Sterile Workers Research Articles

Reproductive activation in honeybee (Apis mellifera) workers protects against abiotic and biotic stress.

Social insect reproductives exhibit exceptional longevity instead of the classic trade-off between somatic maintenance and reproduction. Even normally sterile workers experience a significant increase in life expectancy when they assume a reproductive role. The mechanisms that enable the positive relation between the antagonistic demands of reproduction and somatic maintenance are unclear. To isolate the effect of reproductive activation, honeybee workers were induced to activate their ovaries. These reproductively activated workers were compared to controls for survival and gene expression patterns after exposure to Israeli Acute Paralysis Virus or the oxidative stressor paraquat. Reproductive activation increased survival, indicating better immunity and oxidative stress resistance. After qPCR analysis confirmed our experimental treatments at the physiological level, whole transcriptome analysis revealed that paraquat treatment significantly changed the expression of 1277 genes in the control workers but only two genes in reproductively activated workers, indicating that reproductive activation preemptively protects against oxidative stress. Significant overlap between genes that were upregulated by reproductive activation and in response to paraquat included prominent members of signalling pathways and anti-oxidants known to affect ageing. Thus, while our results confirm a central role of vitellogenin, they also point to other mechanisms to explain the molecular basis of the lack of a cost of reproduction and the exceptional longevity of social insect reproductives. Thus, socially induced reproductive activation preemptively protects honeybee workers against stressors, explaining their longevity. This article is part of the theme issue 'Ageing and sociality: why, when and how does sociality change ageing patterns?'

Novel approach to reduce SARS-CoV-2 transmission during trans-oral robotic surgery.

This study describes a novel approach in the reduction of SARS-CoV-2 transmission during trans-oral robotic surgery (TORS). Eight patients underwent TORS between 01 February 2020 and 07 September 2020. A sterile plastic sheet draped over sterile supports with water-tight seal around each cannula was used to create a sterile working space within which the robotic arms could freely move during operation. This set-up acts as an additional physical barrier against droplet and aerosol transmission. Operative diagnosis; droplet count and distribution on plastic sheet and face shields of console and assistant surgeons, and scrub nurse were documented. TORS tumour excision was performed for patients with suspected tonsillar tumour (n = 3) and tongue base tumour (n = 2). TORS tonsillectomy and tongue base mucosectomy was performed for cervical nodal metastatic carcinoma of unknown origin (n = 3). Droplet contamination was noted on all plastic drapes (n = 8). Droplet contamination was most severe over the central surface at 97.2% (91.7–100.0%), with the highest droplet count along the centre-most column where it overlies the site of operation in the oral cavity 33.3% (n = 31). Droplet count decreased towards the periphery. Contamination rate was 2.8% (0.0–8.3%) over the right lateral surface. There was no droplet contamination over the vertex and left lateral surface of plastic drapes. No droplet contamination was noted on face shields of all parties. The use of sterile plastic drapes with water-tight seal around each robotic cannula can help reduce viral transmission to healthcare providers during TORS.

Dufour\u2019s gland analysis reveals caste and physiology specific signals in Bombus impatiens

Reproductive division of labor in insect societies is regulated through multiple concurrent mechanisms, primarily chemical and behavioral. Here, we examined if the Dufour’s gland secretion in the primitively eusocial bumble bee Bombus impatiens signals information about caste, social condition, and reproductive status. We chemically analyzed Dufour’s gland contents across castes, age groups, social and reproductive conditions, and examined worker behavioral and antennal responses to gland extracts. We found that workers and queens each possess caste-specific compounds in their Dufour’s glands. Queens and gynes differed from workers based on the presence of diterpene compounds which were absent in workers, whereas four esters were exclusive to workers. These esters, as well as the total amounts of hydrocarbons in the gland, provided a separation between castes and also between fertile and sterile workers. Olfactometer bioassays demonstrated attraction of workers to Dufour’s gland extracts that did not represent a reproductive conflict, while electroantennogram recordings showed higher overall antennal sensitivity in queenless workers. Our results demonstrate that compounds in the Dufour’s gland act as caste- and physiology-specific signals and are used by workers to discriminate between workers of different social and reproductive status.

