Infected Control Research Articles

Myristica fragrans Houtt. methanol extract as a promising treatment for Cryptosporidium parvum infection in experimentally immunosuppressed and immunocompetent mice.

Cryptosporidiosis is a major waterborne disease affecting ruminants and humans worldwide. It causes diarrhea and neonatal mortality in buffalo calves, and watery diarrhea and mortality in children and immunodeficient patients. This study aimed to investigate the efficacy of Myristica fragrans methanolic extract in treatment of C. parvum infection in comparison with nitazoxanide (NZX) (a Food and Drug Administration-approved drug control) in immunosuppressed and immunocompetent mice. One hundred laboratory-bred male Swiss albino mice were equally divided into immunocompetent and immunosuppressed groups. Each group was further divided into five subgroups: (1) non-infected and non-treated control, (2) infected and non-treated control (infected with Cryptosporidium parvum oocysts 3 × 103), (3) NZX-treated (100 mg/kg, 200 μL/mouse), (4) M. fragrans Houtt. methanol extract-treated (500 mg/kg), and (5) combination-treated (NZX + M. fragrans extract). Number of oocysts/g of feces, serum immunoglobulin (Ig) G level, and interferon (IFN)-γ, and interleukin (IL)-4 levels were used to evaluate the therapeutic effect. C. parvum oocyst shedding in stool samples was significantly decreased in all treatment groups, with 79.7%, 81.2 %, and 85.5 % reduction in immunocompetent mice treated with NZX, M. fragrans, and their combination, respectively. In immunosuppressed mice, oocyst shedding was reduced by 77.7%, 80.5 %, and 83.7 % upon NZX, M. fragrans, and their combination treatments, respectively. The serum IgG level was lowest in mice treated with a mixture of M. fragrans and NZX, followed by those treated with NZX, and was highest in mice treated with M. fragrans alone. Regarding cytokine levels, all groups treated with M. fragrans had low levels of IFN-γ and IL4 on day 21 post-infection. Collectively, the treatment of cryptosporidiosis with M. fragrans extract was successful in mice, as demonstrated by the measured parameters. M. fragrans reduced C. parvum oocyst shedding and serum IgG, IFN-γ, and IL-4 levels in immunocompetent and immunosuppressed mice.

Harnessing nature's defenders: unveiling the potential of microbial consortia for plant defense induction against Alternaria blight in cumin.

Present study was aimed to develop an efficient microbial consortium for combating Alternaria blight disease in cumin. The research involved isolating biocontrol agents against Alternaria burnsii, characterizing their biocontrol and growth promotion traits, and assessing compatibility. A pot experiment was conducted during rabi season of 2022-2023 to evaluate the bioefficacy of four biocontrol agents (1F, 16B, 31B, and 223B) individually and in consortium, focusing on disease severity, plant growth promotion, and defense responses in cumin challenged with A. burnsii. Microbial isolates 1F, 16B, 31B, and 223B significantly inhibited A. burnsii growth in dual plate assays (~ 86%), displaying promising biocontrol and plant growth promotion activities. They were identified as Trichoderma afroharzianum 1F, Aneurinibacillus aneurinilyticus 16B, Pseudomonas lalkuanensis 31B, and Bacillus licheniformis 223B, respectively. The excellent compatibility was observed among all selected biocontrol agents. Cumin plants treated with consortia of 1F + 16B + 31B + 223B showed least percent disease index (32.47%) and highest percent disease control (64.87%). Consortia of biocontrol agents significantly enhanced production of secondary metabolites (total phenol, flavonoids, antioxidant, and tannin) and activation of antioxidant-defense enzymes (POX, PPOX, CAT, SOD, PAL, and TAL) compared to individual biocontrol treatment and infected control. Moreover, consortium treatments effectively reduced electrolyte leakage over the individual biocontrol agent and infected control treatment. The four-microbe consortium significantly enhanced chlorophyll (154%), carotenoid content (88%), plant height (78.77%), dry weight (72.81%), and seed yield (104%) compared to infected control. Based on these findings, this environmentally friendly four-microbe consortium may be recommended for managing Alternaria blight in cumin.

