Novel microsatellite markers suggest significant genetic isolation in the Eastern Pacific sponge Aplysina gerardogreeni.

BACKGROUND: The Eastern Tropical Pacific (ETP) harbors a great diversity of Porifera. In particular, the Aplysina genus has acquired biotechnological and pharmacological importance. Nevertheless, the ecological aspects of their species and populations have been poorly studied. Aplysina gerardogreeni is the most conspicuous verongid sponge from the ETP, where it is usually found on rocky-coralline ecosystems. We evaluated the polymorphism levels of 18 microsatellites obtained from next-generation sequencing technologies. Furthermore, we tested the null hypothesis of panmixia in A. gerardogreeni population from two Mexican-Pacific localities. METHODS AND RESULTS: A total of 6,128,000 paired reads were processed of which primer sets of 18 microsatellites were designed. The loci were tested in 64 specimens from Mazatlan, Sinaloa (N = 32) and Isabel Island, Nayarit (N = 32). The microsatellites developed were moderately polymorphic with a range of alleles between 2 and 11, and Ho between 0.069 and 0.785. Fifteen loci displayed significant deviation from the Hardy-Weinberg equilibrium. No linkage disequilibrium was detected. A strong genetic structure was confirmed between localities using hierarchical Bayesian analyses, principal coordinates analyses, and fixation indices (FST = 0.108*). All the samples were assigned to their locality; however, there was a small sign of mixing between localities. CONCLUSIONS: Despite the moderate values of diversity in microsatellites, they showed a strong signal of genetic structure between populations. We suggest that these molecular markers can be a relevant tool to evaluate all populations across the ETP. In addition, 17 of these microsatellites were successfully amplified in the species A. fistularis and A. lacunosa, meaning they could also be applied in congeneric sponges from the Caribbean Sea. The use of these molecular markers in population genetic studies will allow assessment of the connectivity patterns in species of the Aplysina genus.

Isolation and characterization of novel microsatellite loci for the Eastern Pacific marine sponge Mycale cecilia by Illumina MiSeq sequencing.

BACKGROUND: Mycale cecilia is an abundant Eastern Tropical Pacific sponge living in a wide variety of habitats, including coral reefs where it may directly interact with corals. It is also known to possess secondary metabolites of pharmacological value. These aspects highlight the importance of having a better understanding of its biology, and genetic and population diversity. METHODS AND RESULTS: In the present study, we isolated and characterized twelve novel microsatellite loci by Illumina MiSeq sequencing. The loci were tested in 30 specimens collected from two coral reef localities (La Paz, Baja California Sur and Isabel Island, Nayarit) from the Mexican Pacific using M13(-21) labeling. All loci were polymorphic, with two to nine alleles per locus. Expected heterozygosities varied from 0.616 to 0.901. Eleven loci were tested and successfully amplified in M. microsigmatosa from the Gulf of Mexico. CONCLUSION: Here we report the first microsatellite loci developed for a sponge species from the Eastern Pacific coast. These molecular markers will be used for population genetic studies of M. cecilia, and potentially in other congeneric species; particularly in vulnerable marine areas that require protection, such as coral reefs.

The Role of Diversity in Mediating Microbiota Structural and Functional Differences in Two Sympatric Species of Abalone Under Stressed Withering Syndrome Conditions.

Withering syndrome (WS) is a gastro-intestinal (GI) infectious disease likely affecting all abalone species worldwide. Structural and functional changes in abalone GI microbiotas under WS-stressed conditions remain poorly investigated. It is unclear if interspecific microbiota differences, such as the presence of certain microbes, their abundance, and functional capabilities, may be involved in the occurrence of this disease. Bacterial microbiotas of healthy Haliotis fulgens and Haliotis corrugata are mainly composed by Tenericutes, Proteobacteria, Fusobacteria, and Spirochaetes. We previously reported species-specific structural and functional profiles of those communities and suggested that they are of consequence to the different susceptibility of each species to WS. Here, we address this question by comparing the structure and function of healthy and dysbiotic microbiota through 454 pyrosequencing and PICRUSt 2, respectively. Our findings suggest that the extent to which WS-stressed conditions may explain structural and functional differences in GI microbiota is contingent on the microbiota diversity itself. Indeed, microbiota differences between stressed and healthy abalone were marginal in the more complex bacterial communities of H. corrugata, in which no significant structural or functional changes were detected. Conversely, significant structural changes were observed in the less complex bacterial microbiota of H. fulgens. Moreover, structural alterations led to a significant downregulation of some metabolic activities conducted by GI bacteria. Accordingly, results suggest that gastro-intestinal bacterial diversity appears to be related with both the health of abalone and the etiology of WS.