The Biology of the Cape Honey Bee, Apis mellifera capensis (Hymenoptera: Apidae): A Review of Thelytoky and Its Influence on Social Parasitism and Worker Reproduction

Abstract Cohesion in social insect colonies is maintained by use of chemical signals produced by the queen, workers, and brood. In honey bees in particular, signals from the queen and brood are crucial for the regulation of reproductive division of labor, ensuring that the only reproductive female individual in the colony is the queen, whereas the workers remain reproductively sterile. However, even given this strict level of control, workers can, in principle, activate their ovaries and lay eggs. Although much is known about the behavioral and physiological traits that accompany the switch from worker sterility to being reproductively active, much less is known regarding the molecular changes that accompany this switch. This review will explore what is known about the genes and molecular pathways involved in the making of laying workers/false queens in the Cape honey bee, Apis mellifera capensis Eschscholtz, through an analysis of the basis for thelytoky in this subspecies, the exocrine glandular chemistry of reproductively dominant workers and what is known about the biosynthesis of their pheromone components. This review will contribute to our understanding of the genetic regulation of thelytoky and the molecular mechanisms that govern reproductive division of labor in honey bees and provide generalizations that may be applicable to other social hymenoptera using this evolutionary fascinating example of worker reproduction.

Tissue-specific transcription patterns support the kinship theory of intragenomic conflict in honey bees (Apis mellifera).

Kin selection may act differently on genes inherited from parents (matrigenes and patrigenes), resulting in intragenomic conflict. This conflict can be observed as differential expression of matrigenes and patrigenes, or parent-specific gene expression (PSGE). In honey bees (Apis mellifera), intragenomic conflict is hypothesized to occur in multiple social contexts. Previously, we found that patrigene-biased expression in reproductive tissues was associated with increased reproductive potential in worker honey bees, consistent with the prediction that patrigenes are selected to promote selfish behaviour in this context. Here, we examined brain gene expression patterns to determine if PSGE is also found in other tissues. As before, the number of transcripts showing patrigene expression bias was significantly greater in the brains of reproductive vs. sterile workers, while the number of matrigene-biased transcripts was not significantly different. Twelve transcripts out of the 374 showing PSGE in either tissue showed PSGE in both brain and reproductive tissues; this overlap was significantly greater than expected by chance. However, the majority of transcripts show PSGE only in one tissue, suggesting the epigenetic mechanisms mediating PSGE exhibit plasticity between tissues. There was no significant overlap between transcripts that showed PSGE and transcripts that were significantly differentially expressed. Weighted gene correlation network analysis identified modules which were significantly enriched in both types of transcripts, suggesting that these genes may influence each other through gene networks. Our results provide further support for the kin selection theory of intragenomic conflict, and provide valuable insights into the mechanisms which may mediate this process.

RNA m6A Modification Functions in Larval Development and Caste Differentiation in Honeybee (Apis mellifera)

Genetically identical female honeybee larvae with different diets develop into sterile workers or fertile queens. It remains unknown whether the reversible RNA N6-methyladenosine (m6A) mark functionally impact this "caste differentiation." Here, we profile the transcriptome-wide m6A methylome of honeybee queen and worker larvae at three instar stages and discover that m6A methylation dynamics are altered by differential feeding. Multiple methylome comparisons show an obvious increase in m6A marks during larval development and reveal a negative correlation between gene expression and m6A methylation. Notably, we find that worker larvae contain more hypermethylated m6A peaks than do queen larvae, and many caste-differentiation-related transcripts are differentially methylated. Chemical suppression of m6A methylation in worker larvae by 3-deazaadenosine (DAA) reduces overall m6A methylation levels and triggers worker larvae to develop queen caste features. Thus, our study demonstrates that m6A functionally impacts caste differentiation and larval development, yet it does not exclude potential contributions from other factors.

