Viable Progeny Research Articles

Influence of Genetic Selection for Growth and Broodstock Diet n-3 LC-PUFA Levels on Reproductive Performance of Gilthead Seabream, Sparus aurata.

Simple SummaryGilthead seabream (GSB) broodstock were genetically selected based on their growth trait either high growth (HG) or low growth (LG) to evaluate the reproductive performance of these both traits under either fish oil (FO) or vegetable oil (VO) based diets feeding regime. The egg and larval quality were significantly improved by the broodstock selected for HG trait fed under FO based diet. This indicates that broodstock selected with HG trait has positive influence on the improving sperm, egg and larval quality in gilthead seabream.Genetic selection in gilthead seabream (GSB), Sparus aurata, has been undertaken to improve the growth, feed efficiency, fillet quality, skeletal deformities and disease resistance, but no study is available to delineate the effect of genetic selection for growth trait on GSB reproductive performance under mass spawning condition. In this study, high growth (HG) or low growth (LG) GSB broodstock were selected to evaluate the sex steroid hormones, sperm, egg quality and reproductive performance under different feeding regime of commercial diet or experimental broodstock diet containing either fish oil (FO) or vegetable oil (VO) based diet. Under commercial diet feeding phase, broodstock selected for either high growth or low growth did not show any significant changes in the egg production per kg female whereas egg viability percentage was positively (p = 0.014) improved by the high growth trait broodstock group. The experimental diet feeding results revealed that both growth trait and dietary fatty acid composition influenced the reproductive performance of GSB broodstock. In the experimental diet feeding phase, we observed high growth trait GSB males produced a higher number of sperm cells (p < 0.001) and also showed a higher sperm motility (p = 0.048) percentage. The viable egg and larval production per spawn per kg female were significantly improved by the broodstock selected for high growth trait and fed with fish oil-based diet. This present study results signifies that gilthead seabream broodstock selected on growth trait could have positive role in improvement of sperm and egg quality to produce viable progeny.

Use of Synonymous Deoptimization to Derive Modified Live Attenuated Strains of Foot and Mouth Disease Virus.

Foot-and-mouth disease (FMD) is one of the most economically important viral diseases that can affect livestock. In the last 70 years, use of an inactivated whole antigen vaccine has contributed to the eradication of disease from many developed nations. However, recent outbreaks in Europe and Eastern Asia demonstrated that infection can spread as wildfire causing economic and social devastation. Therefore, it is essential to develop new control strategies that could confer early protection and rapidly stop disease spread. Live attenuated vaccines (LAV) are one of the best choices to obtain a strong early and long-lasting protection against viral diseases. In proof of concept studies, we previously demonstrated that “synonymous codon deoptimization” could be applied to the P1 capsid coding region of the viral genome to derive attenuated FMDV serotype A12 strains. Here, we demonstrate that a similar approach can be extended to the highly conserved non-structural P2 and P3 coding regions, providing a backbone for multiple serotype FMDV LAV development. Engineered codon deoptimized P2, P3 or P2, and P3 combined regions were included into the A24Cruzeiro infectious clone optimized for vaccine production, resulting in viable progeny that exhibited different degrees of attenuation in cell culture, in mice, and in the natural host (swine). Derived strains were thoroughly characterized in vitro and in vivo. Our work demonstrates that overall, the entire FMDV genome tolerates codon deoptimization, highlighting the potential of using this technology to derive novel improved LAV candidates.

Factors enforcing the species boundary between the human pathogens Cryptococcus neoformans and Cryptococcus deneoformans

Hybridization has resulted in the origin and variation in extant species, and hybrids continue to arise despite pre- and post-zygotic barriers that limit their formation and evolutionary success. One important system that maintains species boundaries in prokaryotes and eukaryotes is the mismatch repair pathway, which blocks recombination between divergent DNA sequences. Previous studies illuminated the role of the mismatch repair component Msh2 in blocking genetic recombination between divergent DNA during meiosis. Loss of Msh2 results in increased interspecific genetic recombination in bacterial and yeast models, and increased viability of progeny derived from yeast hybrid crosses. Hybrid isolates of two pathogenic fungal Cryptococcus species, Cryptococcus neoformans and Cryptococcus deneoformans, are isolated regularly from both clinical and environmental sources. In the present study, we sought to determine if loss of Msh2 would relax the species boundary between C. neoformans and C. deneoformans. We found that crosses between these two species in which both parents lack Msh2 produced hybrid progeny with increased viability and high levels of aneuploidy. Whole-genome sequencing revealed few instances of recombination among hybrid progeny and did not identify increased levels of recombination in progeny derived from parents lacking Msh2. Several hybrid progeny produced structures associated with sexual reproduction when incubated alone on nutrient-rich medium in light, a novel phenotype in Cryptococcus. These findings represent a unique, unexpected case where rendering the mismatch repair system defective did not result in increased meiotic recombination across a species boundary. This suggests that alternative pathways or other mismatch repair components limit meiotic recombination between homeologous DNA and enforce species boundaries in the basidiomycete Cryptococcus species.

