Amanda Carroll-Portillo, October Barnes, Cristina N. Coffman
et al.
Interactions between bacteriophages with mammalian immune cells are of great interest and most phages possess at least one molecular pattern (nucleic acid, sugar residue, or protein structure) that is recognizable to the immune system through pathogen associated molecular pattern (PAMP) receptors (i.e., TLRs). Given that phages reside in the same body niches as bacteria, they share the propensity to stimulate or quench immune responses depending on the nature of their interactions with host immune cells. While most in vitro research focuses on the outcomes of direct application of phages to immune cells of interest, the potential impact of their transcytosis through the intestinal barrier has yet to be considered. As transcytosis through intestinal cells is a necessary step in healthy systems for access by phage to the underlying immune cell populations, it is imperative to understand how this step may play a role in immune activation. We compared the activation of macrophages (as measured by TNFα secretion) following direct phage application to those stimulated by incubation with phage transcytosed through a polarized Caco2 epithelial barrier model. Our results demonstrate that phages capable of activating TNFα secretion upon direct contact maintain the stimulatory capability following transcytosis. Furthermore, activation of macrophages by a transcytosed phage is enhanced as compared to that occurring with an equivalent multiplicity of directly applied phage.
Abstract Mosquitoes are notorious insects that transmit a wide range of infectious diseases, including zika, malaria, chikungunya, filariasis, and dengue. The overuse and incorrect application of synthetic pesticides to control mosquitoes has resulted in resistance development and environmental contamination, both of which have had a negative impact on human health. To address this issue, the larvicidal and pupicidal potential of acetone extract from Casearia tomentosa fruits was investigated. The extract was evaluated in a lab setting against all larval instars and pupa of Culex quinquefasciatus and Aedes albopictus, as well as against third instar larvae in a semi-field condition. Purified compounds through TLC were also tested against 3rd instar larvae of both mosquito and non-target organisms. The FT-IR and GC-MS analyses were used to characterise the extract. Morphological aberration caused by the acetone extract was observed using FESEM. The anal gills and respiratory siphon of both mosquitoes showed significant deformation from their normal state. 100 ppm was found to cause 100% larval mortality at 24 h of exposure in case of Cx. quinquefasciatus and at 72 h of exposure in Ae. albopictus larvae. After 72 h of exposure under in vitro conditions, the extract demonstrated considerable larvicidal activity with LC50 values of 38.33 and 47.56 against 3rd instar larvae of Culex quinquefasciatus and Aedes albopictus, respectively. The acetone extract can be considered as a highly effective mosquito larvicidal agent that is safe for the environment. Graphical abstract
Emmanuel Ehinmitan, Turoop Losenge, Edward Mamati
et al.
The extensive use of chemical pesticides and fertilizers in conventional agriculture has raised significant environmental and health issues, including the emergence of resistant pests and pathogens. Plant growth-promoting rhizobacteria (PGPR) present a sustainable alternative, offering dual benefits as biofertilizers and biocontrol agents. This review delves into the mechanisms by which PGPR enhance plant growth, including nutrient solubilization, phytohormone production, and pathogen suppression. PGPR’s commercial viability and application, particularly under abiotic stress conditions, are also examined. PGPR improves plant growth directly by enhancing nutrient uptake and producing growth-promoting substances and indirectly by inhibiting phytopathogens through mechanisms such as siderophore production and the secretion of lytic enzymes. Despite their potential, the commercialization of PGPR faces challenges, including strain specificity, formulation stability, and regulatory barriers. The review highlights the need for ongoing research to deepen our understanding of plant-microbe interactions and develop more robust PGPR formulations. Addressing these challenges will be crucial for integrating PGPR into mainstream agricultural practices and reducing reliance on synthetic agrochemicals. The successful adoption of PGPR could lead to more sustainable agricultural practices, promoting healthier crops and ecosystems.
Jared Van Blair, Alison Lacombe, Beatrice L. Harvey
et al.
