Biology subjects

Hakimi, H.

Publications and source records attributed to Hakimi, H..

5 recordsLinked to original sources

bioRxiv2025

Comparative transcriptome profiles of first-stage larvae and adult female Dracunculus medinensis (Guinea worm)

Dracunculus medinensis, also called the Guinea worm, is a nematode that causes dracunculiasis, a debilitating neglected tropical disease in humans. The parasite is currently targeted by the global Guinea Worm Eradication Program (GWEP). Historically, GWEP in endemic countries have focused on interrupting transmission of the disease through intervention such as isolation and management of patients, health education, provision of improved water sources and promotion of filtering drinking water to avoid ingestion of the copepod intermediate host (IH) that may contain infectious third-stage larvae. The recent shift of Guinea worm infections in animals - particularly domestic dogs - has introduced an additional challenge to the eradication program, underscoring the urgent need for diagnostics and therapeutics. Understanding the parasite biology and survival strategies in the mammalian host, the copepod IH, and fresh water is pivotal to identifying new control measures. Comparative transcriptomic analysis provides a powerful tool to uncover the molecular mechanisms underlying parasite survival and adaptations. Here, we compared the transcriptome of adult gravid female and first-stage larvae (L1), the stage infective for the copepod IH. Comparative transcriptomic analysis of two adult females and their L1 revealed an upregulation of genes involved in translation, transcription, and DNA repair in L1, likely reflecting adaptations essential for survival in freshwater and subsequent infection of copepods. Additionally, genes involved in cuticle formation were upregulated in adult females highlighting the role of cuticle integrity in retaining millions of L1 until the gravid female worm emerges. We identified highly expressed genes in the adult female that may represent promising candidates for diagnostic markers. This study provides novel insights into the biology of the Guinea worm by examining the transcriptome of L1 and adult female stages. These findings could support the development of novel diagnostics and therapeutics to advance the ongoing eradication effort. Authors summaryGuinea worm disease is caused by the nematode Dracunculus medinensis, a parasitic worm responsible for a debilitating neglected tropical disease in humans and targeted for global eradication. Infection occurs through the consumption of contaminated drinking water with the infective larval stage harbored within fresh water crustacean copepods - the intermediate host of the parasite. The high number of Guinea worm infections in animals specially dogs poses a significant challenge to eradication, as infected animals act as reservoirs, contributing to the parasites persistence in the environment. Furthermore, the absence of early diagnostic tools and effective therapeutics complicates disease control. In this study, we performed a comparative transcriptome analysis of adult female Guinea worms and their first-stage larvae. We identified highly expressed genes in the adult female that may represent promising candidates for diagnostic markers. Additionally, we found genes and pathways upregulated in first-stage larvae which are likely essential for survival in freshwater and subsequent infection of copepods. Our study provides insight into the molecular mechanisms underlying Guinea worm survival across life stages and environments. These findings aim to support the development of novel diagnostics and therapeutics to advance ongoing eradication efforts of this neglected tropical disease.

genomics↗

bioRxiv2024

Assessing the performance of TRX and DUF148 antigens for detection of prepatent Guinea worm (Dracunculus medinensis) infection in dogs

Guinea worm (GW, Dracunculus medinensis) is a nematode that causes a painful and debilitating neglected tropical disease in humans. The GW Eradication Program has decreased human infections by >99% over the last 40 years. However, GW emergence in animal hosts, particularly dogs, has hampered eradication efforts. Currently, there is no method for diagnosing GW infection in animals during the prepatent period, before the adult female worms emerge. Previous works have identified two GW proteins, TRX and DUF148, as immunoreactive antigens with GW-positive human and dog sera. This study developed and validated indirect enzyme-linked immunosorbent assays (ELISA) using each antigen alone or in a combination of both antigens. Using serum samples from experimentally exposed dogs, TRX and DUF148 showed reactivity at 9- and 11-weeks post-exposure, respectively. In an experimentally infected ferret, TRX and DUF148 showed reactivity at 13- and 15-weeks post-exposure, respectively. These antigens were further validated using sera of dogs from endemic villages in Chad (n=47) and shelter dogs from the non-endemic United States (n=492). DUF148 showed better reactivity and sensitivity of 76.6.% in detecting GW infection in prepatent sera compared to TRX. However, DUF148 cross-reacted with one serum sample from Brugia pahangi experimental infection and several shelter dog sera. The anti-DUF148 titer was significantly higher in the shelter dogs positive for gastrointestinal nematodes than in negative dogs. To mitigate this cross-reaction, we produced 3 peptides of DUF148. Peptide 3 from the C-terminal was more reactive with prepatent sera and had a sensitivity of 83%; however, the specificity was not superior to DUF148 whole antigen. The antibody response to DUF148 in Chad dogs with the history of GW emergence waned overtime but was detectable until two years post-GW-emergence. Our findings could facilitate the development of diagnostic methods for early detection of GW infection in dogs in endemic countries. Authors summaryDracunculiasis or Guinea worm (GW) disease is a neglected tropical disease caused by the nematode Dracunculus medinensis and is targeted for eradication by the World Health Organization. The main challenge for the eradication program is the emergence of animal infections, especially dogs. Diagnostic tests are needed to find infected dogs during the prepatent period to better control infection and prevent the spread of GW. Previous studies have found two immunoreactive GW proteins, TRX and DUF148. In this study, we validated these antigens to detect infection in before GW emergence. Using sera of dogs from endemic areas of Chad, we found that DUF148 was more reactive and had promising sensitivity to detect the prepatent infection in an indirect ELISA assay. However, DUF148 also showed cross-reaction with some sera of dogs from the Unites States, a non-endemic area for GW. To mitigate this cross-reactivity, we performed ELISA using shorter peptides. We found peptide 3 that covers the C-terminal of the protein is the immunogenic part of DUF148. However, peptide 3 ELISA did not outperform whole antigen ELISA. This study confirms the applicability of DUF148 ELISA in detecting prepatent infections in dogs and could assist the GW Eradication Program.

