Revisiting the phylogeny and taxonomy of the genus Sidera (Hymenochaetales, Basidiomycota) with particular emphasis on S.vulgaris.

The genus Sidera (Hymenochaetales, Basidiomycota) comprises white-rot, mono- or dimitic fungi with poroid or hydnoid hymenophore. It has a worldwide distribution albeit with fewer species present in the Southern Hemisphere. Although recent studies revealed the existence of several new Sidera species, there are still taxonomic inconsistencies and obscure phylogenetic relationships amongst certain taxa of the genus. In this work, a large number of Sidera collections were used to obtain an updated phylogeny, based on ITS and 28S rDNA sequences by including new material from Mediterranean Europe. The monophyly of the genus was strongly supported and all species with poroid hymenophore formed a highly-supported lineage with two major subclades. In total, 23 putative species were recognised. Amongst those, five are considered to possibly represent entities new to science, but further work is required since they are represented by single specimens or environmental sequences. Examined collections originally named S.lenis from southern Europe were grouped within S.vulgaris. Similarly, several collections under various names were hereby identified as S.vulgaris, including those of the recently described species S.tibetica. Furthermore, a critical discussion (based on morphoanatomical findings) is made on the key features that could be used to distinguish S.lenis from S.vulgaris.

Development of a novel sensitive single-tube nested PCR assay for the detection of African swine fever virus.

African swine fever (ASF) is a highly fatal and contagious viral disease caused by African swine fever virus (ASFV). It has caused significant economic losses to the swine industry and poses a serious threat to food security worldwide. Diagnostic tests with high sensitivity are essential for the effective management of ASF. Here, we describe a single-tube nested PCR (STN-PCR) assay for the detection of ASFV in which two consecutive amplification steps are carried out within a single tube. Two pairs of primers (outer and inner) were designed to target the p72 gene of ASFV. The primer concentrations, annealing temperatures, and number of amplification cycles were optimized to ensure the consecutive utilization of outer and inner primer pairs during amplification while minimizing the likelihood of amplicon contamination. In comparison with two conventional endpoint PCR assays (one of which is recommended by the World Organization for Animal Health), the newly developed STN-PCR assay demonstrated a 100-fold improvement in the limit of detection (LOD), detecting 100 copies of ASFV genomic DNA, whereas the endpoint PCR assays could detect no fewer than 10,000 copies. The clinical performance of the STN-PCR assay was validated using 95 tissue samples suspected of being positive for ASFV, and the assay showed 100% specificity. A Cohen's kappa value of 0.91 indicated perfect agreement between the assays. This new STN-PCR assay is a potentially valuable tool that will facilitate the control of ASF.

Professional values at the beginning of medical school: a quasi-experimental study.

BACKGROUND: Teaching professionalism in medical schools is central to medical education and society. We evaluated how medical students view the values of the medical profession on their first day of medical school and the influence of a conference about the competences of this profession on these students' levels of reflection. METHODS: We studied two groups of medical students who wrote narratives about the values of the medical profession and the influence of the COVID-19 pandemic on these values. The first group wrote the narratives after a conference about the competences of the medical profession (intervention group), and the second group wrote the same narratives after a biochemistry conference (control group). We also compared the levels of reflection of these two groups of students. RESULTS: Among the 175 medical students entering in the 2022 academic year, 159 agreed to participate in the study (response rate = 90.8%). There were more references to positive than negative models of doctor‒patient relationships experienced by the students (58.5% and 41.5% of responses, respectively). The intervention group referred to a more significant number of values than the control group did. The most cited values were empathy, humility, and ethics; the main competences were technical competence, communication/active listening, and resilience. The students' perspectives of the values of their future profession were strongly and positively influenced by the pandemic experience. The students realized the need for constant updating, basing medical practice on scientific evidence, and employing skills/attitudes such as resilience, flexibility, and collaboration for teamwork. Analysis of the levels of reflection in the narratives showed a predominance of reflections with a higher level in the intervention group and of those with a lower level in the control group. CONCLUSIONS: Our study showed that medical students, upon entering medical school, already have a view of medical professionalism, although they still need to present a deeper level of self-reflection. A single, planned intervention in medical professionalism can promote self-reflection. The vision of medical professional identity was strongly influenced by the COVID-19 pandemic, positively impacting the formation of a professional identity among the students who decided to enter medical school.