Agricultural water is commonly treated with chlorine-based disinfectants, which are impacted by water quality. Understanding how water quality influences disinfectants such as chlorine dioxide (ClO2) against pathogenic bacteria is important for creating efficacious sanitation regimens. In this study, the minimum inhibitory concentration (MIC) of ClO2 needed to achieve a 3-Log reduction against Shiga toxin-producing Escherichia coli (STEC) and Listeria monocytogenes was compared across agricultural water samples. Sterile ddH2O served as a control to compare with environmental samples from Salinas Valley, CA, and laboratory standards. To test different dosages and water qualities, stock ClO2 was diluted in 24-well plates with target concentrations of 10, 5, 2.5, and 1.25 mg/L. Well plates were inoculated with pathogens and treated with sanitizer for 5 min. Following treatment, surviving pathogens were enumerated using viable cell counts. The results demonstrate that groundwater samples had the highest water quality of the environmental samples and required the lowest concentration of disinfectant to achieve 3-Log reduction against both bacteria, with MIC between 1.4 and 2.0 mg/L. Open-source samples had lower water quality and required a higher concentration of ClO2 for 3-Log reduction, with MIC between 2.8 and 5.8 mg/L for both pathogens. There was no correlation between pH, turbidity, or conductivity/TDS and reduction for either STEC or L. monocytogenes, suggesting no individual water metric was driving reduction. A lower dosage was required to achieve 3-Log reduction against STEC, while L. monocytogenes required greater concentrations to achieve the same level of reduction. Overall, these results help guide growers in using ClO2 as a broad-spectrum disinfectant and demonstrate its efficacy in reaching 3-Log reduction across agricultural water samples.
Pankratov Fedor, Panasenkov Dmitry, Kartashov Aleksandr
et al.
The article presents a research of the lubricity of the elements of the mechatronic transmission of an energy efficient truck. Particular attention is paid to the processes associated with ensuring reliable operation of the transmission and reducing friction between the components. Analysis of the distribution of fluids, performed using the SPH method, allows us to determine the lubricity conditions to improve performance characteristics, as well as reduce the temperature of the mechanisms to increase the service life and improve the overall efficiency of the system. The results of the research demonstrate the importance of choosing lubricants and their application modes, which directly affects the durability of transmission elements and the energy efficiency of the entire vehicle. These findings can serve as a basis for developing recommendations for optimizing the operation of mechatronic transmissions of cars.
The matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) has been widely applied in routine clinical microbiology laboratories as an efficient and reliable technique for diagnostic purpose. In this work, we evaluated the performance of the newly developed Zybio EXS3000 (Zybio Inc., China) in microbial identification and compared it with VITEK MS (bioMérieux, France). For this study, a total of 1340 isolates from various clinical specimens were collected. These isolates were analyzed simultaneously on both EXS3000 and VITEK MS. The inconsistent or unidentifiable data were further identified using the help of either 16S rRNA gene or ITS region sequencing. During the study, we observed that EXS3000 and VITEK MS provided positive confirmatory diagnostics for 95.0% and 96.5% of the isolates, respectively, which were consistent with the sequencing results. However, it is worth noting that the EXS3000 system needs to improve the identification performance of Candida albicans in the follow-up. There are no significant differences between the two devices in terms of microbial identification performance. The advantage of EXS3000 over VITEK MS is in its ability to perform in significantly lesser time period. In conclusion, the results of this investigation showed that EXS3000 can be used to identify microorganisms in clinical microbiology laboratories.
To investigate the proliferation cycle of a virus, virus-host interaction, and pathogenesis of a virus, virion particles must be concentrated from the media of virus cell culture or the sera of virus-infected patients. Ultracentrifugation of the culture media is a standard method for concentrating virion particles. However, this method is time-consuming and requires special equipment (ultracentrifuge). Moreover, a large number of infectious viruses are lost during enrichment. We developed a new method of hepatitis C virus (HCV) concentration to overcome the issues associated with traditional methods of virus concentration. We used an aqueous two-phase system (ATPS) to concentrate the virus. HCV, which causes various liver diseases, such as liver fibrosis, cirrhosis, and hepatocellular carcinoma, was used as a model virus to test the efficacy and reliability of the ATPS. The efficiency of HCV concentration by the ATPS was approximately three times higher than that by ultracentrifugation. Moreover, the infectivity of the concentrated HCV, which is a labile virus, remained the same after concentration of the virus by the ATPS. Considering the simplicity and effectiveness of the ATPS, it is the method of choice for concentrating viruses.