Temperature stressed Caenorhabditis elegans males fail to mate successfully and successful males produce very few viable cross progeny.

Exposure to moderate temperature stress can have profoundly negative effects on an organism’s reproductive capacity at temperatures where there are minimal or indiscernible effects on the organism as a whole. These negative effects are often more pronounced in males of the species that produce sperm. Previously we showed that few males of Caenorhabditis elegans wild type strains are able to successfully produce any cross progeny after experiencing temperature stress. However, these experiments did not assess the number of progeny from temperature stressed males. To understand if temperature stress can reduce the number of progeny a male sires, we crossed temperature stressed males of three wild type strains of C. elegans: JU1171, LKC34, and N2, to strain matched hermaphrodites of their own genetic background or to uncoordinated hermaphrodites in the N2 background. We found that significantly fewer males exposed to moderate temperature stress can successfully mate and that the small number of males in the population that do successfully mate produce significantly fewer viable cross progeny than unstressed controls. Our results suggest that exposure to moderate temperature stress significantly reduces male C. elegans chances at reproducing similar to what is seen in other organisms.

Production of common carp donor-derived offspring from goldfish surrogate broodstock

Common carp (Cyprinus carpio) is the fourth most-produced species in worldwide aquaculture. Significant efforts are invested in breeding and preservation of genetic integrity of this important species. However, maintaining a carp gene bank in situ can be demanding due to the large body size of the fish. Moreover, ex situ gene banking in common carp is as in other fish species limited to sperm cryopreservation. Recent progress in reproductive biotechnology in fishes has allowed the transfer of germ stem cells (gamete precursors) between individuals or even species. After maturation, the recipients are producing gametes of the donor. Surrogacy can serve as a valuable tool in germplasm banking to recover cryopreserved germ stem cells. Some unfavourable characteristics of the donor species such as long maturation time or large body size hampering its reproduction can be overcome by choosing a surrogate with more convenient characteristics. Efficient protocols for cryopreservation of common carp male and female germ stem cells have been recently developed in our laboratory. The next step has been to assess the potential of smaller goldfish (Carassius auratus) surrogates to produce donor-derived gametes of common carp after intraperitoneal transplantation of testicular cells.High transplantation success was achieved when 44% of the surviving goldfish produced pure donor-derived gametes of common carp at the age of 3 years giving rise to viable progeny. Donor-derived identity of the offspring was confirmed by genotyping and the production of a typical phenotype corresponding to the donor species. Reproductive performance of chimeras was similar to goldfish controls. Assessment of gamete characteristics showed that the size of donor-derived eggs is between control common carp and goldfish eggs. Interestingly, flagellum length in donor-derived spermatozoa was comparable to the common carp flagellum and significantly shorter than goldfish flagellum. Genotyping of the Y chromosome locus in chimeric goldfish provided a novel insight on the fate of transplanted cells, where only genotypic females were capable of producing eggs, while both genotypic males and females were capable of producing sperm. The present technology can be combined with cryopreservation procedures to ease needs for common carp breeds preservation and their recovery using many times smaller goldfish surrogates but also serve as a convenient model to study interspecific surrogacy in cyprinids.

Transgenerational metabolic disorders and reproduction defects induced by benzo[a]pyrene in Xenopus tropicalis.