Prevalence of computer vision syndrome: A systematic review and meta-analysis

Purpose This review aimed to estimate the prevalence of computer vision syndrome (CVS) in the general population and subgroups. Methods A search was conducted in the following the databases: PubMed, SCOPUS, EMBASE, and Web of Science until February 13, 2023. We included studies that assessed the prevalence of CVS in any population. The Joanna Briggs Institute's critical appraisal tool was used to evaluate the methodological quality. A meta-analysis of the prevalence of CVS was done using a random-effects model, assessing the sources of heterogeneity using subgroup and meta-regression analyses. Results A total of 103 cross-sectional studies with 66 577 participants were included. The prevalence of CVS was 69.0% (95% CI: 62.3 to 75.3; I2: 99.7%), ranging from 12.1 to 97.3% across studies. Point prevalence was higher in women than in men (71.4 vs. 61.8%), university students (76.1%), Africa (71.2%), Asia (69.9%), contact lens wearers (73.1% vs. 63.8%) in studies conducted before the COVID-19 pandemic (72.8%), and in those that did not use the CVS-Q questionnaire (75.4%). In meta-regression, using the CVS-Q scale was associated with a lower prevalence of CVS. Conclusion Seven out of ten people suffer from CVS. Preventive strategies and interventions are needed to decrease the prevalence of this condition which can affect productivity and quality of life. Future studies should standardize a definition of CVS. (AU)

Seroepidemological investigation of Toxoplasma gondii and Trichinella spp. in pigs reared by tribal communities and small-holder livestock farmers in Northeastern India.

Toxoplasma gondii and Trichinella spp. are critical tissue-dwelling foodborne zoonotic parasites associated with pork consumption and pig rearing. Despite being a major pig-rearing region in the country, Northeastern India has not undergone any investigation regarding the presence of T. gondii and Trichinella spp. in pigs. Therefore, this study aims to determine the seroprevalence of T. gondii and Trichinella spp. and identify associated risk factors in pigs reared by tribal communities and small-holder livestock farmers in the northeastern region of India. In a cross-sectional serological survey, 400 pigs from 400 households across five northeastern states of India underwent testing for the seroprevalence of porcine toxoplasmosis and trichinellosis. Serum samples (80 from each state) were analyzed using commercially available ELISA assays. Data on backyard farm characteristics and various management aspects were collected, and risk factors linked with prevalence were analyzed through univariate and multivariate logistic regression analysis. The findings revealed that the apparent and true prevalence of anti-T. gondii antibodies were 45% (40.12-49.88, 95% CI) and 45.7% (40.7-50.69, 95% CI), respectively. As for anti- Trichinella antibodies, both the apparent and true prevalence were 0.75% (-0.1-1.6, 95% CI). The univariate and multivariate analyses indicated that age above 24 months (OR 7.20, 95% CI 2.45-23.71), exposure to cats (OR = 5.87, 95% CI 2.55-14.05), and farms operating for breeding purposes (OR = 5.60, 95% CI 3.01-11.04) were significant risk factors associated with the seroprevalence of T. gondii. This study marks the initial documentation of the seroprevalence of T. gondii and Trichinella spp. in pigs reared by tribal communities in Northeastern India. The results emphasize the significance of these parasites as foodborne zoonotic threats in the region, potentially posing substantial public health risks, especially within tribal and rural communities. The insights derived from this research could be valuable in formulating targeted preventive and control strategies against T. gondii and Trichinella spp. in pigs, not only in this region but also in areas with similar rearing practices.

Selective transcriptomic dysregulation of metabolic pathways in liver and retina by short- and long-term dietary hyperglycemia.

