PfAP2-EXP2, an Essential Transcription Factor for the Intraerythrocytic Development of Plasmodium falciparum

Plasmodium falciparum undergoes a series of asexual replications in human erythrocytes after infection, which are effective targets for combatting malaria. Here, we report roles of an ApiAP2 transcription factor PfAP2-EXP2 (PF3D7_0611200) in the intraerythrocytic developmental cycle of P. falciparum. PfAP2-EXP2 conditional knockdown resulted in an asexual growth defect but without an appreciable effect on parasite morphology. Further ChIP-seq analysis revealed that PfAP2-EXP2 targeted genes related to virulence and interaction between erythrocytes and parasites. Especially, PfAP2-EXP2 regulation of euchromatic genes does not depend on recognizing specific DNA sequences, while a CCCTAAACCC motif is found in its heterochromatic binding sites. Combined with transcriptome profiling, we suggest that PfAP2-EXP2 is participated in the intraerythrocytic development by affecting the expression of genes related to cell remodeling at the schizont stage. In summary, this study explores an ApiAP2 member plays an important role for the P. falciparum blood-stage replication, which suggests a new perspective for malaria elimination.

MicroRNA and cDNA-Microarray as Potential Targets against Abiotic Stress Response in Plants: Advances and Prospects

Abiotic stresses, such as temperature (heat and cold), salinity, and drought negatively affect plant productivity; hence, the molecular responses of abiotic stresses need to be investigated. Numerous molecular and genetic engineering studies have made substantial contributions and revealed that abiotic stresses are the key factors associated with production losses in plants. In response to abiotic stresses, altered expression patterns of miRNAs have been reported, and, as a result, cDNA-microarray and microRNA (miRNA) have been used to identify genes and their expression patterns against environmental adversities in plants. MicroRNA plays a significant role in environmental stresses, plant growth and development, and regulation of various biological and metabolic activities. MicroRNAs have been studied for over a decade to identify those susceptible to environmental stimuli, characterize expression patterns, and recognize their involvement in stress responses and tolerance. Recent findings have been reported that plants assign miRNAs as critical post-transcriptional regulators of gene expression in a sequence-specific manner to adapt to multiple abiotic stresses during their growth and developmental cycle. In this study, we reviewed the current status and described the application of cDNA-microarray and miRNA to understand the abiotic stress responses and different approaches used in plants to survive against different stresses. Despite the accessibility to suitable miRNAs, there is a lack of simple ways to identify miRNA and the application of cDNA-microarray. The elucidation of miRNA responses to abiotic stresses may lead to developing technologies for the early detection of plant environmental stressors. The miRNAs and cDNA-microarrays are powerful tools to enhance abiotic stress tolerance in plants through multiple advanced sequencing and bioinformatics techniques, including miRNA-regulated network, miRNA target prediction, miRNA identification, expression profile, features (disease or stress, biomarkers) association, tools based on machine learning algorithms, NGS, and tools specific for plants. Such technologies were established to identify miRNA and their target gene network prediction, emphasizing current achievements, impediments, and future perspectives. Furthermore, there is also a need to identify and classify new functional genes that may play a role in stress resistance, since many plant genes constitute an unexplained fraction.

A Novel Flow Cytometric Approach for the Quantification and Quality Control of Chlamydia trachomatis Preparations

Chlamydia trachomatis is an obligate intracellular pathogenic bacterium with a biphasic developmental cycle manifesting two distinct morphological forms: infectious elementary bodies (EBs) and replicative intracellular reticulate bodies (RBs). Current standard protocols for quantification of the isolates assess infectious particles by titering inclusion-forming units, using permissive cell lines, and analyzing via immunofluorescence. Enumeration of total particle counts is achieved by counting labeled EBs/RBs using a fluorescence microscope. Both methods are time-consuming with a high risk of observer bias. For a better assessment of C. trachomatis preparations, we developed a simple and time-saving flow cytometry-based workflow for quantifying small particles, such as EBs with a size of 300 nm. This included optimization of gain and threshold settings with the addition of a neutral density filter for small-particle discrimination. The nucleic acid dye SYBR® Green I (SGI) was used together with propidium iodide and 5(6)-carboxyfluorescein diacetate to enumerate and discriminate between live and dead bacteria. We found no significant differences between the direct particle count of SGI-stained C. trachomatis preparations measured by microscopy or flow cytometry (p > 0.05). Furthermore, we completed our results by introducing a cell culture-independent viability assay. Our measurements showed very good reproducibility and comparability to the existing state-of-the-art methods, indicating that the evaluation of C. trachomatis preparations by flow cytometry is a fast and reliable method. Thus, our method facilitates an improved assessment of the quality of C. trachomatis preparations for downstream applications.

