes. In contrast, Proteobacteria, and Actinobacteria integrated the second one. Cyanobacteria were occasionally found in all samples of adults and larvae, with high dominance in SW samples. Overall, adult sponges, brooding larvae, and early free-swimming larvae collected inshore shared more OTUs than adult sponges and free-swimming pelagic larvae sampled offshore 1 and 3 km.

Bacteria alpha diversity

The number of bacterial phyla of adult sponges, brooding larvae, and early free-swimming larvae collected nearby showed similar bacterial richness (≥ 22 phyla). However, permutational ANOVA found significant differences (51.3% of total variance explained) among free-swimming pelagic larvae collected offshore to 3 km (12 phyla) regarding other sponge life stages (Suppl. Table S2, Fig. 5). In the SW samples collected in and offshore, 26 and 22 different phyla were found, respectively. Pairwise comparisons also showed significant differences between the average richness of bacterial phylum of free-swimming pelagic larvae collected offshore to 3 km versus bacterioplankton of SW inshore and offshore (Fig. 5). In contrast, the average number of OTUs and Shannon diversity did not show statistical differences among the brooding larvae, adult sponges, early free-swimming larvae collected inshore, and the free-swimming pelagic larvae collected offshore to 1 and 3 km, as well as between SW inshore and SW offshore (Suppl. Table S2, Fig. 5). All community metrics are fully described in suppl. Table S1.

Bacteria beta diversity

PERMANOVA revealed that bacterial assemblage structure differed significantly among life-stage grouping and SW samples, accounting for 63.7% of the total variation (Suppl. Table S2). Pairwise comparisons showed that bacterial assemblages of early free-swimming larvae collected inshore, free-swimming pelagic larvae collected offshore to 1 and 3 km, and SW samples were significantly different, except between adult sponges vs. brooding larvae and the bacterial assemblage of early free-swimming larvae collected inshore, free-swimming pelagic larvae collected offshore to 1 km vs. SW samples, and between seawater samples (Suppl. Table S3). SIMPER results showed bacteria phyla that mainly contributed to average dissimilarities among sponge life stages, and SW samples were Cyanobacteria, Chloroflexi, Bacteroidetes, Nitrospinota, Acidobacteria, Proteobacteria, Poribacteria, and Dadabacteria (Suppl. Table S4). PCO ordination showed an evident overlap among microbiome similarity from adult sponges, brooding larvae, and early larvae collected inshore (Fig. 6).

The bacteria phyla that contributed to these similarities were Chloroflexi, Acidobacteria, Poribacteria, Dadabacteria, Spirochaetes, and Nitrospinota. Interestingly, bacterial assemblage of free-swimming pelagic larvae collected offshore to 1 km were more similar to seawater samples in and offshore, and Cyanobacteria, Proteobacteria, and Actinobacteria were the phyla with a major contribution. In contrast, the bacterial community of free-swimming pelagic larvae collected offshore to 3 km exhibited different compositions that evidenced a strong separation of this group of larvae concerning all samples, resulting in an independent cluster, being Bacteroidetes and Firmicutes, the phyla with significant contribution (Fig. 6).

Finally, a specific analysis between adult sponges vs. all larval-stage revealed the presence of dominant symbiont OTUs in these groups (Fig. 8). Adult and brooding larvae harbor the phylum Chloroflexi (60.7% and 48.8%), which resulted as the most common symbiont in Thoosa mismalolli, except in free-swimming pelagic larvae collected offshore to 3 km. The same pattern was observed at the class level, where Dehalococcoidia was the most abundant bacteria Class in adult sponges (59.9%) and brooding larvae (46.2%). Interestingly, six minor families (< 10 sequences) of Chloroflexi were exclusively found in the adult sponges and shared with brooding larvae as well as early larvae collected inshore. Also, specific OTUs with low abundance matched with members of candidate phylum Poribacteria and were exclusively found in adult sponges (1.7%), brooding larvae (1.7%), and early free-swimming larvae collected inshore (0.2%) but not in free-swimming pelagic larvae collected offshore to 1 and 3 km (Fig. 7).

Discussion

The sponge Thoosa mismalolli hosts a highly diverse bacterial community, which undergoes significant structural changes throughout its life cycle. This microbiome comprises 29 non-redundant bacterial phyla, with Proteobacteria and Chloroflexi being the most dominant, followed by Bacteroidetes, Actinobacteria, and Cyanobacteria. Low abundance or unclassified lineages were also associated with the ‘rare biosphere’. Chloroflexi was the most abundant bacterial phylum (48–60%) at each life stage of T. mismalolli, similar to the observed in other high-microbial-abundance (HMA) sponges such as Xestospongia testudinaria, Hyrtios erectus, and Aaptos lobata^33,34,35.

In order of abundance, the more dominant clades were the candidate families SAR202 and Caldilineae (Chloroflexi), which, along with the candidate phylum Poribacteria, were exclusively shared between adult sponges and brooding larvae. These results are consistent with previous studies of T. mismalolli adults, which report Chloroflexi and Poribacteria as the dominant phyla in the microbiome of this sponge³⁶. Chloroflexi and Poribacteria are aerobic and heterotrophic bacteria that participate in the metabolizing of organic phosphorus and in producing energy through pathways such as glycolysis³⁷, the tricarboxylic acid (TCA) cycle, the pentose phosphate pathway (PPP), and the respiratory chain³⁸. The metabolic functions of these bacterial groups in T. mismalolli are currently unknown. However, given their cryptic sponge habits, it can be inferred that these bacterial phyla participate in the metabolic denitrification pathway during the adult and larval phase³⁶.