Saurabh Dubey, Eirill Ager-Wick, Jitendra Kumar
et al.
Aeromonas species are Gram-negative bacteria that infect various living organisms and are ubiquitously found in different aquatic environments. In this study, we used whole genome sequencing (WGS) to identify and compare the antimicrobial resistance (AMR) genes, integrons, transposases and plasmids found in Aeromonas hydrophila, Aeromonas caviae and Aeromonas veronii isolated from Indian major carp (Catla catla), Indian carp (Labeo rohita), catfish (Clarias batrachus) and Nile tilapia (Oreochromis niloticus) sampled in India. To gain a wider comparison, we included 11 whole genome sequences of Aeromonas spp. from different host species in India deposited in the National Center for Biotechnology Information (NCBI). Our findings show that all 15 Aeromonas sequences examined had multiple AMR genes of which the Ambler classes B, C and D β-lactamase genes were the most dominant. The high similarity of AMR genes in the Aeromonas sequences obtained from different host species point to interspecies transmission of AMR genes. Our findings also show that all Aeromonas sequences examined encoded several multidrug efflux-pump proteins. As for genes linked to mobile genetic elements (MBE), only the class I integrase was detected from two fish isolates, while all transposases detected belonged to the insertion sequence (IS) family. Only seven of the 15 Aeromonas sequences examined had plasmids and none of the plasmids encoded AMR genes. In summary, our findings show that Aeromonas spp. isolated from different host species in India carry multiple AMR genes. Thus, we advocate that the control of AMR caused by Aeromonas spp. in India should be based on a One Health approach.
As hypercholesterolemia is a primary risk factor for coronary artery disease and stroke, there is now an increasing demand for cholesterol-lowering drugs. Statins are a group of extremely successful drugs that lower the cholesterol level in the blood. Natural statins are produced by fermentation using different species of microorganisms. The overall aim of the present study was to identify statin-producing microfungi, which were isolated from different types of little-explored mangrove and oil palm plantation soils. Isolated fungal cultures were characterized on the basis of morphological, physiological, biochemical, and molecular features. Morphological variability was detected amongst the fungal isolates in regard to colony morphology, conidiophores structures, and conidia coloration. Based on their physiological properties and enzyme assays, rapid differentiation of statin-producing isolates was achieved. Further molecular characterization allowed reliable identification of the selected Penicillium microfungi up to the species level. The identified Penicillium cintrinum ESF2M, Penicillium brefeldianum ESF21P, and Penicillium janthinellum ESF26P strains have a scientific interest as novel wild-type producers of natural statins.
Taylor G. Glausen, Gabriela L. Carrillo, Richard M. Jin
et al.
Inflammation in the brain caused by infections lead to seizures and other neurological symptoms. But the microbial products that induce seizures as well as the host pathways downstream of these factors are largely unknown.
The presence of latently infected cells and reservoirs in HIV-1 infected patients constitutes a significant obstacle to achieve a definitive cure. Despite the efforts dedicated to solve these issues, the mechanisms underlying viral latency are still under study. Thus, on the one hand, new strategies are needed to elucidate which factors are involved in latency establishment and maintenance. On the other hand, innovative therapeutic approaches aimed at eradicating HIV infection are explored. In this context, advances of the versatile CRISPR-Cas gene editing technology are extremely promising, by providing, among other advantages, the possibility to target the HIV-1 genome once integrated into cellular DNA (provirus) and/or host-specific genes involved in virus infection/latency. This system, up to now, has been employed with success in numerous in vitro and in vivo studies, highlighting its increasing significance in the field. In this review, we focus on the progresses made in the use of different CRISPR-Cas strategies to target the HIV-1 provirus, and we then discuss recent advancements in the use of CRISPR screens to elucidate the role of host-specific factors in viral latency.
Foteini Roumani, Sarah Azinheiro, Hugo Sousa
et al.