microbiology↗

bioRxiv2024

ves1α genes expression is the major determinant of Babesia bovis-infected erythrocytes cytoadhesion to endothelial cells

Babesia bovis causes the most pathogenic form of babesiosis in cattle, resulting in high mortality in naive adults. This parasite invades red blood cells (RBCs) within the bovine hosts where they multiply and produce clinical disease. Babesia bovis exports numerous proteins into invaded RBCs changing its properties. Thus, the infected RBCs (iRBCs) are capable to cytoadhere in the microvasculature of internal organs and brain, leading to respiratory distress, neurologic signs, and mortality. Variant Erythrocyte Surface Antigen 1 (VESA1) is one of those exported proteins by B. bovis which represents a major virulence factor due to its central role in immune evasion by antigenic variation and intravascular parasite sequestration. VESA1 is a heterodimer protein encoded by ves1 and ves1{beta} multigene family and localized on the ridges, the focal point for cytoadhesion. To gain further insights into the molecular mechanisms of cytoadhesion of B. bovis, we panned the parasites with bovine brain microvasculature endothelial cells, which resulted in obtaining several clones with different cytoadherence abilities. The transcriptome analysis of 2 high and 2 low cytoadherent clones revealed that ves1 sequences were diversified, likely resulting from genomic recombination. On the other hand, ves1{beta} sequences were almost identical among these 4 clones. Insertion and expression of ves1 of a clone with high binding into ef-1 locus of a low binging clone increased cytoadherence confirming the role of ves1 suggested by our transcriptome data. Whole genome sequencing of cytoadherent clones revealed active locus of ves1 on chromosome 2. These results suggest that VESA1a proteins encoded by ves1 genes determine the cytoadherence specificity and/or cytoadherence strength of B. bovis and they are in the active site for recombination. Author summaryBabesia bovis is an apicomplexan intraerythrocytic protozoan parasite which causes the most pathogenic form of babesiosis in cattle. This pathogenicity is the result of parasite multiplication and cytoadherence of infected red blood cells (iRBCs) in the microvasculature of brain and internal organs and is mediated by B. bovis surface exposed ligand, Variant Erythrocyte Surface Antigen 1 (VESA1). Here using parasite panning assay, transcriptomics, and genetic tools, we showed that VESA1a is the main determinant of B. bovis cytoadhesion. VESA1 are large hypervariable proteins (>100kDa) consisting of VESA1a and VESA1b subunits encoded by ves1 and ves1{beta} multigene family. Panning B. bovis with bovine brain endothelial cells resulted in obtaining cytoadherent parasite clones with different binding abilities. Comparative transcriptome analysis revealed diversification of ves1 sequences. Insertion and expression of ves1 of a clone with high-binding ability in the genome of a low-binding clone increased cytoadherence confirming the role of ves1. Mapping RNA-seq on the genome of cytoadherent clones revealed the locus of active transcription and this locus was suggested to be the active site for recombination which promoted the production of variants of ves1 with different binding abilities. Altogether, our results provide new insights into B. bovis cytoadhesion and VESA1 biology.

microbiology↗

bioRxiv2024

Critical role of Babesia bovis spherical body protein 3 in ridge formation on infected red blood cells

