Infection Levels Research Articles

Exploring pathogen population density as a metric for understanding post-COVID infectious disease surges.

After the easing of COVID-19 restrictions, peaks of common infectious diseases surpassed pre-pandemic levels, raising questions about causes and ways to monitor these changes. A proposed measure, the Pathogen Population Density (PPD) score, could help track these shifts. PPD refers to the concentration of infectious agents within a population at a given time and location, serving as a potential indicator of infection levels in susceptible individuals at the population level. It is likely that PPD remains relatively stable within a specific community, as an equilibrium forms between infections and susceptibility. During the pandemic, nonpharmaceutical interventions (NPIs) led to a reduction in infectious diseases, possibly lowering population immunity and decreasing the PPD score. Once NPIs were lifted, the PPD score likely increased sharply due to a larger pool of susceptible individuals, causing more primary infections and stronger recurrent infections, faster transmission, and more severe pathogenic outcomes at the individual level. Monitoring the PPD score over time could help predict when infection peaks will occur. PPD is influenced by factors such as public health strategies, vaccination programs, and the behavior of high-risk individuals. As a quantitative measure, PPD has the potential to serve as a valuable predictive and monitoring tool, helping public health officials anticipate and track changes in infectious disease dynamics. It could be an effective tool for managing future outbreaks or pandemics and serve as a communication tool between scientists and the public to understand the emergence of new disease peaks.

Healthy lifestyle and HPV infection risk: results from a cross-sectional study.

Human Papillomavirus (HPV) infection constitutes a significant global public health challenge despite the widespread implementation of vaccination programs, with infection rates persistently high. Recent studies suggest that lifestyle factors including diet quality (DQ) and physical activity (PA) could play a pivotal role in the mitigation of HPV infections. This investigation explored the influence of DQ, PA, and a healthy lifestyle on the incidence of HPV infection in adult women. Data from 5,308 women aged 20-59 from the National Health and Nutrition Examination Survey (NHANES) 2007-2016. DQ and PA were measured using the Healthy Eating Index (HEI-2015) and the Global Physical Activity Questionnaire (GPAQ), respectively, creating four lifestyle groups based on how well they matched dietary and activity guidelines. The associations between key factors and HPV infection were explored using multivariate logistic regression, trend tests, and interaction tests. Fully adjusted multivariable logistic regression models revealed an inverse association between the risk of HPV infection and higher levels of PA (OR = 0.914; 95% CI: 0.854-0.979) as well as DQ (OR = 0.993; 95% CI: 0.989-0.998). Individuals in the highest tertile of DQ displayed a reduced risk of HPV infection relative to those in the lowest tertile (OR = 0.846; 95% CI: 0.726-0.986). Belonging to the third quintile of PA was linked to a lower risk of HPV infection than the lowest quintile (OR = 0.823; 95% CI: 0.681-0.995). Subgroups adhering to a healthy DQ were linked to a lower risk of HPV infection irrespective of PA meeting guideline recommendations. Our findings underscore the importance of a healthy diet in conjunction with appropriate PA in preventing HPV infection, offering new insights for public health policies and interventions.

Leveraging machine learning to discriminate wheat scab infection levels through hyperspectral reflectance and feature selection methods

Real-time or pre-symptomatic wheat scab (WS) detection is inevitable for precision agriculture to secure yield and quality at the critical grain formation stage. For this, feature selection (FS) techniques and machine learning (ML) have demonstrated their capabilities. However, for the same type and size of dataset, all FS and ML techniques behave differently due to their diverse primary constituents. This study attempts to leverage ML for WS classification and prediction employing different FS techniques on hyperspectral data of wheat spikes. The spectral features were selected and assessed to regress and classify disease occurrence. Relief-F-neural net (NN) manifested the best results with classification accuracy (CA) of 67 % and 89 % at the pre-symptomatic scale and 3 days after inoculation (DAI), respectively. Followed by continuous wavelet transform (CWT)-NN with 63 % CA at the pre-symptomatic scale and CWT-Xgboost with 89 % CA at 3DAI. For prediction, random forest regression revealed best accuracy of R2 = 0.94 and RMSE = 7.70, followed by partial least squares regression with R2 = 0.90 and RMSE = 10.37. The results offer a precise quantitative benchmark for future investigations into the capacity of hyperspectral data and FS for the real-time quantification of plant diseases.

