Tissue Degradation Research Articles

Neutrophils and their friends.

Neutrophils and their friends.

Involvement of lignin deposition and cell wall degradation in stem senescence of Chinese flowering cabbage during storage

Chinese flowering cabbage is susceptible to leaf yellowing, however, little is known about stem senescence after harvest. Herein, changes in the microstructures, lignin and cell wall polysaccharide contents, and expression of their corresponding genes during the stem senescence of Chinese flowering cabbage were investigated. Anatomical and chemical analyses revealed that an increase in cell wall thickness was accompanied by lignin deposition around the vascular cells in the stem. Concurrently, the collapse of parenchyma cells developed gradually, which was probably related to the degradation of cell wall polysaccharides including pectin, hemicellulose, and cellulose, among others. The outer stem tissues, including epidermal cells to vascular cells, could be directly peeled, and the middle of the stem was found to contain prominent parenchyma cells as pith tissues. Activity of lignin biosynthesis-related enzymes of phenylalanine ammonia lyase (PAL), cinnamyl alcohol dehydrogenase (CAD), and peroxidase (POD), and the expression of their corresponding genes BrPAL2/3/6, Br4CL1/4, BrCAD2/4/5/11, and BrPOD31/37 were higher in outer peeled tissues than in pith tissues. The enzyme activity of polygalacturonase and pectin methylesterase, and expression levels of the polysaccharide degradation-related genes BrXET2/3, BrCEL3, and BrMAN1/5 were higher in pith than in outer peeled tissues. These results suggest that lignin deposition around the vascular tissue, and cell wall degradation in pith tissues are involved in the stem senescence of Chinese flowering cabbage after harvest.

Infection Biology of Bipolaris oryzae in Rice and Defence Responses in Compatible and Less Compatible Interactions

The infection biology of Bipolaris oryzae and the defence responses of its host rice were studied using the rice cv. MTL 189 inoculated with two isolates of B5 (highly compatible with MTL 189) and K2 (less compatible). In the former interaction, the hyphal progress was accompanied by tissue degradation and extensive sporulation after 8 days, whereas in the latter interaction, only very limited tissue degradation and sporulation was observed. Quantitative microscopy of the infection showed that the percentages of conidia and appressoria causing penetration and fluorescent epidermal cells (FEC) were lower for isolate K2 than for isolate B5 at 12 and 24 hours after inoculation (hai). Fluorescent papillae (FP) were only seen in the less compatible interaction and the percentage of conidia causing single FEC was highest in the less compatible interaction at 12 hai, but not at 24 hai. Qualitative examination of other defence responses showed that H2O2 started to accumulate at 4 hai in the less compatible interaction, whereas it appeared in the compatible interaction only at 12 hai. The level of H2O2 was generally higher in the less compatible than in the compatible interaction. Cross sections of leaves showed that H2O2 accumulated in the outer walls of epidermal cells. Likewise, accumulation of callose and polyphenolic substances was most pronounced in the less compatible interaction and occurred at the same places as H2O2. To our knowledge, this is the first report implicating H2O2 as an early defence response against the hemibiotrophic pathogen B. oryzae during early infection stages in rice. Understanding defence reactions may aid in resistance breeding.

Inflammation-targeting polyamine nanomedicines for the treatment of ulcerative colitis.

Inflammatory bowel disease (IBD) is a chronic inflammatory disorder characterized by immune system dysfunction. Despite the availability of various anti-inflammatory drugs, they exhibit low therapeutic efficacy with systemic side effects. In this study, we developed oral anti-inflammatory polyamine-based nanomedicines for the treatment of ulcerative colitis. Polyamine-bearing nanoparticles were prepared by the self-assembly of hyaluronic acid in organic solvents and crosslinking with branched oligoethyleneimine. Polyamine nanoparticles were found to suppress excessive inflammatory responses by scavenging the reactive oxygen species (ROS). Moreover, these nanoparticles inhibited enzymatic degradation and targeting of inflamed intestinal tissues. Additionally, they suppressed the inflammatory responses and recovered the pathological disorders in the colon of an ulcerative colitis mouse model. Therefore, polyamine-based nanomedicines exhibit great potential as biocompatible ROS-scavenging drugs for the treatment of IBD.