A high glycemic index (HGI) diet induces hyperglycemia, a risk factor for diseases affecting multiple organ systems. Here, we evaluated tissue-specific adaptations in the liver and retina after feeding HGI diet to mice for 1 or 12 month. In the liver, genes associated with inflammation and fatty acid metabolism were altered within 1 month of HGI diet, whereas 12-month HGI diet-fed group showed dysregulated expression of cytochrome P450 genes and overexpression of lipogenic factors including Srebf1 and Elovl5. In contrast, retinal transcriptome exhibited HGI-related notable alterations in energy metabolism genes only after 12 months. Liver fatty acid profiles in HGI group revealed higher levels of monounsaturated and lower levels of saturated and polyunsaturated fatty acids. Additionally, HGI diet increased blood low-density lipoprotein, and diet-aging interactions affected expression of mitochondrial oxidative phosphorylation genes in the liver and disease-associated genes in retina. Thus, our findings provide new insights into retinal and hepatic adaptive mechanisms to dietary hyperglycemia.

Prevalence of computer vision syndrome: A systematic review and meta-analysis.

PURPOSE: This review aimed to estimate the prevalence of computer vision syndrome (CVS) in the general population and subgroups. METHODS: A search was conducted in the following the databases: PubMed, SCOPUS, EMBASE, and Web of Science until February 13, 2023. We included studies that assessed the prevalence of CVS in any population. The Joanna Briggs Institute's critical appraisal tool was used to evaluate the methodological quality. A meta-analysis of the prevalence of CVS was done using a random-effects model, assessing the sources of heterogeneity using subgroup and meta-regression analyses. RESULTS: A total of 103 cross-sectional studies with 66 577 participants were included. The prevalence of CVS was 69.0% (95% CI: 62.3 to 75.3; I2: 99.7%), ranging from 12.1 to 97.3% across studies. Point prevalence was higher in women than in men (71.4 vs. 61.8%), university students (76.1%), Africa (71.2%), Asia (69.9%), contact lens wearers (73.1% vs. 63.8%) in studies conducted before the COVID-19 pandemic (72.8%), and in those that did not use the CVS-Q questionnaire (75.4%). In meta-regression, using the CVS-Q scale was associated with a lower prevalence of CVS. CONCLUSION: Seven out of ten people suffer from CVS. Preventive strategies and interventions are needed to decrease the prevalence of this condition which can affect productivity and quality of life. Future studies should standardize a definition of CVS.

Novel helix loop-mediated isothermal amplification (HAMP) assay for colorimetric detection of Staphylococcus aureus in milk.

Staphylococcus aureus is an important and leading cause of foodborne diseases worldwide. Prompt detection and recall of contaminated foods are crucial to prevent untoward health consequences caused by S. aureus. Helix loop-mediated isothermal amplification (HAMP) is an exciting recent addition to the array of available isothermal-based nucleic acid amplification techniques. This study aimed to develop and evaluate a HAMP assay for detecting S. aureus in milk and milk products. The assay is completed in 75 minutes of isothermal temperature incubation (64 ˚C) and dye-based visual interpretation of results based on colour change. The specificity of the developed assay was ascertained using 27 S. aureus and 17 non S. aureus bacterial strains. The analytical sensitivity of the developed HAMP assay was 9.7 fg/µL of pure S. aureus DNA. The detection limit of the HAMP assay in milk (86 CFU/mL) was 1000x greater than the routinely used endpoint PCR (86 × 103 CFU/mL). The practicality of applying the HAMP assay was also assessed by analysing milk and milk product samples (n = 95) obtained from different dairy farms and retail outlets. The developed test is a more rapid, sensitive, and user-friendly method for the high-throughput screening of S. aureus in food samples and may therefore be suitable for field laboratories. To our knowledge, this is the first study to develop and evaluate the HAMP platform for detecting S. aureus.

Biofilm-forming antimicrobial-resistant pathogenic Escherichia coli: A one health challenge in Northeast India.

