scholarly journals In Vitro, Ex Vivo, and In Vivo Activities of Diamidines against Trypanosoma congolense and Trypanosoma vivax

2017 ◽  
Vol 61 (5) ◽  
Author(s):  
Kirsten Gillingwater ◽  
Christina Kunz ◽  
Christiane Braghiroli ◽  
David W. Boykin ◽  
Richard R. Tidwell ◽  
...  
Keyword(s):  
Ex Vivo ◽  
Trypanosoma Congolense ◽  
Tsetse Fly ◽  
Sub Saharan Africa ◽  
Content Type ◽  
Lead Compounds ◽  
Single Bolus Dose ◽  
Sub Saharan

ABSTRACT African animal trypanosomosis (AAT) is caused by the tsetse fly-transmitted protozoans Trypanosoma congolense and T. vivax and leads to huge agricultural losses throughout sub-Saharan Africa. Three drugs are available to treat nagana in cattle (diminazene diaceturate, homidium chloride, and isometamidium chloride). With increasing reports of drug-resistant populations, new molecules should be investigated as potential candidates to combat nagana. Dicationic compounds have been demonstrated to have excellent efficacy against different kinetoplastid parasites. This study therefore evaluated the activities of 37 diamidines, using in vitro and ex vivo drug sensitivity assays. The 50% inhibitory concentrations obtained ranged from 0.007 to 0.562 μg/ml for T. congolense and from 0.019 to 0.607 μg/ml for T. vivax. On the basis of these promising results, 33 of these diamidines were further examined using in vivo mouse models of infection. Minimal curative doses of 1.25 mg/kg of body weight for both T. congolense- and T. vivax-infected mice were seen when the diamidines were administered intraperitoneally (i.p.) over 4 consecutive days. From these observations, 15 of these 33 diamidines were then further tested in vivo, using a single bolus dose for administration. The total cure of mice infected with T. congolense and T. vivax was seen with single i.p. doses of 5 and 2.5 mg/kg, respectively. This study identified a selection of diamidines which could be considered lead compounds for the treatment of nagana.

scholarly journals Physiological Differences in Cryptococcus neoformans Strains In Vitro versus In Vivo and Their Effects on Antifungal Susceptibility

2016 ◽  
Vol 61 (3) ◽  
Author(s):  
Nina T. Grossman ◽  
Arturo Casadevall
Keyword(s):  
Cryptococcus Neoformans ◽  
Cryptococcal Meningitis ◽  
Antifungal Susceptibility ◽  
Sub Saharan Africa ◽  
Content Type ◽  
Laboratory Conditions ◽  
Physiological Differences ◽  
Sub Saharan

ABSTRACT Cryptococcus neoformans is an environmentally ubiquitous fungal pathogen that primarily causes disease in people with compromised immune systems, particularly those with advanced AIDS. There are estimated to be almost 1 million cases per year of cryptococcal meningitis in patients infected with human immunodeficiency virus, leading to over 600,000 annual deaths, with a particular burden in sub-Saharan Africa. Amphotericin B (AMB) and fluconazole (FLC) are key components of cryptococcal meningitis treatment: AMB is used for induction, and FLC is for consolidation, maintenance and, for occasional individuals, prophylaxis. However, the results of standard antifungal susceptibility testing (AFST) for AMB and FLC do not correlate well with therapeutic outcomes and, consequently, no clinical breakpoints have been established. While a number of explanations for this absence of correlation have been proffered, one potential reason that has not been adequately explored is the possibility that the physiological differences between the in vivo infection environment and the in vitro AFST environment lead to disparate drug susceptibilities. These susceptibility-influencing factors include melanization, which does not occur during AFST, the size of the polysaccharide capsule, which is larger in infecting cells than in those grown under normal laboratory conditions, and the presence of large polyploid “titan cells,” which rarely occur under laboratory conditions. Understanding whether and how C. neoformans differentially expresses mechanisms of resistance to AMB and FLC in the AFST environment compared to the in vivo environment could enhance our ability to interpret AFST results and possibly lead to the development of more applicable testing methods.

scholarly journals Schistosoma mansoni eggs induce Wnt/β-catenin signaling and activate the protooncogene c-Jun in human and hamster colon

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Jakob Weglage ◽  
Friederike Wolters ◽  
Laura Hehr ◽  
Jakob Lichtenberger ◽  
Celina Wulz ◽  
...  
Keyword(s):  
Signaling Pathways ◽  
Colorectal Carcinogenesis ◽  
Sub Saharan Africa ◽  
Interleukin 4 ◽  
Health Burden ◽  
Sub Saharan ◽  
Considerable Morbidity ◽  
First Time