A-273 Rapid, Portable Invasive Fungal Infection Diagnostic and Candida Antimicrobial Susceptibility Testing

Abstract Background Traumatic wounds are associated with a higher risk of invasive fungal infections (IFI), which lead to a significant increase in amputation or death. Actionable treatment decisions are hampered because true infections can be difficult to differentiate from external wound contaminates, and tissue biopsy and quantification of fungal load is required to determine if treatment is necessary, which currently takes several days. GeneCapture is developing the CAPTURE 60-minute, portable infection identification platform to detect infectious levels of common fungi including the genera Aspergillus, Fusarium, and Scedosporium, the family Herpotrichiellaceae, and the order Mucorales. The company is also pursuing rapid identification and antimicrobial susceptibility testing for Candida species from multiple infection types. Methods This study seeks to determine the Limit of Detection (LoD), sensitivity and specificity of the CAPTURE ID assay for fungal pathogens. Since tissue biopsy samples must be used to assess the presence of IFI in wounds, GeneCapture has developed a manual process for the crude homogenization of muscle tissue to allow this matrix to be used as the starting sample for the ID assay. Mock samples were tested for inclusivity and exclusivity on a series of IFI ID panels against multiple strains of the included fungi, against fungi not yet specifically detected on the panel, and against bacteria that commonly infect wounds. While the LoD for yeasts may be determined in CFU/mL, there is no similar, easy-to-define metric for molds. GeneCapture has developed a quantitative PCR method to detect the single copy beta-tubulin gene to roughly correlate fungal dry mass and nuclei number to assay signal for molds cultured in Potato Dextrose Broth. Determining beta-tubulin copy numbers for extracts from mock samples then allowed detection of the progression of IFI over the course of infection. Lastly, antimicrobial susceptibility testing (AST) was performed on two species of Candida to validate utilizing GeneCapture’s rapid phenotypic AST method for fungal infections. Results To date, the CAPTURE ID panel has been tested against 27 included or excluded strains of fungi or bacteria with 97% sensitivity and 100% specificity. The current LoD for mold organisms varies from approximately 1,000 nuclei for Aspergillus species, to 100,000 nuclei for Fusarium species. Additional work to understand this range is underway. Lastly, we have correctly determined the susceptibility of Candida species using our novel approach within 4-6 hours. Conclusions GeneCapture’s ID and AST technology offer a unique solution to address the increasing prevalence of fungal infections. Our platform can identify the most common fungal components in a wound within 60 minutes of receiving a tissue biopsy sample. Candida infections occur across multiple matrices and rapidly identifying the species of Candida and then determining its phenotypic susceptibility profile will have a large impact on treating these difficult pathogens.

B-351 Genotoxic Producing Escherichia coli Protects and Promotes Breast Adenocarcinoma

Abstract Background Recent studies have linked certain colibactin producing subphylas of E. coli to colorectal cancer. The bacterium synthesizes colibactin via the pks genomic island containing a series of clbA to clbS genes. The genotoxin causes inter-strand DNA cross-links, which lead to double strand breaks and promotion of colorectal adenocarcinoma. The published studies have prioritized the colorectal regions of the body where E. coli is commonly found, but none have identified the same genotoxic relationship with other types of adenocarcinoma. In this study, we evaluate DNA damage and cellular changes of mammary breast tissue based on high and low levels of colibactin producing E. coli exposure. The goal is to evaluate if colibactin can affect cells in the mammary breast tissue promoting breast adenocarcinoma growth. Methods Two cell lines were acquired from ATCC: primary mammary epithelial cells (PCS-600-010) and MCF-7 breast adenocarcinoma cells (HTB-22). Both cell lines were split into six culture flasks and maintained until ∼80% confluence was reached using the ATCC cell culture protocols. Once all the flasks had reached the target confluence, they were divided into exposure groups of high and low concentrations of colibactin producing E. coli, a group of non-colibactin producing E. coli, a group exposed only to phosphate buffered saline, and a control group with no exposure. Cellular morphology was observed using light microscopy before, after exposure, and every 5 minutes up to 15 minutes. E. coli and colibactin DNA concentrations were quantified using quantitative polymerase chain reaction. Genomic DNA was extracted from each group of cells. The DNA from each group was whole genome sequenced and analyzed for genomic variation. Results The normal cells exposed to E. coli showed immediate lysing and continued to lyse after a 5 minute incubation; however, the adenocarcinoma positive cells showed no lysing up to 30 minutes incubation. Sequencing results show genomic variation between the groups and specific break points associated with the E. coli exposed groups. Conclusions Our results show that the presence of E. coli with mammary breast tissue can destroy normal tissue, promote lysis resistance, and allow adenocarcinoma cells to continue to grow. The colibactin producing E. coli does cause double stranded breaks and can lead to cell death. This new information points out a critical need to monitor the microbiome and infection levels of people at risk or in early stages of cancer to prevent adenocarcinoma growth and destruction of normal tissue. This study indicates that colibactin producing E. coli may play a larger role in tissue damage and promotion of other types of cancer. More research is needed to elucidate the molecular pathway and mechanisms of colibactin, which will lead to new diagnostics or therapeutics that monitor E. coli levels or inhibit colibactin’s toxic effects.