Notably, our study also found differences in the microbial assemblage composition between adult and larval stages. Specifically, Actinobacteria were more abundant in the microbiome of almost all larval stages (BL; 3.3%, LIS; 11.0%, LO1;5.6%, except in LO3;0.03%) compared to adult sponges (2.5%). Actinobacteria are crucial in providing nutrition during embryonic and larval stages and defending against potential predators by producing bioactive secondary metabolites with defensive properties^39,40.

Moreover, the bacterial assemblage transmitted to brooding and early free-swimming larvae is a subset of the maternal microbiome in the adult stage. The 16S rRNA gene data consistently demonstrated that the richness and composition of the bacterial assemblage were similar between adult sponges, brooding larvae, and early free-swimming larvae collected inshore. These findings corroborate the VT hypothesis and confirm the ability of T. mimalolli to inherit, in fidelity, a bacterial consortium that constitutes an essential part of its core microbiome.

Non-random patterns of active transmission of the complete core microbiome or selective passenger of specific symbionts across within generations from adult sponges have been observed in viviparous sponges such as Clathia prolifera, Halichondria bowerbanki⁴¹, Oscarella lobularis, Mycale laxissima^21,22,42 and in most HMA sponges^33,43,44,45. Apparently, the phagocytosis of bacteria by gametes⁴⁶, embryos or brooding larvae⁴⁷, specialized nurse cells⁴⁸, as well as a bi-parental mechanism of transmission intervened by “umbilici”¹⁹, are the principal transfer pathways by which symbionts are acquired by the germ line directly from maternal pool⁴⁹; therefore analogous pathways of selective symbiont transmission may also occur across generations in T. mismalolli between adult parents and their progeny, particularly during embryogenesis when the embryos are provided with a subset of maternal symbionts.

One of the most notable findings of this study was that the microbiome of free-swimming pelagic larvae collected 1 km offshore had an 85% similarity to the bacterioplankton in the surrounding seawater. From the holobiont’s perspective, recent research on coral and sponges suggests that holobionts can modulate their microbial composition, as a tolerance response to cope and resist or, in long-term acclimatize to stressful or fluctuating environmental conditions⁵⁰.

Indeed, the bacterial composition associated with adult sponges, brooding larvae, and early free-swimming larvae collected inshore differed significantly in richness and composition from those of free-swimming pelagic larvae collected 3 km offshore, far from their progenitor parents. During the dispersal life-phase of the larvae, the microbial community composition underwent significant changes in dominance and diversity. Additionally, there was a species-specific enrichment of four different phyla: Bacteroidetes, Proteobacteria, Tenericutes, and Firmicutes. In particular, the Bacteroidetes phylum represents the most abundant group of bacteria in marine environments, exhibiting a global distribution that closely follows that of Proteobacteria and Cyanobacteria. They have been observed in various locations, including in the water column in a free-floating state, attached to particles, and as frequent symbionts associated with sponges¹³. Bacteroidetes play a regulatory role in the development of marine invertebrates, triggering and enhancing the transition from swimming larvae to promoting their recruitment⁵¹. Members of the Bacteroidetes can migrate from the water column to the larvae, resulting in a notable increase in their abundance during the free-swimming stage. These bacteria function as competent larval-stage-specific associates, releasing hydrosoluble molecules and extracellular vesicles that regulate the morphogenetic processes⁵². The abrupt increase in the proportion of Bacteroidetes in the microbiome of free-swimming hoplitomella larvae may be indicative of the onset of their metamorphosis or even to enhance the settlement success of competent larvae.

It is interesting to note that the bacterial assemblages of the free-swimming pelagic larvae of T. mismalolli in the dispersal life-phase experienced an evident depletion in inherited bacteria such as Chloroflexi, Acidobacteria, Actinobacteria, Gemmatimonadetes, Poribacteria, Nitrospirota, and Spirochaetes compared with adult sponges, brooding larvae, and early free-swimming larvae collected in the same habitat. Microbial consumption of symbionts during the larval stage and their potential use as a nutrition source has been previously described via TEM and fluorescent in situ hybridization (FISH) in several marine larvae sponges such as Corticium candelabrum, Ircinia oros, and Amphimedon queenslandica^47,53,54. TEM observations throughout the different larval phases of T. mismalolli, provide compelling evidence that the hoplitomella larva is capable of phagocytosing its bacterial symbionts, which are vertically inherited via germline. Therefore, it can be hypothesized that 'during the initial phase of the larval stage, the inherited bacterial phyla (i.e. Chloroflexi and Poribacteria) can facilitate the translocation of nutrients, vitamin production, and nutrient utilization, as previously suggested in adult sponges^55,56.

Nevertheless, the phagocytosis of symbiotic bacteria observed during the larval dispersal phase of T. mismalolli, may be a pathway for the acquisition of additional energy or a response to expel microbes obtained through mixed patterns of vertical transmission and horizontal transmission, both mechanisms that enhance their survival and possibility to disperse and contribute to subsidiary recruitment in other niches. Previous research has documented the phagocytosis of symbionts during the larval stage, as well as the utilization of bacteria as a nutritional source by invertebrates, including polychaetes⁵⁷, echinoderms^58,59, crustaceans⁶⁰, and sponges, which have been shown to enhance the settlement success of competent larvae^{47,48,49,50,51,52,53}. However, further testing particularly applied to T. mismalolli is necessary to prove this hypothesis.

The regulation of symbiont composition in adult sponges and larvae is not yet well understood. Some studies have proposed host defense strategies, such as molecular mimicry⁶¹ and a repertoire of key genes associated with immune recognition and defense⁶². For instance, candidate mechanisms that modulate the molecular crosstalk between sponge hosts and symbiotic bacteria include the production of diverse types of eukaryotic-like ankyrin-repeat domains and tetratricopeptide repeat proteins⁶³, and autophagy-related proteins⁶⁴. We theorize that similar host-bacteria interactions may occur in T. mismalolli throughout their larval life stages.

