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Maintaining genetic diversity in cultured shellfish can be challenging due to high variance in individual reproductive success, founder effects, and rapid genetic drift, but is important to retain adaptive potential and avoid inbreeding depression. To support broodstock management and selective breeding in cultured Pacific oysters (Crassostrea (Magallana) gigas), we developed an amplicon panel targeting 592 genomic regions and SNP variants with an average of 50 amplicons per chromosome. Target SNPs were selected based on elevated observed heterozygosity or differentiation in Pacific oyster populations in British Columbia, Canada. The use of the panel for parentage applications was evaluated using multiple generations of oysters from a breeding program on Vancouver Island, Canada (n = 181) and families selected for Ostreid herpesvirus-1 resistance from the Molluscan Broodstock Program in Oregon, USA (n = 136). Population characterization was evaluated using wild, naturalized, farmed, or hatchery oysters sampled throughout the Northern Hemisphere (n = 189). Technical replicates showed high genotype concordance (97.5%; n = 68 replicates). Parentage analysis found suspected pedigree and sample handling errors, demonstrating the panel's value for quality control in breeding programs. Suspected null alleles were identified and found to be largely population dependent, suggesting population-specific variation impacting target amplification. Null alleles were identified using existing data without the need for pedigree information, and once they were removed, assignment rates increased to 93.0 and 86.0% of possible assignments in the two breeding program datasets. A pipeline for analyzing the amplicon sequence data from sequencer output, amplitools, is also provided., Competing Interests: Conflicts of interest B.J.G.S. is affiliated with Sutherland Bioinformatics. The author has no competing financial interests to declare. Some authors affiliated with ThermoFisher Scientific have potential conflicts considering that the AgriSeq Targeted Genotyping by Sequencing solutions and associated Oligo panels that were designed and validated in the study are offered by ThermoFisher Scientific. However, the selection of markers, and data analysis was primarily conducted by other authors. The authors declare that the research was conducted in a scientific manner without any commercial considerations that could be construed as potential conflict of interest and further declare no other conflicts of interest. The other authors declare no competing interests., (© The Author(s) 2024. Published by Oxford University Press on behalf of The Genetics Society of America.)

Rare events ( oddballs ) produce a variety of enhanced physiological responses relative to frequent events ( standards ), including the P3b component of the event-related potential (ERP) waveform. Previous research has suggested that the P3b is related to working memory, which implies that working memory representations will be enhanced for rare stimuli. To test this hypothesis, we devised a modified oddball paradigm in which the target was a disk presented at one of 16 different locations, which were divided into a rare set and a frequent set. Participants made a binary response on each trial to report whether the target appeared in the rare set or the frequent set. As expected, the P3b was much larger for stimuli appearing at a location within the rare set. We also included occasional probe trials in which the subject reported the exact location of the target. We found that these reports were more accurate for locations within the rare set than for locations within the frequent set. Moreover, the mean accuracy of these reports was correlated with the mean amplitude of the P3b. We also applied multivariate pattern analysis to the ERP data to "decode" the remembered location of the target. Decoding accuracy was greater for locations within the rare set than for locations within the frequent set. We then replicated and extended our behavioral findings in a follow-up experiment. These behavioral and electrophysiological results demonstrate that although both frequent and rare events are stored in working memory, the representations are enhanced for rare oddball events., Competing Interests: The authors declare no competing financial interests.

Multivariate pattern analysis (MVPA) approaches can be applied to the topographic distribution of event-related potential (ERP) signals to "decode" subtly different stimulus classes, such as different faces or different orientations. These approaches are extremely sensitive, and it seems possible that they could also be used to increase effect sizes and statistical power in traditional paradigms that ask whether an ERP component differs in amplitude across conditions. To assess this possibility, we leveraged the open-source ERP CORE data set and compared the effect sizes resulting from conventional univariate analyses of mean amplitude with two MVPA approaches (support vector machine decoding and the cross-validated Mahalanobis distance, both of which are easy to compute using open-source software). We assessed these approaches across seven widely studied ERP components (N170, N400, N2pc, P3b, lateral readiness potential, error related negativity, and mismatch negativity). Across all components, we found that multivariate approaches yielded effect sizes that were as large or larger than the effect sizes produced by univariate approaches. These results indicate that researchers could obtain larger effect sizes, and therefore greater statistical power, by using multivariate analysis of topographic voltage patterns instead of traditional univariate analyses in many ERP studies., (© 2024 Society for Psychophysiological Research.)