Influence of Wheat Cultivars, Infection Level, and Climate after Anthesis on Efficacy of Fungicide for Control of Fusarium Head Blight in the Huang-Huai-Hai Plain of China

Fusarium head blight (FHB), caused by the Fusarium graminearum species complex, causes significant losses in grain yield and quality of wheat (Triticum aestivum) by inducing floret sterility. Grains become contaminated with mycotoxins, especially deoxynivalenol (DON), making them unsuitable for consumption. To clarify the impact of wheat cultivar resistance, infection level, and climate after anthesis on the efficacy of a fungicide for the control of FHB, we treated two moderately susceptible cultivars and 11 susceptible cultivars with fungicide (48% phenamacril + tebuconazole) at anthesis over two years. FHB incidence (INC), disease severity index (DSI), Fusarium-damaged kernels, DON contamination, thousand-kernel weight, and yield were evaluated under artificially inoculated and naturally infected field trials in 2018 and 2021. The results of multi-factor variance analysis show that the control efficacy with respect to INC and DSI is affected by cultivar, fungicide, infection level, and climatic conditions including the average daily temperature, average daily relative humidity, and total rainfall from anthesis to 21 days after anthesis (p < 0.01). Notably, cultivar resistance (deviance = 13.34, 9.55, and 11.22) is more important than fungicide (deviance = 5.77, 6.66, and 6.69) to control the efficacy of INC, DSI, and DON. The results also suggest that infection level appears to be more important than cultivars and fungicide to control the efficacy of INC, and more important than fungicide to control the efficacy of DSI. Total rainfall is more important than other climatic factors. Our results reveal that fungicide is more effective in moderately susceptible cultivars (‘Zhengmai 9023’ and ‘Xinong 979’, 89.5%~98.9%) and some susceptible cultivars than in other susceptible cultivars (‘Zhengmai 7698’ and ‘Zhoumai 27’, 51.9%~67.2%). Thus, integrating cultivar resistance with fungicide application can be an effective strategy for the management of FHB and DON in winter wheat in the Huang-huai-hai Plain of China.

Optical quality changes of the eye during peak SARS-CoV-2 pandemic in young adults

IntroductionTo determine whether severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection affects corneal morphology and optical quality. MethodsIn this cross-sectional study, ophthalmological indicators were examined during the peak of SARS-CoV-2 infection after adjusting for public health control measures. Participants were divided into control (remained uninfected), A (infected during follow-up), and B (infected prior to the first consultation) groups. Effects of varying SARS-CoV-2 infection levels were determined using reverse transcription polymerase chain reaction. Changes in corneal morphology, backscatter, and aberrations were measured. Corneal parameters, such as flat keratometry, steep keratometry, and surface variance, vertical asymmetry, height asymmetry, and height decentration indices were considered. ResultsOverall, 110 participants (208 eyes, 42.7% male; age 17–31 years) were enrolled. Eighteen (16.3%) were infection-free during the outbreak with unchanged corneal morphology, backscatter, and aberration. In group A, 34.73 ± 9.30 days after infection, the backscatter of the anterior corneal layer and central layer (both p=0.000) decreased. Total low-order aberration, defocus, horizontal coma, and spherical aberration of the cornea increased (p<0.05), while corneal morphology after infection did not change (p>0.05). In group B, a decrease in backscattering in the corneal middle layer and an increase in horizontal coma (p<0.05) were noted. ConclusionBackscattering of the anterior and intermediate layers of the cornea decreased and corneal aberrations increased after SARS-CoV-2 infection, which affected corneal optical quality. However, corneal morphology and thickness remained unchanged. Ophthalmological indicators and optical quality should be monitored during SARS-CoV-2 infection.

