CircZNF652 accelerates the proliferation and migration of primary lung carcinoma cells by downregulating miR-766

Purpose: To explore the biological functions and molecular mechanism of circZNF652 involvement in primary lung carcinoma.Methods: CircZNF652 levels in primary lung carcinoma cases and controls were determined using quantitative real-time polymerase chain reaction (qRT-PCR). Its prognostic value in primary lung carcinoma was examined by depicting it with Kaplan-Meier curves. The biological functions of circZNF652 in regulating proliferative and migratory capacities in A549 and SPC-A-1 cells were analyzed from the curves. Interaction between circZNF652 and its downstream gene, miR-766, wasassessed, and their co-regulation on primary lung carcinoma was determined by rescue experiments.Results: CircZNF652 was abnormally and significantly upregulated in primary lung carcinoma cases (p< 0.05), resulting in a poor prognosis. The knockdown effect of circZNF652 attenuated the proliferative and migratory capacities of A549 and SPC-A-1 cells, and downregulated epithelial-mesenchymal transition (EMT)-associated genes. CircZNF652 bound and negatively regulated miR-766, a keydownstream gene involved in circZNF652-induced aggravation of primary lung carcinoma.Conclusion: CircZNF652 serves as an oncogene, triggering the aggravation of primary lung carcinoma by negatively regulating miR-766. The results of this study may provide new insights into the treatment of lung carcinoma.

DNA/RNA heteroduplex oligonucleotide technology for regulating lymphocytes in vivo

Manipulating lymphocyte functions with gene silencing approaches is promising for treating autoimmunity, inflammation, and cancer. Although oligonucleotide therapy has been proven to be successful in treating several conditions, efficient in vivo delivery of oligonucleotide to lymphocyte populations remains a challenge. Here, we demonstrate that intravenous injection of a heteroduplex oligonucleotide (HDO), comprised of an antisense oligonucleotide (ASO) and its complementary RNA conjugated to α-tocopherol, silences lymphocyte endogenous gene expression with higher potency, efficacy, and longer retention time than ASOs. Importantly, reduction of Itga4 by HDO ameliorates symptoms in both adoptive transfer and active experimental autoimmune encephalomyelitis models. Our findings reveal the advantages of HDO with enhanced gene knockdown effect and different delivery mechanisms compared with ASO. Thus, regulation of lymphocyte functions by HDO is a potential therapeutic option for immune-mediated diseases.

Lutzomyia longipalpis Antimicrobial Peptides: Differential Expression during Development and Potential Involvement in Vector Interaction with Microbiota and Leishmania

Antimicrobial peptides (AMPs) are produced to control bacteria, fungi, protozoa, and other infectious agents. Sand fly larvae develop and feed on a microbe-rich substrate, and the hematophagous females are exposed to additional pathogens. We focused on understanding the role of the AMPs attacin (Att), cecropin (Cec), and four defensins (Def1, Def2, Def3, and Def4) in Lutzomyia longipalpis, the main vector of visceral leishmaniasis in the Americas. Larvae and adults were collected under different feeding regimens, in addition to females artificially infected by Leishmania infantum. AMPs’ gene expression was assessed by qPCR, and gene function of Att and Def2 was investigated by gene silencing. The gene knockdown effect on bacteria and parasite abundance was evaluated by qPCR, and parasite development was verified by light microscopy. We demonstrate that L. longipalpis larvae and adults trigger AMPs expression during feeding, which corresponds to an abundant presence of bacteria. Att and Def2 expression were significantly increased in Leishmania-infected females, while Att suppression favored bacteria growth. In conclusion, L. longipalpis AMPs’ expression is tuned in response to bacteria and parasites but does not seem to interfere with the Leishmania cycle.

Reversibility of the knockdown effect at application of aerosols against pyrethroid-resistant house flies

Knockdown resistance to pyrethroids and pyrethrins is the first sign of insecticide resistance, suggesting a decrease in insect nervous system sensitivity. On the Russian market of insecticides, there are many products in the aerosol package intended for the destruction of flying insects, which include pyrethroids, which provide a quick knockdown effect in insecticide susceptible insect strains. In resistant populations, in many cases, the reversibility of paralysis is observed, which indicates insufficient effectiveness of insecticidal products. The insecticidal activity of four products in aerosol package containing pyrethroids or pyrethrins in various concentrations on houseflies in natural populations was studied in comparison with the insecticide susceptible against laboratory on strain S-NIID. The most effective formulation was on the basis of natural pyrethrins. The reversibility of paralysis of resistant populations was detected using all aerosols and amounted to 0–37 % for the Kaluga strain, 10–93 % for the Krasnogorsk, KSK-1 0–87 %, KSK-2 20–99 %, S-NIID 0 %.