Babesia bovis, an apicomplexan intraerythrocytic protozoan parasite, causes serious economic loss to cattle industries around the world. Infection with this parasite leads to accumulation of infected red blood cells (iRBCs) in the brain microvasculature that results in severe clinical complications known as cerebral babesiosis. Throughout its growth within iRBCs, the parasite exports various proteins to the iRBCs that lead to the formation of protrusions known as "ridges" on the surface of iRBCs, which serve as sites for cytoadhesion to endothelial cells. Spherical body proteins (SBPs; proteins secreted from spherical bodies, which are organelles specific to Piroplasmida) are exported into iRBCs, and four proteins (SBP1-4) have been reported to date. In this study, we elucidated the function of SBP3 using an inducible gene knockdown (KD) system. Localization of SBP3 was assessed by immunofluorescence assay, and only partial colocalization was detected between SBP3 and SBP4 inside the iRBCs. In contrast, colocalization was observed with VESA-1, which is a major parasite ligand responsible for the cytoadhesion. Immunoelectron microscopy confirmed localization of SBP3 at the ridges. SBP3 KD was performed using the glmS system, and effective KD was confirmed by Western blotting, immunofluorescence assay, and RNA-seq analysis. The SBP3 KD parasites showed severe growth defect suggesting its importance for parasite survival in the iRBCs. VESA-1 on the surface of iRBCs was scarcely detected in SBP3 KD parasites, whereas SBP4 was still detected in the iRBCs. Moreover, abolition of ridges on the iRBCs and reduction of iRBCs cytoadhesion to the bovine brain endothelial cells were observed in SBP3 KD parasites. Immunoprecipitation followed by mass spectrometry analysis detected the host Band 3 multiprotein complex, suggesting an association of SBP3 with iRBC cytoskeleton proteins. Taken together, this study revealed the vital role of SBP3 in ridge formation and its significance in the pathogenesis of cerebral babesiosis. Author summaryBabesia bovis causes a high-mortality complication called cerebral babesiosis in cattle, similar to cerebral malaria in humans. Both complications are caused by the cytoadhesion of infected red blood cells (iRBCs) to the host brain endothelial cells. These parasites export numerous proteins to the host iRBCs and produce protrusions on the iRBCs that are called ridges for B. bovis and knobs for Plasmodium falciparum. Ridges and knobs play an important role in cytoadhesion as they are the sites of adherence; however, the molecules responsible for ridge formation remain unknown. In this study, we showed that SBP3 is a crucial protein for ridge formation. The SBP3 knockdown parasites showed severe growth defects and abolition of ridges on the iRBCs, and cytoadhesion of iRBCs to the bovine brain endothelial cells was significantly reduced. An immunoprecipitation experiment suggested an association of SBP3 with the host Band 3 multiprotein complex. Although there is no similarity in amino acid sequences, we suggest SBP3 to be a functional ortholog of KAHRP in P. falciparum. In summary, our results shed light on the molecular mechanism of ridge formation and the pathogenesis of B. bovis.

microbiology↗

bioRxiv2019

Plasmodium caprae infection is negatively correlated with infection of Theileria ovis and Anaplasma ovis in goats

Theileria, Babesia, and Anaplasma are tick-borne pathogens affecting livestock industries worldwide. In this study, we surveyed the presence of Babesia ovis, Theileria ovis, Theileria lestoquardi, Anaplasma ovis, Anaplasma phagocytophilum, and Anaplasma marginale in 200 goats from 3 different districts in Sistan and Baluchestan province, Iran. Species-specific diagnostic PCR and sequence analysis revealed that 1.5%, 12.5%, and 80% of samples were positive for T. lestoquardi, T. ovis, and A. ovis, respectively. Co-infections of goats with up to 3 pathogens were seen in 22% of the samples. We observed a positive correlation between A. ovis and T. ovis infection. In addition, by analyzing the data with respect to Plasmodium caprae infection in these goats, a negative correlation was found between P. caprae and A. ovis and between P. caprae and T. ovis. This study contributes to understanding the epidemiology of vector-borne pathogens and their interplay in goats.\n\nImportanceTick-borne pathogens include economically important pathogens restricting livestock farming worldwide. In endemic areas livestock are exposed to different tick species carrying various pathogens which could result in co-infection with several tick-borne pathogens in a single host. The co-infection and interaction among pathogens are important in determining the outcome of disease. Little is known about pathogen interactions in the vector and the host. In this study, we show for the first time that co-infection of P. caprae, a mosquito transmitted pathogen, with T. ovis and A. ovis. Analysis of goat blood samples revealed a positive correlation between A. ovis and T. ovis. Moreover, a negative correlation was seen between P. caprae, a mosquito transmitted pathogen, and the tick-borne pathogens T. ovis or A. ovis.

epidemiology↗