Metabolic disorders induced by endocrine disruptors (ED) may contribute to amphibian population declines but no transgenerational studies have evaluated this hypothesis. Here we show that Xenopus tropicalis, exposed from the tadpole stage, to the ED benzo[a]pyrene (BaP, 50ng.L-1) produced F2 progeny with delayed metamorphosis and sexual maturity. At the adult stage, F2-BaP females displayed fatty liver with inflammation, tissue disorganization and metabolomic and transcriptomic signatures typical of nonalcoholic steato-hepatitis (NASH). This phenotype, similar to that observed in F0 and F1 females, was accompanied by a pancreatic insulin secretory defect. Metabolic disrupted F2-BaP females laid eggs with metabolite contents significantly different from the control and these eggs did not produce viable progeny. This study demonstrated that an ED can induce transgenerational disruption of metabolism and population collapse in amphibians under laboratory conditions. These results show that ED benzo[a]pyrene can impact metabolism over multiple generations and support epidemiological studies implicating environmental EDs in metabolic diseases in humans.

Ploidy Levels and Fitness-Related Traits in Purebreds and Hybrids Originating from Sterlet (Acipenser ruthenus) and Unusual Ploidy Levels of Siberian Sturgeon (A. baerii).

The present study aimed to investigate and compare fitness-related traits and ploidy levels of purebreds and hybrids produced from sturgeon broodstock with both normal and abnormal ploidy levels. We used diploid Acipenser ruthenus and tetraploid A. baerii males and females to produce purebreds and reciprocal hybrids of normal ploidy levels. Likewise, we used diploid A. ruthenus and tetraploid A. baerii females mated to pentaploid and hexaploid A. baerii males to produce hybrids of abnormal ploidy levels. Fertilization of ova of A. ruthenus and A. baerii of normal ploidy with the sperm of pentaploid and hexaploid A. baerii produced fully viable progeny with ploidy levels that were intermediate between those of the parents as was also found in crosses of purebreds and reciprocal hybrids of normal ploidy levels. The A. ruthenus × pentaploid A. baerii and A. ruthenus × hexaploid A. baerii hybrids did not survive after 22 days post-hatch (dph). Mean body weight and cumulative survival were periodically checked at seven-time intervals. The recorded values of mean body weight were significantly higher in A. baerii × pentaploid A. baerii hybrids than other groups at three sampling points (160, 252 and 330 dph). In contrast, the highest cumulative survival was observed in A. baerii × A. ruthenus hybrids at all sampling points (14.47 ± 5.70 at 497 dph). Overall, most of the studied sturgeon hybrids displayed higher mean BW and cumulative survival compared to the purebreds. The utilization of sturgeon hybrids should be restricted to aquaculture purposes because they can pose a significant genetic threat to native populations through ecological interactions.

Resumption of donor-origin spermatogenesis in senescent goldfish Carassius auratus (Linnaeus, 1758) following spermatogonial cell therapy

Stem cell research has come into prominence because of its applications in assisted reproductive technology and the treatment of deadly diseases. In teleost fishes, spermatogonial stem cells have been effectively used to produce surrogate gametes and progeny through germ cell transplantation technique. The present study is the first report of an innovative application of stem cell therapy in fish species for revitalising the reproductive competence of senescent individuals. Senescent male goldfish, Carassius auratus, approximately 10 years of age were procured from a fish-breeding farm and were reared locally in the lab for an additional two years. The senescence of the individuals was then evaluated and confirmed using histological analysis, gonadal index assessment, and germ-cell specific vasa gene expression. Analyses revealed absence of spermatogonial cells and other germ cells in the testes of the senescent fish (n = 5). Spermatogonial cells from sexually immature C. auratus male donor were isolated using discontinuous percoll gradients, labelled with the fluorescent dye PKH-26, and transplanted into the gonads of senescent C. auratus males through urogenital papilla. Six months after the transplant, spermatozoa were collected through applying gentle manual pressure on the abdomen and were observed under a microscope. All C. auratus males with the transplant had produced spermatozoa from the transplanted cells. This was confirmed by the retention of PKH-26 in the spermatozoa and diagnostic SSR locus. Gravid C. auratus females were artificially inseminated with the spermatozoa of those senescent males and natural spawning was allowed. As a result viable progeny were produced. These observation suggests that the reproductive competence of senescent male fishes can be revitalised through spermatogonial stem cell therapy to produce functional gametes.

Cellular Vimentin Interacts with Foot-and-Mouth Disease Virus Nonstructural Protein 3A and Negatively Modulates Viral Replication.