Salivary Biomarkers (Vitamin D, Calcium, and Estrogen Hormone) in Postmenopausal Women with Osteoporosis

Background: Osteoporosis is a systemic skeletal disease that weakens bones by lowering mineral density and causing micro-architectural degradation of bony tissue. Aim: This study was conducted to examine the use of vitamin D, calcium, and estrogen levels in saliva as biomarkers for early detection of osteoporosis. Materials and Methods: Ninety postmenopausal women were divided into three groups based on T-scores from dual-energy X-ray absorptiometry scans. The groups consist of 30 women with osteoporosis, 30 women with osteopenia, and 30 women serving as controls. The enzyme-linked immunosorbent assay was used to look for vitamin D and estrogen in the saliva by specific vitamin D3 and estradiol kits. Calcium concentrations were measured by a colorimetric method. Results: The level of salivary biomarkers (vitamin D, calcium, and estrogen) showed a clear distinction between the groups analyzed. Salivary estrogen and vitamin D levels were significantly lower in patients with osteoporosis compared to those with osteopenia and controls. The highest concentration of calcium was found in the osteoporosis group’s saliva compared to the other groups. Conclusion: The results highlight the significance of vitamin D, calcium, and estrogen in saliva as a simple, noninvasive biomarker in diagnosing postmenopausal osteoporosis.

Fibrillin-1 and asprosin, novel players in metabolic syndrome.

Fibrillin-1 is a major component of the extracellular microfibrils, where it interacts with other extracellular matrix proteins to provide elasticity to connective tissues, and regulates the bioavailability of TGFβ family members. A peptide consisting of the C-terminal 140 amino acids of fibrillin-1 has recently been identified as a glucogenic hormone, secreted from adipose tissue during fasting and targeting the liver to release glucose. This fragment, called asprosin, also signals in the hypothalamus to stimulate appetite. Asprosin levels are correlated with many of the pathologies indicative of metabolic syndrome, including insulin resistance and obesity. Previous studies and reviews have addressed the therapeutic potential of asprosin as a target in obesity, diabetes and related conditions without considering mechanisms underlying the relationship between generation of asprosin and expression of the much larger fibrillin-1 protein. Profibrillin-1 undergoes obligatory cleavage at the cell surface as part of its assembly into microfibrils, producing the asprosin peptide as well as mature fibrillin-1. Patterns of FBN1 mRNA expression are inconsistent with the necessity for regulated release of asprosin. The asprosin peptide may be protected from degradation in adipose tissue. We present evidence for an alternative possibility, that asprosin mRNA is generated independently from an internal promoter within the 3' end of the FBN1 gene, which would allow for regulation independent of fibrillin-synthesis and is more economical of cellular resources. The discovery of asprosin opened exciting possibilities for treatment of metabolic syndrome related conditions, but there is much to be understood before such therapies could be introduced into the clinic.

Ендотоксикоз і запалення як предиктори поглиблення функціональної недостатності підшлункової залози при хронічному панкреатиті у поєднанні із цукровим діабетом 2 типу

According to scientists, a significant role in the progression of chronic pancreatitis (CP) and diabetes mellitus (DM), as well as in the occurrence of complications of these diseases, is played by inflammation and endogenous intoxication by the products of protein breakdown as a result of the inflammatory process, fibrosis, and degradation of connective tissue in the patient’s body, in particular, and in the pancreas. The aim is to analyze the state of parameters of inflammation, endotoxicosis, and their influence on the functional capacity of the pancreas in the comorbid course of chronic pancreatitis and type 2 diabetes mellitus (DM2). Materials and methods. 112 patients with CP in the phase of mild therapeutic exacerbation in combination with DM2 in the stage of subcompensation were examined. The results. Moderate and moderate inverse correlations were established between CRP and fecal α-elastase in patients with CP and comorbidity of CP and DM2 (respectively r=-0.423 and r=-0.565, p<0,05). An increase in the strength of reliable direct moderate correlations between the levels of HbA1c and CRP in patients with CP in combination with DM2 compared to those with isolated CP (respectively r=0.313 and r=0.410, p<0,05) was proved. Conclusions: We proved a reliable influence of the index of endogenous intoxication on the level of pancreatic functional insufficiency according to the content of fecal α-elastase, which was more significant in the combined course of CP and DM2 compared to that in isolated CP. An increase in the strength of reliable direct, moderate, and moderate correlations between HbA1c levels and the index of endogenous intoxication in patients with isolated CP and CP in combination with DM2 has been proven.