"Ant Queen Aria"

"Ant Queen Aria" Anna Lindemann (bio) "Ant Queen Aria" looks to the ant colony as a means to reflect on gendered roles and social hierarchy. With original lyrics by Emma Komlos-Hrobsky, the aria depicts the transformation of a woman into an ant queen laying eggs. As the sole reproducer for the ant colony, the ant queen has the formidable task of laying twenty thousand to two hundred thousand eggs. Meanwhile, a handful of males appear fleetingly, and a half million sterile female workers— sisters of the queen—find food and tend to the young. The ant colony's division of labor prompts a reflection on gendered divisions of labor in human society and the struggle many women face when balancing family and work. "Ant Queen Aria" is scored for soprano and digital instruments played on keyboard and was composed for virtuosic musicians Lucy Fitz Gibbon and Ryan MacEvoy McCullough. The digital instruments include custom Logic Pro EXS24 Sampler percussion and "prepared piano" timbres, along with distorted Synful Orchestra synthesized sul ponticello strings. "Ant Queen Aria" is from the concert-length art science performance, The Colony, a work of multimedia opera-theater exploring sisterhood and the evolution of communication in both ants and humans. www.thecolony.show [End Page 153] Click for larger view View full resolution Fig 1a. Soprano Lucy Fitz Gibbon performs "Ant Queen Aria" in 2019. Click for larger view View full resolution Fig 1b. Through shadow performance and digital animation, she transforms from a woman into a chimerical ant queen about to lay eggs. Her abdominal segments stretch apart, and her abdomen swells to accommodate the next generation growing within her. [End Page 154] Click for larger view View full resolution [End Page 155] Click for larger view View full resolution [End Page 156] Click for larger view View full resolution [End Page 157] Click for larger view View full resolution [End Page 158] Click for larger view View full resolution [End Page 159] Anna Lindemann anna lindemann calls herself an evo-devo artist. Her work as a composer, animator, and performer explores the emerging field of evolutionary developmental biology (evo-devo). Her evo-devo art, including the opera-theater performances The Colony and Theory of Flight, has been featured internationally at theaters, concert halls, digital art conferences, planetariums, biology conferences, film festivals, and natural history museums. Anna received an MFA in integrated electronic arts from Rensselaer Polytechnic Institute and a BS in ecology and evolutionary biology from Yale. She is an assistant professor in the Digital Media & Design department at the University of Connecticut. www.annalindemann.com. Copyright © 2021 Suzanne G. Cusick

HoloPointer: a virtual augmented reality pointer for laparoscopic surgery training

PurposeIn laparoscopic surgery training, experts guide novice physicians to desired instrument positions or indicate relevant areas of interest. These instructions are usually given via verbal communication or using physical pointing devices. To facilitate a sterile work flow and to improve training, new guiding methods are needed. This work proposes to use optical see-through augmented reality to visualize an interactive virtual pointer on the laparoscopic.MethodsAfter an interdisciplinary development, the pointer’s applicability and feasibility for training was evaluated and it was compared to a standard condition based on verbal and gestural communication only. In this study, ten surgical trainees were guided by an experienced trainer during cholecystectomies on a laparoscopic training simulator. All trainees completed a virtual cholecystectomy with and without the interactive virtual pointer in alternating order. Measures included procedure time, economy of movement and error rates.ResultsResults of standardized variables revealed significantly improved economy of movement (p = 0.047) and error rates (p = 0.047), as well as an overall improved user performance (Total z-score; p = 0.031) in conditions using the proposed method.ConclusionThe proposed HoloPointer is a feasible and applicable tool for laparoscopic surgery training. It improved objective performance metrics without prolongation of the task completion time in this pre-clinical setup.