Prevalence of knowledge and awareness about Helicobacter pylori infection among urban population of Kingdom of Saudi Arabia

IntroductionThis cross-sectional study investigates the prevalence of knowledge and awareness of Helicobacter pylori (H. pylori) infection among the urban population of the Kingdom of Saudi Arabia (KSA). The study is motivated by the need to understand the specific demographic segments within the urban setting and their awareness levels of this prevalent bacterial infection affecting the digestive system. MethodologyThe cross-sectional study, employing a purposive sampling method and involving 840 participants from major cities in the KSA, utilized a structured survey questionnaire to gather data on demographic factors, awareness levels, attitudes, and experiences related to H. pylori infection, with subsequent statistical analysis and adherence to ethical guidelines for participant confidentiality and privacy protection. ResultThe study involved 840 participants from major cities, revealing that 59.16% were aware of H. pylori. Demographic variations included higher participation from Jeddah and Riyadh, with the majority being females aged 18-25, Saudi nationals, and individuals with undergraduate education. About 57.02% reported experiencing suspected H. pylori symptoms, and 49.16% had prior infections, with varying severity. Hesitancy toward screening (42.61%), but 72.97% believed in preventability. Limited awareness of treatments (15.35%) and identification of factors like water and food were noted. Participants perceived the seriousness of H. pylori in the mid-range (5 to 7). ConclusionThe findings shed light on variations in awareness, knowledge gaps, and attitudes, emphasizing the need for targeted public health interventions, educational campaigns, and policy measures to address the challenges associated with H. pylori infections in urban areas.

Antioxidant and anti-glycation activities of Mandevilla velutina extract and effect on parasitemia levels in Trypanosoma cruzi experimental infection: in vivo, in vitro and in silico approaches

Ethnopharmacological relevanceMandevilla velutina (Mart. Ex Stadelm.) Woodson, known in Brazil as "infalível" and "jalapa", is a medicinal plant native from the Cerrado region (Brazilian Savannah). The underground organ (xylopodium) of this species is prepared as ethanolic extract or infusion and it is commonly used in traditional medicine to treat snake venom. Although, locals and indigenous populations from Cerrado have used M. velutina for the treatment of infection by Trypanosoma cruzi (Chagas’ disease). Aim of the studyThis study aimed to evaluate the in vitro antioxidant and anti-glycation activities of the crude hydroethanolic extract of M. velutina xylopodium. Besides, it aimed to evaluate its effect on parasitemia levels in in vivo T. cruzi experimental infection. In addition, this study aimed to determine possible interactions between the main compound of the extract and molecular targets associated with survival and virulence of T. cruzi in in silico approaches. Materials and MethodsDetermination of total polyphenols, flavonoids and steroidal aglycones content were performed. In addition, high performance liquid chromatography (HPLC) was carried out to identify main compounds of the extract. Antioxidant activity was determined by DPPH radical scavenging, ferric ion reducing power (FRAP), Thiobarbituric acid reactive species (TBARS) and Oxygen Radical Absorbance Capacity (ORAC) methods. Anti-glycation activity was demonstrated through relative mobility in electrophoresis (RME), determination of free amino groups and inhibition of AGEs formation. Determination of the action of extract in parasitemia levels was performed by T. cruzi experimental infection of mice and nitrite levels were measured in the serum of animals evaluated in this study. Molecular docking analyses of the main compound (Velutinol A) with DNA and molecular targets associated with survival and virulence of T. cruzi. ResultsPhytoconstituents evaluation exhibited the presence polyphenols, flavonoids and steroidal aglycone, and HPLC identified the major presence of Velutinol A. Antioxidant and anti-glycation evaluations showed that the extract present significant activity in all methods evaluated. In addition, extract reduced the number of trypomastigotes and increased the survival of treated animals. The treatment using extract showed an interference in the synthesis of physiological nitric oxide as an immune response to infection. In silico assays demonstrated interaction between Velutinol A and DNA and molecular targets of T. cruzi. ConclusionsThe results showed that the hydroethanolic extract of M. velutina xylopodium contains bioactive compounds including polyphenols, flavonoids and steroidal aglycones (mainly Velutinol A) of which may be responsible for the antioxidant, anti-glycation and anti-parasitic activity against T. cruzi. Trypanocidal activity of M. velutina compounds may be linked to their influence on NO synthesis during infection and/or their capacity to bind and inhibit molecules associated to virulence and survival of T. cruzi.