Localized Peptidoglycan Biosynthesis in Chlamydia trachomatis Conforms to the Polarized Division and Cell Size Reduction Developmental Models

Cell size regulation in bacteria is a function of two basic cellular processes: the expansion of the cell envelope and its constriction at spatially defined points at what will eventually become the division plane. In most bacterial species, both cell wall expansion and restriction are dependent on peptidoglycan (PG), a structural polymer comprised of sugars and amino acids that imparts strength and rigidity to bacterial membranes. Pathogenic Chlamydia species are unique in that their cell walls contain very little PG, which is restricted almost entirely to the apparent division plane of the microbe’s replicative forms. Very little is known about the degree to which PG affects the size and shape of C. trachomatis during its division process, and recent studies suggest the process is initiated via a polarized mechanism. We conducted an imaging study to ascertain the dimensions, orientation, and relative density of chlamydial PG throughout the organism’s developmental cycle. Our analysis indicates that PG in replicating C. trachomatis can be associated with four, broad structural forms; polar/septal disks, small/thick rings, large rings, and small/thin rings. We found that PG density appeared to be highest in septal disks and small/thick rings, indicating that these structures likely have high PG synthesis to degradation ratios. We also discovered that as C. trachomatis progresses through its developmental cycle PG structures, on average, decrease in total volume, indicating that the average cell volume of chlamydial RBs likely decreases over time. When cells infected with C. trachomatis are treated with inhibitors of critical components of the microbe’s two distinct PG synthases, we observed drastic differences in the ratio of PG synthesis to degradation, as well as the volume and shape of PG-containing structures. Overall, our results suggest that C. trachomatis PG synthases differentially regulate the expansion and contraction of the PG ring during both the expansion and constriction of the microbe’s cell membrane during cell growth and division, respectively.

Drought Stress in Grain Legumes: Effects, Tolerance Mechanisms and Management

Grain legumes are important sources of proteins, essential micronutrients and vitamins and for human nutrition. Climate change, including drought, is a severe threat to grain legume production throughout the world. In this review, the morpho-physiological, physio-biochemical and molecular levels of drought stress in legumes are described. Moreover, different tolerance mechanisms, such as the morphological, physio-biochemical and molecular mechanisms of legumes, are also reviewed. Moreover, various management approaches for mitigating the drought stress effects in grain legumes are assessed. Reduced leaf area, shoot and root growth, chlorophyll content, stomatal conductance, CO2 influx, nutrient uptake and translocation, and water-use efficiency (WUE) ultimately affect legume yields. The yield loss of grain legumes varies from species to species, even variety to variety within a species, depending upon the severity of drought stress and several other factors, such as phenology, soil textures and agro-climatic conditions. Closure of stomata leads to an increase in leaf temperature by reducing the transpiration rate, and, so, the legume plant faces another stress under drought stress. The biosynthesis of reactive oxygen species (ROS) is the most detrimental effect of drought stress. Legumes can adapt to the drought stress by changing their morphology, physiology and molecular mechanism. Improved root system architecture (RSA), reduced number and size of leaves, stress-induced phytohormone, stomatal closure, antioxidant defense system, solute accumulation (e.g., proline) and altered gene expression play a crucial role in drought tolerance. Several agronomic, breeding both conventional and molecular, biotechnological approaches are used as management practices for developing a drought-tolerant legume without affecting crop yield. Exogenous application of plant-growth regulators (PGRs), osmoprotectants and inoculation by Rhizobacteria and arbuscular mycorrhizal fungi promotes drought tolerance in legumes. Genome-wide association studies (GWASs), genomic selection (GS), marker-assisted selection (MAS), OMICS-based technology and CRISPR/Cas9 make the breeding work easy and save time in the developmental cycle to get resistant legumes. Several drought-resistant grain legumes, such as the chickpea, faba bean, common bean and pigeon pea, were developed by different institutions. Drought-tolerant transgenic legumes, for example, chickpeas, are developed by introgressing desired genes through breeding and biotechnological approaches. Several quantitative trait loci (QTLs), candidate genes occupying drought-tolerant traits, are identified from a variety of grain legumes, but not all are under proper implementation. Hence, more research should be conducted to improve the drought-tolerant traits of grain legumes for avoiding losses during drought.