Integrating eDNA metabarcoding and simultaneous underwater visual surveys to describe complex fish communities in a marine biodiversity hotspot.

Marine biodiversity can be surveyed using underwater visual censuses and recently with eDNA metabarcoding. Although a promising tool, eDNA studies have shown contrasting results related to its detection scale and the number of species identified compared to other survey methods. Also, its accuracy relies on complete reference databases used for taxonomic assignment and, as other survey methods, species detection may show false-negative and false-positive errors. Here, we compared results from underwater visual censuses and simultaneous eDNA metabarcoding fish surveys in terms of observed species and community composition. We also assess the effect of a custom reference database in the taxonomic assignment, and evaluate occupancy, capture and detection probabilities, as well as error rates of eDNA survey data. We amplified a 12S rRNA fish barcode from 24 sampling sites in the gulf of California. More species were detected with eDNA metabarcoding than with UVC. Because each survey method largely detected different sets of species, the combined approach doubled the number of species registered. Both survey methods recovered a known biodiversity gradient and a biogeographic break, but eDNA captured diversity over a broader geographic and bathymetric scale. Furthermore, the use of a modest-sized custom reference database significantly increased taxonomic assignment. In a subset of species, occupancy models revealed eDNA surveys provided similar or higher detection probabilities compared to UVC. The occupancy value of each species had a large influence on eDNA detectability, and in the false positive and negative error. Overall, these results highlight the potential of eDNA metabarcoding in complementing other established ecological methods for studies of marine fishes.

The complete mitogenome of excavating sponge Thoosa mismalolli (Demospongiae, Tetractinellida, Thoosidae) from Northeastern Tropical Pacific.

The complete mitogenome of Thoosa mismalolli Carballo, Cruz-Barraza & Gómez, 2004 (Tetractinellida, Thoosidae) was sequenced. This is the first complete mitogenome of the suborden Thoosina and the third Tetractinellid so far. The mitochondrial genome of T. mismalolli was assembled based on reads obtained with the Illumina HiSeq platform. The length of complete mitogenome is 19,019 bp long and contained 14 protein-coding genes and 23 tRNA, with two tRNA genes. Phylogenetic reconstruction (maximum-likelihood) based on mitogenome of Tetractinellids, supports T. mismalolli as a sister group. This result is congruent with those obtained with molecular markers (CO1, 18S, and 28S), supporting the monophyletic status of Thoosa and providing additional molecular data in favor of the suborder Thoosina.

Echinoderes unispinosus (Kinorhyncha: Cyclorhagida), a new record from deep-sea sediments in the Gulf of Mexico.

Kinorhyncha is a phylum of exclusively marine, meiofaunal invertebrates (Sørensen Pardos 2008). Currently, the knowledge about the worldwide distribution of most of its species is considerably patchy, as several taxa have been reported from a single or few localities, usually within a limited geographic area (Sørensen Pardos 2008; Yamasaki et al. 2018a). This even becomes more evident for deep-sea kinorhynchs, as noticed by Sørensen et al. (2018). Given the particular value of new records from the deep-sea to increase our understanding in the distribution of meiofaunal organisms plus the observed relationship of morphological variation through the bathymetric gradient, we aim to report the first record of Echinoderes unispinosus Yamasaki, Neuhaus George, 2018 in the southern edge of the Gulf of Mexico (GoM), and contribute with new information about the morphological intraspecific variation of this species. In Mexico, studies of kinorhynch biodiversity are limited to the northern Gulf of California (50-1570m depth) (Álvarez-Castillo et al. 2015, 2018; Cepeda et al. 2019) and the Yucatán Peninsula (Sánchez Martínez 2019). The GoM is a semi-closed basin located in a transition zone with both subtropical and tropical weather. Deep-sea muddy sediment samples of the present study were collected during the expedition XIXIMI-5 (10-24 June 2016) and processed according to Cisterna-Céliz et al. (2019). Kinorhynch specimens were mounted, measured and photographed according to Cepeda et al. (2019). Two adult males were identified as E. unispinosus, one from station B11 (2298 m depth) and another from station B15 (3708m depth). Kinorhynch specimens were deposited at the Aquatic Invertebrates Collection of the Facultad de Ciencias, Universidad Nacional Autónoma de México (UNAM), Mexico, under accession numbers: KGM.001 and KGM.002.