Principal component analysis (PCA) has been widely employed for dimensionality reduction prior to multivariate pattern classification (decoding) in EEG research. The goal of the present study was to provide an evaluation of the effectiveness of PCA on decoding accuracy (using support vector machines) across a broad range of experimental paradigms. We evaluated several different PCA variations, including group-based and subject-based component decomposition and the application of Varimax rotation or no rotation. We also varied the numbers of PCs that were retained for the decoding analysis. We evaluated the resulting decoding accuracy for seven common event-related potential components (N170, mismatch negativity, N2pc, P3b, N400, lateralized readiness potential, and error-related negativity). We also examined more challenging decoding tasks, including decoding of face identity, facial expression, stimulus location, and stimulus orientation. The datasets also varied in the number and density of electrode sites. Our findings indicated that none of the PCA approaches consistently improved decoding performance related to no PCA, and the application of PCA frequently reduced decoding performance. Researchers should therefore be cautious about using PCA prior to decoding EEG data from similar experimental paradigms, populations, and recording setups., Competing Interests: Declaration of competing interest None., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

In nitrogen-limiting conditions, approximately 10% of the vegetative cells in filaments of the cyanobacterium Anabaena (Nostoc) sp. strain PCC 7120 differentiate into nitrogen-fixing heterocysts. During the late stages of heterocyst differentiation, three DNA elements, each embedded within an open reading frame, are programmed to excise from the chromosome by site-specific recombination. The DNA elements are named after the genes that they interrupt: nifD, fdxN, and hupL. The nifD and fdxN elements each contain a gene, xisA or xisF, respectively, that encodes the site-specific recombinase required for programmed excision of the element. Here, we show that the xisC gene (alr0677), which is present at one end of the 9,435-bp hupL element, is required for excision of the hupL element. A strain in which the xisC gene was inactivated showed no detectable excision of the hupL element. hupL encodes the large subunit of uptake hydrogenase. The xisC mutant forms heterocysts and grows diazotrophically, but unlike the wild type, it evolved hydrogen gas under nitrogen-fixing conditions. Overexpression of xisC from a plasmid in a wild-type background caused a low level of hupL rearrangement even in nitrogen-replete conditions. Expression of xisC in Escherichia coli was sufficient to produce rearrangement of an artificial substrate plasmid bearing the hupL element recombination sites. Sequence analysis indicated that XisC is a divergent member of the phage integrase family of recombinases. Site-directed mutagenesis of xisC showed that the XisC recombinase has functional similarity to the phage integrase family.

The fdxN element, along with two other DNA elements, is excised from the chromosome during heterocyst differentiation in Anabaena sp. strain PCC 7120. Previous work showed that rearrangement of the fdxN element requires the xisF gene, which encodes a site-specific recombinase, and suggested that at least one other heterocyst-specific factor is involved. Here we report that the xisH and xisI genes are necessary for the heterocyst-specific excision of the fdxN element. Deletion of a 3.2 kb region downstream of the xisF gene blocked the fdxN-element rearrangement in heterocysts. The 3.2 kb deletion was complemented by the two overlapping genes xisH and xisI. Interestingly, extra copies of xisHI on a replicating plasmid resulted in the xisF-dependent excision of the fdxN element in vegetative cells. Therefore, xisHI are involved in the control of cell-type specificity of the fdxN rearrangement. The xisHI genes had no effect on the two other DNA rearrangements. The xisHI-induced excision of the fdxN element produced strains lacking the element and demonstrates that the 55 kb element contains no essential genes. xisH and xisI do not show similarity to any known genes.

Two site-specific DNA rearrangements occur during heterocyst differentiation in the cyanobacterium Anabaena sp. strain PCC 7120: the deletion of an 11 kb element from within the nifD gene and the deletion of a 55 kb element from within the fdxN gene. Three Nostoc and six Anabaena strains were screened for the presence of the nifD and fdxN elements by Southern hybridization with Anabaena PCC 7120 DNA probes. Eight of the nine strains contained DNA sequences that were similar to the nifD element. Three strains, Nostoc sp. strain Mac, Anabaena cylindrica and Anabaena sp. strain M131, also showed significant similarity to portions of the 55 kb fdxN element. Anabaena sp. strain CA lacked both the nifD and fdxN elements. Southern analysis of vegetative cell and heterocyst DNA from A. cylindrica and a Fox+ revertant of Nostoc Mac (isolate R2) showed rearrangement of the nifD and fdxN elements in heterocysts. We found no RFLPs between Anabaena M131 and Anabaena PCC 7120 suggesting that strain M131 is a Het- derivative of strain PCC 7120.