Nonstructural protein 3A of foot-and-mouth disease virus (FMDV) is a partially conserved protein of 153 amino acids that is in most FMDVs examined to date, and it plays important roles in virus replication, virulence, and host range. To better understand the role of 3A during FMDV infection, we used coimmunoprecipitation followed by mass spectrometry to identify host proteins that interact with 3A in FMDV-infected cells. Here, we report that cellular vimentin is a host binding partner for 3A. The 3A-vimentin interaction was further confirmed by coimmunoprecipitation, glutathione S-transferase (GST) pull down, and immunofluorescence assays. Alanine-scanning mutagenesis indicated that amino acid residues 15 to 21 at the N-terminal region of the FMDV 3A are responsible for the interaction between 3A and vimentin. Using reverse genetics, we demonstrate that mutations in 3A that disrupt the interaction between 3A and vimentin are also critical for virus growth. Overexpression of vimentin significantly suppressed the replication of FMDV, whereas knockdown of vimentin significantly enhanced FMDV replication. However, chemical disruption of the vimentin network by acrylamide resulted in a significant decrease in viral yield, suggesting that an intact vimentin network is needed for FMDV replication. These results indicate that vimentin interacts with FMDV 3A and negatively regulates FMDV replication and that the vimentin-3A interaction is essential for FMDV replication. This study provides information that should be helpful for understanding the molecular mechanism of FMDV replication.IMPORTANCE Foot-and-mouth disease virus (FMDV) nonstructural protein 3A plays important roles in virus replication, host range, and virulence. To further understand the role of 3A during FMDV infection, identification of host cell factors that interact with FMDV 3A is needed. Here, we found that vimentin is a direct binding partner of FMDV 3A, and manipulation of vimentin has a negative effect on virus replication. We also demonstrated that amino acid residues 15 to 21 at the N-terminal region of the FMDV 3A are responsible for the interaction between 3A and vimentin and that the 3A-vimentin interaction is critical for viral replication since the full-length cDNA clone harboring mutations in 3A, which were disrupt 3A-vimentin reactivity, could not produce viable virus progeny. This study provides information that not only provides us a better understanding of the mechanism of FMDV replication but also helps in the development of novel antiviral strategies in the future.

Assessment of cross compatibility in three strains of &lt;i&gt;Pleurotus species &lt;/i&gt;and yield attributes of the surviving progenies

Despite the upsurge in mushroom consumption in Nigeria, breeding of new strains for improved quality, early maturity and biological efficiency is a new concept. This study investigated cross-compatibility among three mushroom parental strains (Pleurotus ostreatus, P. tuber-regium and Lentinus edodes) and assessed yield attributes in parents and the progenies of compatible strains – (P. ostreatus and L. edodes). The crosses were carried out at the Department of Biology, Kogi State University, Anyigba, Kogi State, Nigeria using the dual culture technique. Among the twenty-seven combinations, only five (5) hybridized successfully. Crosses between P. tuber-regium and the other two strains (P. ostreatus and L. edodes) failed to yield viable progenies while the cross between P. ostreatus (po) and L. edodes (le) yielded five (5) new strains. Data collected on morphological and yield attributes of the compatible parents and the new strains revealed that three (3) strains (po1 x le1, po3 x le2 and po3 x le3) produced white fluffy colony growth morphology which represented the characters of P. ostreatus while the remaining two strains (po2 x le1 and po2 x le3) had milky, appressed morphology, representing the attributes of L. edodes. The strain from cross combination po3 x le3 completed spawn run in 37 days which was the fastest (ρ &lt; 0.05) among the strains and also exhibited superiority (ρ &lt; 0.05 or ρ &lt; 0.01) for all other attributes including fresh yield and biological efficiency over other strains and the two parents. Two other strains viz. po2 x le3 and po3 x le2 also exhibited significantly (ρ &lt; 0.05) higher yield and biological efficiency than either of the parental strains. These new strains can be exploited for commercial production of improved mushrooms in Nigeria.&#x0D; Key words: Cross Compatibility, Pleurotus, Dual Culture, Yield, Biology efficiency

Parasitism of Locally Recruited Egg Parasitoids of the Fall Armyworm in Africa.