AbstractSchistosomiasis (bilharzia) is a neglected tropical disease caused by parasitic flatworms of the genus Schistosoma, with considerable morbidity in parts of the Middle East, South America, Southeast Asia, in sub-Saharan Africa, and particularly also in Europe. The WHO describes an increasing global health burden with more than 290 million people threatened by the disease and a potential to spread into regions with temperate climates like Corsica, France. The aim of our study was to investigate the influence of S. mansoni infection on colorectal carcinogenic signaling pathways in vivo and in vitro. S. mansoni infection, soluble egg antigens (SEA) and the Interleukin-4-inducing principle from S. mansoni eggs induce Wnt/β-catenin signaling and the protooncogene c-Jun as well as downstream factor Cyclin D1 and markers for DNA-damage, such as Parp1 and γH2a.x in enterocytes. The presence of these characteristic hallmarks of colorectal carcinogenesis was confirmed in colon biopsies from S. mansoni-infected patients demonstrating the clinical relevance of our findings. For the first time it was shown that S. mansoni SEA may be involved in the induction of colorectal carcinoma-associated signaling pathways.

scholarly journals Ex VivoMaturation Assay for Testing Antimalarial Sensitivity of Rodent Malaria Parasites

2016 ◽  
Vol 60 (11) ◽  
pp. 6859-6866 ◽  
Author(s):  
Zi Wei Chang ◽  
Benoit Malleret ◽  
Bruce Russell ◽  
Laurent Rénia ◽  
Carla Claser
Keyword(s):  
Ex Vivo ◽  
Single Step ◽  
Parasite Development ◽  
Ring Stage ◽  
Malaria Parasites ◽  
Rodent Malaria ◽  
Content Type ◽  
Parasite Biology

ABSTRACTEx vivoassay systems provide a powerful approach to studying human malaria parasite biology and to testing antimalarials. For rodent malaria parasites, short-termin vitroculture andex vivoantimalarial susceptibility assays are relatively cumbersome, relying onin vivopassage for synchronization, since ring-stage parasites are an essential starting material. Here, we describe a new approach based on the enrichment of ring-stagePlasmodium berghei,P. yoelii, andP. vinckei vinckeiusing a single-step Percoll gradient. Importantly, we demonstrate that the enriched ring-stage parasites develop synchronously regardless of the parasite strain or species used. Using a flow cytometry assay with Hoechst and ethidium or MitoTracker dye, we show that parasite development is easily and rapidly monitored. Finally, we demonstrate that this approach can be used to screen antimalarial drugs.

scholarly journals Sulfamethoxazole Susceptibility of Mycobacterium tuberculosis Isolates from HIV-Infected Ugandan Adults with Tuberculosis Taking Trimethoprim-Sulfamethoxazole Prophylaxis

2015 ◽  
Vol 59 (9) ◽  
pp. 5844-5846 ◽  
Author(s):  
Sam Ogwang ◽  
Caryn E. Good ◽  
Brenda Okware ◽  
Mary Nsereko ◽  
Michael R. Jacobs ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Multidrug Resistant ◽  
Sub Saharan Africa ◽  
Content Type ◽  
Pulmonary Tb ◽  
Resistant Tuberculosis ◽  
Multidrug Resistant Tuberculosis ◽  
Sub Saharan

ABSTRACTAdditional drugs are needed for the treatment of multidrug-resistant tuberculosis (TB). Sulfamethoxazole has been shown to havein vitroactivity againstMycobacterium tuberculosis; however, there is concern about resistance given the widespread use of trimethoprim-sulfamethoxazole prophylaxis among HIV-infected patients in sub-Saharan Africa. Thirty-eight of 40Mycobacterium tuberculosisisolates (95%) from pretreatment sputum samples from Ugandan adults with pulmonary TB, including HIV-infected patients taking trimethoprim-sulfamethoxazole prophylaxis, were susceptible with MICs of ≤38.4 μg/ml.

scholarly journals Intestinal IgA Regulates Expression of a Fructan Polysaccharide Utilization Locus in Colonizing Gut Commensal Bacteroides thetaiotaomicron

mBio ◽  
2019 ◽  
Vol 10 (6) ◽  
Author(s):  
Payal Joglekar ◽  
Hua Ding ◽  
Pablo Canales-Herrerias ◽  
Pankaj Jay Pasricha ◽  
Justin L. Sonnenburg ◽  
...  
Keyword(s):  
Gut Microbiota ◽  
Ex Vivo ◽  
Microbial Colonization ◽  
Bacteroides Thetaiotaomicron ◽  
Content Type ◽  
Microbial Utilization ◽  
Polysaccharide Utilization Locus ◽  
The Impact

