Biology subjects

Lustigman, S.

Publications and source records attributed to Lustigman, S..

8 recordsLinked to original sources

bioRxiv2025

Fexinidazole and Corallopyronin A target Wolbachia-infected sheath cells present in filarial nematodes

The discovery of the endosymbiotic bacteria Wolbachia as an obligate symbiont of filarial nematodes has led to antibiotic-based treatments for filarial diseases. While lab and clinical studies have yielded promising results, recent animal studies reveal that Wolbachia levels may rebound following treatment with suboptimal doses of the antibiotic rifampicin. Previous work showed that a likely source of the bacterial rebound in females were dense clusters of Wolbachia in ovarian tissue. The number, size, and density of these Wolbachia clusters were not diminished despite antibiotic treatment. Here we define the cellular characteristics of the Wolbachia clusters in Brugia pahangi (wBp) and identify drugs that also target them. We have evidence that the Wolbachia clusters originate from newly formed sheath cells adjacent to the ovarian Distal Tip Cells. The dramatically enlarged volume of an infected sheath cell is strikingly similar to endosymbiont-induced bacteriocytes found in many insect species. Ultrastructural analysis reveals that the clustered Wolbachia present within the sheath cells exhibit a distinct morphology and form direct connections with the oocyte membrane and possibly the cytoplasm. This includes membrane-based channels providing a connection between Wolbachia-infected sheath cells and oocytes. We also determined that the Wolbachia within the sheath cells are either quiescent or replicating at a very low rate. Screens of known antibiotics and other drugs revealed that two drugs, Fexinidazole and Corallopyronin A, significantly reduced the number of clustered Wolbachia located within the sheath cells.

pathology↗

bioRxiv2023

Discovery of New Broad-Spectrum Anti-Infectives for Eukaryotic Pathogens Using Bioorganometallic Chemistry

Drug resistance observed with many anti-infectives clearly highlights the need for new broad-spectrum agents to treat especially neglected tropical diseases (NTDs) caused by eukaryotic parasitic pathogens including fungal infections. Since these diseases target the most vulnerable communities who are disadvantaged by health and socio-economic factors, new agents should be, if possible, easy-to-prepare to allow for commercialization based on their low cost. In this study, we show that simple modification of one of the most well-known antifungal drugs, fluconazole, with organometallic moieties not only improves the activity of the parent drug but also broadens the scope of application of the new derivatives. These compounds were highly effective in vivo against pathogenic fungal infections and potent against parasitic worms such as Brugia, which causes lymphatic filariasis and Trichuris, one of the soil-transmitted helminths that infects millions of people globally. Notably, the identified molecular targets indicate a mechanism of action that differs greatly from the parental antifungal drug, including targets involved in biosynthetic pathways that are absent in humans, offering great potential to expand our armamentarium against drug-resistant fungal infections and NTDs targeted for elimination by 2030. Overall, the discovery of these new compounds with broad-spectrum activity opens new avenues for the development of treatments for several current human infections, either caused by fungi or by parasites, including other NTDs, as well as newly emerging diseases. ONE-SENTENCE SUMMARYSimple derivatives of the well-known antifungal drug fluconazole were found to be highly effective in vivo against fungal infections, and also potent against the parasitic nematode Brugia, which causes lymphatic filariasis and against Trichuris, one of the soil-transmitted helminths that infects millions of people globally.

pharmacology and toxicology↗

bioRxiv2021

Localization and RNAi-driven inhibition of a Brugia malayi encoded Interleukin-5 Receptor Binding protein

A molecule termed BmIL5Rbp (aka Bm8757) was identified from Brugia malayi filarial worms and found to competitively inhibit human IL-5 binding to its human receptor. After the expression and purification of a recombinant BmIL5Rbp and generation of BmIL5Rbp-specific rabbit antibody, we localized the molecule on B. malayi worms through immunohistochemistry and immunoelectron microscopy. RNA interference was used to inhibit BmIL5Rbp mRNA and protein production. BmIL5Rbp was shown to localize to the cuticle of Brugia malayi and to be released in their excretory/secretory products. RNAi inhibited BmIL5Rbp mRNA production by 33% and reduced the surface protein expression by [~]50% and suppressed the release of BmIL5Rbp in the excretory/secretory products. RNAi has been used successfully to knock down the mRNA and protein expression of BmIL5Rbp in the early larval stages of B. malayi and provided a proof-of-principle for the local inhibition of the human IL5 receptor. These findings provide evidence that a parasite encoded IL5R antagonist could be utilized therapeutically.