Potentially zoonotic pathogens and parasites in opportunistically sourced urban brown rats (Rattus norvegicus) in and around Helsinki, Finland, 2018 to 2023.

BackgroundBrown rats (Rattus norvegicus) are synanthropic rodents with worldwide distribution, which are known to harbour many zoonotic pathogens and parasites. No systematic zoonotic surveys targeting multiple pathogens and parasites have previously been conducted in urban rats in Finland.AimIn Helsinki, Finland, we explored the presence and prevalence in brown rats of certain pathogens and parasites (including helminths, viruses and bacteria) across potentially zoonotic taxa.MethodsWe opportunistically received rat carcasses from pest management operators and citizens from 2018 to 2023. We searched for heart- or lungworms, performed rat diaphragm digestion to check for Trichinella and morphologically identified intestinal helminths. We assessed virus exposure by immunofluorescence assay or PCR, and detected bacteria by PCR (Leptospira) or culture (Campylobacter).ResultsAmong the rats investigated for helminths, no heart- or lungworms or Trichinella species were detected and the most common finding was the cestode Hymenolepis nana (in 9.7% of individuals sampled, 28/288). For some of the surveyed virus taxa, several rats were seropositive (orthopoxviruses, 5.2%, 11/211; arenaviruses, 2.8%, 6/211; hantaviruses 5.2%, 11/211) or tested positive by PCR (rat hepatitis E virus, 1.8%, 4/216). Campylobacter jejuni (6.6%, 17/259) and Leptospira interrogans (1.2%, 2/163) bacteria were also present in the rat population examined.ConclusionsPrevalences of potentially zoonotic pathogens and parasites in brown rats in Helsinki appeared low. This may explain low or non-existent diagnosis levels of rat-borne pathogen and parasite infections reported in people there. Nevertheless, further assessment of under-diagnosis, which cannot be excluded, would enhance understanding the risks of zoonoses.

Universal versus taxon-specific drivers of helminth prevalence and intensity of infection.

Two key epidemiological parameters, prevalence and mean intensity of infection, together capture the abundance of macroparasite populations, the strength of density-dependent effects they experience, their potential impact on host population dynamics and the selective pressures they exert on their hosts. Yet, the drivers of the extensive variation observed in prevalence and mean intensity of infection, even among related parasite taxa infecting related hosts, remain mostly unknown. We performed phylogenetically grounded Bayesian modelling across hundreds of amphibian populations to test the effects of various predictors of prevalence and intensity of infection by six families of helminth parasites. We focused on the potential effects of key host traits and environmental factors pertinent to focal host populations, i.e. the local diversity of the amphibian community and local climatic variables. Our analyses revealed several important determinants of prevalence or intensity of infection in various parasite families, but none applying to all families. Our study uncovered no universal driver of parasite infection levels, even among parasite taxa from the same phylum, or with similar life cycles and transmission modes. Although local variables not considered here may have effects extending across taxa, our findings suggest the need for a taxon-specific approach in any attempt to predict disease dynamics and impacts in the face of environmental and climatic changes.

MOF-Encapsulated Copper-Doped Carbon Dots Nanozymes with Excellent Biological Activity Promote Diabetes Wound Healing

Abstract Poor wound healing in diabetics is primarily caused by persistently high levels of inflammation and recurrent bacterial infections. The catalytic therapy technique based on nanozyme medicine has emerged as a beacon of hope for patients with diabetic wounds. However, the use of a single-atom nanozyme may still have limitations, including nanozyme burst release, immunological clearance, and insufficient antibacterial activity. To address the aforementioned problems, we provide a new nano-catalytic therapeutic agent for diabetic skin ulcers that incorporates a single-atom nanozyme with high antioxidant activity into a metal-organic framework (ZIF-Cu/C-dots). First, a Cu single-atom nanozyme supported by ultra-small carbon dots (Cu/C-dots) with high antioxidant activity was created. A nanozyme-integrated metal-organic framework was then created, utilizing Cu/C-dots as ligands and Zn2+ as the core metal. Cu/C-dots have good oxidase-like activity, shielding the biological system from ROS damage and reducing the expression of TNF-α and IL-1β. Zn2+ also has good antibacterial activity (the antibacterial rate was more than 90%). This integrated technique prevents nanozyme aggregation, improves nanozyme biocompatibility, slows down the breakdown of ZIF, and allows for the regulated release of Cu/C-dots and Zn2+ as needed. Finally, in vivo studies have shown that ZIF-Cu/C-dots can effectively alleviate inflammation at the site of diabetic wounds, accelerate vascular regeneration, promote collagen deposition, and enhance tissue remodeling, serving as a novel nano-catalytic platform for the treatment of wounds that are difficult to heal.