Overall, this study represents the first comparative analysis of the bacterial assemblages associated with the excavating sponge Thoosa mismalolli (Suborder Thoosina), which is one of the most common and important borers of corals along the tropical Pacific Ocean^26,27. Our results reveal a high bacterial diversity in the microbiome of T. mismalolli, both in the adult sponge and the larval phase, comparable to that observed in HMA sponges⁴⁵.

While further studies are still needed, our TEM and molecular results (NGS) indicate variability in the microbiome throughout the different life stages of T. mismalolli and the evidence that the host shapes part of the bacterial assemblage during the planktonic phase and may contribute to their successful settlement. Nevertheless, further high-throughput metagenomics and transcriptomic studies are required to elucidate the metabolic pathways to utilize the maternal inheritance or indigenous bacterioplankton as a nutritional source for energy and the species-specific factors that shape symbiont communities and the molecular mechanisms of bacterial recognition in this holobiont sponge. This will facilitate a holistic understanding of the functional roles of these symbiotic associations in each ontogenetic phase of Thoosa mismalolli and other marine sponges.

Methods

Sampling locations

Samples were collected on the same day (July 2017) in the coral ecosystem at Isla Isabel National Park (IINP), Nayarit, Mexico (21°50′41.0" N, 105°53′02.2" W) (Fig. 8, top panel). IINP is an insular natural protected area with a shallow coral ecosystem and the presence of Pocilloporid and Poritid corals.

Adult sponges and brooding larvae collection

Branches of the coral Pocillopora spp. invaded by the excavating sponge, Thoosa mismalolli were collected by SCUBA diving and kept in plastic bags containing seawater on ice in a cooler until processing. Each coral branch was fragmented to extract the sponge tissue, manually dissected with an aseptic needle, and transferred to filtered seawater. The adult sponges brooding larvae were aseptically dissected with a sterile scalpel to extract mature larvae. All samples were promptly fixed in triplicate with ethanol (96%) for subsequent DNA extraction and in glutaraldehyde (2.5%) in seawater for transmission electron microscopy (TEM) analysis and stored at -20 °C until laboratory processing.

Planktonic larvae collection

Recently released larvae (early free-swimming larvae; EFL) were collected in the surrounding water near coral colonies invaded by T. mismalolli in Bahía Tiburoneros (core population; 21°50′41.0" N, 105°53′02.2" W). In addition, planktonic larvae in the dispersal process (free-swimming pelagic larvae; FPL) were collected at 1 km (21°50′ 4.34" N, 105°52′49.44" W) and 3 km (21°49′8.02" N, 105°52′17.41" W) offshore far away from the core population (Fig. 8, bottom panel). In all cases, the planktonic larvae were collected by three hauls performed at noon for 10 min, using two 22 cm diameter plankton nets with a mesh size of 150 μm. Samples were transferred to sealed plastic containers (1 L) and individually filtered through a 100-μm mesh net and subsequently observed under a microscope stereoscope (Zeiss, Oberkochen, Germany) to determine the presence of larvae. All larvae were individually transferred to Petri dishes filled with filtered seawater and fixed in 96% ethanol for morphological identification and DNA extraction.

Seawater collection

To analyze the bacterioplankton assemblage, a triplicate of seawater samples (SW) surrounding the coral colonies invaded by sponges was collected simultaneously to compare sponge symbionts with 1.0 L sterilized flasks. Additionally, 1 L of surface SW (n = 3) was collected offshore the reef. Samples were kept on ice and transported to the laboratory to be vacuum-filtered through two 0.2-μm pore size nitrocellulose filters, 47 mm in diameter (Millipore), and fixed in 96% ethanol for DNA extraction. Water samples were collected to compare the bacterioplankton of surrounding seawater with symbionts from adult sponges and larvae from each habitat (inshore and offshore).

Taxonomic and molecular identification of adult sponges and larvae

Taxonomical identification of adults and larvae was performed following the taxonomical criteria established by Carballo et al.²⁸ and Bautista-Guerrero et al.³⁰, respectively. To establish the nucleotide relationship among all samples (adults and hoplitomella larvae from different habitats) and to avoid misidentification with the sympatric Thoosa calpulli species, sequences of the mitochondrial gene encoding for the cytochrome c oxidase subunit 1 (COI mtDNA) were amplified and sequenced according to the barcode methodology as described in Carballo et al.²⁷. Briefly, species identity analysis was performed using the NCBI BLASTn, and once the identity was confirmed, a graphic representation of ML tree (suppl. Fig. S1) was reconstructed using 19 sequences (661 selected sites) of cox-1 mtDNA, MEGA v. X⁶⁵. All sequences were submitted to the GenBank nucleotide database with the following accession numbers: OK598105, OK598106, OK598107, OK598108, OK598109, OK598110, OK598111, OK598112, OK598113, OK598114, OK598115, OK598116, OK598117, OK598118, OK598119.

Transmission electron microscopy and scanning electron microscopy

Small pieces tissues of three adult sponges brooding larvae and three free-swimming pelagic larvae were immediately fixed in glutaraldehyde 2.5% in seawater and post-fixed in 2% osmium tetroxide in PBS for 30 min, followed by 20–25 min rinses in distilled water. All samples were rinsed in ethanol (100%) and embedded in Araldite resin for semi-thin and ultrathin sections. Ultrathin sections were stained with uranyl acetate, contrasted with lead citrate, and observed in a Zeiss EM 900 transmission electron microscope (TEM). For scanning electron microscopy (SEM), Small pieces of tissues of three adult sponges brooding larvae and ten larvae were fixed in glutaraldehyde 5%. All larvae and tissue of adult sponges were rinsed in ethanol, mounted on glass stubs, critical-point dried, and sputter-coated with gold–palladium before viewing with a scanning electron microscopy (SEM) a JEOL JSM-5300 operating at 15.0 kV (Akishima, Tokyo, Japan).