Gene-regulatory context of honey bee worker sterility

The highly organized societies of the Western honey bee Apis mellifera feature a highly reproductive queen at the center of attention and a large cohort of daughters that suppress their own reproduction to help rear more sisters, some of whom become queens themselves. This reproductive altruism is peculiar because in theory it evolves via indirect selection on genes for altruism that are expressed in the sterile workers but not in the reproductive queens. In this study we attempt to situate lists of genes previously implicated in queenright worker sterility into a broader regulatory framework. To do so we use a model bee brain transcriptional regulatory network as a template to infer how sets of genes responsive to ovary-suppressing queen pheromone are functionally interconnected over the model's topology. We predict that genes jointly involved in the regulation of worker sterility should be tightly networked, relative to genes whose functions are unrelated to each other. We find that sets of mapped genes – ranging in size from 17 to 250 – are well dispersed across the network's substructural scaffolds, suggesting that ovary de-activation involves genes that reside within more than one transcriptional regulatory module. For some sets, however, this dispersion is biased into certain areas of the network's substructure. Our analysis identifies the regions enriched for sterility genes and likewise identifies local hub genes that are presumably critical to subnetwork function. Our work offers a glimpse into the gene regulatory context of honey bee worker sterility and uses this context to identify new candidate gene targets for functional analysis. Finally, to the extent that any sterility-related modules identified here have evolved via selection for worker altruism, we can assume that this selection was indirect and of the type specifically invoked by inclusive fitness theory.

Reproductive physiology corresponds to adult nutrition and task performance in a Neotropical paper wasp: a test of dominance-nutrition hypothesis predictions

The dominance-nutrition hypothesis predicts that nutritional intake and energetic costs in adulthood interact to drive behavioral and physiological differences between females in primitively eusocial insects, and thereby affect reproductive caste. We tested predictions of this hypothesis in independent-founding Mischocyttarus pallidipectus paper wasps. We measured stable isotope tissue composition to compare nutritional status before adult emergence and among adult females. Adult tissue δ15N content (an indicator of feeding at a higher trophic level because it is enriched in animal prey relative to plant-based foods) was significantly higher in adults than at the end of pupal development, suggesting adult nutrition affects δ15N content after the end of pupal development. We then asked if behavior and nutritional status predicted ovary development. We measured ovary development, nitrogen and carbon stable isotope ratios, dominance behavior, and task performance (foraging, as indicated by time spent on the nest) for adult female wasps. We used social network analysis to quantify differences in social status between females with developed and undeveloped (filamentous) ovaries. Dominant females spent more time on the nest and were significantly more enriched in δ15N than subordinate females. These data support the dominance-nutrition hypothesis: adult behavior and energy expenditure, and access to animal-based diets, correspond to female reproductive physiology, and may play a role in adult-stage caste determination. Within animal social groups, differential access to resources can affect differences in reproductive success. This may affect caste determination in eusocial colonies, where reproductive tasks are divided between female castes, with a queen (or multiple queens) capable of egg-laying and helped by sterile workers. Female sterile workers and reproductives of some social Hymenoptera are morphologically similar at adult emergence, suggesting that adult experience may impact caste status. We asked whether patterns of nutritional physiology in Mischocyttarus pallidipectus paper wasps matched adult-stage caste determination by testing predictions of the dominance-nutrition hypothesis, which states that adult nutrition and energetic costs of tasks shape reproductive female development through behavioral and physiological changes.