This study aimed to investigate the prevalence of Shiga toxin-producing Escherichia coli (STEC), Enteropathogenic E. coli (EPEC), and Enterotoxigenic E. coli (ETEC) in common food animals (cattle, goats, and pigs) reared by tribal communities and smallholder farmers in Northeast India. The isolates were characterized for the presence of virulence genes, extended-spectrum beta-lactamases (ESBL) production, antimicrobial resistance, and biofilm production, and the results were statistically interpreted. In pathotyping 141 E. coli isolates, 10 (7.09%, 95% CI: 3.45%-12.66%) were identified as STEC, 2 (1.42%, 95% CI: 0.17%-5.03%) as atypical-EPEC, and 1 (0.71%, 95% CI: 0.02%-3.89%) as typical-EPEC. None of the isolates were classified as ETEC. Additionally, using the phenotypic combination disc method (ceftazidime with and without clavulanic acid), six isolates (46.1%, 95% CI: 19.22%-74.87%) were determined to be ESBL producers. Among the STEC/EPEC strains, eleven (84.6%, 95% CI: 54.55%-98.08%) and one (7.7%, 95% CI: 0.19%-36.03%) strains were capable of producing strong or moderate biofilms, respectively. PFGE analysis revealed indistinguishable patterns for certain isolates, suggesting clonal relationships. These findings highlight the potential role of food animals reared by tribal communities and smallholder farmers as reservoirs of virulent biofilm-forming E. coli pathotypes, with implications for food contamination and zoonotic infections. Therefore, monitoring these pathogens in food animals is crucial for optimizing public health through one health strategy.

First Seroepidemiological Investigation of Hepatitis E Virus Infection in Backyard Pigs from Northeastern India: Prevalence and Associated Risk Factors.

Hepatitis E virus (HEV) is the leading cause of acute viral hepatitis globally, with zoonotic potential, and pigs are considered the major reservoir. To determine the seroprevalence of HEV infection in pigs reared in backyard conditions in the northeastern region of India, blood samples were collected from 400 pigs from five northeastern states (80 samples from each state) and tested for IgG antibodies against HEV using an ELISA assay. Questionnaires on farm characteristics and management practices were completed, and risk factors associated with HEV were studied using univariate and multivariate analysis. The apparent seroprevalence of HEV infection was 51% (46.1-55.9, 95% CI), with a true prevalence of 52.98% (47.22-58.75, 95% CI). The risk factors significantly associated with higher HEV seropositivity were as follows: lack of disinfection (OR 4.65), feeding swill (restaurant and bakery waste) (OR 2.55), failure to follow the all-in-all-out production system (OR 3.47), and medium holding size (OR 9.83), which refers to mixed rearing of younger and older age groups. This study demonstrates that HEV is widespread among pigs reared in northeastern India. The risk factor analysis conducted in this study provides valuable insights into the prevalence of HEV in the region.

Obesity decreases energy and protein intake in hospitalized COVID-19 patients undergoing continuous positive airway pressure treatment.

Patients with SARS-CoV generally require noninvasive treatments including continuous positive airway pressure (CPAP), and appropriate nutritional therapy. Our hypothesis was that a higher body mass index (BMI) would result in greater respiratory difficulties during CPAP treatment and lead to a reduction in energy and protein intake. The study was conducted prospectively and retrospectively in a Brazilian public hospital. The sample comprised all patients (n = 70) hospitalized in COVID-19 wards and using the Elmo System (ELMO) helmet. To ensure proper nutritional support, the hospital established a standard ELMO diet and questionnaire for estimating dietary adherence. The median length of stay was 11 days (interquartile range, 9-20), and 84.3% of the patients received hospital discharge. The median duration of helmet use was 1.5 days (interquartile range, 1-4). From mid-upper arm circumference adequacy measurements, 35 patients (50%) were classified as being overweight or living with obesity and 36 adult patients (66.7%) were considered living with obesity based on their BMI. The prevalence of overweight elderly patients was 81.3%. The median energy and protein intake adequacy percentages were 65% and 72.5%, respectively. Patients with higher BMIs exhibited lower oxygen flows while wearing the ELMO helmet. Younger age and lower energy and protein intake characterized the obesity group. A correlation between oxygen therapy and food intake in COVID-19 patients undergoing CPAP treatment was observed. Patients with higher BMIs and/or mid-upper arm circumference adequacies were more susceptible to energy and protein intake declines, and patients with obesity presented the lowest intake values.

Long wavelength-sensing cones of zebrafish retina exhibit multiple layers of transcriptional heterogeneity.