Unraveling the Intricacies: Pioneering Insights into the Interplay of the Gut and Oral Microbiome in Diabetes and Periodontal Disease

Exploring the intricate interplay between two prevalent chronic conditions, diabetes mellitus and periodontal disease, reveals a bidirectional relationship. Recent evidence underscores the pivotal influence of the microbiome, particularly the gut and oral microbiome, in shaping the pathogenesis of both diabetes and periodontal disease. The present comprehensive review aimed to elucidate the current understanding of how these microbial communities contribute to the development and progression of diabetes, especially when compounded by periodontal disease. However, emerging evidence suggests a complex bidirectional relationship between these two conditions. The microbiome’s involvement in these conditions unfolds through multifaceted mechanisms, with microbial dysbiosis influencing systemic inflammation, insulin resistance, and periodontal tissue degradation. The authors explored the dynamic crosstalk between the gut and oral microbiome, shedding light on how alterations in these microbial ecosystems may exacerbate the interconnected manifestations of diabetes and periodontal disease. Furthermore, present review unraveled the potential therapeutic implications for targeted interventions. By dissecting the microbiome-driven pathways, authors identified the promising avenues for precision medicine and tailored therapies. This exploration opens new vistas for developing strategies that leverage the microbiome to mitigate the impact of diabetes with periodontal disease. As the authors navigated this complex terrain, the manuscript underscores the urgency of a holistic understanding and targeted modulation of the microbiome to revolutionise treatment paradigms for these intertwined chronic conditions. The present manuscript aimed to review the current understanding of the role of the gut and oral microbiome in the development and progression of diabetes with periodontal disease.

Combined effects of high fat diet and exercise on autophagy in white adipose tissue of mice

AimThe effects of nutrition and exercise on autophagy are not well studied. This study aimed to investigate the combined effects of high-fat diets (HFD) and exercise training (ET) on autophagy in white adipose tissue of mice. Materials and methodsMale C57BL/6 mice were assigned into four groups of 7 mice per group: (1) Control, (2) high-fat diet-induced obesity (HFD-Ob), (3) exercise training (ET), and (4) high-fat diet with exercise training (HFD-ET). The HFD-Ob group was fed a high-fat diet for 14 weeks, while the ET group continuously ran on a treadmill for five sessions per week for seven weeks, and the HFD-ET group had both HFD and exercise training. qReal-time–PCR and western blot were used to measure the mRNA and protein levels of autophagy markers in white adipose tissue. ResultsMice from the HFD group showed higher levels in autophagy-related gene5 (ATG5, p = 0.04), ATG7 (p = 0.002), cathepsin B (CTSB, p = 0.0004), LC3-II (p = 0.03) than control. Mice in the ET group displayed higher levels of genes for ATG7 (p = 0.0003), microtubule-associated protein1-light chain 3 (LC3, p = 0.05), lysosome-associated membrane protein 2 (LAMP2, p = 0.04) and cathepsin L (CTSL, p = 0.03) than control. Mice from the HFD-ET group had higher levels of genes for ATG7 (p = 0.05) and CTSL (p = 0.043) and lower levels of genes for CTSB (p = 0.045) compared to the HFD group and lower levels of LAMP2 (p = 0.02) compared to the ET group. ConclusionThere were increases in autophagosome formation in the white adipose tissue from mice in the HFD and ET groups. A combination of HFD and ET enhances autophagosome formation and modulates lysosomal degradation in white adipose tissue.