A RELEVÂNCIA DOS PROGRAMAS DE AUTOCONTROLE NA SAÚDE PÚBLICA EM INDÚSTRIAS DE PRODUTOS DE ORIGEM ANIMAL

Self-Control Programs (PACs) are fundamental for the agroindustry, aiming at food safety and reducing risks to public health. Along with Good Manufacturing Practices (GMP), Standard Operational Hygiene Procedures (PPHO) and the Hazard Analysis and Critical Control Points Program (HACCP). The research aims to demonstrate the importance of PAC as a tool capable of minimizing and controlling levels of contamination, infection and/or food poisoning in the food industry, discussing the regulations issued by supervisory authorities on the quality, health and safety of food, as well as good practices in the production of products of animal origin. The importance of studying this topic is that it has great relevance due to the fact that it addresses the importance of the hygienic-sanitary and technological quality of raw materials, animal health and public health. The study was carried out using articles, books and academic sources through electronic searches in SciELO, Google Scholar, PUBMED, PubVet of relevance in addition to Laws and books, bibliographical research and insights into the need to highlight and demonstrate the importance of programs in the food industry. It was concluded that PACs are essential to reduce risks to public health and enhance food quality and safety for consumers.

Do diet and Fumagillin treatment impact Vairimorpha (Nosema) spp. (Microspora: Nosematidae) infections in honey bees (Hymenoptera: Apidae) and improve survival and growth of colonies overwintered in cold storage?

Vairimorpha (Microsporidia: Nosematidae)is a microsporidian that infects honey bees especially in winter. Fumagillin can reduce infections, but whether overwintering survival is improved is unclear. The diet also may influence the severity of Nosema infections. We examined the relationship between Nosema and colony size and survival in hives overwintered in cold storage facilities. In year 1, no Fumagillin treatments were applied. Colony size and survival after cold storage and almond bloom were comparable between groups with high and low pre-cold storage infections. In year 2, size and survival were compared among colonies with and without Fumagillin treatment that were fed either pollen or protein supplement prior to overwintering. Colonies treated with Fumagillin had lower spore numbers than untreated, but colony sizes and survival were similar among the treatments. However, more colonies with zero spores per bee could be rented for almond pollination and were alive after bloom than those averaging >1 million spores per bee. Fat body metrics can affect overwintering success. In both years, fat body weights and protein concentrations increased, and lipid concentrations decreased while bees were in cold storage. Fat body metrics did not differ with Nosema infection levels. However, Fumagillin negatively affected pre-cold storage fat body protein concentrations and colony sizes after cold storage and almond bloom. Treating with Fumagillin before overwintering in cold storage might result in greater colony survival if spore numbers are high, but undetectable or even negative effects when spore numbers are low.

Revealing real-time 3D in vivo pathogen dynamics in plants by label-free optical coherence tomography

Microscopic imaging for studying plant-pathogen interactions is limited by its reliance on invasive histological techniques, like clearing and staining, or, for in vivo imaging, on complicated generation of transgenic pathogens. We present real-time 3D in vivo visualization of pathogen dynamics with label-free optical coherence tomography. Based on intrinsic signal fluctuations as tissue contrast we image filamentous pathogens and a nematode in vivo in 3D in plant tissue. We analyze 3D images of lettuce downy mildew infection (Bremia lactucae) to obtain hyphal volume and length in three different lettuce genotypes with different resistance levels showing the ability for precise (micro) phenotyping and quantification of the infection level. In addition, we demonstrate in vivo longitudinal imaging of the growth of individual pathogen (sub)structures with functional contrast on the pathogen micro-activity revealing pathogen vitality thereby opening a window on the underlying molecular processes.

Diversity of microbial, biocontrol agents and nematode abundance on a susceptible Prunus rootstock under a Meloidogyne root gradient infection.