Evaluation of muscular apoptotic changes and myogenin gene expression in experimental trichinosis after stem cells and atorvastatin added to ivermectin treatment

Trichinosis is a common parasitic disease that affects the striated skeletal muscles, causing apoptotic and degenerative changes associated with myogenin expression in the affected myocytes. Hence, this study aimed to assess the ameliorative effects of stem cells and atorvastatin added to ivermectin on the infected myocytes during the muscular phase of murine trichinosis. 120 laboratory Swiss albino male mice were divided into 10 groups, and each group was subdivided into intestinal and muscular phases (each n = 6); uninfected control; untreated infected control; infected received ivermectin monotherapy; infected received atorvastatin monotherapy; infected received stem cells monotherapy; infected received ivermectin and atorvastatin dual therapy; infected received ivermectin and stem cells dual therapy; infected received atorvastatin and stem cells dual therapy; infected received ivermectin 0.2, atorvastatin 40, and stem cells triple therapy; and infected received ivermectin 0.1, atorvastatin 20, and stem cells triple therapy. Intestinal phase mice were sacrificed on the 5th day post-infection, while those of the muscular phase were sacrificed on the 35th day post-infection. Parasitological, histopathological, ultrastructural, histochemical, biochemical, and myogenin gene expression assessments were performed. The results revealed that mice that received ivermectin, atorvastatin, and stem cell triple therapies showed the maximum reduction in the adult worm and larvae burden, marked improvement in the underlying muscular degenerative changes (as was noticed by histopathological, ultrastructural, and histochemical Feulgen stain assessment), lower biochemical levels of serum NK-κB and tissue NO, and lower myogenin expression. Accordingly, the combination of stem cells, atorvastatin, and ivermectin affords a potential synergistic activity against trichinosis with considerable healing of the underlying degenerative sequel.

Genetic variants of IL-10 promoter influence susceptibility to HIV-1 infection and disease progression in the Polish population: IL-10 polymorphisms and HIV-1

The risk of HIV-1 infection and the rate of disease progression vary considerably among individuals and the genetic makeup of the host may be one of the possible reasons for this. We aimed to determine association of functional single nucleotide polymorphism (SNPs), −1082A/G (rs1800896), −819C/T (rs1800871), and −592C/A (rs1800872) in IL-10 gene, with the susceptibility to HIV-1 infection and clinical parameters expressed as a baseline CD4+ T cell count, CD8+ T cell count, and viral load. Therapy naïve HIV-1 infected individuals and HIV-1 seronegative controls from Poland were recruited for this study. Genotyping results revealed significantly higher frequency of −1082G/G genotype (28.1 % vs 16.1 %; p = 0.0019, OR=0.49) and −1082G allele (47.6 % vs 38.8 %; p = 0.0028, OR = 0.70) as well as lower frequency of −592 and −819 heterozygosity (45.0 % vs 34.4 %; p = 0.0266, OR = 1.47) in controls compared to seropositive subjects. High producing haplotype GCC was associated with increased risk of HIV-1 infection (p = 0.0018, OR = 1.52). Individuals possessing −592 and −819 minor allele had significantly higher CD8+ T cell count compared to the wild type allele carriers (p = 0.0303). Moreover, presence of −1082G allele was related with lower viral load as well as CD4+ and CD8+ T cells counts among patients infected with R5 HIV-1 variant. Thus, IL-10 gene promoter variants may be a risk factor for HIV-1 transmission and may modulate disease progression in the Polish population.

Correlates of disease severity in bluetongue as a model of acute arbovirus infection.

Most viral diseases display a variable clinical outcome due to differences in virus strain virulence and/or individual host susceptibility to infection. Understanding the biological mechanisms differentiating a viral infection displaying severe clinical manifestations from its milder forms can provide the intellectual framework toward therapies and early prognostic markers. This is especially true in arbovirus infections, where most clinical cases are present as mild febrile illness. Here, we used a naturally occurring vector-borne viral disease of ruminants, bluetongue, as an experimental system to uncover the fundamental mechanisms of virus-host interactions resulting in distinct clinical outcomes. As with most viral diseases, clinical symptoms in bluetongue can vary dramatically. We reproduced experimentally distinct clinical forms of bluetongue infection in sheep using three bluetongue virus (BTV) strains (BTV-1IT2006, BTV-1IT2013 and BTV-8FRA2017). Infected animals displayed clinical signs varying from clinically unapparent, to mild and severe disease. We collected and integrated clinical, haematological, virological, and histopathological data resulting in the analyses of 332 individual parameters from each infected and uninfected control animal. We subsequently used machine learning to select the key viral and host processes associated with disease pathogenesis. We identified and experimentally validated five different fundamental processes affecting the severity of bluetongue: (i) virus load and replication in target organs, (ii) modulation of the host type-I IFN response, (iii) pro-inflammatory responses, (iv) vascular damage, and (v) immunosuppression. Overall, we showed that an agnostic machine learning approach can be used to prioritise the different pathogenetic mechanisms affecting the disease outcome of an arbovirus infection.