ABSTRACT Gut-derived immunoglobulin A (IgA) is the most abundant antibody secreted in the gut that shapes gut microbiota composition and functionality. However, most of the microbial antigens targeted by gut IgA remain unknown, and the functional effects of IgA targeting these antigens are currently understudied. This study provides a framework for identifying and characterizing gut microbiota antigens targeted by gut IgA. We developed a small intestinal ex vivo culture assay to harvest lamina propria IgA from gnotobiotic mice, with the aim of identifying antigenic targets in a model human gut commensal, Bacteroides thetaiotaomicron VPI-5482. Colonization by B. thetaiotaomicron induced a microbe-specific IgA response that was reactive against diverse antigens, including capsular polysaccharides, lipopolysaccharides, and proteins. IgA against microbial protein antigens targeted membrane and secreted proteins with diverse functionalities, including an IgA specific against proteins of the polysaccharide utilization locus (PUL) that are necessary for utilization of fructan, which is an important dietary polysaccharide. Further analyses demonstrated that the presence of dietary fructan increased the production of fructan PUL-specific IgA, which then downregulated the expression of fructan PUL in B. thetaiotaomicron, both in vivo and in vitro. Since the expression of fructan PUL has been associated with the ability of B. thetaiotaomicron to colonize the gut in the presence of dietary fructans, our work suggests a novel role for gut IgA in regulating microbial colonization by modulating their metabolism. IMPORTANCE Given the significant impact that gut microbes have on our health, it is essential to identify key host and environmental factors that shape this diverse community. While many studies have highlighted the impact of diet on gut microbiota, little is known about how the host regulates this critical diet-microbiota interaction. In our present study, we discovered that gut IgA targeted a protein complex involved in the utilization of an important dietary polysaccharide: fructan. While the presence of dietary fructans was previously thought to allow unrestricted growth of fructan-utilizing bacteria, our work shows that gut IgA, by targeting proteins responsible for fructan utilization, provides the host with tools that can restrict the microbial utilization of such polysaccharides, thereby controlling their growth.

scholarly journals Serum Bactericidal Assays To Evaluate Typhoidal and Nontyphoidal Salmonella Vaccines

2014 ◽  
Vol 21 (5) ◽  
pp. 712-721 ◽  
Author(s):  
Mary Adetinuke Boyd ◽  
Sharon M. Tennant ◽  
Venant A. Saague ◽  
Raphael Simon ◽  
Khitam Muhsen ◽  
...  
Keyword(s):  
Sub Saharan Africa ◽  
Content Type ◽  
Vaccine Candidates ◽  
Live Attenuated Vaccines ◽  
Bactericidal Antibody ◽  
Paratyphi B ◽  
Sub Saharan ◽  
Functional Antibodies ◽  
New Vaccines

ABSTRACTInvasiveSalmonellainfections for which improved or new vaccines are being developed include enteric fever caused bySalmonella entericaserovars Typhi, Paratyphi A, and Paratyphi B and sepsis and meningitis in young children in sub-Saharan Africa caused by nontyphoidalSalmonella(NTS) serovars, particularlyS. entericaserovars Typhimurium and Enteritidis. Assays are needed to measure functional antibodies elicited by the new vaccines to assess their immunogenicities and potential protective capacities. We developedin vitroassays to quantify serum bactericidal antibody (SBA) activity induced byS. Typhi,S. Paratyphi A,S. Typhimurium, andS. Enteritidis vaccines in preclinical studies. Complement from various sources was tested in assays designed to measure antibody-dependent complement-mediated killing. Serum from rabbits 3 to 4 weeks of age provided the best complement source compared to serum from pigs, goats, horses, bovine calves, or rabbits 8 to 12 weeks of age. ForS. Enteritidis,S. Typhimurium, andS. Typhi SBA assays to be effective, bacteria had to be harvested at log phase. In contrast,S. Paratyphi A was equally susceptible to killing whether it was grown to the stationary or log phase. The typhoidal serovars were more susceptible to complement-mediated killing than were the nontyphoidal serovars. Lastly, the SBA endpoint titers correlated with serum IgG anti-lipopolysaccharide (LPS) titers in mice immunized with mucosally administeredS. Typhimurium,S. Enteritidis, andS. Paratyphi A but notS. Typhi live attenuated vaccines. The SBA assay described here is a useful tool for measuring functional antibodies elicited bySalmonellavaccine candidates.