The fall armyworm (FAW), Spodoptera frugiperda (J.E. Smith) (Lepidoptera: Noctuidae), is an insect native to the tropical and subtropical Americas that has recently spread to Africa, where it predominately attacks maize, sorghum and other plant species. Biological control is an environmentally friendly way of combatting the pest and contributes to an integrated pest management approach. In Africa, several trichogrammatid parasitoids and Telenomus remus Nixon (Hymenoptera: Platygastridae) have been found parasitizing eggs of the FAW. In Niger, the egg parasitoids encountered include Trichogrammatoidea sp. (Hymenoptera: Trichogrammatidae) and Telenomus remus Nixon. Parasitism of the FAW eggs by the two egg parasitoids was assessed in the laboratory, followed by field testing on sentinel eggs. In the laboratory, T. remus parasitized on average 78% of FAW eggs, compared to 25% for Trichogrammatoidea sp. Telenomus remus was able to parasitize egg masses that were fully covered with scales, while Trichogrammatoidea sp. parasitized only uncovered egg masses. On-farm releases of T. remus in sorghum fields caused up to 64% of FAW egg parasitism. Parasitized eggs yielded viable progeny, which can contribute to FAW egg parasitism build-up during the cropping season. Our findings lay the groundwork for the use of T. remus in augmentative releases against FAW in Africa.

A sexual hybrid and autopolyploids detected in seed from crosses between Neslia paniculata and Camelina sativa (Brassicaceae)

It is important to understand the probability of hybridization and potential for introgression of transgenic crop alleles into wild populations as part of pre-release risk assessment. Here we completed bidirectional crosses between the emerging crop, camelina [Camelina sativa (L.) Crantz] and its weedy relative, ball mustard [Neslia paniculata (L.) Desv.]. Ball mustard is a self-compatible annual that produces hard ball-like seeds similar to canola or mustard seed in size and shape. A total of 1593 crosses were completed and collected with camelina as the maternal parent, while 3253 crosses were successfully collected in the reverse direction. Putatively hybrid seedlings were screened with flow cytometry and species-specific nuclear ribosomal internal transcribed spacer (ITS) markers. Three plants had DNA contents close to expectations for hybrids, but only one of these, formed on camelina, had the expected ITS markers. This hybrid exhibited low fertility, and neither self-pollination nor backcrossing produced viable progeny. The other two plants, formed on ball mustard, had high pollen and seed fertility and were identified as ball mustard neoautotetraploids. Therefore, the hybridization rate between camelina and ball mustard is relatively low at one in 20 000 ovules pollinated when camelina is the maternal parent. However, autotetraploids may form frequently in ball mustard, and tetraploid populations may exist in nature.

Hybrid between Danio reriofemaleandDanio nigrofasciatus male produces aneuploid sperm with limited fertilization capacity.

In the Danio species, interspecific hybridization has been conducted in several combinations. Among them, only the hybrid between a zebrafish (D. rerio) female and a spotted danio (D. nigrofasciatus) male was reported to be fertile. However, beyond these investigations, by means of reproductive biology, gametes of the hybrid have also not been investigated genetically. For this study, we induced a hybrid of the D. rerio female and D. nigrofasciatus male in order to study its developmental capacity, reproductive performance and gametic characteristics. Its hybrid nature was genetically verified by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analysis of the rhodopsin gene. Almost all the hybrids (36/37) were males, and only one was female. Developing oocytes were observed in the hybrid female, but ovulated eggs have not been obtained thus far. Microscopic observation revealed various head sizes of sperm in the hybrid males. Flow cytometry showed that the hybrid males generated aneuploid sperm with various ploidy levels up to diploidy. In backcrosses between D. rerio females and hybrid males, fertilization rates were significantly lower than the control D. rerio, and most resultant progeny with abnormal appearance exhibited various kinds of aneuploidies ranging from haploidy to triploidy, but only one viable progeny, which survived more than four months, was triploid. This suggested the contribution of fertile diploid sperm of the hybrid male to successful fertilization and development.