Effect of <i>Bacillus subtilis</i> in combination with chitosan salicylate on peroxidase and catalase activity in <i>B. sorokiniana</i> infected wheat

The aim of the work was to study the effect of Bacillus subtilis strains VKM B‑2604D and VKM B‑2605D which form the basis of the Vitaplan biological product, and their combinations with chitosan salicylate on antioxidant defense enzymes in the process of formation of resistance of wheat plants to infection with the dark brown blotch pathogen Bipolaris sorokiniana. In wheat plants treated with B. subtilis strains and their combinations with chitosan salicylate, upon subsequent infection, catalase and peroxidase are activated, which regulate the intensity of oxidative stress induced by the introduction of the pathogen. Taking into account the data obtained by us, it can be assumed that the increase in plant resistance to the pathogen is realized through the control of the activity of antioxidant enzymes (inparticular, catalase, peroxidase), which maintain the concentration of H2O2 at the level necessary for the neutralization of the phytopathogen in tissues, including the direct destruction of the invading pathogen and / or activation of crosslinking and lignification of the cell wall. These processes strengthen the cell wall and help contain the spread of the pathogen in the plant. At the same time, protective reactions are also switched on, leading to the development of induced resistance in wheat plants to dark brown spotting, which manifests itself in a decrease in the development of the disease by 25–45% relative to the infected control in the treated plants. The obtained results indicate that the combination of active strains of microorganisms-antagonists of plant pathogens and chitosan salicylate is promising for increasing the biological efficiency and expanding the spectrum of action of the developed prescription forms of biological products.

Efficacy of Commonly Used Anthelmintics Against Naturally Occurring Gastrointestinal Nematodes in Goats

Efficacy of four commonly used anthelmintics have been evaluated against naturally occurring gastrointestinal nematodes in goats following the method of faecal egg count reduction test (FECRT) in the present study. Fifty goats in the age group of 1 to 2 years old and of either sex, having infection with strongyle group of nematodes were selected in the sheep and goat farm under project ‘Biotech Kisan Hub”, West Bengal University of Animal and Fishery sciences and the goats were equally divided into five groups; group I (infected control), group II (albendazole treated), group III (fenbendazole treated), group IV (ivermectin treated) and group V (levamisole treated). Faecal egg count (FEC) in terms of eggs per gram of faeces (EPG) were estimated in all the experimental goats (n = 50) on 0-day post treatment (0 DPT) and again on 10 DPT. Efficacy of albendazole, fenbendazole, ivermectin and levamisole were 78.15%, 80.24%, 86.38% and 95.46%, respectively. Comparatively less efficacy of albendazole and fenbendazole indicated the emergence of anthelmintic resistance and lower efficacy of ivermectin might be due selection of resistant worms. Efficacy of levamisole was comparatively higher and it can be used strategically to control GI helminth parasites of goats in that farm.

Antiplasmodial evidence, host mitochondrial biology and possible mechanisms of action of a composite extract of Azadiractha indica and Curcuma longa in Plasmodium berghei-infected mice.

Azadirachta indica A. Juss (Meliaceae) (AI) and Curcuma longa L. (Zingiberaceae) (CL) are used for malaria treatment but their anti-glycolytic and host mitochondrial effects have not been studied. The AI stem-bark and CL rhizomes were extracted with methanol. Methanol extract of CL (Turmeric) was partitioned to yield methanol fraction (MF). Swiss mice infected with Plasmodium berghei (NK 65 strain) were treated with 200 and 400mg/kg of AI and turmeric for seven days. Turmeric and MF (200 and 400mg/kg) were combined with 400mg/kg AI to treat mice infected with Plasmodium berghei (ANKA strain) for four days. Drug and infected controls mice were treated with artemether lumefantrine (10mg/kg) and distilled water (10mL/kg), respectively. Serum lactate dehydrogenase (LDH) and aldolase activities were determined. Liver mitochondria were obtained for mitochondrial permeability transition (mPT) pore opening and FoF1 ATPase assays. The curcumin content of turmeric was determined using HPLC while LD50 of Turmeric and AI was also determined. The AI, and its combination with turmeric decreased parasite load and increased chemosuppression in both sensitive and resistant studies while MF and its combinations with AI induced mPT pore opening. In the resistant experiment, AI + Turmeric 400mg/kg decreased FoF1 ATPase, LDH and aldolase activities against the infected control. The LD50 values of both extracts were above 2000mg/kg while the MF had the highest curcumin content. Antiplasmodial mechanisms of action of AI, CL and their combinations involve anti-glycolytic effects. Their composite formulations are more potent in malaria treatment.