pathology↗

bioRxiv2021

Insights into the L3 to L4 developmental program through proteomics

The establishment of infection with the lymphatic dwelling filarial parasites is dependent on the infectivity and subsequent development of the infective larvae (L3) within the human host to later stages (L4, adults) that require several developmental molts. The molecular mechanisms underlying the developmental processes in parasitic nematodes are not clearly defined. We report the proteomic profiles throughout the entire L3 to L4 molt using an established in vitro molting process for the human pathogen B. malayi. A total of 3466 proteins of B. malayi and 54 from Wolbachia were detected at one or more time points. Based on the proteomic profiling, the L3 to L4 molting proteome can be broadly divided into an early, middle and late phase. Enrichment of proteins, protein families and functional categories between each time point or between phases primarily relate to energy metabolism, immune evasion through secreted proteins, protein modification, and extracellular matrix-related processes involved in the development of new cuticle. Comparative analyses with somatic proteomes and transcriptomes highlighted the differential usage of cysteine proteinases (CPLs), BmCPL-1, -4 and -5 in the L3-L4 molt compared to the adults and microfilariae. Inhibition of the CPLs effectively blocked the in-vitro L3 to L4 molt. Overall, only 4 Wolbachia proteins (Wbm0495, Wbm0793, Wbm0635, and Wbm0786) were detected across all time points and suggest that they play an inconsequential role in the early developmental process. ImportanceThe neglected tropical diseases of lymphatic filariasis, onchocerciasis (or river blindness), and loiasis are the three major filarial infections of humans that cause long-term disability, impaired childhood growth, reduced reproductive capacity. Global efforts to control and/or eliminate these infections as a public health concern are based on strategies and tools to strengthen the diagnostics, therapeutic and prophylactic measures. A deeper understanding of the genes, proteins and pathways critical for the development of the parasite is needed to help further investigate the mechanisms of parasite establishment and disease progression, because not all the transmitted infective larvae get to develop successfully and establish infections. The significance of this study is in identifying the proteins and the pathways that are needed by the parasite for successful developmental molts, that in turn will allow for investigating targets of therapeutic and prophylactic potential.

microbiology↗

bioRxiv2021

Sex-specific glycosylation of secreted immunomodulatory proteins in the filarial nematode Brugia malayi

The extended persistence of filarial nematodes within a host suggests immunomodulatory mechanisms that allow the parasites to resist or evade the host immune response. There is increasing evidence for immunomodulatory glycans expressed by a diversity of parasitic worms. In this study, we integrate multiple layers of the host-parasite interface to investigate the glycome of a model filarial parasite, Brugia malayi. We report a significant overrepresentation of terminal GalNAc moieties in adult female worms coupled with an overall upregulation in O-glycosylation, T-antigen expression, and a bias for galactose containing glycans. Adult males preferentially displayed a bias for terminal GlcNAc containing glycans, and fucosylated epitopes. Subsequent proteomic analysis confirmed sex-biases in protein glycosylation and highlighted the sex-specific glycosylation of well characterized immunomodulators expressed and secreted by B. malayi. We identify sex-specific effectors at that interface and suggest approaches to selectively interfere with the parasitic life cycle and potentially control transmission.

microbiology↗

bioRxiv2020

Optimizing 3-dimensional human skin models to facilitate long-term development of Onchocerca volvulus, a debilitating obligatory human parasite