Centromere scission drives chromosome shuffling and reproductive isolation

A fundamental characteristic of eukaryotic organisms is the generation of genetic variation via sexual reproduction. Conversely, significant large-scale genome structure variations could hamper sexual reproduction, causing reproductive isolation and promoting speciation. The underlying processes behind large-scale genome rearrangements are not well understood and include chromosome translocations involving centromeres. Recent genomic studies in the Cryptococcus species complex revealed that chromosome translocations generated via centromere recombination have reshaped the genomes of different species. In this study, multiple DNA double-strand breaks (DSBs) were generated via the CRISPR/Cas9 system at centromere-specific retrotransposons in the human fungal pathogen Cryptococcus neoformans The resulting DSBs were repaired in a complex manner, leading to the formation of multiple interchromosomal rearrangements and new telomeres, similar to chromothripsis-like events. The newly generated strains harboring chromosome translocations exhibited normal vegetative growth but failed to undergo successful sexual reproduction with the parental wild-type strain. One of these strains failed to produce any spores, while another produced ∼3% viable progeny. The germinated progeny exhibited aneuploidy for multiple chromosomes and showed improved fertility with both parents. All chromosome translocation events were accompanied without any detectable change in gene sequences and thus suggest that chromosomal translocations alone may play an underappreciated role in the onset of reproductive isolation and speciation.

Ploidy Level Influences Pollen Tube Growth and Seed Viability in Interploidy Crosses of Hydrangea macrophylla.

All Hydrangea macrophylla cultivars tested to date are diploid or triploid and triploid H. macrophylla have thicker stems, larger flowers, and larger stoma compared to related diploids. It is unknown whether interploidy crosses between diploid and triploid hydrangeas can be used to develop triploid varieties. The objective of this study was to compare pollen tube development, fruit formation, and seed viability among intra- and interploidy pollinations of H. macrophylla and evaluate the genome size and pollen viability of resultant progeny. By 24 h post-pollination, pollen tubes had reached the ovaries of diploid flowers in 48.7% of samples while pollen tubes reached the ovaries in only 8.7% of triploid flowers (χ 2 = 30.6, p < 0.001). By 48 h post-pollination pollen tubes reached the ovaries of diploid and triploid flowers in 72.5% and 53.8% of samples, respectively (χ 2 = 26.5, p = 0.001). There was no difference in percentage of flowers with pollen tubes reaching the ovaries in diploid and triploid flowers at 72 h after pollination (χ 2 = 7.5, p = 0.60). Analysis of covariance showed that pollen tube length at 24 and 48 h post-pollination was significantly influenced by ploidy and flower length of the female parent. Progeny of interploidy crosses was diploid and aneuploid; no triploid progeny were recovered from crosses using triploid parents. Mean genome sizes of offspring from each cross type ranged from 4.56 pg for 2x × 2x offspring to 5.17 pg for 3x × 3x offspring. Estimated ploidy of offspring ranged from 2x for 2x × 2x crosses to 2.4x for 3x × 3x crosses. Pollen stainability rates of flowering offspring using a modified Alexander's stain ranged from 69.6% to 76.4%.

Shaping the genome of plants

Fertilization of an egg cell by more than one sperm cell can produce viable progeny in a flowering plant.

Mathematical Modeling and Simulations of Centriole Positioning During Mitosis of Cells in Confined Environments

In order to properly undergo mitosis, epithelial cells pass through various biological checkpoints that cause cells to round up, stiffen and divide into two daughter cells. These checkpoints are influenced by biochemical and mechanical factors. Contrary to normal cell division, confined oncogenic cells are able to forgo mitotic rounding and proceed to divide into three to five viable progeny. Why cancerous cells are able to undergo mitosis in confined compact environments is not well understood. We hypothesize that success with confined cell division is at least partly driven by changes in the mechanistic properties and cortical rigidity of cancer cells. To test this hypothesis, we simulate microtubule-motor assembly driven mitotic spindle formation and the alignment of chromosomes along the metaphase plate as the cell rounds up during mitosis. We simulate the cell rounding process with and without confinement and identify the differences in spindle formation dynamics and spindle structure in the two scenarios. These findings will provide insights into the mechanistic pathways driving cancer cell proliferation during tumor growth.

First Field Release of a Genetically Engineered, Self-Limiting Agricultural Pest Insect: Evaluating Its Potential for Future Crop Protection.