Melatonin and copper oxide nanoparticles synergistically mitigate clubroot disease and enhance growth dynamics in Brassica rapa

Clubroot, a devastating soil borne disease affecting 30%∼50% of Brassicaceae crops worldwide, lacks effective control measures. In the present study, we explored the potential of melatonin (MT) and copper oxide nanoparticle (CuO-NPs) in mitigating clubroot severity in the Brassica rapa ssp. pekinensis. Following 18 h priming with MT, CuO-NPs, or both seeds were grown in controlled environment using synthetic potting mix. Inoculated with Plasmodiophora brassicae spores on 5th day, followed by a soil drench phyto-nano treatment with a week interval. Plants were assessed for various health and growth indices including disease, biometrics, photosynthesis, reactive oxygen species (ROS), antioxidant enzyme activity, hormones and genes expression at onset of secondary clubroot infection using established protocols. Statistical analysis employed ANOVA with Fisher's LSD for significance assessment (P < 0.05). Our results revealed that seed priming with both MT (50 μMol/L) and CuO-NPs (200 mg/L), followed by soil drenching significantly reduced clubroot incidence (38%) and disease index (57%), compared to control treatments. This synergistic effect was associated with enhanced plant growth (shoots: 48% and roots: 59%). Plants treated with both MT and CuO-NPs showed robust antioxidant defenses, significantly increased superoxide dismutase (SOD (25/29%)), catalase (CAT (83/55%)), and ascorbate peroxidase (APX (83/46%)) activity in both shoots/roots, respectively, compared to infected control. Notably, salicylic acid and jasmonic acid levels doubled in treated plants, while stress hormone abscisic acid (ABA) decreased by 80% in roots and 21% in shoots. Gene expression analysis corroborated these findings, showing that the combined treatment activated antioxidant defense genes (SOD, APX and CAT) by 1.9–7.2-fold and upregulated hormone signaling genes JAZ1 (7.8-fold), MYC2 (3.9-fold) and SABP2 (36-fold). Conversely, ABA biosynthesis genes (ABA1 and NCED1) were downregulated up to 7.2-fold, while plant resistance genes NPR1, PRB1 and PDF1.2 were dramatically increased by up to 6.3-fold compared to infected plants. Overall, our combined treatment approach significantly reduces clubroot severity in B. rapa via enhanced antioxidant defenses, improved ROS scavenging, coordinated hormonal regulation and increased pathogen response genes. This study offers promising strategy for developing effective control measures against clubroot in susceptible cruciferous crops.

Antagonistic interactions between maize seeds microbiome species and the late wilt disease agent, Magnaporthiopsis maydis.

Magnaporthiopsis maydis is a maize pathogen that causes severe damage to commercial corn fields in the late growth stages. Late wilt disease (LWD) has spread since its discovery in the 1960s in Egypt and is now reported in about 10 countries. The pathogen has a hidden endophytic lifecycle in resistant corn plants and secondary hosts such as green foxtail, watermelon lupin and cotton. At the same time, it could be an opportunist and hinder the host development under the right conditions. This study uncovered M. maydis interactions with newly identified maize endophytes. To this end, six fungi were isolated from the seeds of three sweet corn cultivars having varying susceptibility to LWD. These isolates were identified using colony morphology and microscopic characterization, universal internal transcribed spacer (ITS) molecular targeting and phylogenetic analysis. Most of them belonged to pathogenic species. Compared to three previously identified bioprotective microorganisms, the new species were tested for their ability to secrete metabolites that repress M. maydis in vitro and to antagonize it in a solid media confront test and a seedlings pathogenicity assay. The opportunistic fungal species Aspergillus flavus (ME1), Aspergillus terreus (PE3) and the reference biocontrol bacteria Bacillus subtilis (R2) achieved the highest M. maydis inhibition degree in the plates tests (74-100% inhibition). The seedlings' pathogenicity assay that predicts the seeds' microflora resistance to M. maydis highlighted the bio-shielding potential of most species (23% or more epicotyl elongation over the infected control). Fusarium sp. (ME2) was the leading species in this measure (43% enhancement), and B. subtilis gave the best protection in terms of seeds' germination (50%) and sprouts' biomass (34%). The results of this study could enhance our understanding of the pathobiome's role in the context of LWD and represent a first step in using the seeds' natural protective microflora to develop novel management strategies.