DNA extraction and next-generation sequencing analysis

DNA extraction was performed from 21 samples (individually), corresponding to three adult sponges (AD), three brooding larvae (BL), three early planktonic larvae collected inshore (LIS), three free-swimming pelagic larvae collected offshore to 1 km (LO1), and three free-swimming pelagic larvae collected offshore to 3 km (LO3) and their respective three seawater samples collected inshore (WIS) and three at offshore (WOS) from each site.

Total DNA was extracted using the Promega Wizard Genomic DNA Purification kit following the manufacturer’s instructions. The DNA concentration was measured using a Qubit 4 Fluorometer (Invitrogen, Thermo Fisher Scientific), and the quality was assessed in a 1% agarose gel electrophoresis. The DNA samples were preserved at -20 °C until subsequent sequencing analysis. The bacterial assemblages associated with each sample were estimated by sequencing the V3 region of the 16S rRNA gene. Briefly, PCR amplifications were done with diluted DNA (1:100) using the universal bacterial primers proposed by Huse et al.⁶⁶ 16S-V3_338f. (ACYCCTACGGGRGGCAGCAG) and 16S-V3_533r (TTACCGCGGCTGCTGGCAC) to generate amplicons of ∼230 bp. To amplify PCR products, 1 μL of 20 ng metagenomic DNA was used in a 11.5 μL total reaction volume containing 7.1 μL H₂O, 0.8 μL MgCl₂ (25 mM), 0.7 μL dNTP (2.5 mM), 0.13 μL forward primers (25 mM), 0.13 μL reverse primers (25 mM), and 1 μL HotStarTaq Plus Master Mix Kit (Qiagen, USA).

PCR protocol conditions were as follows: one cycle at 94 °C 5 min; then 30 cycles at 94 °C 1 min; 48 °C 1 min, 72 °C 1 min, and a final extension of one cycle at 72 °C 5 min. All positive PCR products were confirmed in a 2% (w/v) agarose gel electrophoresis and gel purified using the Wizard SV Gel and PCR Clean-Up System kit (Promega). Triplicate amplicon products of previous PCR reactions were pooled and were used to prepare the DNA library following the Ion Torrent library protocol using an Ion Torrent reagent kit V3 (2 × 300 pb) according to the manufacturer’s instructions. After normalizing all samples at a concentration of 2 nM, an equimolar pool of DNA was sequenced into a 316 Chip Kit with the Ion Torrent Personal Genome Machine system (PGM) (Life Technologies, USA). NGS sequencing was performed at Centro de Investigación en Alimentación y Desarrollo, A.C., Mazatlán, Sinaloa, México (CIAD).

16S rRNA gene sequencing analysis

FASTQ sequences were filtered to remove low-quality and polyclonal sequences with metagenomic accepted tools and pipelines (https://github.com/GenomicaMicrob/metagenomic_ pipeline). Briefly, all high-quality sequences were filtered and depleted of barcodes (https://github.com/GenomicaMicrob/pair- end_cleaner), small sequences (< 150 bp), bases with a Phred quality lower than Q20, de-replication with ambiguous bases (n's), and chimeric sequences were removed with chimera_detector v.0.1.1 (https://github.com/GenomicaMicrob/chimera_detector). The identification of operational taxonomic units (OTUs) was performed by clustering at 3% divergence (97% similarity) with mg_classifier v.1.7.0 pipeline (https://github.com/GenomicaMicrob/mg_classifier). Taxonomy assignment was performed against the SILVA database in the alignment step. No target sequences, such as singleton DNA, chloroplast, mitochondria, and Eukarya, were completely removed from the dataset. Thresholds to delimit prokaryotic taxonomic levels were based on the proposal of Yarza et al.⁶⁷.

Statistical analyses

Bacterial alpha-diversity was analyzed with sample-based rarefactions to contrast the total observed Phyla richness vs. expected richness estimated by non-parametric procedures as Chao2, Jackknife 1, and Jackknife 1, considering the incidence-based rarity of Phyla as unique (one record on all samples) and duplicates (two records in all samples). All rarefaction curves were built with 10,000 permutations. Likewise, the bacterial richness (S, at Phylum level), abundance (N), and Shannon diversity (H', nats) were estimated among all samples, considering the number of OTUs. The variation of these community metrics among sponge life stages and collected seawater was compared with a permutational one-way analysis of variance (ANOVA), based on the different sponge life stages (i.e., adult sponges [AD:], brooding larvae [BL], larvae collected inshore [LIS], free-swimming pelagic larvae collected offshore at 1 km [LO1], free-swimming pelagic larvae collected offshore at 3 km [LO3]) and collected seawater (inshore [WIS] and offshore [WOS]) as a factor. This permutational ANOVA was performed with Euclidean distance matrices and Anderson et al.⁶⁸ criteria. Statistical significance was tested with 10,000 permutations under an unrestricted permutation of the raw data method and the sum of squares type III (partial).