The complete mitogenome of the Eastern Pacific sponge Aplysina gerardogreeni (Demospongiae, Verongida, Aplysinidae).

We report the first mitochondrial genome of a Verongid sponge, Aplysina gerardogreeni from the Pacific Ocean. This has 19,620 bp and includes 14 protein-coding genes, 2 rRNAs genes, and 25 tRNAs genes. The gene arrangement was similar to the one found in two Caribbean Aplysina mitogenomes previously reported. Comparative analyses revealed a few substitutions among congeneric mitogenomes. The mitogenome of A. gerardogreeni could be useful to study the evolution of Verongimorpha group and also to identify adequate genes for its molecular systematics.

Structure, dynamics and predicted functional role of the gut microbiota of the blue (Haliotis fulgens) and yellow (H. corrugata) abalone from Baja California Sur, Mexico.

The GI microbiota of abalone contains a highly complex bacterial assemblage playing an essential role in the overall health of these gastropods. The gut bacterial communities of abalone species characterized so far reveal considerable interspecific variability, likely resulting from bacterial interactions and constrained by the ecology of their abalone host species; however, they remain poorly investigated. Additionally, the extent to which structural changes in the microbiota entail functional shifts in metabolic pathways of bacterial communities remains unexplored. In order to address these questions, we characterized the gut microbiota of the northeast Pacific blue (Haliotis fulgens or HF) and yellow (Haliotis corrugata or HC) abalone by 16S rRNA gene pyrosequencing to shed light on: (i) their gut microbiota structure; (ii) how bacteria may interact among them; and (iii) predicted shifts in bacterial metabolic functions associated with the observed structural changes. Our findings revealed that Mycoplasma dominated the GI microbiome in both species. However, the structure of the bacterial communities differed significantly in spite of considerable intraspecific variation. This resulted from changes in predominant species composition in each GI microbiota, suggesting host-specific adaptation of bacterial lineages to these sympatric abalone. We hypothesize that the presence of exclusive OTUs in each microbiota may relate to host-specific differences in competitive pressure. Significant differences in bacterial diversity were found between species for the explored metabolic pathways despite their functional overlap. A more diverse array of bacteria contributed to each function in HC, whereas a single or much fewer OTUs were generally observed in HF. The structural and functional analyses allowed us to describe a significant taxonomic split and functional overlap between the microbiota of HF and HC abalone.

Eco-Evolutionary Processes Generating Diversity Among Bottlenose Dolphin, Tursiops truncatus, Populations off Baja California, Mexico.

For highly mobile species that nevertheless show fine-scale patterns of population genetic structure, the relevant evolutionary mechanisms determining structure remain poorly understood. The bottlenose dolphin (Tursiops truncatus) is one such species, exhibiting complex patterns of genetic structure associated with local habitat dependence in various geographic regions. Here we studied bottlenose dolphin populations in the Gulf of California and Pacific Ocean off Baja California where habitat is highly structured to test associations between ecology, habitat dependence and genetic differentiation. We investigated population structure at a fine geographic scale using both stable isotope analysis (to assess feeding ecology) and molecular genetic markers (to assess population structure). Our results show that there are at least two factors affecting population structure for both genetics and feeding ecology (as indicated by stable isotope profiles). On the one hand there is a signal for the differentiation of individuals by ecotype, one foraging more offshore than the other. At the same time, there is differentiation between the Gulf of California and the west coast of Baja California, meaning that for example, nearshore ecotypes were both genetically and isotopically differentiated either side of the peninsula. We discuss these data in the context of similar studies showing fine-scale population structure for delphinid species in coastal waters, and consider possible evolutionary mechanisms.

Isolation by environment in the highly mobile olive ridley turtle (Lepidochelys olivacea) in the eastern Pacific.