Antimicrobial and therapeutic potentials of the blackberry extracts against Escherichia coli infection in male albino rats.

Blackberries have garnered attention recently due to their high concentration of bioactive components like anthocyanin and their health advantages. Therefore, this study aims to determine the bioactive profile, antioxidant and antibacterial effects of blackberry extracts (BBEs). Then, evaluate the protective effect of BBEs (20%, 30% and 40%) in a rat model of 2 mL of 4-8 × 106 Escherichia coli ATTC 25922 strain colony-forming unit mL-1 oral infection on the seventh day of the experiment. Rats were divided into six groups: G1: control (C-: normal or negative group), G2: (C+: infected or positive group), G3: infected-treated group by 20% BBE, G4: infected-treated group by 30% BBE, G5: infected-treated group by 40% of BBE and G6: infected-treated group by Gentamicin. The results showed that BBE had a high content of total phenolic compounds, flavonoid, anthocyanin contents, and different vitamins (vitamins A, E and C), reaching 450, 186, 58.83 mg 100 g-1, 2.68, 2.14 and 107.46 mg 100g-1 fresh weight, respectively, which showed great antioxidant and antibacterial effects. Therefore, liver enzymes, kidney function and lipid profiles were significantly higher in the infected group than in the control or infected-treated groups. Furthermore, BBE ameliorated inflammation of the intestine and hepatocyte damage compared to the infected control group. These results suggest that consistent intake of BBE might alleviate hepatic inflammation and the gut microbiota in ways that could significantly impact human health. © 2024 Society of Chemical Industry.

Lining the small intestine with mycobacteriophages protects from Mycobacterium avium subsp. paratuberculosis and eliminates fecal shedding

Johne's disease (JD), a chronic, infectious enteritis caused by Mycobacterium avium subsp. paratuberculosis (MAP), affects wild and domestic ruminants. There is no cure or effective prevention, and current vaccines have substantial limitations, leaving this disease widespread in all substantial dairy industries causing economic, and animal welfare implications. Mycobacteriophages (MPs) have been gaining interest in recent years and are proposed as a promising solution to curtailing MAP infection. Using a well-validated infection model, we have demonstrated the preventative potential of MPs to protect dairy calves against MAP infection. Calves were supplemented daily with a phage cocktail from birth till weaning at 2 m of age and inoculated with MAP at 2 wk of age. Infection status was measured for 4.5 mo through blood, fecal, and postmortem tissue samples. Our findings highlight the remarkable efficacy of orally administered MPs. Notably, fecal shedding of MAP was entirely eliminated within 10 wk, in contrast to the infected control group where shedding continued for the entirety of the trial period. Postmortem tissue culture analysis further supported the effectiveness of MPs, with only 1 out of 6 animals in the phage-treated group testing positive for MAP colonized tissues compared to 6 out of 6 animals in the infected control group. Additionally, plaque assay results demonstrated the ability of phages to persist within the intestinal tract. Collectively, these results underscore the potential of orally administered MP cocktails as a highly effective intervention strategy to combat JD in dairy calves and by extension in the dairy industry.

Corneal nerve fiber morphology following COVID-19 infection in vaccinated and non-vaccinated population

To examine corneal subbasal nerve changes in patients who received vaccination against SARS-CoV-2 virus and underwent COVID-19 infection compared to infected non-vaccinated patients and healthy controls. Twenty-nine eyes of 29 vaccinated patients (mean age: 36.66 ± 12.25 years) within six months after PCR or Ag test proven COVID-19 infection and twenty-eight eyes of 28 age-matched infected, non-vaccinated patients (mean age: 42.14 ± 14.17 years) were enrolled. Twenty-five age-matched healthy individuals (mean age: 47.52 ± 18.45 years) served as controls. In vivo confocal microscopy (Heidelberg Retina Tomograph II Rostock Cornea Module, Germany) was performed in each group. Corneal subbasal nerve plexus morphology and corneal dendritic cells (DC) were evaluated. Significantly higher corneal nerve fiber density (P < 0.001), nerve branch density (P < 0.001), nerve fiber length (P < 0.001), total branch density (P = 0.007), nerve fiber area (P = 0.001) and fractal dimension (P < 0.001) values were observed in vaccinated patients after COVID-19 infection compared to the non-vaccinated group. Significantly higher DC density was observed in the non-vaccinated group compared to the control group (P = 0.05). There was a statistically significant difference in the size of mature DCs (P < 0.0001) but the size of immature DCs did not differ significantly among the 3 groups (P = 0.132). Our results suggest that SARS-CoV-2 vaccination may have a protective effect against the complications of COVID-19 disease on the corneal subbasal nerve fibers.