Bacterial beta-diversity analysis considered the relative abundance of OTUs and taxonomic composition changes at Phylum level plotted as stacked bar plots. Likewise, the most dominant bacterial Phylum was also assessed with ordinary dominance plots by sponge life stage and collected seawater. Bacterial dissimilitude among sponge life stages were also analyzed using different methods: 1) A shade plot was performed with a matrix of Phylum level composition and their number of OTUs. For bacterial Phyla (R mode) classification, a Whittaker association coefficient was estimated based on previous data pretreatment using a standardizing method (i.e., relative abundance percentages). In contrast, a Bray–Curtis similarity and a square-root transformation were used for the samples (Q mode) ordinated per sponge life stages. Group average linkage method was utilized in both R and Q modes classification analyses; 2) A permutational multidimensional analysis of variance (PERMANOVA) was used to compare the bacterial structure assemblage at Phylum level across the sponge life stages and seawater (inshore and offshore). PERMANOVA model was built with a pre-fourth-root data transformation and Bray–Curtis similarity, following the same permutational ANOVA experimental design; 3) A similarity percentage analysis (SIMPER) was used to estimate the contribution of Phyla towards average dissimilarity among pair groups by sponge life stages and collected seawater. The SIMPER results were presented based on the criteria de Cáceres et al.⁶⁹; 4) Besides, a principal coordinates analysis (PCO) was used to visualize the similarity relationships of bacterial assemblage among sponge life stages and seawater. SIMPER and PCO were built with the same PERMANOVA data pretreatment and resemblance coefficient. Rarefaction, boxplots, permutational ANOVA, stacked bar plots, ordinary dominance plots, PERMANOVA, SIMPER, and PCO were performed with PRIMER 7⁷⁰.

Data availability

Metagenome datasets (FASTQ files) that support the findings of this study have been deposited in the NCBI Sequence Read Archive under the BioProject accession No: PRJNA768530 (https://www.ncbi.nlm.nih.gov/bioproject/PRJNA768530/). The source data underlying Supplementary Figs. S1–S7, are provided as a Source Data file.