The levels of hypoxia- and angiogenesis-related regulators and matrix metalloproteinase 9 activity in tear fluid of patients with non-penetrating ocular traumas

This article was focused on the evaluation of protein biomarkers related to thrombosis, hypoxia, angiogenesis, and tissue remodeling in tear fluid of patients with non-penetrating corneal trauma. 32 patients with non-penetrating corneal injures were enrolled in the study, the control group consisted of 15 healthy patients. Samples of tear fluid were collected from the patients and control volunteers with the use of a disposable end micropipette. Protein levels of D-dimer, hypoxia-inducible factor 1α (HIF-1α), angiostatins, and matrix metalloproteinase 9 (MMP-9) in tear fluids were determined by western blot analysis. Proteolytic activity values of MMP-9 were measured by gelatin zymography. Results of western blot and zymography assay were calculated by densitometry analysis and expressed as arbitrary units. Significant increase of D-dimer and HIF-1α levels in tear fluid of patients with injured cornea by 7.3 (p<0.05) and 56 (p<0.001) folds, respectively was shown, compared with control, indicating thrombotic events and hypoxia condition to be involved in pathogenesis of ocular trauma. Dramatically elevated levels/activity of MMP-9 enzyme (by 105 folds vs. control, p<0.001) suggests intense tissue remodeling and degradation of extracellular matrix in the damaged cornea. Up-regulation of angiostatin level, products of proteolytical cleavage of plasminogen, in tear fluid collected from patients with traumatic eye in comparison with healthy volunteers (by 7.3 folds, p<0.05), could represent an adaptive mechanism, which counteracts excessive hypoxia-induced neovascularization of injured cornea. It is summarized that there was a strong association between elevation of D-dimer, HIF-1α, angiostatins, and MMP-9 levels suggesting thrombosis- and hypoxia-mediated mechanisms triggering wound healing of injured cornea. The findings of this study are novel and provide a basis for further investigations of the reparation mechanisms during non-penetrating ocular trauma. Studied proteins of tear fluid can serve as relevant biomarkers of corneal wound healing and are appropriate for diagnostic and prognostic purposes.

Modulatory Mechanisms of Pathogenicity in Porphyromonas gingivalis and Other Periodontal Pathobionts.

The pathogenesis of periodontitis depends on a sustained feedback loop where bacterial virulence factors and immune responses both contribute to inflammation and tissue degradation. Periodontitis is a multifactorial disease that is associated with a pathogenic shift in the oral microbiome. Within this shift, low-abundance Gram-negative anaerobic pathobionts transition from harmless colonisers of the subgingival environment to a virulent state that drives evasion and subversion of innate and adaptive immune responses. This, in turn, drives the progression of inflammatory disease and the destruction of tooth-supporting structures. From an evolutionary perspective, bacteria have developed this phenotypic plasticity in order to respond and adapt to environmental stimuli or external stressors. This review summarises the available knowledge of genetic, transcriptional, and post-translational mechanisms which mediate the commensal-pathogen transition of periodontal bacteria. The review will focus primarily on Porphyromonas gingivalis.

Показатели качества Agaricus bisporus после обработки УФ-излучением

Ultraviolet treatment increases the shelf life of plant products. It inhibits the changes in the quality indicators of raw materials that are responsible for storage capacity. The research objective was to establish qualitative indicators for champignons (Agaricus bisporus) after ultraviolet treatment.
 Fresh champignons were placed in plastic trays and sealed in plastic bags. The samples were treated with ultraviolet radiation in ranges A, B, and C. After that, they were stored in a refrigerator at 4 ± 2°C for 16 days, i.e., until the end of storage period. The quality indicators underwent a paired two-sample test, which defined the equality/inequality of variances in replicates and equality/inequality of means at a given error probability (α).
 The obtained indicators characterized the degradation of mushroom plant tissue after ultraviolet treatment during storage: texture, moisture content, weight loss, soluble solids, pH, lightness, and color. The authors developed a new approach to determine significant changes in the quality indicators and estimated probability values. For each range, they established the most effective dose at α = 0.05 and 0.1. All the indicators proved significant for range A samples at 327.8–800.0 J/m2 and α = 0.05; all indicators were significant except weight loss in the range from 219.5 to 800.0 J/m2 at α = 0.1. For range B samples, pH, color, and lightness were significant at 104.6–200.0 J/m2 and α = 0.05; all but mass loss and texture were significant at 172.2–200.0 J/m2 and α = 0.1. For range C samples, pH, color, and lightness were significant at 412.4–439.5 J/m2 and 755.9–800.0 J/m2 at α = 0.05; all indicators were significant at 363.3–486.2 J/m2 and 728.2–800.0 J/m2 at α = 0.1.
 The new analytical method made it possible to determine the ranges of external effect intensity, in which the changes in the indicator were significant or insignificant.