Evaluation of Infected and Non-infected Wounds Healing Activity of Eriosema robustum Hydroethanolic Leaves Extract Ointments in Streptozotocin Induced Diabetic Rats

Untreated diabetic wounds provide an optimal environment for bacterial growth, which, over time, can develop resistance to common antibiotics and ultimately result in amputation. Therefore, it is necessary to search for new sources of antimicrobial molecules with wound healing activity owing to the presence of different secondary metabolites in medicinal plants. Aims: This study was to evaluate the in vivo antibacterial and diabetic wound healing capabilities of 70° hydroethanolic extract of Eriosema robustum leaves on non-infected and infected diabetic wound. Methodology: To do this, obese albino Wistar male rats (200–280 g) were divided into eleven groups and were made diabetic by intraperitoneal injection with a low dose of streptozotocin at 45 mg/kg of body weight. An excision wound with a surface area of 314 mm2 was created on the dorsal area of each animal, except in the uninjured diabetic group (UDG). The 70° hydroethanolic extract was used to prepare 1%, 5%, and 10% ointments, with L-Mesitran serving as the reference ointment. Healing potential was assessed by measuring wound contraction rates and determining serum and tissue hydroxyproline, serum lactate deydrogenase (LDH) and total protein levels (TP). The antibacterial power evaluated in vivo of Eriosema robustum leaves was also assessed by culturing the skin after healing. Results: The results demonstrated a significantly faster healing rate in the non-infected groups (5%,10% and L-Mesitran) compared to the infected groups. The levels of tissue hydroxyproline and total proteins were significantly (p &lt; 0.05) elevated in all treated groups compared to infected and negative controls, unlike serum hydroxyproline levels. LDH levels were significantly (p &lt; 0.05) elevated in both negative control group compared to the treated groups. The culture of different skin samples on previously injured areas on the 20th day of treatment showed no growth of S. aureus on completely healed areas and a low rate in the groups treated during the healing process. Conclusion: 70° Hydroethanolic leaves extract of Eriosema robustum possess in vivoantibacterial activities and diabetic wound healing potential.

Benznidazole/miltefosine combination improves the treatment of chagas disease

Chagas disease, caused by Trypanosoma cruzi, is one of the most prevalent parasitic diseases in Latin America. Currently, the available treatments, benznidazole and nifurtimox, have high toxicity, low solubility and reduced efficacy. This study investigates the effectiveness of combining miltefosine with benznidazole as a strategy to improve existing treatments. The research was conducted with 78 Swiss mice infected with 5000 trypomastigotes of T. cruzi (strain Y). The animals were divided into groups and treated with doses of 40mg/kg of miltefosine, administered orally for 10 consecutive or alternate days, and doses of 50 or 100mg/kg of benznidazole, administered for 20 consecutive days. Infected and uninfected control groups were included. After 150 days, the animals were sacrificed for analysis of parasitological cure. The combination of miltefosine and benznidazole demonstrated superior efficacy compared to isolated treatments. Alternating administration of 40mg/kg miltefosine with 50mg/kg benznidazole resulted in 62.5% cure, comparable to treatment with 100mg/kg benznidazole alone. The combination of 40 mg/kg of miltefosine with 100 mg/kg of benznidazole increased the cure rate to 87.5% (consecutive) and 100% (alternating). qPCR indicated a significant reduction in parasite DNA in cardiac and colonic tissues in the groups treated with the combinations. The results confirm that the combination of miltefosine with benznidazole enhances the effectiveness of the treatment, especially when miltefosine is administered every other day. This strategy can reduce the dose of benznidazole required, reducing toxicity and improving treatment adherence. Thus, the combination of drugs represents a promising approach to improving Chagas disease therapy, worthy of additional clinical investigation.