Evaluation Of Filter Cake Oil As Antifoam In Yeast Production. New Use For This By-Product Of The Sugarcane Derivatives Industry

Purpuse: Fodder yeast is obtained in an aerobic fermentation process where foaming is a major problem to be solved. In this article, the antifoam property of crude and purified filter cake oil is evaluated in order to use this residual as an alternative to replace the import of commercial antifoam agents and to reduce the production costs of fodder yeastMethod: Knock down test and the comparisons with two commercial antifoam agents were done. Blackstrap molasses medium at 20 and 40 g/L of total reducing sugar was used. All products were studied in their pure form and commercial ones also in dilutions 1:2 and 1:5 v/v. Hansen's solubility parameters (HSPs) to analyze the affinity of each defoamer for yeast were determined.Results: It was obtained the crude and purified filter cake oil showed similar behavior to commercial defoamers with an immediate antifoam effect, removing between 40 and 60% of the initial foam at both sugar concentrations in the first 5 minutes. The regression model showed purified filter cake oil has the greatest knockdown effect (Ca = 55.85 and 74.32) and with greater foam suppression stability the commercial defoamer Quimifoam Máster (Cb = -0.69 and -1.38). Ra values obtained in HSPs test, indicated the affinity of defoamers to the medium. Conclusion: Purified filter cake oil is an effective product for its use as an antifoam with the best knock down effect for both concentrations of sugars in the medium. The determination of HSPs corroborates the effectiveness of this product to suppress foam.

CRISPR Interference to Inducibly Repress Gene Expression in Chlamydia trachomatis

The ability to inducibly repress gene expression is critical to the study of organisms, like Chlamydia, with reduced genomes wherein the majority of genes are likely to be essential. We recently described the feasibility of a CRISPR interference system to inducibly repress gene expression in Chlamydia trachomatis. However, the initial system suffered from some drawbacks, primarily leaky expression of the anhydrotetracycline (aTc) inducible dCas9 ortholog and plasmid instability, that prevented population-wide studies (e.g. transcript analyses) of the effects of knockdown. Here, we describe various modifications to the original system that have allowed us to measure gene expression changes within a transformed population of C. trachomatis serovar L2. These modifications include (i) a change in the vector backbone, (ii) the introduction of a weaker ribosome binding site driving dCas9 translation, and (iii) the addition of a degradation tag to the dCas9 itself. With these changes, we demonstrate the ability to inducibly repress a target gene sequence as measured by the absence of protein by immunofluorescence analysis and by decreased transcript levels. Importantly, the expression of dCas9 alone (i.e. without a gRNA) had minimal impact on chlamydial growth or development. We also describe complementation of the knockdown effect by introducing a transcriptional fusion of the target gene 3’ to the dCas9. Finally, we demonstrate the functionality of a second CRISPRi system based on a dCas12 system that expands the number of potential chromosomal targets. These tools should provide the ability to study essential gene function in Chlamydia.

Knockdown of calreticulin, laccase, and Snf7 Genes Through RNAi Is Not Effective to Control the Asian Citrus Psyllid (Hemiptera: Livideae)

The Asian citrus psyllid, Diaphorina citri Kuwayama, transmits the bacteria Candidatus Liberibacter associated with huanglongbing (HLB), a devastating disease of the citrus industry. The use of genetically modified plants is an alternative to control this vector. Conversely, technology based on RNA interference (RNAi) for silencing specific genes of a target insect could be attempted. This work evaluated the knockdown effect of the target genes calreticulin (DcCRT), laccase (DcLAC), and Snf7 (DcSnf7) by RNAi through feeding D. citri in Murraya paniculata leaves after the uptake of an aqueous solution with dsRNA homologous to each vector target gene. Confocal microscopy revealed the uptake of the fluorescent-labeled dsRNA by detached leaves and the symplastic movement, allowing the ingestion by the feeding insect. A reduction in the survival rate was observed only 144 h after the beginning of feeding with dsRNA targeting DcSnf7; however, no reduction in transcript accumulation. The knockdown of the DcCRT and DcLAC genes was detected only 12 and 96 h after insect feeding, respectively. Additionally, a reduction in amino acid excretion from insects fed with dsRNA targets to DcCRT and DcLAC was observed 120 h after the beginning of feeding. However, the effects of the dsRNAs tested here appear to be minimal, both at the transcriptional and phenotype levels. For most concentrations and time points, no effects were observed. Therefore, the knockdown of genes DcCRT, DcLAC, and DcSnf7 do not appear to have the potential to control of D. citri through RNAi-mediated gene silencing.