Possible Involvement of Hsp90 in the Regulation of Telomere Length and Telomerase Activity During the Leishmania amazonensis Developmental Cycle and Population Proliferation

The Leishmania developmental cycle comprises three main life forms in two hosts, indicating that the parasite is continually challenged due to drastic environmental changes. The disruption of this cycle is critical for discovering new therapies to eradicate leishmaniasis, a neglected disease that affects millions worldwide. Telomeres, the physical ends of chromosomes, maintain genome stability and cell proliferation and are potential antiparasitic drug targets. Therefore, understanding how telomere length is regulated during parasite development is vital. Here, we show that telomeres form clusters spread in the nucleoplasm of the three parasite life forms. We also observed that amastigotes telomeres are shorter than metacyclic and procyclic promastigotes and that in parasites with continuous in vitro passages, telomere length increases over time. These observed differences in telomere length among parasite’s life stages were not due to lack/inhibition of telomerase since enzyme activity was detected in all parasite life stages, although the catalysis was temperature-dependent. These data led us to test if, similar to other eukaryotes, parasite telomere length maintenance could be regulated by Hsp83, the ortholog of Hsp90 in trypanosomatids, and Leishmania (LHsp90). Parasites were then treated with the Hsp90 inhibitor 17AAG. The results showed that 17AAG disturbed parasite growth, induced accumulation into G2/M phases, and telomere shortening in a time-dependent manner. It has also inhibited procyclic promastigote’s telomerase activity. Besides, LHsp90 interacts with the telomerase TERT component as shown by immunoprecipitation, strongly suggesting a new role for LHsp90 as a parasite telomerase component involved in controlling telomere length maintenance and parasite life span.

The efficiency of methods for catching insects - vectors of vector-borne diseases of animals and their species composition

The article provides the results of study of the effectiveness of collection methods and the taxonomic identification of insects in the regions of Russia. During the research three methods of collecting insects were used: a UV trap, fly strips and a liquid gadfly trap (“death puddle”). The following blood-sucking insects play a key role in the epizootology of transmissible infections - houseflies (Muscidae), black flies (Simuliidae), mosquitoes (Culicidae), biting midges (Ceratopogonidae), gadflies (Tabanidae). There has been obtained new information on the species, territorial and temporal dynamics of the distribution of vectors of lumpy skin disease in cattle in the subjects of the Russian Federation. Data collection of the spatial and temporal spread of the disease visualized using GIS-technologies have been generated on the basis of the statistical reports. As the result of the research, it has been established that in the Nizhny Novgorod region representatives of the Psychodidae family, the percentage of which was 40.9 %, predominate in the nocturnal entomological complex. They are followed by mosquitoes (genus Culex) - 21.6 %, biting midges - 16.4 % manure flies - 7.0 % and black flies - 3.0 %, respectively. In the Saratov region, the nocturnal entomological complex was represented by manure flies (family Sphaeroceridae), black flies (family Simuliidae) and mosquitoes (genus Culex), the percentages were 56,0, 32,0 and 12,0 %, respectively. At the same time, in the Saratov region there were collected 239 times less mosquitoes than in the Nizhny Novgorod region, which was due to an increase in temperature in the conditions of the southern regions. This caused the water bodies to dry out and reduced the pool of insects whose developmental cycle is related to water. It has been established that all-year keeping of cattle in winter cow yards provides the diversity and rise in the number of insect vectors, which increases the risk of lumpy skin disease as compared to the grazing system of cattle keeping. For collection daytime insect vectors, it is recommended to use fly strips covered with rosin and mineral oil. For collection insects of the nocturnal entomocomplex, which are the main transmitters of the lumpy skin disease virus, one should use ultraviolet traps.