Testosterone leads to Trypanosoma cruzi glycoprotein synthesis and increased of inflammatory mediators in bone marrow-derived macrophages

Despite all the scientific progress in recent decades to unravel the immune processes and the way the parasite bypasses the immune system, Chagas disease is still a major public health problem, affecting an estimated 3.5 million people. Among the components that may participate in the response against the parasite, testosterone has been gaining more and more visibility. Studies indicate that the parasite itself seems to carry out steroidogenesis, in which, in co-culture with androgen precursors, T. cruzi has been shown to produce TS, but the purpose of the TS synthesized by the parasite and how this can influence its invasion glycoproteins is still unclear unknown. The aim of this study was to evaluate the influence of testosterone in Trypanosoma cruzi infection on the immune response of bone marrow-derived macrophages. Bone marrow from male rats was extracted and cultured with RMPI medium containing 30% L929 cell supernatant for macrophage differentiation. The cells were incubated for 10 days and, after this period, they were seeded in 96 wells in the amount of 1 x 105 cells per well. TS was added at different concentrations of 20 μM, 10 μM, 5 μM and 1 μM and then infected with the Y strain of T. cruzi, at a rate of 10 parasites per cell, with the culture remaining for six, 12 and 24 h. The supernatant was collected and the production of nitric oxide (NO), tumor necrosis factor (TNF) and the number of cell parasites was assessed by staining with 4′-6′-diamino-2-phenylindole (DAPI) and ranked by high Content Screening (HSC). The parasite was then cultured with the addition of TS, at the mentioned concentrations, leaving it for six and 12 h and then performing the RT-PCR of the mucins. DAPI staining revealed a significant increase in the number of parasites in cells containing TS. The exception was observed when 1 μM of hormone/well was used. A reduction in TNF production was found with 20 and 10 μM of TS for 6 h stimulation, although increased levels were observed with 5 and 1 μM, similar to the infected control. However, there was an increase in TNF production and not after 12 h. The relative expression of parasite glycoprotein 82 was increased with the presence of TS in the medium, regardless of time. Our data suggest that TS may contribute to cellular immunosuppression, increasing parasite infection in the cell, as well as inflammatory mediators that lead to cell and tissue damage in infected individuals, as well as the possible use of TS to allow their invasion into the cell hosts.

Study of the Effect of Tumor Necrosis Factor Alpha (TNF-α) Antagonist (Adalimumab) on Giardiasis in Mice

Abstract Giardia lamblia (G. lamblia) is the most common worldwide intestinal protozoal. Serum TNF-α contributes to the recovery from Giardia lamblia infection especially in the first and second week of infection. TNF-α antagonists are biologics used to treat many inflammatory diseases as rheumatoid arthritis. The present study aimed to study the effect of TNF-α antagonist (Adalimumab) on experimental giardiasis in mice by parasitological and histopathological examination. A total of 45 parasite-free laboratory-bred Swiss albino mice included into 2 main groups: test group and control group were used. Test group included mice infected with G. lamblia and receiving three intraperitoneal Adalimumab injections from days 9 till 15 post infection (pi). This group was further subdivided into 3 subgroups (Ia, Ib, Ic) according to Adalimumab doses which were 1.5, 3 and 6 mg/kg BW, received, respectively. Control group (non-Adalimumab-treated group) included a non-infected healthy group and Giardia -infected group. Monitoring the serum TNF-α level in all mice was done on two occasions (day 16 and 28 pi) by ELISA to follow variation in its level with and without administration of the drug. Infection outcome was assessed by following cyst shedding in stools of infected mice on intermittent days, counting trophozoites in intestinal wash, registering weight of animals, and recording pathological changes occurring in histopathological sections from small intestine. Highly significant increase in serum TNF-α was recorded in infected control mice on day 16 post-infection than in normal control group. Similarly, a significant increase in TNF-α level in the 3 test subgroups but attaining significant lower levels. On day 28 post infection, serum TNF- α level decreased in all infected mice with no significant differences being higher in infected control group than in non-infected control group. On assessing the effect of decreased serum TNF-α by the drug on infection outcome, highly significant increases in cyst shedding were recorded in the test group specifically group receiving 3 mg/kg BW than in control infected group on day 16 pi, and in trophozoite counts on day 28 pi. Mice weight gain was highly significant being more in control non-infected group than in control infected group. It was significantly lower in test group than normal infection control. The intestinal sections of test subgroups showed moderate to severe inflammatory reaction with more inflammatory cellular infiltration in the lamina propria causing villous shortening and blunting more than that observed in infection control group. Furthermore, lymphoid follicles and sloughing of the epithelium were only observed in test subgroups on day 16 p.i. It was concluded that TNF-α antagonist (Adalimumab) led to increased parasitic load as evidenced by cyst shedding, trophozoite count, less weight gains with moderate to severe histopathological damage in test subgroups.

Therapeutic potential of Garcinia kola against experimental toxoplasmosis in rats.