References

Dunn, C. et al. Broad phylogenomic sampling improves resolution of the animal tree of life. Nature 452, 745–749 (2008).
Article ADS CAS PubMed MATH Google Scholar
de Voogd, N. J. et al. World Porifera Database. Accessed at https://www.marinespecies.org/porifera on 2024-01-19. https://doi.org/10.14284/359 (2024).
Bell, J. J. The functional roles of marine sponges. Estuar. Coast. Shelf. Sci. 79, 341–353 (2008).
Article ADS MATH Google Scholar
De Goeij, J. M. et al. Surviving in a marine desert: The sponge loop retains resources within coral reefs. Science 342, 108–110 (2013).
Article ADS PubMed Google Scholar
Rix, L. et al. Reef sponges facilitate the transfer of coral-derived organic matter to their associated fauna via the sponge loop. Mar. Ecol. Prog. Ser. 589, 85–96 (2018).
Article ADS CAS MATH Google Scholar
Vacelet, J. & Donadey, C. Electron microscope study of the association between some sponges and bacteria. J. Exp. Mar. Biol. Ecol. 30, 301–314 (1977).
Article Google Scholar
Garson, M. J., Flowers, A. E., Webb, R. I., Charan, R. D. & McCaffrey, E. J. A. Sponge/dinoflagellate association in the haplosclerid sponge Haliclona sp.: Cellular origin of cytotoxic alkaloids by Percoll density gradient fractionation. Cell. Tissue Res. 293, 365–373 (1998).
Article CAS PubMed Google Scholar
Webster, N. S., Wilson, K. J., Blackall, L. L. & Hill, R. T. Phylogenetic diversity of bacteria associated with the marine sponge Rhopaloeides odorabile. Appl. Environ. Microbiol. 67, 434–444 (2001).
Article ADS CAS PubMed PubMed Central Google Scholar
Hentschel, U. et al. Microbial diversity of marine sponges. In Sponges (Porifera): Progress in Molecular and Subcellular Biology (ed. Mueller, W. E. G.) 59–88 (Springer, 2003).
Chapter MATH Google Scholar
Thacker, R. W. & Starnes, S. Host specificity of the symbiotic cyanobacterium Oscillatoria spongeliae in marine sponges, Dysidea spp.. Mar. Biol. 142, 643–648 (2003).
Article CAS MATH Google Scholar
Lee, O. O. et al. Pyrosequencing reveals highly diverse and species-specific microbial communities in sponges from the Red Sea. ISME J. 5, 650–664 (2011).
Article CAS PubMed MATH Google Scholar
Webster, N. S. & Thomas, T. The sponge hologenome. mBio 7, 10–1128 (2016).
Article MATH Google Scholar
Pita, L., Rix, L., Slaby, B. M., Franke, A. & Hentschel, U. The sponge holobiont in a changing ocean: From microbes to ecosystems. Microbiome 6, 1–18 (2018).
Article Google Scholar
Webster, N. S. & Taylor, M. W. Marine sponges and their microbial symbionts: love and other relationships. Environ. Microbiol. 14, 335–346 (2012).
Article CAS PubMed MATH Google Scholar
Hentschel, U., Piel, J., Degnan, S. M. & Taylor, M. W. Genomic insights into the marine sponge microbiome. Nat. Rev. Microbiol. 10, 641–654 (2012).
Article CAS PubMed Google Scholar
Lackner, G. et al. Insights into the lifestyle of uncultured bacterial natural product factories associated with marine sponges. Proc. Natl. Acad. Sci. USA 114, 347–356 (2017).
Article MATH Google Scholar
Schmidt, E. W., Obraztsova, A. Y., Davidson, S. K., Faulkner, D. J. & Haygood, M. G. Identification of the antifungal peptide-containing symbiont of the marine sponge Theonella swinhoei as a novel δ-proteobacterium, “Candidatus Entotheonella palauensis”. Mar. Biol. 136, 969–977 (2000).
Article CAS Google Scholar
Tang, S. L. et al. Bacteria associated with an encrusting sponge (Terpios hoshinota) and the corals partially covered by the sponge. Environ. Microbiol. 13, 1179–1191 (2011).
Article PubMed MATH Google Scholar
Kaye, H. R. Sexual reproduction in four Caribbean commercial sponges. II. Oogenesis and transfer of bacterial symbionts. Invertebr. Repro. Dev. 19, 13–24 (1991).
Article MATH Google Scholar
Baldacconi, R., Nonnis-Marzano, C., Gaino, E. & Corriero, G. Sexual reproduction, larval development and release in Spongia officinalis L. (Porifera, Demospongiae) from the Apulian coast. Mar. Biol. 152, 969–979 (2007).
Article Google Scholar
Ereskovsky, A. V. & Bouryesnault, N. Cleavage pattern in Oscarella species (Porifera, Demospongiae, Homoscleromorpha): transmission of maternal cells and symbiotic bacteria. J. Nat. Hist. 36, 1761–1775 (2002).
Article Google Scholar
Enticknap, J. J., Kelly, M., Peraud, O. & Hill, R. T. Characterization of a culturable alphaproteobacterial symbiont common to many marine sponges and evidence for vertical transmission via sponge larvae. Appl. Environ. Microbiol. 72, 3724–3732 (2006).
Article ADS CAS PubMed PubMed Central Google Scholar
Mohamed, N. M. et al. Diversity and quorum-sensing signal production of Proteobacteria associated with marine sponges. Environ. Microbiol. 10, 75–86 (2008).
Article CAS PubMed MATH Google Scholar
Turon, M., Cáliz, J., Garate, L., Casamayor, E. O. & Uriz, M. J. Showcasing the role of seawater in bacteria recruitment and microbiome stability in sponges. Sci. Rep. 8, 15201 (2018).
Article ADS PubMed PubMed Central Google Scholar
Thacker, R. W. & Freeman, C. J. Sponge-microbe symbioses: recent advances and new directions. Adv. Mar. Biol. 62, 57–111 (2012).
Article PubMed MATH Google Scholar
Carballo, J. L., Bautista-Guerrero, E., Nava-Bravo, H., Cruz-Barraza, J. A. & Chávez, J. A. Boring sponges, an increasing threat for coral reefs affected by bleaching events. Ecol. Evol. 3, 872–886 (2013).
Article PubMed PubMed Central Google Scholar
Carballo, J. L., Bautista-Guerrero, E., Cárdenas, P., Cruz-Barraza, J. A. & Aguilar-Camacho, J. M. Molecular and morphological data from Thoosidae in favour of the creation of a new suborder of Tetractinellida. Syst. Biodivers. 16, 512–521 (2018).
Article Google Scholar
Carballo, J. L., Cruz-Barraza, J. A. & Gómez, P. Taxonomy and description of clionaid sponges (Hadromerida, Clionaidae) from the Pacific Ocean of Mexico. Zool. J. Linn. Soc. 141, 353–387 (2004).
Article Google Scholar
Vacelet, J. Planktonic armoured propagules of the excavating sponge Alectona (Porifera: Demospongiae) are larvae: evidence from Alectona wallichii and A. mesatlantica sp. nov.. Mem. Qld. Mus. 44, 627–642 (1999).
Google Scholar
Bautista-Guerrero, E., Carballo, J. L. & Maldonado, M. Reproductive cycle of the coral-excavating sponge Thoosa mismalolli (Clionaidae) from Mexican Pacific coral reefs. Invertebr. Biol. 129, 285–296 (2010).
Article Google Scholar
Uriz, M. J., Turon, X. & Mariani, S. Ultrastructure and dispersal potential of sponge larvae: Tufted versus evenly ciliated parenchymellae. Mar. Ecol. 29, 280–329 (2008).
Article ADS Google Scholar
Trégouboff, G. Contribution à la connaissance des larves planctoniques d’éponges. Arch. Zool. Exp. Gén. 82, 357–399 (1942).
Google Scholar
Gloeckner, V. et al. The HMA-LMA dichotomy revisited: an electron microscopical survey of 56 sponge species. Biol. Bull. 227, 78–88 (2014).
Article PubMed MATH Google Scholar
Thomas, T. et al. Diversity, structure and convergent evolution of the global sponge microbiome. Nat. Commun. 7, 11870 (2016).
Article ADS CAS PubMed PubMed Central MATH Google Scholar
Moitinho-Silva, L. et al. Predicting the HMA-LMA status in marine sponges by machine learning. Front. Microbiol. 8, 258275 (2017).
Article Google Scholar
González-Castillo, A., Carballo, J. L. & Bautista-Guerrero, E. Genomics and phylogeny of the proposed phylum “Candidatus Poribacteria” associated with the excavating sponge Thoosa mismalolli. Antonie Van Leeuwenhoek 114, 2163–2174 (2021).