SARS-CoV-2 is the coronavirus responsible for COVID-19, which has spread worldwide, affecting more than 200 countries, infecting over 140 million people in one year. The gold standard to identify infected people is RT-qPCR, which is highly sensitive, but needs specialized equipment and trained personnel. The demand for these reagents has caused shortages in certain countries. Isothermal nucleic acid techniques, such as loop-mediated isothermal amplification (LAMP) have emerged as an alternative or as a complement to RT-qPCR. In this study, we developed and evaluated a multi-target RT-LAMP for the detection of SARS-CoV-2. The method was evaluated against an RT-qPCR in 152 clinical nasopharyngeal swab samples. The results obtained indicated that both assays presented a “good concordance” (Cohen’s k of 0.69), the RT-LAMP was highly specific (99%) but had lower sensitivity compared to the gold standard (63.3%). The calculated low sensitivity was associated with samples with very low viral load (RT-qPCR Cq values higher than 35) which may be associated with non-infectious individuals. If an internal Cq threshold below 35 was set, the sensitivity and Cohen’s k increased to 90.9% and 0.92, respectively. The interpretation of the Cohen’s k for this was “very good concordance”. The RT-LAMP is an attractive approach for frequent individual testing in decentralized setups.
<b>Background and purpose:</b> Identifying the macromolecular targets of drug molecules is a fundamental aspect of drug discovery and pharmacology. Several drugs remain without known targets (orphan) despite large-scale in silico and in vitro target prediction efforts. Ligand-centric chemical-similarity-based methods for in silico target prediction have been found to be particularly powerful, but the question remains of whether they are able to discover targets for target-orphan drugs. <b>Experimental Approach:</b> We used one of these in silico methods to carry out a target prediction analysis for two orphan drugs: actarit and malotilate. The top target predicted for each drug was carbonic anhydrase II (CAII). Each drug was therefore quantitatively evaluated for CAII inhibition to validate these two prospective predictions. <b>Key Results:</b> Actarit showed in vitro concentration-dependent inhibition of CAII activity with submicromolar potency (IC<sub>50</sub> = 422 nM) whilst no consistent inhibition was observed for malotilate. Among the other 25 targets predicted for actarit, RORγ (RAR-related orphan receptor-gamma) is promising in that it is strongly related to actarit’s indication, rheumatoid arthritis (RA). <b>Conclusion and Implications:</b> This study is a proof-of-concept of the utility of MolTarPred for the fast and cost-effective identification of targets of orphan drugs. Furthermore, the mechanism of action of actarit as an anti-RA agent can now be re-examined from a CAII-inhibitor perspective, given existing relationships between this target and RA. Moreover, the confirmed CAII-actarit association supports investigating the repositioning of actarit on other CAII-linked indications (e.g., hypertension, epilepsy, migraine, anemia and bone, eye and cardiac disorders).
Amit Aggarwal, Ritu Singhal, Manpreet Bhalla
et al.
Introduction: Lowenstein Jensen (LJ) culture contamination is one of the most frequent problems encountered during Mycobacterium tuberculosis (M. tuberculosis) culture. Contaminated cultures are repeated at an additional cost and thus hinder diagnosis. This problem is of more significant concern in Extrapulmonary (EP) samples which have very fewer bacilli loads. Unfortunately, our current contaminant knowledge from past studies is minimal, which is entirely based on pulmonary samples, and not on EP samples. Development of newer culture methods will remain incomplete unless we have a good knowledge about contaminant profiles from both types of samples. Aim: To isolate and identify bacterial and fungal contaminants growing on LJ media during M. tuberculosis complex culture in both pulmonary and EP samples. Materials and Methods: LJ media pairs (5074) were inoculated, of which 2030 were inoculated with pulmonary samples and 3044 with EP samples. Mycobacterial, non-Mycobacterial and fungal growth were differentiated based on characteristics like colony morphology (on Chocolate agar, blood agar, Mackonkey Agar and Sabouraud’s Dextrose Agar), staining (Gram stain and Ziehl Neelsen) and biochemical reactions (Indole, Urea, Citrate and Triple Sugar Iron). Statistical analysis was done using SPSS version 20 (IBM®, New York, NY, USA) and Chi-square test was performed. Results: Overall Contamination Rate (CR) was 2.2%. Individually, CR was 2.9% (60/2030) in pulmonary samples and 1.7% (52/3044) in EP samples (p<0.05). Of the total 303 organisms isolated in the study, 87.8% (266/303) were of bacterial origin and remaining 12.2% (37/303) were fungal. Bacteria isolated belonged to 12 different genera amongst which aerobic spore bearers (Bacillus spp) were the most common. All the 37 fungal isolates were moulds, of which 22 were Aspergillus spp (A. flavus, A. fumigatus and A. niger). Bacterial contaminants were more in pulmonary samples, whereas fungal in EP samples (p<0.05). All the contaminants were breakthrough as none grew as mixture along with acid-fast organisms. Conclusion: Breakthrough nature of contaminants indicates that they probably act by completely inhibiting the growth of any acid-fast bacilli present in the sample. This observation becomes quite relevant in EP samples wherein M. tuberculosis bacilli load is already very less. This M. tuberculosis growth masking can thus decrease the overall sensitivity of LJ culture.