Cerebral toxoplasmosis, the most common opportunistic infection in immunocompromised individuals, is increasingly reported in immunocompetent individuals due to mutant strains of Toxoplasma gondii, which, furthermore, are reported to be resistant to available treatments. We assessed the therapeutic potential of Garcinia kola, a medicinal plant reported to have antiplasmodial and neuroprotective properties, against experimental toxoplasmosis in rats. Severe toxoplasmosis was induced in male Wistar rats (156.7 ± 4.1 g) by injecting them with 10 million tachyzoites in suspension in 500 µl of saline (intraperitoneal), and exclusive feeding with a low-protein diet [7% protein (weight by weight)]. Then, animals were treated with hexane, dichloromethane, and ethyl acetate fractions of Garcinia kola. Footprints were analysed and open-field and elevated plus maze ethological tests were performed when symptoms of severe disease were observed in the infected controls. After sacrifice, blood samples were processed for Giemsa staining, organs were processed for haematoxylin and eosin staining, and brains were processed for Nissl staining and cell counting. Compared with non-infected animals, the infected control animals had significantly lower body weights (30.27%↓, P = 0.001), higher body temperatures (P = 0.033) during the sacrifice, together with signs of cognitive impairment and neurologic deficits such as lower open-field arena centre entries (P < 0.001), elevated plus maze open-arm time (P = 0.029) and decreased stride lengths and step widths (P < 0.001), as well as neuronal loss in various brain areas. The ethyl acetate fraction of Garcinia kola prevented or mitigated most of these signs. Our data suggest that the ethyl acetate fraction of Garcinia kola has therapeutic potential against cerebral toxoplasmosis.

Characterization of urinary metabolites associated with malaria infection using infra-red spectroscopy and liquid chromatography–mass spectrometry in South Western Uganda

Early malaria diagnosis improves outcomes during malaria treatment; routine diagnostic techniques rely on blood samples obtained invasively. Therefore, this study used infra-red (IR) spectroscopy coupled with Principle Component Analysis (PCA) to study the urinary profile of malaria patients and that of controls aimed at understanding metabolite perturbation during malaria infection so as to contribute towards development of non-invasive malaria diagnosis methods. Freeze dried human urine samples form malaria infected individuals (cases) and controls were screened in the IR region of 4000 cm−1 to 600 cm−1 and overall spectral differences were observed at wave numbers 1618 cm−1, 1679 cm−1 (amino acids). Peaks at 3030 cm−1 (NH4+) and 940 cm−1 (O–H of carboxylic acids) showed high absorbance in patients compared to controls. Liquid-chromatography–mass spectrometry (LC–MS/MS) was used to quantify amino acids in the urine samples and the results indicated a significant increase of amino acid cystine (P = 0.012). Lysine and tyrosine also increased in patients compared to controls. The use of IR-PCA differentiated clusters of urine samples from patients with malaria from control and the demonstrated amino acid perturbation is consistent with malaria infection. This data provides baseline information for application in development of a non-invasive diagnostic tests for malaria.

Upregulation of inflammatory genes and pathways links obesity to severe COVID-19

Obesity is a risk factor for developing severe COVID-19. However, the mechanism underlying obesity-accelerated COVID-19 remains unclear. Here, we report results from a study in which 2–3-month-old K18-hACE2 (K18) mice were fed a western high-fat diet (WD) or normal chow (NC) over 3 months before intranasal infection with a sublethal dose of SARS-CoV2 WA1 (a strain ancestral to the Wuhan variant). After infection, the WD-fed K18 mice lost significantly more body weight and had more severe lung inflammation than normal chow (NC)-fed mice. Bulk RNA-seq analysis of lungs and adipose tissue revealed a diverse landscape of various immune cells, inflammatory markers, and pathways upregulated in the infected WD-fed K18 mice when compared with the infected NC-fed control mice. The transcript levels of IL-6, an important marker of COVID-19 disease severity, were upregulated in the lung at 6–9 days post-infection in the WD-fed mice when compared to NC-fed mice. Transcriptome analysis of the lung and adipose tissue obtained from deceased COVID-19 patients found that the obese patients had an increase in the expression of genes and the activation of pathways associated with inflammation as compared to normal-weight patients (n = 2). The K18 mouse model and human COVID-19 patient data support a link between inflammation and an obesity-accelerated COVID-19 disease phenotype. These results also indicate that obesity-accelerated severe COVID-19 caused by SARS-CoV-2 WA1 infection in the K18 mouse model would be a suitable model for dissecting the cellular and molecular mechanisms underlying pathogenesis.