scholarly journals CLIPB10 is a Terminal Protease in the Regulatory Network That Controls Melanization in the African Malaria Mosquito Anopheles gambiae

2021 ◽  
Vol 10 ◽  
Author(s):  
Xin Zhang ◽  
Miao Li ◽  
Layla El Moussawi ◽  
Sally Saab ◽  
Shasha Zhang ◽  
...  
Keyword(s):  
Anopheles Gambiae ◽  
Serine Proteases ◽  
Ex Vivo ◽  
Protein A ◽  
Tumor Formation ◽  
Sub Saharan Africa ◽  
Humoral Immune ◽  
Wide Range ◽  
Sub Saharan

Humoral immune responses in animals are often tightly controlled by regulated proteolysis. This proteolysis is exerted by extracellular protease cascades, whose activation culminates in the proteolytic cleavage of key immune proteins and enzymes. A model for such immune system regulation is the melanization reaction in insects, where the activation of prophenoxidase (proPO) leads to the rapid formation of eumelanin on the surface of foreign entities such as parasites, bacteria and fungi. ProPO activation is tightly regulated by a network of so-called clip domain serine proteases, their proteolytically inactive homologs, and their serpin inhibitors. In Anopheles gambiae, the major malaria vector in sub-Saharan Africa, manipulation of this protease network affects resistance to a wide range of microorganisms, as well as host survival. However, thus far, our understanding of the molecular make-up and regulation of the protease network in mosquitoes is limited. Here, we report the function of the clip domain serine protease CLIPB10 in this network, using a combination of genetic and biochemical assays. CLIPB10 knockdown partially reversed melanotic tumor formation induced by Serpin 2 silencing in the absence of infection. CLIPB10 was also partially required for the melanization of ookinete stages of the rodent malaria parasite Plasmodium berghei in a refractory mosquito genetic background. Recombinant serpin 2 protein, a key inhibitor of the proPO activation cascade in An. gambiae, formed a SDS-stable protein complex with activated recombinant CLIPB10, and efficiently inhibited CLIPB10 activity in vitro at a stoichiometry of 1.89:1. Recombinant activated CLIPB10 increased PO activity in Manduca sexta hemolymph ex vivo, and directly activated purified M. sexta proPO in vitro. Taken together, these data identify CLIPB10 as the second protease with prophenoloxidase-activating function in An. gambiae, in addition to the previously described CLIPB9, suggesting functional redundancy in the protease network that controls melanization. In addition, our data suggest that tissue melanization and humoral melanization of parasites are at least partially mediated by the same proteases.

scholarly journals Listeria monocytogenes MenI Encodes a DHNA-CoA Thioesterase Necessary for Menaquinone Biosynthesis, Cytosolic Survival, and Virulence

2021 ◽  
Vol 89 (5) ◽  
Author(s):  
Hans B. Smith ◽  
Tin Lok Li ◽  
Man Kit Liao ◽  
Grischa Y. Chen ◽  
Zhihong Guo ◽  
...  
Keyword(s):  
Listeria Monocytogenes ◽  
Ex Vivo ◽  
Strong Activity ◽  
Content Type ◽  
Transport Chain ◽  
Intermediate Metabolites ◽  
Bacterial Replication ◽  
Functional Analyses

ABSTRACT Listeria monocytogenes is a Gram-positive, intracellular pathogen that is highly adapted to invade and replicate in the cytosol of eukaryotic cells. Intermediate metabolites in the menaquinone biosynthesis pathway are essential for the cytosolic survival and virulence of L. monocytogenes, independent of the production of menaquinone (MK) and aerobic respiration. Determining which specific intermediate metabolite(s) are essential for cytosolic survival and virulence has been hindered by the lack of an identified 1,4-dihydroxy-2-naphthoyl-coenzyme A (DHNA-CoA) thioesterase essential for converting DHNA-CoA to DHNA in the MK synthesis pathway. Using the recently identified Escherichia coli DHNA-CoA thioesterase as a query, homology sequence analysis revealed a single homolog in L. monocytogenes, LMRG_02730. Genetic deletion of LMRG_02730 resulted in an ablated membrane potential, indicative of a nonfunctional electron transport chain (ETC) and an inability to aerobically respire. Biochemical kinetic analysis of LMRG_02730 revealed strong activity toward DHNA-CoA, similar to its E. coli homolog, further demonstrating that LMRG_02730 is a DHNA-CoA thioesterase. Functional analyses in vitro, ex vivo, and in vivo using mutants directly downstream and upstream of LMRG_02730 revealed that DHNA-CoA is sufficient to facilitate in vitro growth in minimal medium, intracellular replication, and plaque formation in fibroblasts. In contrast, protection against bacteriolysis in the cytosol of macrophages and tissue-specific virulence in vivo requires the production of 1,4-dihydroxy-2-naphthoate (DHNA). Taken together, these data implicate LMRG_02730 (renamed MenI) as a DHNA-CoA thioesterase and suggest that while DHNA, or an unknown downstream product of DHNA, protects the bacteria from killing in the macrophage cytosol, DHNA-CoA is necessary for intracellular bacterial replication.