Programmed DNA rearrangements that occur during cellular differentiation are uncommon and have been described in only two prokaryotic organisms. Here, we identify the developmentally regulated rearrangement of a hydrogenase gene in heterocysts of the cyanobacterium Anabaena sp. strain PCC 7120. Heterocysts are terminally differentiated cells specialized for nitrogen fixation. Late during heterocyst differentiation, a 10.5-kb DNA element is excised from within the hupL gene by site-specific recombination between 16-bp direct repeats that flank the element. The predicted HupL polypeptide is homologous to the large subunit of [NiFe] uptake hydrogenases. hupL is expressed similarly to the nitrogen-fixation genes; hupL message was detected only during the late stages of heterocyst development. An open reading frame, named xisC, identified near one end of the hupL DNA element is presumed to encode the element's site-specific recombinase. The predicted XisC polypeptide is homologous with the Anabaena sp. strain PCC 7120 site-specific recombinase XisA. Neither XisC nor XisA shows sequence similarity to other proteins, suggesting that they represent a different class of site-specific recombinase.

Two DNA elements are excised from the chromosome during Anabaena heterocyst differentiation. We have identified the gene xisF which encodes the site-specific recombinase responsible for the excision of a 55-kb element from within the fdxN gene. The cloned xisF gene is sufficient to cause site-specific rearrangement of an artificial substrate in Escherichia coli. Inactivation of xisF in the Anabaena chromosome prevents excision of the fdxN element and growth in nitrogen-deficient medium but does not alter the development of heterocysts. Forced transcription of xisF in vegetative cells did not result in excision of the fdxN element, suggesting that other factors may be involved in cell-type specificity. The predicted XisF protein shows significant similarity to the Bacillus subtilis SpoIVCA recombinase.

The filamentous cyanobacterium Anabaena sp. strain PCC 7120 produces terminally differentiated heterocysts in response to a lack of combined nitrogen. Heterocysts are found approximately every 10th cell along the filament and are morphologically and biochemically specialized for nitrogen fixation. At least two DNA rearrangements occur during heterocyst differentiation in Anabaena sp. strain PCC 7120, both the result of developmentally regulated site-specific recombination. The first is an 11-kilobase-pair (kb) deletion from within the 3' end of the nifD gene. The second rearrangement occurs near the nifS gene but has not been completely characterized. The DNA sequences found at the recombination sites for each of the two rearrangements show no similarity to each other. To determine the topology of the rearrangement near the nifS gene, cosmid libraries of vegetative-cell genomic DNA were constructed and used to clone the region of the chromosome involved in the rearrangement. Cosmid clones which spanned the DNA separating the two recombination sites that define the ends of the element were obtained. The restriction map of this region of the chromosome showed that the rearrangement was the deletion of a 55-kb DNA element from the heterocyst chromosome. The excised DNA was neither degraded nor amplified, and its function, if any, is unknown. The 55-kb element was not detectably transcribed in either vegetative cells or heterocysts. The deletion resulted in placement of the rbcLS operon about 10 kb from the nifS gene on the chromosome. Although the nifD 11-kb and nifS 55-kb rearrangements both occurred under normal aerobic heterocyst-inducing conditions, only the 55-kb excision occurred in argon-bubbled cultures, indicating that the two DNA rearrangements can be regulated differently.

The 3' region of the Anabaena variabilis nifD gene contains an 11-kilobase-pair element which is excised from the chromosome during heterocyst differentiation. We have sequenced the recombination sites which border the element in vegetative cells and the rearranged heterocyst sequences. In vegetative cells, the element was flanked by 11-base-pair direct repeats which were identical to the repeats present at the ends of the nifD element in Anabaena sp. strain PCC 7120 (Anabaena strain 7120). Although Anabaena strain 7120 and A. variabilis are quite distinct in many ways, the overall sequence similarity between the two strains for the regions sequenced was 96%. Like the Anabaena strain 7120 element, the A. variabilis element was excised in heterocysts to produce a functional nifD gene and a free circularized element which was neither amplified nor degraded. The Anabaena strain 7120 xisA gene is located at the nifK-proximal end of the nifD element and is required for excision of the element in heterocysts. The A. variabilis element also contained an xisA gene which could complement a defective Anabaena strain 7120 xisA gene. A. variabilis did not contain the equivalent of the Anabaena strain 7120 fdxN 55-kilobase-pair element.