Root-knot nematodes (RKNs) of the genus Meloidogyne are one of the most damaging genera to cultivated woody plants with a worldwide distribution. The knowledge of the soil and rhizosphere microbiota of almonds infested with Meloidogyne could help to establish new sustainable and efficient management strategies. However, the soil microbiota interaction in deciduous woody plants infected with RKNs is scarcely studied. This research was carried out in six commercial almond groves located in southern Spain and infested with different levels of Meloidogyne spp. within each grove. Several parameters were measured: nematode assemblages, levels and biocontrol agents in Meloidogyne's eggs, levels of specific biocontrol agents in rhizoplane and soil, levels of bacteria and fungi in rhizoplane and soil, fungal and bacterial communities by high-throughput sequencing of internal transcribed spacer (ITS), and 16S rRNA gene in soil and rhizosphere of the susceptible almond hybrid rootstock GF-677 infested with Meloidogyne spp. The studied almond groves showed soil degradation by nematode assemblies and fungi:bacterial ratio. Fungal parasites of Meloidogyne eggs were found in 56.25% of the samples. However, the percentage of parasitized eggs by fungi ranged from 1% to 8%. Three fungal species were isolated from Meloidogyne eggs, specifically Pochonia chlamydosporia, Purpureocillium lilacinum, and Trichoderma asperellum. The diversity and composition of the microbial communities were more affected by the sample type (soil vs rhizosphere) and by the geographical location of the samples than by the Meloidogyne density, which could be explained by the vigorous hybrid rootstock GF-677 and a possible dilution effect. However, the saprotrophic function in the functional guilds of the fungal ASV was increased in the highly infected roots vs the low infected roots. These results indicate that the presence of biocontrol agents in almond fields and the development of new management strategies could increase their populations to control partially RKN infection levels.

Non-invasive, pre-symptomatic detection of potato cyst nematode infection in tomato using chlorophyll fluorescence analysis.

Potato cyst nematodes (PCN) are notorious pathogens in all major potato production areas worldwide. Mainly due to the low mobility of this soil pathogen, PCN infestations are mostly observed as patches ('foci') that only cover a fraction of the acreage. In-field pre-symptomatic localization of this pathogen is valuable as it would allow for the localized application of control measures. Although the mapping of foci is technically feasible, it is unpractical as it would take the analysis of numerous soil samples. We investigated whether chlorophyll fluorescence (Chl-F) could be suitable as a rapid, non-destructive method for early PCN detection. To this end, the impact of four Globodera pallida densities on the Chl-F of tomato was investigated in a phenotyping greenhouse for 26 days. Furthermore, classical plant performance indicators biomass and root surface area were compared with Chl-F. Thermal dissipation ('NPQ_Lss') and actual photosynthetic rate ('QY_Lss') responded at 1 DPI, while QY_Lss was most sensitive to low PCN infection levels. Chl-F parameters responded more readily to PCN infection than biomass and root surface area. The efficiency of photosystem II (QY_max) and the potential activity of photosystem II (Fv/Fo) initially increased at low PCN infection levels, whereas a sharp decrease was observed at higher infestation levels. Hence, our data suggest that low PCN levels promoted plant performance before becoming detrimental at higher levels. While Chl-F allowed for early and sensitive PCN detection, it remains to be investigated whether these signals can be distinguished from those produced by other below-ground stressors in the field.

PRODUCTION OF RECOMBINANT SALMONELLA ABORTUS EQUI PROTEINS

The cause of mare abortions of salmonellosis etiology is pathogen Salmonella abortusequi. The economic damage caused by this infection is due to the loss of reproductive capacity of the sires, litter failure, reduced productivity and veterinary costs. High level of infection of horses is registered in many countries, including Kazakhstan. The main method of diagnosis is bacteriological, but its disadvantages are low sensitivity and duration of results. The OIE recommends the use of PCR and ELISA for diagnosis of infection, however, due to the high cost of test systems, the use of PCR in veterinary laboratories is not always justified, and the effectiveness of the ELISA method depends largely on the structure and purity of the antigen. Currently, genetic engineering methods make it possible to obtain recombinant antigens of animal pathogens containing separate proteins, which excludes the occurrence of nonspecific reactions. These antigens can be used in tests for serological diagnostics, immunization of laboratory animals, and in the development of vaccines. According to literature data, it is known that diagnostically significant antigens in bacteria of the genus Salmonella are outer membrane proteins. Therefore, the outer membrane protein OmpX of S. abortusequi, which has high immunogenicity and conservativeness was selected as a target. The aim of this work is to obtain recombinant OmpX antigens of S. abortusequi and study their properties. As a result, E. coli strains producing recombinant OmpX protein of S. abortusequi were created. The synthesized genes were cloned into expression vectors and transformed into competent E. coli BL21 cells. The rOmpX of Salmonella abortusequi was produced and purified; electrophoretic analysis showed the purity of the preparation with a molecular mass of 23 kDa. The recombinant protein was analyzed by LC/MC-MC spectrometry, and peptides containing QTTDYPTYKHDTSDYGFSYGAGLQFN amino acid sequences were identified. A search of the peptide spectrum in the SwissProt database showed that they corresponded to the OmpX protein of Salmonella abortusequi. To determine the quality of protein refolding and the conservation of antigenic epitopes of the protein, Western blot was used with control positive sera that specifically reacted with a protein of molecular mass 23kDa. The resulting recombinant OmpX protein was then used as an antigen in an indirect ELISA to detect antibodies in serum samples from aborted mares. The results of n-ELISA showed the presence of antibodies in serum at a titer of 1:1600 (+26.6; - 21.1). Thus, rOmpX of S. abortusequiwith a molecular mass of 23 kDa was obtained and its main properties were studied. The correctness of protein refolding was confirmed by Western blot and ELISA with positive control sera. The specificity of recombinant proteins was proved by interaction with positive serum samples of horses from unfavorable farms. The obtained recombinant antigens can be used in the creation of tests for the diagnosis of salmonellosis abortion of horses in the Republic of Kazakhstan. WPM с добавлением БАП 1,0 мг/л, ИМК 0,1 мг/л, ГК 0,2 мг/л, где было в среднем образовано 7,36 микропобегов на один эксплант.