Alternative, biologically-based approaches for pest management are sorely needed and one approach is to use genetically engineered insects. Herein we describe a series of integrated field, laboratory and modeling studies with the diamondback moth, Plutella xylostella, a serious global pest of crucifers. A “self-limiting” strain of Plutella xylostella (OX4319L), genetically engineered to allow the production of male-only cohorts of moths for field releases, was developed as a novel approach to protect crucifer crops. Wild-type females that mate with these self-limiting males will not produce viable female progeny. Our previous greenhouse studies demonstrated that releases of OX4319L males lead to suppression of the target pest population and dilution of insecticide-resistance genes. We report results of the first open-field release of a non-irradiated, genetically engineered self-limiting strain of an agricultural pest insect. In a series of mark-release-recapture field studies with co-releases of adult OX4319L males and wild-type counterparts, the dispersal, persistence and field survival of each strain were measured in a 2.83 ha cabbage field. In most cases, no differences were detected in these parameters. Overall, 97.8% of the wild-type males and 95.4% of the OX4319L males recaptured dispersed <35 m from the release point. The predicted persistence did not differ between strains regardless of release rate. With 95% confidence, 75% of OX4319L males released at a rate of 1,500 could be expected to live between 3.5 and 5.4 days and 95% of these males could be expected to be detected within 25.8–34.9 m from the release point. Moth strain had no effect on field survival but release rate did. Collectively, these results suggest similar field behavior of OX4319L males compared to its wild-type counterpart. Laboratory studies revealed no differences in mating competitiveness or intrinsic growth rates between the strains and small differences in longevity. Using results from these studies, mathematical models were developed that indicate release of OX4319L males should offer efficacious pest management of P. xylostella. Further field studies are recommended to demonstrate the potential for this self-limiting P. xylostella to provide pest suppression and resistance management benefits, as was previously demonstrated in greenhouse studies.

Su(var)2-10 and the SUMO Pathway Link piRNA-Guided Target Recognition to Chromatin Silencing.

Regulation of transcription is the main mechanism responsible for precise control of gene expression. Whereas the majority of transcriptional regulation ismediated by DNA-binding transcription factors that bind to regulatory gene regions, an elegant alternative strategy employs small RNA guides, Piwi-interacting RNAs (piRNAs) to identify targets of transcriptional repression. Here, we show that inDrosophila the small ubiquitin-like protein SUMOand the SUMO E3 ligase Su(var)2-10 are required for piRNA-guided deposition of repressive chromatin marks and transcriptional silencing of piRNA targets. Su(var)2-10 links the piRNA-guided target recognition complex to the silencing effector by binding the piRNA/Piwi complex and inducing SUMO-dependent recruitment of the SetDB1/Wde histone methyltransferase effector. We propose that in Drosophila, the nuclear piRNA pathway has co-opted a conserved mechanism of SUMO-dependent recruitment of the SetDB1/Wde chromatin modifier to confer repression of genomic parasites.

Systematic studies of Anopheles (Cellia) kochi (Diptera: Culicidae): Morphology, cytogenetics, cross-mating experiments, molecular evidence and susceptibility level to infection with nocturnally subperiodic Brugia malayi

Anopheles kochi Dӧnitz (Diptera: Culicidae) is a malaria vector in some countries in South and Southeast Asia. This is the first report to provide clear evidence that two different cytological forms of An. kochi are conspecific based on systematic studies. Two karyotypic forms, i.e., Form A (X1, X2, Y1) and a novel Form B (X1, X2, Y2) were obtained from a total of 15 iso-female lines collected from five provinces in Thailand. Form A was common in all provinces, whereas Form B was restricted to Ubon Ratchathani province. This study determined whether the two karyotypic variants of An. kochi exist as a single or cryptic species by performing cross-mating experiments in association with the sequencing of the second internal transcribed spacer (ITS2) of ribosomal DNA (rDNA), and cytochrome c oxidase subunit I (COI) of mitochondrial DNA (mtDNA). Cross-mating experiments between the two karyotypic forms revealed genetic compatibility by providing viable progenies through F2 generations. The two forms showed a high sequence similarity of those two DNA regions (average genetic distances: ITS2 = 0.002–0.005, COI = 0.000–0.009). The phylogenetic trees based on ITS2 and COI sequences also supported that four strains (from Bhutan, Cambodia, Indonesia, and Thailand) were all of the same species. Five sensilla types housed on the antennae of female An. kochi were observed under scanning electron microscopy (SEM). In addition, this study found that An. kochi was a refractory vector, revealed by 0% susceptibility rates to infection with nocturnally subperiodic Brugia malayi. The cibarial armature was a resistant mechanism, as it killed the microfilariae in the foregut before they penetrated into the developmental site.