Ginsenoside Rg1 inhibits nucleus pulposus cell apoptosis, inflammation and extracellular matrix degradation via the YAP1/TAZ pathway in rats with intervertebral disc degeneration

PurposeIntervertebral disc degeneration (IDD) is one of the main causes of low back pain, which not only affects patients’ life quality, but also places a great burden on the public health system. Recently, ginsenoside Rg1 has been found to act in IDD; however, the mechanism is still unclear. The purpose of this study is to explore the function of ginsenoside Rg1 and its molecular mechanism in IDD.MethodsThe rat model of IDD and nucleus pulposus (NP) experimental groups treated with ginsenoside Rg1 was constructed for investing the role of ginsenoside Rg1 in IDD rats. In the in vitro and in vivo study, the histological morphological changes, motor threshold (MT), inflammatory factors, oxidative stress, apoptosis and expression of the YAP1/TAZ signaling pathway-related proteins of the intervertebral discs (IVD) were measured by histological staining, mechanical and thermal stimulation, ELISA, qRT-PCR, flow cytometry, and western blot, respectively.ResultsGinsenoside Rg1 significantly increased the threshold for mechanical and thermal stimulation and alleviated histological changes in IDD rats. Ginsenoside Rg1 had a significant inhibitory effect on the secretion level of inflammatory factors, redox activity, extracellular matrix (ECM) degradation in IVD tissue and NP cells, and apoptosis in NP cells. Further investigation revealed that ginsenoside Rg1 significantly inhibited the expression of YAP1/TAZ signaling pathway-related proteins. Additionally, the above inhibitory effect of ginsenoside Rg1 on IDD progression was concentration-dependent, that is, the highest concentration of ginsenoside Rg1 was most effective.ConclusionGinsenoside Rg1 inhibits IDD progression by suppressing the activation of YAP1/TAZ signaling pathway. This means that ginsenoside Rg1 has the potential to treat IDD.

Structure-Based Discovery of Selective Histone Deacetylase 8 Degraders with Potent Anticancer Activity.

Inducing protein degradation by proteolysis targeting chimeras has gained tremendous momentum as a promising novel therapeutic strategy. Here, we report the design, synthesis, and biological characterization of highly potent proteolysis targeting chimeric small molecules targeting the epigenetic regulator histone deacetylase 8 (HDAC8). We developed potent and effective HDAC8 degraders, as exemplified by SZUH280 (16e), which effectively induced HDAC8 protein degradation and inhibited cancer cell growth even at low micromolar concentrations. Our preliminary mechanistic studies revealed that SZUH280 hampers DNA damage repair in cancer cells, promoting cellular radiosensitization. In mice, a single SZUH280 dose induced rapid and prolonged HDAC8 protein degradation in xenograft tumor tissues. Moreover, SZUH280 alone or in combination with irradiation resulted in long-lasting tumor regression in an A549 tumor mouse model. Our findings qualify a new chemical tool for HDAC8 knockdown and may lead to the development of a new class of cancer therapeutics.

Modified fibrin hydrogel for sustained delivery of RNAi lipopolyplexes in skeletal muscle.