Rapid real-time quantitative colorimetric LAMP methodology for field detection of Verticillium dahliae in crude olive-plant samples

BackgroundVerticilium dahliae is the most important wilt pathogen of olive trees with a broad host range causing devastating diseases currently without any effective chemical control. Traditional detection methodologies are based on symptoms-observation or lab-detection using time consuming culturing or molecular techniques. Therefore, there is an increasing need for portable tools that can detect rapidly V. dahliae in the field.ResultsIn this work, we report the development of a novel method for the rapid, reliable and on-site detection of V. dahliae using a newly designed isothermal LAMP assay and crude extracts of olive wood. For the detection of the fungus, LAMP primers were designed targeting the internal transcribed spacer (ITS) region of the rRNA gene. The above assay was combined with a purpose-built prototype portable device which allowed real time quantitative colorimetric detection of V. dahliae in 35 min. The limit of detection of our assay was found to be 0.8 fg/μl reaction and the specificity 100% as indicated by zero cross-reactivity to common pathogens found in olive trees. Moreover, detection of V. dahliae in purified DNA gave a sensitivity of 100% (Ct < 30) and 80% (Ct > 30) while the detection of the fungus in unpurified crude wood extracts showed a sensitivity of 80% when multisampling was implemented. The superiority of the LAMP methodology regarding robustness and sensitivity was demonstrated when only LAMP was able to detect V. dahliae in crude samples from naturally infected trees with very low infection levels, while nested PCR and SYBR qPCR failed to detect the pathogen in an unpurified form.ConclusionsThis study describes the development of a new real time LAMP assay, targeting the ITS region of the rRNA gene of V. dahliae in olive trees combined with a 3D-printed portable device for field testing using a tablet. The assay is characterized by high sensitivity and specificity as well as ability to operate using directly crude samples such as woody tissue or petioles. The reported methodology is setting the basis for the development of an on-site detection methodology for V. dahliae in olive trees, but also for other plant pathogens.

An Investigation on Inception V3 Network Models for Disease Severity Identification

Due to the high incubation period of COVID-19 and the use of restricted data, it is difficult to anticipate the number of casualties and the rate at which the virus would spread in the COVID-19 scenario. Monitoring those who are impacted and those who have interacted with them is perhaps the most difficult procedure. Since COVID-19 does require 14 days to incubate, it is doubtful that patients have been detected earlier. Patients have the ability to influence everyone they interact with while they are in the designated incubation period. Typically, the various machine learning algorithms are given the COVID-19 deep properties in order to classify the illness severity levels. Consequently, this may help in the early diagnosis of COVID-19. The created a deep feature extraction and classification model that successfully classified the COVID-19 severity levels using a variety of deep and machine learning algorithms. It has the ability to classify COVID-19 severity levels for early, accurate diagnosis. Ultimately, the results showed that the Inception V3 classifier outperformed all other models in both feature extraction and classificational, successfully extracting deep features from CT images and classifying the severity levels of COVID-19 infection. The testing results also showed that the classification of deep characteristics achieves promising use in COVID-19 severity prediction and diagnosis.