Spatial and temporal scales at which processes modulate genetic diversity over the landscape are usually overlooked, impacting the design of conservation management practices for widely distributed species. We examine processes shaping population divergence in highly mobile species by re-assessing the case of panmixia in the iconic olive ridley turtle from the eastern Pacific. We implemented a biophysical model of connectivity and a seascape genetic analysis based on nuclear DNA variation of 634 samples collected from 27 nesting areas. Two genetically distinct populations largely isolated during reproductive migrations and mating were detected, each composed of multiple nesting sites linked by high connectivity. This pattern was strongly associated with a steep environmental gradient and also influenced by ocean currents. These findings relate to meso-scale features of a dynamic oceanographic interface in the eastern tropical Pacific (ETP) region, a scenario that possibly provides different cost-benefit solutions and selective pressures for sea turtles during both the mating and migration periods. We reject panmixia and propose a new paradigm for olive ridley turtles where reproductive isolation due to assortative mating is linked to its environment. Our study demonstrates the relevance of integrative approaches for assessing the role of environmental gradients and oceanographic currents as drivers of genetic differentiation in widely distributed marine species. This is relevant for the conservation management of species of highly mobile behaviour, and assists the planning and development of large-scale conservation strategies for the threatened olive ridley turtles in the ETP.

Monomorphic pathogens: The case of Candidatus Xenohaliotis californiensis from abalone in California, USA and Baja California, Mexico.

Withering syndrome (WS) is a chronic wasting disease affecting abalone species attributed to the pathogen Candidatus Xenohaliotis californiensis (CXc). Wild populations of blue (Haliotis fulgens) and yellow (H. corrugata) abalone have experienced unusual mortality rates since 2009 off the peninsula of Baja California and WS has been hypothesized as a possible cause. Currently, little information is available about the genetic diversity of CXc and particularly the possible existence of strains differing in pathogenicity. In a recent phylogenetic analysis, we characterized five coding genes from this rickettsial pathogen. Here, we analyze those genes and two additional intergenic non-coding regions following multi-locus sequence typing (MLST) and multi-spacer typing (MST) approaches to assess the genetic variability of CXc and its relationship with blue, yellow and red (H. rufescens) abalone. Moreover, we used 16S rRNA pyrosequencing reads from gut microbiomes of blue and yellow abalone to complete the genetic characterization of this prokaryote. The presence of CXc was investigated in more than 150 abalone of the three species; furthermore, a total of 385 DNA sequences and 7117 16S rRNA reads from Candidatus Xenohaliotis californiensis were used to evaluate its population genetic structure. Our findings suggest the absence of polymorphism in the DNA sequences of analyzed loci and the presence of a single lineage of CXc infecting abalone from California (USA) and Baja California (Mexico). We posit that the absence of genetic variably in this marine rickettsia may be the result of evolutionary and ecological processes.

Phenotypic variation in dorsal fin morphology of coastal bottlenose dolphins (Tursiops truncatus) off Mexico.

Geographic variation in external morphology is thought to reflect an interplay between genotype and the environment. Morphological variation has been well-described for a number of cetacean species, including the bottlenose dolphin (Tursiops truncatus). In this study we analyzed dorsal fin morphometric variation in coastal bottlenose dolphins to search for geographic patterns at different spatial scales. A total of 533 dorsal fin images from 19 available photo-identification catalogs across the three Mexican oceanic regions (Pacific Ocean n = 6, Gulf of California n = 6 and, Gulf of Mexico n = 7) were used in the analysis. Eleven fin shape measurements were analyzed to evaluate fin polymorphism through multivariate tests. Principal Component Analysis on log-transformed standardized ratios explained 94% of the variance. Canonical Discriminant Function Analysis on factor scores showed separation among most study areas (p < 0.05) with exception of the Gulf of Mexico where a strong morphometric cline was found. Possible explanations for the observed differences are related to environmental, biological and evolutionary processes. Shape distinction between dorsal fins from the Pacific and those from the Gulf of California were consistent with previously reported differences in skull morphometrics and genetics. Although the functional advantages of dorsal fin shape remains to be assessed, it is not unlikely that over a wide range of environments, fin shape may represent a trade-off among thermoregulatory capacity, hydrodynamic performance and the swimming/hunting behavior of the species.

Multigenetic characterization of 'Candidatus Xenohaliotis californiensis'.