Permethrin Resistance Status and Associated Mechanisms in Aedes albopictus (Diptera: Culicidae) From Chiapas, Mexico

There are major public health concerns regarding the spread of mosquito-borne diseases such as dengue, Zika, and chikungunya, which are mainly controlled by using insecticides against the vectors, Aedes aegypti (Linnaeus) and Aedes albopictus (Skuse). Pyrethroids are the primary class of insecticides used for vector control, due to their rapid knockdown effect and low toxicity to vertebrates. Unfortunately, continued use of pyrethroids has led to widespread insecticide resistance in Ae. aegypti; however, we lack information for Ae. albopictus—a sympatric species in Chiapas since 2002. In this study, we evaluated the permethrin resistance status of Ae. albopictus collected from Mexico and Texas. We also selected for permethrin resistance in the laboratory and investigated the potential mechanisms conferring resistance in this species. Knockdown resistance mutations, specifically F1534C, in the voltage-gated sodium channel gene, and increased activity of detoxifying enzymes were evaluated. Low levels of permethrin resistance (&lt;2.4-fold) were observed in our field populations of Ae. albopictus and the F1534C mutation was not detected in any of the sites. Low levels of resistance were also observed in the artificially selected strain. There was significantly higher cytochrome P450 activity in our permethrin-selected and nonselected strains from Mexico compared to the control strain. Our results suggest the Ae. albopictus sampled from 2016 are mostly susceptible to pyrethroids. These results contrast with the high levels of permethrin resistance (&gt;58-fold) found in Ae. aegypti from the same sites in Mexico. This research indicates the importance of continued monitoring of Ae. albopictus populations to prevent resistance from developing in the future.

Comparative Studies on the Efficacy and Mode of Action of Chlorpyrifos, Cypermethrin and ‘Tofeto’ Against Anopheles spp (Diptera: Culicidae)

The efficacy of ‘Tofeto’ and two synthetic insecticides, chlorpyrifos and cypermettrin were compared as fumigants and repellants against Anopheles spp. Knockdown, mortality and repellency responses were noted and compared. ‘Tofeto’ had a significantly higher knockdown effect on mosquitoes, while chlorpyrifos had the least. Cypermethrin had the fastest mortality time. ‘Tofeto’ compared favourably with cypermethrin and chlorpyrifos as a repellant having the highest percentage repellence and repellency index. Keywords: Synthetic insecticides, Repellants, Haematophagus, Anthropophilic, Knockdown

Oxidase, Esterase, and KDR-Associated Pyrethroid Resistance in Culex quinquefasciatus Field Collections of Collier County, Florida

ABSTRACT In several insect species, resistance to pyrethroids and DDT (dichlorodiphenyltrichloroethane) is linked to point mutations in the voltage-gated sodium channel (VGSC) gene. Pyrethroid-based insecticides prolong the opening of sodium channels, causing paralysis known as a “knockdown” effect before mortality occurs. Point mutations in the VGSC gene result in decreased pyrethroid binding and reduced sensitivity to the insecticide—this resistance mechanism is known as knockdown resistance (kdr) as insects do not die but recover from paralysis with time. In Culex mosquito species loss of target site sensitivity to pyrethroids is linked to a number of substitutions, one of which is leucine (L) to phenylalanine (F) at residue 1014 (L1014F) in the VGSC gene. Here we report the identification of kdr-associated pyrethroid resistance and developing resistance in Cx. quinquefasciatus field collections from Collier County, FL. Evaluation of position 1014 of the VGSC in Cx. quinquefasciatus collections from 7 locations in Collier County, FL, revealed a wide range of genotypes from one part of the district to the other. Centers for Disease Control and Prevention bottle bioassay, linear regression analysis, and cage trial evaluations suggest that the L1014F mutation plays a role, at least in part, to the pyrethroid resistance status of Cx. quinquefasciatus collected in Collier County, FL. Furthermore, we identified resistance attributed to both oxidase and esterase activity, indicating that multiple mechanisms are responsible for pyrethroid resistance in Collier County Cx. quinquefasciatus.