Worker Queens? Behavioral Flexibility of Queens in the Little Fire Ant Wasmannia auropunctata

Many species of social hymenoptera demonstrate behavioral flexibility, where older workers that typically forage can revert to younger worker tasks, such as nursing, when these are absent. This flexibility is typical of the sterile worker class, yet rare in queens. In the little fire ant (Wasmannia auropunctata) queens have been reported to perform only egg laying. We examined behavior of queens of W. auropunctata after demographic manipulation. When half of the workers were removed from the colony, queens were observed caring for eggs, larvae and pupae as well as eating outside of the nest, like forager workers. We examined the relationship between these atypical queen behaviors and their juvenile hormone binding protein (JHbp) and vitellogenin (Vg) expression via QRT-PCR method. JHbp and Vg expression decreased when queens were performing worker tasks, resembling the expected expression pattern of typical sterile workers. Flexibility in queen behaviors in the little fire ant may be an important adaptation to changing environments. As a significant invasive species, such adaptation may increase the probability of colony survival during propagation. Our results not only present new insights in behavioral flexibility in social insects, but also increases our understanding of the success of this significant invasive species.

Identification of a queen pheromone mediating the rearing of adult sexuals in the pharaoh ant Monomorium pharaonis.

The division of labour between reproductive queens and mostly sterile workers is among the defining characteristics of social insects. Queen-produced chemical signals advertising her presence and fertility status, i.e. queen pheromones, are normally used to assert the queen's reproductive dominance in the colony. Most queen pheromones identified to date are chemicals that stop the daughter workers from reproducing. Nevertheless, it has long been suggested that queen pheromones could also regulate reproduction in different ways. In some multiple-queen ants with obligately sterile workers, for example-such as fire ants and pharaoh ants-queen pheromones are thought to regulate reproduction by inhibiting the rearing of new sexuals. Here, we identify the first such queen pheromone in the pharaoh ant Monomorium pharaonis and demonstrate its mode of action via bioassays with the pure biosynthesized compound. In particular, we show that the monocyclic diterpene neocembrene, which in different Monomorium species is produced solely by fertile, egg-laying queens, strongly inhibits the rearing of new sexuals (queens and males) and also exerts a weakly attractive 'queen retinue' effect on the workers. This is the first time that a queen pheromone with such a dual function has been identified in a social insect species with obligately sterile workers.

Association between host wing morphology polymorphism and Wolbachia infection in Vollenhovia emeryi (Hymenoptera: Myrmicinae).

Many eusocial insects, including ants, show complex colony structures, distributions, and reproductive strategies. In the ant Vollenhovia emeryi Wheeler (Hymenoptera: Myrmicinae), queens and males are produced clonally, while sterile workers arise sexually, unlike other ant species and Hymenopteran insects in general. Furthermore, there is a wing length polymorphism in the queen caste. Despite its evolutionary remarkable traits, little is known about the population structure of this ant species, which may provide insight into its unique reproductive mode and polymorphic traits. We performed in‐depth analyses of ant populations from Korea, Japan, and North America using three mitochondrial genes (COI, COII, and Cytb). The long‐winged (L) morph is predominant in Korean populations, and the short‐winged (S) morph is very rare. Interestingly, all L morphs were infected with Wolbachia, while all Korean S morphs lacked Wolbachia, demonstrating a association between a symbiont and a phenotypic trait. A phylogenetic analysis revealed that the S morph is derived from the L morph. We propose that the S morph is associated with potential resistance to Wolbachia infection and that Wolbachia infection does not influence clonal reproduction (as is the case in other ant species).

The potential for a CRISPR gene drive to eradicate or suppress globally invasive social wasps

CRISPR gene drives have potential for widespread and cost-efficient pest control, but are highly controversial. We examined a potential gene drive targeting spermatogenesis to control the invasive common wasp (Vespula vulgaris) in New Zealand. Vespula wasps are haplodiploid. Their life cycle makes gene drive production challenging, as nests are initiated by single fertilized queens in spring followed by several cohorts of sterile female workers and the production of reproductives in autumn. We show that different spermatogenesis genes have different levels of variation between introduced and native ranges, enabling a potential ‘precision drive’ that could target the reduced genetic diversity and genotypes within the invaded range. In vitro testing showed guide-RNA target specificity and efficacy that was dependent on the gene target within Vespula, but no cross-reactivity in other Hymenoptera. Mathematical modelling incorporating the genetic and life history traits of Vespula wasps identified characteristics for a male sterility drive to achieve population control. There was a trade-off between drive infiltration and impact: a drive causing complete male sterility would not spread, while partial sterility could be effective in limiting population size if the homing rate is high. Our results indicate that gene drives may offer viable suppression for wasps and other haplodiploid pests.