RNA interference is a promising therapeutical approach presently hindered by delivery concerns such as rapid RNA degradation and targeting of individual tissues. Injectable hydrogels are one potentially simple and direct route towards overcoming these barriers. Here we report on the utility of a combination of a mildly modified form of the clinically utilised fibrin hydrogel with Invivofectamine® 3.0, a lipid nonviral transfection vector, for local and sustained release. PEGylation of fibrin allowed for controlled release of small interfering RNA (siRNA)-lipopolyplexes for at least 10 days and greatly increased the stability of fibrin in vitro and in vivo. A 3D cell culture model and a release study showed transfection efficacy of siRNA-lipopolyplexes was retained for a minimum of 7 days. Injection in conjunction with PEGylated-fibrinogen significantly increased retention of siRNA-lipopolyplexes in mouse skeletal muscle and enhanced knockdown of myostatin mRNA that correlated with muscle growth. Thus, the increased efficacy observed here for the combination of a lipid nanoparticle, the only type of nonviral vector approved for the clinic, with fibrin, might allow for more rapid translation of injectable hydrogel-based RNA interference.

Percutaneous delivery and degradation of a shape memory elastomer poly(glycerol dodecanedioate) in porcine pulmonary arteries

Shape memory biodegradable elastomers are an emergent class of biomaterials well-suited for percutaneous cardiovascular repair requiring nonlinear elastic materials with facile handling. We have previously developed a chemically crosslinked shape memory elastomer, poly (glycerol dodecanedioate) (PGD), exhibiting tunable transition temperatures around body temperature (34–38 °C), exhibiting nonlinear elastic properties approximating cardiac tissues, and favorable degradation rates in vitro. Degree of tissue coverage, degradation and consequent changes in polymer thermomechanical properties, and inflammatory response in preclinical animal models are unknown material attributes required for translating this material into cardiovascular devices. This study investigates changes in the polymer structure, tissue coverage, endothelialization, and inflammation of percutaneously implanted PGD patches (20 mm × 9 mm x 0.5 mm) into the branch pulmonary arteries of Yorkshire pigs for three months. After three months in vivo, 5/8 samples exhibited (100%) tissue coverage, 2/8 samples exhibited 85–95% tissue coverage, and 1/8 samples exhibited limited (<20%) tissue coverage with mild-moderate inflammation. PGD explants showed a (60–70%) volume loss and (25–30%) mass loss, and a reduction in polymer crosslinks. Lumenal and mural surfaces and the cross-section of the explant demonstrated evidence of degradation. This study validates PGD as an appropriate cardiovascular engineering material due to its propensity for rapid tissue coverage and uneventful inflammatory response in a preclinical animal model, establishing a precedent for consideration in cardiovascular repair applications.

Advanced glycation end-product accumulation differs by location and sex in aged osteoarthritic human menisci

There is a clear link between increasing age and meniscus degeneration, leading to increased injury, osteoarthritis (OA) progression, and often total knee replacement. Advanced glycation end-products (AGEs) are non-enzymatic crosslinks and adducts that accumulate in collagen with age, altering tissue mechanics and cell function, ultimately leading to increased injury and inflammation. AGEs, both fluorescent and non-fluorescent, play a central role in age-related degradation of tissues throughout the body; however, little is known about their role in meniscus degeneration. The objective of this study was to characterize changes in aged OA menisci, specifically evaluating zonal AGE accumulation, to gain a better understanding of changes that may lead to age-related meniscal degeneration. Deidentified human menisci (N=48, 52-84 years old) were obtained from subjects undergoing total knee replacement. Changes in extracellular matrix (ECM) were assessed by gross morphology, confocal analysis, and biochemical assays. Deoxyribonucleic acid (DNA), glycosaminoglycan (GAG), collagen, and AGE accumulation were compared with patient age, zonal region, and patient sex. There were minimal changes in DNA, GAG, and collagen concentration with age or zone. However, collagen fraying and AGEs increased with age, with more AGEs accumulating in the meniscal horns compared to the central body and in male menisci compared to females. Overall, this work provides greater insights into regional changes that occur in human menisci with age and OA. These results suggest AGEs may play a role in the degeneration of the meniscus, with AGEs being a possible target to reduce age-related tears, degeneration, and OA progression.