Article PubMed Google Scholar
Kamke, J. et al. Single-cell genomics reveals complex carbohydrate degradation patterns in poribacterial symbionts of marine sponges. ISME. J. 7, 2287–2300 (2013).
Article CAS PubMed PubMed Central MATH Google Scholar
Bayer, K., Schmitt, S. & Hentschel, U. Microbial nitrification in Mediterranean sponges: possible involvement of ammonium-oxidizing Betaproteobacteria. Museu. Nacional. 165–171 (2008).
Noyer, C., Casamayor, E. O. & Becerro, M. A. Environmental heterogeneity and microbial inheritance influence sponge-associated bacterial composition of Spongia lamella. Microb. Ecol. 68, 611–620 (2014).
Article ADS PubMed Google Scholar
Montalvo, N. F., Mohamed, N. M., Enticknap, J. J. & Hill, R. T. Novel actinobacteria from marine sponges. Antonie Van Leeuwenhoek 87, 29–36 (2005).
Article CAS PubMed Google Scholar
Sacristán-Soriano, O. et al. Ontogeny of symbiont community structure in two carotenoid-rich, viviparous marine sponges: comparison of microbiomes and analysis of culturable pigmented heterotrophic bacteria. Environ. Microbiol. Rep. 11, 249–261 (2019).
Article PubMed Google Scholar
Lévi, C. & Porte, A. Etude au microscope électronique de l’éponge Oscarella lobularis Schmidt et de sa larve amphiblastula. Cah. Biol. Mar. 3, 307–315 (1962).
MATH Google Scholar
Steger, D. et al. Diversity and mode of transmission of ammonia-oxidizing Archaea in marine sponges. Environ. Microbiol. 10, 1087–1094 (2008).
Article CAS PubMed MATH Google Scholar
Lee, O. O., Chui, P. Y., Wong, Y. H., Pawlik, J. R. & Qian, P. Y. Evidence for vertical transmission of bacterial symbionts from adult to embryo in the Caribbean Sponge Svenzea zeai. Appl. Environ. Microbiol. 75, 6147–6156 (2009).
Article ADS CAS PubMed PubMed Central Google Scholar
Webster, N. S. et al. Deep sequencing reveals exceptional diversity and modes of transmission for bacterial sponge symbionts. Environ. Microbiol. 12, 2070–2082 (2010).
Article MathSciNet CAS PubMed PubMed Central MATH Google Scholar
Usher, K. M., Sutton, D. C., Toze, S., Kuo, J. & Fromont, J. Inter-generational transmission of microbial symbionts in the marine sponge Chondrilla australiensis (Demospongiae). Mar. Fresh. Res. 56, 125–131 (2005).
Article Google Scholar
de Caralt, S., Uriz, M. J. & Wijffels, R. J. Vertical transmission and successive location of symbiotic bacteria during embryo development and larva formation in Corticium candelabrum (Porifera: Demospongiae). J. Mar. Biol. UK 87, 1693–1699 (2007).
Article Google Scholar
Sharp, K. H., Eam, B., Faulkner, D. J. & Haygood, M. G. Vertical transmission of diverse microbes in the tropical sponge Corticium sp. Appl. Environ. Microbiol. 73, 622–629 (2007).
Article ADS CAS PubMed MATH Google Scholar
Carrier, T. J. et al. Maternal provisioning of an obligate symbiont in a sponge. Ecol. Evol. 13, e10012 (2023).
Article PubMed PubMed Central MATH Google Scholar
Stévenne, C., Micha, M., Plumier, J. C. & Roberty, S. Corals and sponges under the light of the Holobiont concept: How microbiomes underpin our understanding of marine ecosystems. Front. Mar. Sci. 8, 698853 (2021).
Article Google Scholar
Wu, S., Ou, H., Liu, T., Wang, D. & Zhao, J. Structure and dynamics of microbiomes associated with the marine sponge Tedania sp during its life cycle. FEMS Microbiol. Ecol. 94, fiy055 (2018).
Article Google Scholar
Li, M. et al. Bacteroidetes bacteria, important players in the marine sponge larval development process. iScience 24, 102662 (2021).
Article ADS CAS PubMed PubMed Central Google Scholar
Ereskovsky, A. V. & Tokina, D. B. Morphology and fine structure of the swimming larvae of Ircinia oros (Porifera, Demospongiae, Dictyoceratida). Invertebr. Reprod. Dev. 45, 137–150 (2004).
Article Google Scholar
Fieth, R. A., Gauthier, M. E. A., Bayes, J., Green, K. M. & Degnan, S. M. Ontogenetic changes in the bacterial symbiont community of the tropical demosponge Amphimedon queenslandica: metamorphosis is a new beginning. Front. Mar. Sci. 3, 228 (2016).
Article Google Scholar
Thomas, T. et al. Functional genomic signatures of sponge bacteria reveal unique and shared features of symbiosis. ISME. J. 4, 1557–1567 (2010).
Article CAS PubMed MATH Google Scholar
Radax, R. et al. Metatranscriptomics of the marine sponge Geodia barretti: tackling phylogeny and function of its microbial community. Environ. Microbiol. 14, 1308–1324 (2012).
Article CAS PubMed MATH Google Scholar
Gosselin, L. A. & Qian, P. Y. Can bacterivory alone sustain larval development in the polychaete Hydroides elegans and the barnacle Balanus amphitrite?. Mar. Ecol. Prog. Ser. 161, 93–101 (1997).
Article ADS Google Scholar
Rivkin, R. B., Bosch, I., Pearse, J. S. & Lessard, E. J. Bacterivory: a novel feeding mode for asteroid larvae. Science 233, 1311–1314 (1986).
Article ADS CAS PubMed Google Scholar
Ayukai, T. Ingestion of ultraplankton by the planktonic larvae of the crown-of-thorns starfish, Acanthaster planci. Biol. Bull. 186, 90–100 (1994).
Article CAS PubMed Google Scholar
Nevejan, N. et al. Bacteria as food in aquaculture: do they make a difference?. Rev. Aquacult. 10, 180–212 (2018).
Article MATH Google Scholar
Hill, M. S. Production possibility frontiers in phototroph: heterotroph symbioses: trade-offs in allocating fixed carbon pools and the challenges these alternatives present for understanding the acquisition of intracellular habitats. Front. Microbiol. 5, 357 (2014).
Article PubMed PubMed Central MATH Google Scholar
Alex, A. & Antunes, A. Whole genome sequencing of the Symbiont Pseudovibrio sp. from the intertidal marine sponge Polymastia penicillus revealed a gene repertoire for host-switching permissive lifestyle. Genome. Biol. Evol. 7, 3022–3032 (2015).
Article CAS PubMed PubMed Central Google Scholar
Nguyen, M. T., Liu, M. & Thomas, T. Ankyrin-repeat proteins from sponge symbionts modulate amoebal phagocytosis. Mol. Ecol. 23, 1635–1645 (2014).
Article CAS PubMed Google Scholar
Virgin, H. W. & Levine, B. Autophagy genes immunity. Nat. Immunol. 10, 461–470 (2009).
Article CAS PubMed PubMed Central Google Scholar
Kumar, S., Stecher, G., Li, M., Knyaz, C. & Tamura, K. MEGA X: molecular evolutionary genetics analysis across computing platforms. Mol. Biol. Evol. 35, 1547 (2018).
Article CAS PubMed PubMed Central Google Scholar
Huse, S. M. et al. Exploring microbial diversity and taxonomy using SSU rRNA hypervariable tag sequencing. PLoS Genet. 4, 1–10 (2008).
Article MATH Google Scholar
Yarza, P. et al. Uniting the classification of cultured and uncultured bacteria and archaea using 16S rRNA gene sequences. Nat. Rev. Microbiol. 12, 635–645 (2014).
Article CAS PubMed MATH Google Scholar
Anderson, M. J., Gorely, R. N. & Clarke, K. R. PERMANOVA+ Primer: Guide to Software and Statistical Methods, 214. (PRIMER-E Ltd, 2008).
Cáceres, I., Ibarra-García, E. C., Ortiz, M., Ayón-Parente, M. & Rodríguez-Zaragoza, F. A. Effect of fisheries and benthic habitat on the ecological and functional diversity of fish at the Cayos Cochinos coral reefs (Honduras). Mar. Biodivers. 50, 9 (2020).
Article Google Scholar
Clarke, K. R. & Gorley, R. N. PRIMER v7: User Manual/Tutorial. PRIMER-E: Plymouth (2015).