A mathematical model of anarchy in bees

Functional worker sterility in queenright honey bees (Apis mellifera) is fostered by the colony-level costs of this behavior. However, anarchy in the beehive can occur when a fraction of workers lay eggs, which, if reared, will develop into males (drones). Because males do not gather resources but still consume them, they represent an additional energic cost for the colony. This small number of anarchic worker-laid eggs is normally found and eaten by non-anarchic bees via worker policing, a controlling mechanism induced by queen’s pheromones. However, worker policing by anarchic bees is possible by mimicking the pheromone marker of the queen. Here, we show that the intensity of worker policing has an upper-bound limit set by the loss in its population-level benefits. We developed a mathematical model to assess the benefits of worker policing at different intensities and costs of anarchy in the beehive. We found that there is an optimal rate in which workers should police in order to maximize colony-level benefits. The adjustment of worker policing intensity depending on the degree of anarchy in the population can keep the number of drones descending from workers low. However, this control can collapse when worker policing gets saturated due to high frequencies of anarchism. Our findings can help to advance the understanding on the maintenance of eusociality in bees by showing how the balance between anarchic behavior and worker policing regulates productivity in the beehive.

Bumblebee Workers Show Differences in Allele-Specific DNA Methylation and Allele-Specific Expression.

Allele-specific expression is when one allele of a gene shows higher levels of expression compared with the other allele, in a diploid organism. Recent work has identified allele-specific expression in a number of Hymenopteran species. However, the molecular mechanism which drives this allelic expression bias remains unknown. In mammals, DNA methylation is often associated with genes which show allele-specific expression. DNA methylation systems have been described in species of Hymenoptera, providing a candidate mechanism. Using previously generated RNA-Seq and whole-genome bisulfite sequencing from reproductive and sterile bumblebee (Bombus terrestris) workers, we have identified genome-wide allele-specific expression and allele-specific DNA methylation. The majority of genes displaying allele-specific expression are common between reproductive and sterile workers and the proportion of allele-specific expression bias generally varies between genetically distinct colonies. We have also identified genome-wide allele-specific DNA methylation patterns in both reproductive and sterile workers, with reproductive workers showing significantly more genes with allele-specific methylation. Finally, there is no significant overlap between genes showing allele-specific expression and allele-specific methylation. These results indicate that cis-acting DNA methylation does not directly drive genome-wide allele-specific expression in this species.

The nuclear and mitochondrial genomes of Frieseomelitta varia \u2013 a highly eusocial stingless bee (Meliponini) with a permanently sterile worker caste

BackgroundMost of our understanding on the social behavior and genomics of bees and other social insects is centered on the Western honey bee, Apis mellifera. The genus Apis, however, is a highly derived branch comprising less than a dozen species, four of which genomically characterized. In contrast, for the equally highly eusocial, yet taxonomically and biologically more diverse Meliponini, a full genome sequence was so far available for a single Melipona species only. We present here the genome sequence of Frieseomelitta varia, a stingless bee that has, as a peculiarity, a completely sterile worker caste.ResultsThe assembly of 243,974,526 high quality Illumina reads resulted in a predicted assembled genome size of 275 Mb composed of 2173 scaffolds. A BUSCO analysis for the 10,526 predicted genes showed that these represent 96.6% of the expected hymenopteran orthologs. We also predicted 169,371 repetitive genomic components, 2083 putative transposable elements, and 1946 genes for non-coding RNAs, largely long non-coding RNAs. The mitochondrial genome comprises 15,144 bp, encoding 13 proteins, 22 tRNAs and 2 rRNAs. We observed considerable rearrangement in the mitochondrial gene order compared to other bees. For an in-depth analysis of genes related to social biology, we manually checked the annotations for 533 automatically predicted gene models, including 127 genes related to reproductive processes, 104 to development, and 174 immunity-related genes. We also performed specific searches for genes containing transcription factor domains and genes related to neurogenesis and chemosensory communication.ConclusionsThe total genome size for F. varia is similar to the sequenced genomes of other bees. Using specific prediction methods, we identified a large number of repetitive genome components and long non-coding RNAs, which could provide the molecular basis for gene regulatory plasticity, including worker reproduction. The remarkable reshuffling in gene order in the mitochondrial genome suggests that stingless bees may be a hotspot for mtDNA evolution. Hence, while being just the second stingless bee genome sequenced, we expect that subsequent targeting of a selected set of species from this diverse clade of highly eusocial bees will reveal relevant evolutionary signals and trends related to eusociality in these important pollinators.