'Candidatus Xenohaliotis californiensis' (or Ca.Xc) is the aetiological agent of withering syndrome, a chronic wasting disease affecting most if not all North American species of abalone, and has been described as a Rickettsiales-like prokaryote. Genetic data regarding this species are limited to the 16S rRNA gene. The inability to grow it axenically has hindered its genetic and genomic characterization and, in consequence, a thorough analysis of its systematics. Here, we amplified and sequenced five genes (16S rRNA, 23S rRNA, ftsZ, virD4 and virB11) of Ca.Xc from infected abalone to analyse its phylogenetic position. Phylogenies from concatenated DNA and amino acid sequences with representative genera of most Rickettsiales unequivocally place Ca.Xc in the family Anaplasmataceae. Furthermore, the family has two reciprocally monophyletic lineages: one leading to (Neorickettsia, Ca.Xc) and the other to ((Ehrlichia, Anaplasma), Wolbachia)). A molecular-clock Bayesian reconstruction places Ca.Xc as the most basal lineage in Anaplasmataceae. These phylogenetic hypotheses shed light on patterns of host evolution and of ecological transitions. Specifically, Neorickettsia and Ca.Xc inhabit aquatic hosts whereas the remaining Anaplasmataceae are found in terrestrial hosts. Additionally, our evolutionary timeline places the directly transmitted marine Ca.Xc as the basal Anaplasmataceae, ancestral to both freshwater and terrestrial species with adaptations leading to more complex life cycles involving intermediate vectors or reservoir species; this supports the hypothesis of a marine origin for this bacterial family.

Hyperparasitism by the bacteriophage (Caudovirales) infecting Candidatus Xenohaliotis californiensis (Rickettsiales-like prokaryote) parasite of wild abalone Haliotis fulgens and Haliotis corrugata from the Peninsula of Baja California, Mexico.

Candidatus Xenohaliotis californiensis (CXc) is a Rickettsiales-like prokaryote that is considered the causal agent of Withering Syndrome (WS), a chronic disease of abalone, from the west coast of North America and it is listed by the International Organization for Animal Health (OIE) as a reportable agent due to its pathogenicity. This bacterium in red abalone Haliotis rufescens, black abalone Haliotis cracherodii, and yellow abalone Haliotis corrugata from California, US and Baja California, Mexico has been found to be infected by a bacteriophage. To date, there is no information on the epizootiology of CXc and its bacteriophage in natural populations of abalone; furthermore, it is unknown if the bacteriophage was also present in CXc infecting blue abalone Haliotis fulgens. The objective of this study was to determine the distribution, prevalence and intensity of CXc, as well as to determine the distribution and prevalence of the bacteriophage and to study interactions between host sex and hyperparasitism in blue abalone and yellow abalone. Tissue samples were obtained from seven localities where the commercial capture of wild abalone is carried out. Samplings were conducted throughout the 2012-2013 capture seasons and a total of 182 blue abalone and 170 yellow abalone were obtained. The prevalence and intensity of CXc and the prevalence of the bacteriophage were determined by histology. The identity of CXc was confirmed by PCR, product sequence analysis and in situ hybridization while the identity of the bacteriophage was corroborated by TEM. The prevalence of CXc infected and uninfected by the bacteriophage was 80% in blue abalone and 62% in yellow abalone. Low infection intensities were found in 86% of blue abalone and 82% of yellow abalone. Infection intensity was significantly higher in undifferentiated yellow abalone. The bacteriophage in CXc showed a prevalence of 22% and 31% in blue abalone and yellow abalone respectively. These results show that CXc and its bacteriophage are widely distributed in the peninsula of Baja California and that they are well established in natural populations of blue abalone and yellow abalone. Additionally, this data constitutes the first record of a bacteriophage in blue abalone.

The complete mitochondrial genome of the giant electric ray, Narcine entemedor (Elasmobranchii: Torpediniformes).

The complete mitochondrial genome of the giant electric ray is 17,081 bp long (GenBank accession KM386678) and includes 2 ribosomal RNA, 22 transfer RNA, 13 protein-coding genes, an origin of replication, 2 non-coding regions. The mitochondrial gene arrangement is similar to that found in other batoids. The control region possessed a set of tandem repeats. Start codon ATG and stop codon TAA/T were found in most protein-coding genes. The base composition of the genome is 36.2% A, 29.9% T, 21.9% C, and 11.9% G.