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Acknowledgements

This work has been funded by the project ECOLOGIA LARVARIA Y DISPERSIÓN DE ALGUNAS ESPONJAS DESTRUCTORAS DE CORALES [SEP-CONACyT, grant No. CB-254806]. Through the Director’s office, the Institute of Marine Sciences and Limnology (ICML) from the National Autonomous University of México (UNAM), and the Centro Universitario de la Costa, Universidad de Guadalajara (UdeG-CuC), paid the article processing charge (APC), to grant open access for this article. We thank the staff at CONANP for logistic support. We are infinitely grateful Mateo Amillano Cisneros and Benjamín Yáñez Chávez for their logistic support in field samplings Julisa Enciso-Ibarra for her invaluable help in Sample Preparation for NGS, and Lourdes Palma Tirado (INB-UNAM) for the processing of the TEM images. The collection of the biological material was authorized by the Secretaría de Agricultura, Ganadería, Desarrollo Rural, Pesca y Alimentación (SAGARPA) who granted the permission PPF/DGOPA-296/17.

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Authors and Affiliations

Laboratorio de Ecología Marina, Centro de Investigaciones Costeras, Centro Universitario de la Costa, Universidad de Guadalajara, Av. Universidad No. 203 Del, Ixtapa, 48280, Puerto Vallarta, Jalisco, Mexico
Eric Bautista-Guerrero & Alma P. Rodríguez-Troncoso
Laboratorio de Biología Marina, Departamento de Zoología, Facultad de Biología, Universidad de Sevilla, Avenida Reina Mercedes s/n, Seville, Spain
José Luis Carballo
Instituto de Ciencias del Mar y Limnología, Universidad Nacional Autónoma de México (Unidad Académica Mazatlán), Avenida Joel Montes Camarena s/n, PO Box 811, 82000, Mazatlán, SIN, Mexico
José Luis Carballo
Laboratorio de Ecología Molecular, Microbiología y Taxonomía (LEMITAX), Departamento de Ecología Aplicada, Centro Universitario de Ciencias Biológicas y Agropecuarias, Universidad de Guadalajara, Camino Ramón Padilla Sánchez No. 2100 Nextipac, C.P. 45110, Zapopan, Jalisco, Mexico
Fabián Alejandro Rodríguez Zaragoza
Centro de Investigación en Alimentación y Desarrollo, Mazatlán Unit for Aquaculture and Environmental Management, Avenida Sábalo Cerritos s/n, 82112, Mazatlán, Sinaloa, Mexico
Bruno Goméz-Gil
Laboratorio de Biología Molecular, Centro de Investigación en Alimentación y Desarrollo, A.C., Avenida Sábalo-Cerritos s/n, 82010, Mazatlán, Sinaloa, Mexico
Alejandra García-Gasca
Laboratorio de Bioinformática Microbiana, Programa Académico de Ingeniería en Biotecnología, Universidad Politécnica de Sinaloa, Carretera Municipal Libre Km. 3 Mazatlán-Higueras, 82199, Mazatlán, Sinaloa, Mexico
Adrián González-Castillo

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Fabián Alejandro Rodríguez Zaragoza
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Bruno Goméz-Gil
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Alejandra García-Gasca
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Alma P. Rodríguez-Troncoso
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Adrián González-Castillo
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Contributions

E.B-G. and JL.C. planned the research; E.B-G, B.G.G., and F.A.R.Z analyzed and interpreted the data; E.B.G., JL.C., B.G.G., A.G-C, and F.A.R.Z drafted the fits draft of the manuscript and prepared figures and table; A.P.R-T and A.G.G. commented on later versions of the manuscript and provide reagents and material. All authors reviewed and approved the final version of the manuscript.

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Correspondence to José Luis Carballo.

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Bautista-Guerrero, E., Carballo, J.L., Rodríguez Zaragoza, F.A. et al. Changes in microbiome composition during ontogeny and dispersal of the coral boring sponge Thoosa mismalolli. Sci Rep 15, 2355 (2025). https://doi.org/10.1038/s41598-025-85622-x

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Received: 03 July 2024
Accepted: 06 January 2025
Published: 18 January 2025
DOI: https://doi.org/10.1038/s41598-025-85622-x