Molecular Modeling the Proteins from the exo-xis Region of Lambda and Shigatoxigenic Bacteriophages

Despite decades of intensive research on bacteriophage lambda, a relatively uncharacterized region remains between the exo and xis genes. Collectively, exo-xis region genes are expressed during the earliest stages of the lytic developmental cycle and are capable of affecting the molecular events associated with the lysogenic-lytic developmental decision. In Shiga toxin-producing E. coli (STEC) and enterohemorragic E. coli (EHEC) that are responsible for food- and water-borne outbreaks throughout the world, there are distinct differences of exo-xis region genes from their counterparts in lambda phage. Together, these differences may help EHEC-specific phage and their bacterial hosts adapt to the complex environment within the human intestine. Only one exo-xis region protein, Ea8.5, has been solved to date. Here, I have used the AlphaFold and RoseTTAFold machine learning algorithms to predict the structures of six exo-xis region proteins from lambda and STEC/EHEC phages. Together, the models suggest possible roles for exo-xis region proteins in transcription and the regulation of RNA polymerase.

Host–Pathogen Interactions of Chlamydia trachomatis in Porcine Oviduct Epithelial Cells

Chlamydia trachomatis (Ct) causes the most prevalent bacterial sexually transmitted disease leading to ectopic pregnancy and infertility. Swine not only have many similarities to humans, but they are also susceptible to Ct. Despite these benefits and the ease of access to primary tissue from this food animal, in vitro research in swine has been underutilized. This study will provide basic understanding of the Ct host–pathogen interactions in porcine oviduct epithelial cells (pOECs)—the counterparts of human Fallopian tube epithelial cells. Using NanoString technology, flow cytometry, and confocal and transmission-electron microscopy, we studied the Ct developmental cycle in pOECs, the cellular immune response, and the expression and location of the tight junction protein claudin-4. We show that Ct productively completes its developmental cycle in pOECs and induces an immune response to Ct similar to human cells: Ct mainly induced the upregulation of interferon regulated genes and T-cell attracting chemokines. Furthermore, Ct infection induced an accumulation of claudin-4 in the Ct inclusion with a coinciding reduction of membrane-bound claudin-4. Downstream effects of the reduced membrane-bound claudin-4 expression could potentially include a reduction in tight-junction expression, impaired epithelial barrier function as well as increased susceptibility to co-infections. Thereby, this study justifies the investigation of the effect of Ct on tight junctions and the mucosal epithelial barrier function. Taken together, this study demonstrates that primary pOECs represent an excellent in vitro model for research into Ct pathogenesis, cell biology and immunity.

Dynamics of Aedes albopictus invasion. Insights from a spatio-temporal model

France displays a latitudinal range for the expansion of Aedes albopictus invasive populations that is not yet completely colonized providing a critical opportunity to address key invasion processes. We propose a spatio-temporal model (DISTIGRI) to describe and predict current and future expansion at both intra- and inter-annual scales of Aedes albopictus. This process-based model integrates mechanistic descriptions of the developmental cycle and the dispersal process of Aedes albopictus within a reaction-diffusion framework, depending on climatic suitability and photoperiod with a high spatio-temporal resolution. Using this model coupled with a climatic database, we propose several maps describing the current intra-annual distribution of Aedes albopictus, including the date of first emergence and the length of the period with significant adult presence. We also compute its future distribution over the next 10 years under several climatic scenarios, which shows a range expansion with a strong dependence on the climatic scenario. The outputs of the model may constitute a valuable asset for designing control and avoidance strategies, and to anticipate the biting nuisance with a high spatio-temporal resolution. These outputs also emphasize the importance of taking both dispersal and life cycle into account to obtain accurate descriptions of out-of-equilibrium processes such as ongoing invasions.