Introduction: Colistin is associated with dose-dependent nephrotoxicity. N-acetylcycteine (NAC) may reduce the risk of concomitant acute kidney injury (AKI) due to its antioxidant properties. We report a retrospective cohort study evaluating the role of N-acetylcysteine (NAC) in the development of colistin (COL) associated nephrotoxicity.
Methodology: A single centre retrospective cohort study was conducted in a university hospital between January 2014 and June 2015. Nephrotoxicity was defined and staged per the RIFLE (Risk, Injury, Failure, Loss of kidney function, and End-stage kidney disease) criteria. We evaluated the association between NAC use and COL-related nephrotoxicity by comparing the incidence of nephrotoxicity in patients receiving colistin with or without adjunctive NAC.
Results: Forty-six patients received intravenous (IV) COL and 46 patients received IV NAC+COL. The cumulative COL doses did not differ between the two groups (p = 0.802). The initial creatinine value doubled in 29 (63%) patients undergoing NAC+COL therapy and in 27 (58.7%) patients in the COL group (p = 0.669). The median doubling time of baseline creatinine was 6 and 7 days in the NAC+COL and COL groups, respectively. The mean hospital stay, potentially nephrotoxic agent use, and mortality rates were statistically higher for the patients receiving NAC+COL (p < 0.005).
Conclusions: The present study was not able to reveal any beneficial effect of NAC for patients undergoing COL therapy. The NAC+COL group had a higher baseline risk for development of AKI. However, the incidence of AKI was comparable between the groups. The results of the study would not solely exhibit the protective effect of adjunctive NAC therapy.
Daniel A. John, Lisa K. Williams, Venkateswarlu Kanamarlapudi
et al.
Campylobacter remain the major cause of human gastroenteritis in the Developed World causing a significant burden to health services. Campylobacter are pathogens in humans and chickens, although differences in mechanistic understanding are incomplete, in part because phenotypic strain diversity creates inconsistent findings. Here, we took Campylobacter jejuni isolates (n = 100) from multi-locus sequence typed collections to assess their pathogenic diversity, through their inflammatory, cytotoxicity, adhesion, invasion and signaling responses in a high-throughput model using avian and human intestinal epithelial cells. C. jejuni induced IL-8 and CXCLi1/2 in human and avian epithelial cells, respectively, in a MAP kinase-dependent manner. In contrast, IL-10 responses in both cell types were PI 3-kinase/Akt-dependent. C. jejuni strains showed diverse levels of invasion with high invasion dependent on MAP kinase signaling in both cell lines. C. jejuni induced diverse cytotoxic responses in both cell lines with cdt-positive isolates showing significantly higher toxicity. Blockade of endocytic pathways suggested that invasion by C. jejuni was clathrin- and dynamin-dependent but caveolae- independent in both cells. In contrast, IL-8 (and CXCLi1/2) production was dependent on clathrin, dynamin, and caveolae. This study is important because of its scale, and the data produced, suggesting that avian and human epithelial cells use similar innate immune pathways where the magnitude of the response is determined by the phenotypic diversity of the Campylobacter species.