Onchocerciasis also known as river blindness is a neglected tropical disease and the world’s second-leading infectious cause of blindness in humans; it is caused by Onchocerca volvulus. Current therapies kill microfilariae but fail to kill the adult parasites, which reside within subcutaneous nodules. To support a more target-driven drug development that can also cure and thus support the elimination of this disease, an in-depth understanding of O. volvulus biology especially the factors that support the longevity of these worms in the human host (>10 years) is required. However, research is hampered by a lack of access to adult worms. O. volvulus is an obligatory human parasite and no small animal models that can propagate this parasite were successfully developed. The current optimized 2-dimensional (2-D) in vitro culturing method starting with O. volvulus infective larvae does not yet support the development of mature adult worms. To overcome these limitations, we have developed 3-dimensional (3-D) culture systems that simulated the human in vivo niche using in vitro engineered skin and adipose tissue. We show that an optimized indirect co-culture of in vitro skin tissue with fourth-stage larvae supported a significant increase in growth until the pre-adult stage with a median length of 816 – 831 μm as compared to 767 μm of 2-D cultured larvae. Notably, when larvae were co-cultured directly with adipose tissue models, a significant improvement for larval motility and thus fitness was observed; 95 % compared to 26 % in the 2-D system. These promising co-culture concepts are a first step to further improve the long-term development of adult worms in vitro, and thus provide the filarial research community with a valuable source of O. volvulus worms at various developmental stages, which may accelerate innovative unsolved biomedical inquiries into the parasite’s biology.Author summary The filarial nematode Onchocerca volvulus is an obligatory human parasite and the causative agent of onchocerciasis, better known as river blindness. In 2017, more than 20 million infections with O. volvulus were estimated worldwide, 99 % of the patients live in Africa. Current international control programs focus on the reduction of microfilaridermia by mass drug administration of ivermectin. However, to meet the elimination goals, additional therapy strategies are needed that also target the adult worms. As this parasite is obliged to humans, there are no small animal models that sustain the full life cycle of the parasite, thus greatly impeding the research on this filarial nematode. To overcome these drawbacks, we developed co-culture systems based on engineered human skin and adipose tissue that represent the in vivo niche of O. volvulus in which we were able to establish improved conditions of culturing to the pre-adult stages of the parasite. Furthermore, our new culture approach could significantly reduce the use of animal models currently used for drug testing of surrogate larvae.View Full Text

bioengineering↗

bioRxiv2020

Yeast-Expressed SARS-CoV Recombinant Receptor-Binding Domain (RBD219-N1) Formulated with Alum Induces Protective Immunity and Reduces Immune Enhancement

We developed a severe acute respiratory syndrome (SARS) subunit recombinant protein vaccine candidate based on a high-yielding, yeast-engineered, receptor-binding domain (RBD219-N1) of the SARS beta-coronavirus (SARS-CoV) spike (S) protein. When formulated with Alhydrogel®, RBD219-N1 induced high-level neutralizing antibodies against both pseudotyped virus and a clinical (mouse-adapted) isolate of SARS-CoV. Here, we report that mice immunized with RBD219-N1/Alhydrogel® were fully protected from lethal SARS-CoV challenge (0% mortality), compared to ∼ 30% mortality in mice when immunized with the SARS S protein formulated with Alhydrogel®, and 100% mortality in negative controls. An RBD219-N1 formulation Alhydrogel® was also superior to the S protein, unadjuvanted RBD, and AddaVax (MF59-like adjuvant)-formulated RBD in inducing specific antibodies and preventing cellular infiltrates in the lungs upon SARS-CoV challenge. Specifically, a formulation with a 1:25 ratio of RBD219-N1 to Alhydrogel® provided high neutralizing antibody titers, 100% protection with non-detectable viral loads with minimal or no eosinophilic pulmonary infiltrates. As a result, this vaccine formulation is under consideration for further development against SARS-CoV and potentially other emerging and re-emerging beta-CoVs such as SARS-CoV-2.Competing Interest StatementThe authors have declared no competing interest.View Full Text

immunology↗

bioRxiv2019

Prediction pipeline for discovery of regulatory motifs associated with Brugia malayi molting

Filarial nematodes can cause debilitating diseases in humans. They have complicated life cycles involving an insect vector and mammalian hosts, and they go through a number of developmental molts. While whole genome sequences of parasitic worms are now available, very little is known about transcription factor (TF) binding sites and their cognate transcription factors that play a role in regulating development. To address this gap, we developed a novel motif prediction pipeline, Emotif Alpha, that integrates ten different motif discovery algorithms, multiple statistical tests, and a comparative analysis of conserved elements between the filarial worms Brugia malayi and Onchocerca volvulus, and the free-living nematode Caenorhabditis elegans. We identified stage-specific TF binding motifs in B. malayi, with a particular focus on those potentially involved in the L3-L4 molt, a stage important for the establishment of infection in the mammalian host. Using an in vitro molting system, we tested and validated three of these motifs demonstrating the accuracy of the motif prediction pipeline.

bioinformatics↗