scholarly journals Solution structure, glycan specificity and of phenol oxidase inhibitory activity of Anopheles C-type lectins CTL4 and CTLMA2

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Ritika Bishnoi ◽  
Gregory L. Sousa ◽  
Alicia Contet ◽  
Christopher J. Day ◽  
Chun-Feng David Hou ◽  
...  
Keyword(s):  
Malaria Vector ◽  
Solution Structure ◽  
Biochemical Characterization ◽  
Phenol Oxidase ◽  
Sub Saharan Africa ◽  
Scattering Data ◽  
Molecular Features ◽  
Sub Saharan

Abstract Malaria, the world’s most devastating parasitic disease, is transmitted between humans by mosquitoes of the Anopheles genus. An. gambiae is the principal malaria vector in Sub-Saharan Africa. The C-type lectins CTL4 and CTLMA2 cooperatively influence Plasmodium infection in the malaria vector Anopheles. Here we report the purification and biochemical characterization of CTL4 and CTLMA2 from An. gambiae and An. albimanus. CTL4 and CTLMA2 are known to form a disulfide-bridged heterodimer via an N-terminal tri-cysteine CXCXC motif. We demonstrate in vitro that CTL4 and CTLMA2 intermolecular disulfide formation is promiscuous within this motif. Furthermore, CTL4 and CTLMA2 form higher oligomeric states at physiological pH. Both lectins bind specific sugars, including glycosaminoglycan motifs with β1-3/β1-4 linkages between glucose, galactose and their respective hexosamines. Small-angle x-ray scattering data supports a compact heterodimer between the CTL domains. Recombinant CTL4/CTLMA2 is found to function in vivo, reversing the enhancement of phenol oxidase activity in dsCTL4-treated mosquitoes. We propose these molecular features underline a common function for CTL4/CTLMA2 in mosquitoes, with species and strain-specific variation in degrees of activity in response to Plasmodium infection.

scholarly journals Discovery of Safe and Orally Effective 4-Aminoquinaldine Analogues as Apoptotic Inducers with Activity against Experimental Visceral Leishmaniasis

2011 ◽  
Vol 56 (1) ◽  
pp. 432-445 ◽  
Author(s):  
Partha Palit ◽  
Abhijit Hazra ◽  
Arindam Maity ◽  
R. S. K. Vijayan ◽  
Prabu Manoharan ◽  
...  
Keyword(s):  
Visceral Leishmaniasis ◽  
Oral Delivery ◽  
Leishmania Donovani ◽  
Reactive Oxygen Species Generation ◽  
Oral Route ◽  
Apoptotic Pathway ◽  
Content Type ◽  
Lead Compounds

ABSTRACTNovel antileishmanials are urgently required to overcome emergence of drug resistance, cytotoxic effects, and difficulties in oral delivery. Toward this, we investigated a series of novel 4-aminoquinaldine derivatives, a new class of molecules, as potential antileishmanials. 4-Aminoquinaldine derivatives presented inhibitory effects onL. donovanipromastigotes and amastigotes (50% inhibitory concentration range, 0.94 to 127 μM). Of these, PP-9 and PP-10 were the most effectivein vitroand demonstrated strong efficaciesin vivothrough the intraperitoneal route. They were also found to be effective against both sodium antimony gluconate-sensitive and -resistantLeishmania donovanistrains in BALB/c mice when treated orally, resulting in more than 95% protection. Investigation of their mode of action revealed that killing by PP-10 involved moderate inhibition of dihydrofolate reductase and elicitation of the apoptotic cascade. Our studies implicate that PP-10 augments reactive oxygen species generation, evidenced from decreased glutathione levels and increased lipid peroxidation. Subsequent disruption ofLeishmaniapromastigote mitochondrial membrane potential and activation of cytosolic proteases initiated the apoptotic pathway, resulting in DNA fragmentation and parasite death. Our results demonstrate that PP-9 and PP-10 are promising lead compounds with the potential for treating visceral leishmaniasis (VL) through the oral route.

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