Robot and ultrasound assisted needle insertion to the transverse carpal ligament

BackgroundA potential alternative treatment to surgery for carpal tunnel syndrome is to inject enzymes into the transverse carpal ligament to decrease its stiffness and alleviate pressure off the median nerve. An accurate injection is needed for delivery to achieve the effects of tissue degradation. The purposes of this study were to 1) determine injection sites using 3D reconstructed anatomy, and 2) insert the needle to the middle of the transverse carpal ligament thickness in situ. MethodsSix fresh-frozen cadaveric hands were used in this study. Five injection sites were determined in the sagittal plane along the center of the transverse carpal ligament thickness ulnar to the thenar muscle attachment using 3D ultrasonographic reconstruction. Each injection was delivered by rigidly fixing a 27-gauge needle to a six degrees of freedom robot arm programmed to insert the needle tip to the intended target. Ultrasound images were taken of the needle after insertion to measure accuracy and precision of the needle placement. FindingsThe needle tip was successfully delivered to the middle region of the transverse carpal ligament thickness and visualized using ultrasound imaging. The accuracy and precision of the needle insertion were 0.83 and 0.31 mm, respectively. InterpretationMethodology was established for robot-assisted needle insertion to the transverse carpal ligament using 3D ultrasonographic reconstructed anatomy. This methodology can be used in the future to deliver enzymatic injections to the transverse carpal ligament as a potential treatment for carpal tunnel syndrome.

Assessing the Putative Anticryptococcal Properties of Crude and Clarified Extracts from Mollusks.

Cryptococcus neoformans is an encapsulated human fungal pathogen with a global distribution that primarily infects immunocompromised individuals. The widespread use of antifungals in clinical settings, their use in agriculture, and strain hybridization have led to increased evolution of resistance. This rising rate of resistance against antifungals is a growing concern among clinicians and scientists worldwide, and there is heightened urgency to develop novel antifungal therapies. For instance, C. neoformans produces several virulence factors, including intra- and extra-cellular enzymes (e.g., peptidases) with roles in tissue degradation, cellular regulation, and nutrient acquisition. The disruption of such peptidase activity by inhibitors perturbs fungal growth and proliferation, suggesting this may be an important strategy for combating the pathogen. Importantly, invertebrates such as mollusks produce peptidase inhibitors with biomedical applications and anti-microbial activity, but they are underexplored in terms of their usage against fungal pathogens. In this protocol, a global extraction from mollusks was performed to isolate potential peptidase inhibitors in crude and clarified extracts, and their effects against classical cryptococcal virulence factors were assessed. This method supports the prioritization of mollusks with antifungal properties and provides opportunities for the discovery of anti-virulence agents by harnessing the natural inhibitors found in mollusks.

Lactic Acid Bacteria in the Management of Oily Spot Disease of Pomegranate.

The bacterial pathogen of oily spot disease, a major threat to pomegranate growers, was isolated from infected plant parts of pomegranate collected from orchards in Maharashtra, India. The pathogen was identified as Xanthomonas axonopodis pv. punicae (Xap) following phenotypic and molecular characterization by 16S rRNA gene sequencing. It produced pectinase, cellulase, xylanase in medium and in experimentally inoculated tissues with pathogen where pectinase activity was maximum (32.2U/g). Pearson correlation analysis showed a perfect positive correlation (P < 0.05) between enzyme activity and disease rating scale. This indicates the co-synthesis of hydrolytic enzymes that aid in tissue degradation and suggests their role as virulence factors. Out of 150 indigenously isolated lactic acid bacteria (LAB), Lactococcus lactis subsp. cremoris PB6, Lactobacillus brevis PFR77 and L. lactis subsp. cremoris PFL9, the potent antagonists of Xap, were used in the management of bacterial blight. Under laboratory conditions, cell formulation of PB6, PFR77 and PFL9 were equally effective (P > 0.05) and significantly (P < 0.05) reduced the infection in fruits. Under field conditions, the disease severity index for the treatments where plants received a spray of PB6 with streptocycline, was lowest (4.61%) as compared to cells (15.74%), culture supernatant (20.66%) and their integrated treatments (21.38%), and streptocycline (15.37%) treatments. However, no significant difference (P > 0.05) was noticed between cells and streptocycline treatments, thus, indicating the effectiveness of LAB in treating bacterial blight. This is the first report on the use of antagonistic LAB for the control of oily spot disease of pomegranate.