Introduction: Understanding how photoreceptor genes are regulated is important for investigating retinal development and disease. While much is known about gene regulation in cones, the mechanism by which tandemly-replicated opsins, such as human long wavelength-sensitive and middle wavelength-sensitive opsins, are differentially regulated remains elusive. In this study, we aimed to further our understanding of transcriptional heterogeneity in cones that express tandemly-replicated opsins and the regulation of such differential expression using zebrafish, which express the tandemly-replicated opsins lws1 and lws2. Methods: We performed bulk and single cell RNA-Seq of LWS1 and LWS2 cones, evaluated expression patterns of selected genes of interest using multiplex fluorescence in situ hybridization, and used exogenous thyroid hormone (TH) treatments to test selected genes for potential control by thyroid hormone: a potent, endogenous regulator of lws1 and lws2 expression. Results: Our studies indicate that additional transcriptional differences beyond opsin expression exist between LWS1 and LWS2 cones. Bulk RNA-Seq results showed 95 transcripts enriched in LWS1 cones and 186 transcripts enriched in LWS2 cones (FC > 2, FDR < 0.05). In situ hybridization results also reveal underlying heterogeneity within the lws1- and lws2-expressing populations. This heterogeneity is evident in cones of mature zebrafish, and further heterogeneity is revealed in transcriptional responses to TH treatments. Discussion: We found some evidence of coordinate regulation of lws opsins and other genes by exogenous TH in LWS1 vs. LWS2 cones, as well as evidence of gene regulation not mediated by TH. The transcriptional differences between LWS1 and LWS2 cones are likely controlled by multiple signals, including TH.

Prevalence of bovine tuberculosis and analysis of risk factors among the dairy farms in and around Guwahati metropolitan city, India.

Bovine tuberculosis (bTB), a neglected zoonotic disease caused by Mycobacterium bovis is being reported worldwide. The present work was carried out from December 2020 to November 2021 to assess the prevalence and risk factors of bTB in peri-urban and urban dairy farms of Guwahati, Assam, India. A questionnaire was used to collect data on knowledge about bTB on 36 farms, and ten animals per farm were screened by single intradermal comparative cervical tuberculin test (SICCT) to determine the prevalence of bTB, giving a total of 360 animals. The demographic data of the farmers revealed that 61.1% respondents were illiterate, 66.7% had no awareness about bovine tuberculosis and 41.7% consumed unpasteurised milk and milk products. SICCT showed that 38 cattle from 18 of the farms were positive reactors for bTB, yielding an overall animal level prevalence of 10.55% (95% confidence interval (CI = 7.58-14.2%) and a 50% herd prevalence (95% CI 32.9-67.1%). Animals 5 years and above were found to be more likely to be positive for bTB (17.18%). The study highlighted the widespread prevalence of bovine tuberculosis in peri-urban and urban dairy farms of Guwahati which gives a picture also about other major cities of India. Hence, it is of utmost importance to undertake a comprehensive epidemiological study in such cities for effective control and prevention of bTB in a one health approach.

Ultra-rare complement factor 8 coding variants in families with age-related macular degeneration.

Genome-wide association studies have uncovered 52 independent common and rare variants across 34 genetic loci, which influence susceptibility to age related macular degeneration (AMD). Of the 5 AMD-associated complement genes, complement factor H (CFH) and CFI exhibit a significant rare variant burden implicating a major contribution of the complement pathway to disease pathology. However, the efforts for developing AMD therapy have been challenging as of yet. Here, we report the identification of ultra-rare variants in complement factors 8A and 8B, two components of the terminal complement membrane attack complex (MAC), by whole exome sequencing of a cohort of AMD families. The identified C8 variants impact local interactions among proteins of C8 triplex in vitro, indicating their effect on MAC stability. Our results suggest that MAC, and not the early steps of the complement pathway, might be a more effective target for designing treatments for AMD.

Vibrational properties of metastable orthorhombic Bi2Se3.