Novel gene arrangement in the mitochondrial genome of the Cortés Geoduck clam (Panopea globosa).

The mitogenome of the Cortés geoduck clam Panopea globosa (Genbank accession KM580068) has a length of 15,469 bp and contains 13 protein-coding genes, 2 ribosomal RNA and 22 transfer RNA genes, as conventional metazoan mitochondrial genomes. Structural genes start with ATG, ATA and GTG codons; whereas TAG and TAA are used as stop codons. Base composition is: 23.3% A, 40.4% T, 10.1% C and 26.1% G. As is typical of marine bivalves, all genes are coded on the same strand. On the other hand, the gene arrangement is considerably different from those found in other heterodont bivalve mitogenomes.

The complete mitochondrial DNA of the Pacific Geoduck clam (Panopea generosa).

The complete mitochondrial genome of the Pacific Geoduck Panopea generosa (Genbank accession KM580067) is 15,585 bp in size and contains the typical 37 genes (13 protein-coding, 2 ribosomal RNA and 22 transfer RNA) found in metazoan mitogenomes, including the rare ATPase subunit 8 gene (ATP8). All genes are coded in the same strand but the gene arrangement is novel among heterodont bivalves. The base composition was: A 25.0%, T 38.7%, C 11.2% and G 25.0%. The genome is structurally similar to that of its congener the Cortes Geoduck Panopea globosa.

The mitochondrial genome of the banded guitarfish, Zapteryx exasperata (Jordan and Gilbert, 1880), possesses a non-coding duplication remnant region.

The complete mitochondrial genome of the banded guitarfish is 17,310 bp long and includes 2 ribosomal RNA, 22 transfer RNA, and 13 protein-coding genes, a replication origin and a control region (GenBank accession number KM370325). Gene arrangement is similar to that found in other batoids. An extra non-coding region was found between the genes coding for transfer RNA proline and threonine possessing a set of tandem repeat motifs pointing to its origin as a duplication remnant. Start codon ATG and stop codon TAA/T were found in most protein-coding genes. The base composition of the genome is 32.3% A, 30.2% T, 24.3% C and 13.1% G.

Complete mitochondrial genome of the beaubrummel Damselfish, Stegastes flavilatus (Pisces: Perciformes, Pomacentridae).

The mitogenome of the beaubrummel damselfish, Stegastes flavilatus Gill, 1862 (Genebank accession number KP136922), has a total length of 16,816 bp. It encodes 13 protein-coding, two ribosomal RNAs (rRNAs), and 22 transfer RNA (tRNA) genes. Base composition is 28.6% A, 26.0% T, 29.8% C, and 15.7% G and 45.5% GC content. The gene arrangement was found to be the same of other pomacentrid mitogenomes.

Mitochondrial Genetic Structure and Matrilineal Origin of White Sharks, Carcharodon carcharias, in the Northeastern Pacific: Implications for Their Conservation.

White sharks (Carcharodon carcharias, WS henceforth) are globally and regionally threatened. Understanding their patterns of abundance and connectivity, as they relate to habitat use, is central for delineating conservation units and identifying priority areas for conservation. We analyzed mitochondrial data to test the congruence between patterns of genetic connectivity and of individual movements in the Northeastern Pacific (NEP) and to trace the matrilineal origin of immature WS from coastal California and Baja California to adult aggregation areas. We analyzed 186 mitochondrial control region sequences from sharks sampled in Central California (CC; n = 61), Southern California Bight (SCB; n = 25), Baja California Pacific coast (BCPC; n = 9), Bahía Vizcaíno (BV; n = 39), Guadalupe Island (GI; n = 45), and the Gulf of California (GC; n = 7). Significant mitochondrial differentiation between adult aggregation areas (CC, GI) revealed two reproductive populations in the NEP. We found general concordance between movement patterns of young and adult WS with genetic results. Young sharks from coastal California and Baja California were more likely born from females from GI. Mitochondrial differentiation of young-of-the-year from SCB and BV suggests philopatry to nursery areas in females from GI. These results provide a genetic basis of female reproductive behavior at a regional scale and point to a preponderance of sharks from GI in the use of the sampled coastal region as pupping habitat. These findings should be considered in Mexican and US management and conservation strategies of the WS NEP population.