Diet quantity influences caste determination in honeybees (Apis mellifera).

In species that care for their young, provisioning has profound effects on offspring fitness. Provisioning is important in honeybees because nutritional cues determine whether a female becomes a reproductive queen or sterile worker. A qualitative difference between the larval diets of queens and workers is thought to drive this divergence; however, no single compound seems to be responsible. Diet quantity may have a role during honeybee caste determination yet has never been formally studied. Our goal was to determine the relative contributions of diet quantity and quality to queen development. Larvae were reared in vitro on nine diets varying in the amount of royal jelly and sugars, which were fed to larvae in eight different quantities. For the middle diet, an ad libitum quantity treatment was included. Once adults eclosed, the queenliness was determined using principal component analysis on seven morphological measurements. We found that larvae fed an ad libitum quantity of diet were indistinguishable from commercially reared queens, and that queenliness was independent of the proportion of protein and carbohydrate in the diet. Neither protein nor carbohydrate content had a significant influence on the first principle component 1 (PC1), which explained 64.4% of the difference between queens and workers. Instead, the total quantity of diet explained a significant amount of the variation in PC1. Large amounts of diet in the final instar were capable of inducing queen traits, contrary to the received wisdom that queen determination can only occur in the third instar. These results indicate that total diet quantity fed to larvae may regulate the difference between queen and worker castes in honeybees.

A method combining TA cloning and fluorescence screening for rapid acquisition of transgenic seeds.

The establishment of transgenic plants has greatly promoted the progress of plant research. However, traditional selection methods using antibiotics or herbicides may miss any positive transformants with growth defects. Additionally, screening with antibiotics/herbicides requires a huge amount of seeds, sterile work conditions and alarge amount of space to germinate plants, making the selection process time- and labor-consuming. In this study, we constructed a novel stable transformation vector, plasmid of OLE1-GFP T-DNA vector (pOGT), which can shorten the steps of cloning foreign genes into expression vectors by using TA cloning. Additionally, selection of transformed seeds with fluorescence overcomes the difficulties of conventional selection with antibiotics/herbicides and simplifies the screening process for transgenic plants.

Use of Isolation Technologies for Sterility Testing of Medicinal Products

The importance of using isolation technologies, e.g., an isolator, for sterility testing of medicines is demonstrated. The advantages and disadvantages for the Sterility parameter of using isolators in manufacturing processes and pharmaceutical analysis during examination and quality control stages are discussed. The goal of the work was to optimize the chamber biological cleaning mode with a working load in a GPPI isolator (ESCO, Indonesia), which enables this equipment to be used for sterility testing of medicines. The parameters on which the success of the biodecontamination cycle of the isolator chamber depends are analyzed. Potential biological cleaning modes are studied. The optimum mode for the working load using the GPPI isolator as an example is found. The isolator system is proven to provide sterile working conditions corresponding to cleanroom class A, which makes the equipment suitable for sterility testing of medicines.