Chalcogenide materials are being considered as some of the most promising systems for energy harvesting and energy conversion. Among them, the orthorhombic family of compounds X2Y3 (with X = Bi, Sb and Y = S, Se) has attracted special attention due to its interesting atomic structure and thermoelectric and optical properties. While Bi2S3 and Sb2Se3 have already been applied to solar cells, practical application of the new metastable Bi2Se3 is still a challenge due to the lack of data and knowledge on its properties. Here, the vibrational and structural properties of the orthorhombic metastable phase of Bi2Se3 are investigated by using Raman spectroscopy and ab initio calculations. We perform Raman spectroscopy measurements along with in situ thermal treatment on samples grown by electrochemical deposition. We show that by increasing the temperature an improved crystallization occurs in the orthorhombic structure, followed by recrystallization to the usual rhombohedral phase. Our results point out specific Raman modes of the orthorhombic phase. First principles computational results based on the density functional theory support the experimental data and describe three singlet Raman active vibrational modes, such as B(2)3g, B(2)2g and A(6)g.

Quantized Information in Spectral Cyberspace.

The constant-Q Gabor atom is developed for spectral power, information, and uncertainty quantification from time-frequency representations. Stable multiresolution spectral entropy algorithms are constructed with continuous wavelet and Stockwell transforms. The recommended processing and scaling method will depend on the signature of interest, the desired information, and the acceptable levels of uncertainty of signal and noise features. Selected Lamb wave signatures and information spectra from the 2022 Tonga eruption are presented as representative case studies. Resilient transformations from physical to information metrics are provided for sensor-agnostic signal processing, pattern recognition, and machine learning applications.

Evaluation of the lignocellulose degradation potential of Mediterranean forests soil microbial communities through diversity and targeted functional metagenomics.

The enzymatic arsenal of several soil microorganisms renders them particularly suitable for the degradation of lignocellulose, a process of distinct ecological significance with promising biotechnological implications. In this study, we investigated the spatiotemporal diversity and distribution of bacteria and fungi with 16S and Internally Trascribed Spacer (ITS) ribosomal RNA next-generation-sequencing (NGS), focusing on forest mainland Abies cephalonica and insular Quercus ilex habitats of Greece. We analyzed samples during winter and summer periods, from different soil depths, and we applied optimized and combined targeted meta-omics approaches aiming at the peroxidase-catalase family enzymes to gain insights into the lignocellulose degradation process at the soil microbial community level. The microbial communities recorded showed distinct patterns of response to season, soil depth and vegetation type. Overall, in both forests Proteobacteria, Actinobacteria, Acidobacteria were the most abundant bacteria phyla, while the other phyla and the super-kingdom of Archaea were detected in very low numbers. Members of the orders Agaricales, Russulales, Sebacinales, Gomphales, Geastrales, Hysterangiales, Thelephorales, and Trechisporales (Basidiomycota), and Pezizales, Sordariales, Eurotiales, Pleosporales, Helotiales, and Diaporthales (Ascomycota) were the most abundant for Fungi. By using optimized "universal" PCR primers that targeted the peroxidase-catalase enzyme family, we identified several known and novel sequences from various Basidiomycota, even from taxa appearing at low abundance. The majority of the sequences recovered were manganese peroxidases from several genera of Agaricales, Hysterangiales, Gomphales, Geastrales, Russulales, Hymenochaetales, and Trechisporales, while lignin -and versatile-peroxidases were limited to two to eight species, respectively. Comparisons of the obtained sequences with publicly available data allowed a detailed structural analysis of polymorphisms and functionally relevant amino-acid residues at phylogenetic level. The targeted metagenomics applied here revealed an important role in lignocellulose degradation of hitherto understudied orders of Basidiomycota, such as the Hysterangiales and Gomphales, while it also suggested the auxiliary activity of particular members of Proteobacteria, Actinobacteria, Acidobacteria, Verrucomicrobia, and Gemmatimonadetes. The application of NGS-based metagenomics approaches allows a better understanding of the complex process of lignocellulolysis at the microbial community level as well as the identification of candidate taxa and genes for targeted functional investigations and genetic modifications.

Components of TOR and MAP kinase signaling control chemotropism and pathogenicity in the fungal pathogen Verticillium dahliae.

Filamentous fungi can sense useful resources and hazards in their environment and direct growth of their hyphae accordingly. Chemotropism ensures access to nutrients, contact with other individuals (e.g., for mating), and interaction with hosts in the case of pathogens. Previous studies have revealed a complex chemotropic sensing landscape during host-pathogen interactions, but the underlying molecular machinery remains poorly characterized. Here we studied mechanisms controlling directed hyphal growth of the important plant-pathogenic fungus Verticillium dahliae towards different chemoattractants. We found that the homologs of the Rag GTPase Gtr1 and the GTPase-activating protein Tsc2, an activator and a repressor of the TOR kinase respectively, play important roles in hyphal chemotropism towards nutrients, plant-derived signals, and heterologous α-pheromone of Fusarium oxysporum. Furthermore, important roles of these regulators were identified in fungal development and pathogenicity. We also found that the mitogen-activated protein kinase (MAPK) Fus3 is required for chemotropism towards nutrients, while the G protein-coupled receptor (GPCR) Ste2 and the MAPK Slt2 control chemosensing of plant-derived signals and α-pheromone. Our study establishes V. dahliae as a suitable model system for the analysis of fungal chemotropism and discovers new components of chemotropic signaling during growth and host-pathogen interactions of V. dahliae.

Development of a novel visual isothermal amplification assay for rapid detection of Brucella spp.

Brucellosis is an economically important livestock disease worldwide besides having a noteworthy impact on human health. In this study, a rapid, simple, and ultra-sensitive nuclei-acid diagnostic technique was developed for the detection of brucellosis harnessing saltatory rolling circle amplification (SRCA). The diagnostic method was developed using World Organization for Animal Health (WOAH) approved primers targeting the bcsp31 gene of the Brucella genome. The assay can be accomplished within 90 min at a temperature of 65 °C without the requirement of sophisticated instrumentation. The result interpretation can be done with the naked eye with the aid of SYBR green dye. The developed technique displayed 100% specificity by amplifying only 10 reference and field strains of Brucella spp. and there was no cross-reactivity with the other tested pathogens. The lower limit of detections of SRCA and end-point PCR assays were 9.7 fg/µL (2.7 genome copies of Brucella) and 970 fg/µL, respectively. Thus, the developed SRCA assay was found to be 100× more sensitive than the end-point PCR assay. To the best of our knowledge, our study is the first one to develop an SRCA-based assay for the detection of brucellosis and it can be a diagnostic tool for resource-constrained laboratories and veterinary hospitals.

Novel sensitive isothermal-based diagnostic technique for the detection of African swine fever virus.

A rapid, simple, and sensitive diagnostic technique for the detection of African swine fever virus (ASFV) nucleic acid was developed for testing clinical samples in the field or resource-constrained settings. In the current study, the saltatory rolling-circle amplification (SRCA) technique was used for the first time to detect ASFV. The technique was developed using World Organization for Animal Health (WOAH)-approved primers targeting the p72 gene of the ASFV genome. The assay can be performed within 90 minutes at an isothermal temperature of 58°C without a requirement for sophisticated instrumentation. The results can be interpreted by examination with the naked eye with the aid of SYBR Green dye. This assay exhibited 100% specificity, producing amplicons only from ASFV-positive samples, and there was no cross-reactivity with other pathogenic viruses and bacteria of pigs that were tested. The lower limits of detection of SRCA, endpoint PCR, and real-time PCR assays were 48.4 copies/µL, 4.84 × 103 copies/µL, and 4.84 × 103 copies/µL, respectively. Thus, the newly developed SRCA assay was found to be 100 times more sensitive than endpoint and real-time PCR assays. Clinical tissue samples obtained from ASFV-infected domestic pigs and other clinical samples collected during 2020-22 from animals with suspected ASFV infection were tested using the SRCA assay, and a 100% accuracy rate, negative predictive value, and positive predictive value were demonstrated. The results indicate that the SRCA assay is a simple yet sensitive method for the detection of ASFV that may improve the diagnostic capacity of field laboratories, especially during outbreaks. This novel diagnostic technique is completely compliant with the World Health Organization's "ASSURED" criteria advocated for disease diagnosis, as it is affordable, specific, sensitive, user-friendly, rapid and robust, equipment-free, and deliverable. Therefore, this SRCA assay may be preferable to other complex molecular techniques for diagnosing African swine fever.