Toxocariasis is still a neglected parasitic disease worldwide and much about its biology and diagnosis has yet to be understood. The migration of third stage larvae via bloodstream suggests a potential use of molecular t...Toxocariasis is still a neglected parasitic disease worldwide and much about its biology and diagnosis has yet to be understood. The migration of third stage larvae via bloodstream suggests a potential use of molecular tools in diagnosis as well to deepen the knowledge about its migration behaviors. Conventional PCR was applied in serum and tissue samples from BALB/c mice infected with 5 and 500 embryonated eggs. Blood samples were collected at 15, 30, 60, 90 and 120 days post-infection. Organs were excised at 170 days post infection. There was no DNA amplification in serum samples in any group or day post-infection; contrarily, tissue samples showed DNA amplification. These results also support a continuous larval migration after and/or simultaneously with the neurotropic-myotropic phase. Thus, molecular tools might be useful as a differential diagnosis method, but do not replace immunodiagnostics techniques.
Plasmodium the causative agent of malaria is a member of the phylum Apicomplexa, where all invasive forms have a substrate-dependent motility called gliding, key to malaria transmission. Gliding allows parasite host-cell...Plasmodium the causative agent of malaria is a member of the phylum Apicomplexa, where all invasive forms have a substrate-dependent motility called gliding, key to malaria transmission. Gliding allows parasite host-cell recognition, binding, cell entry and trespassing the cytoplasm. In this process Plasmodium releases molecules from micronemes and the cell surface that are deposited on trails left behind on the substratum as the parasite progresses. Previously we identified the heat shock protein 70-1 (HSP 70-1) on the surface and micronemes of P. berghei ookinetes, the parasite form that invades the mosquito midgut. To investigate if this protein is shed of from the parasite during invasion, we searched HSP 70-1 in gliding trails deposited on a solid surface by P. berghei ookinetes.
Brugia malayi asparaginyl-tRNA synthetase (BmAsnRS) has been identified as an immunodominant antigen and a physiocrine that mimics Interleukin-8 (IL-8) to induce chemotaxis and angiogenesis in endothelial cells. Computat...Brugia malayi asparaginyl-tRNA synthetase (BmAsnRS) has been identified as an immunodominant antigen and a physiocrine that mimics Interleukin-8 (IL-8) to induce chemotaxis and angiogenesis in endothelial cells. Computational analyses have shown that the N-terminal region of BmAsnRS has a novel fold, a lysine rich β-hairpin α-helix, (FLIRTKKDGKQIWE) which is similar to that present in IL-8 chemokine, CXCR1. This novel fold is involved in tRNA binding and is integral for the manifestation of the disease, lymphatic filariasis (LF). Drug discovery programmes carried out so far for LF have not been successful because of the target (BmAsnRS) resistance due to the disease-associated mutation. Mutations in AARS targets have been shown to correlate with several diseases. However, no disease-associated mutational studies have been carried out for LF. BmAsnRS has been an established target for LF. It was proposed, therefore, to study the effect of single point mutations in BmAsnRS so as to elucidate the molecular target. An understanding of the molecular consequences of mutations will provide insight into how resistance develops in addition to the identification of the likely resistance-conferring mutations. Three mutants were, therefore, generated by site-directed mutagenesis using CUPSAT server and their angiogenic properties evaluated. Cytometric analysis of the mutants on endothelial cell cycle was also carried out. CUPSAT prediction of protein stability upon point mutations reveal that two mutants generated are likely resistance-conferring mutations. All the three mutants show significant reduction in their angiogenic properties and reduction in the DNA content in the cells of S and G2/M phases thus showing altered function of the gene encoding the drug target. The resistance- conferring mutants, however, show angiogenic properties nearer to the wild type protein, BmAsnRS. Future work on designing newer drugs may take into consideration these drug resistance-conferring mutations.
Lymphatic filariasis is a parasitic disease caused by the worms Wuchereria bancrofti, Brugia malayi and Brugia timori. Three anti-filarial drugs namely Diethylcarbamazine, Ivermectin and Albendazole and their combination...Lymphatic filariasis is a parasitic disease caused by the worms Wuchereria bancrofti, Brugia malayi and Brugia timori. Three anti-filarial drugs namely Diethylcarbamazine, Ivermectin and Albendazole and their combinations are used as the control strategies for filariasis. The disease has received much attention in drug discovery due to the unavailability of vaccines and the toxic pharmaceutical properties of the existing drugs. In Wolbachia endosymbiont Brugia malayi, the UDP-N-acetylmuramoyl-tripeptide-d-alanyl-d-alanine ligase (MurF) plays a key role in peptidoglycan biosynthesis pathway and therefore can be considered as effective drug target against filariasis disease. Therefore, in the present study, MurF was selected as the therapeutic target to identify specific inhibitors against filariasis. Homology modeling was performed to predict the three-dimensional structure of MurF due to the absence of the experimental structure. Further molecular dynamics simulation and structure-based high throughput virtual screening with three different chemical databases (Zinc, Maybridge and Specs) were carried out to identify potent inhibitors and also to check their conformations inside the binding site of MurF, respectively. Top three compounds with high docking score and high relative binding affinity against MurF were selected. Further, validation studies, including predicted ADME (Absorption, Distribution, Metabolism, Excretion) assessment, binding free energy using MM-GBSA (Molecular Mechanics Generalized Born Surface Area) and DFT (Density Functional Theory) calculations were performed for the top three compounds. From the results, it was observed that all the three compounds were predicted to show high reactivity, acceptable range of pharmacokinetic properties and high binding affinity with the drug target MurF. Overall, the results could provide more understanding on the inhibition of MurF enzyme and the screened compounds could lead to the development of new specific anti-filarial drugs.
Mandala WL, Harawa V, Dzinjalamala F
… +1 more, Tembo D
Mol Biochem Parasitol
· 2021 Nov · PMID 34666102
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Plasmodium falciparum malaria still remains a major global public health challenge with over 220 million new cases and well over 400,000 deaths annually. Most of the deaths occur in sub-Saharan Africa which bears 90 % of...Plasmodium falciparum malaria still remains a major global public health challenge with over 220 million new cases and well over 400,000 deaths annually. Most of the deaths occur in sub-Saharan Africa which bears 90 % of the malaria cases. Such high P. falciparum malaria-related morbidity and mortality rates pose a huge burden on the health and economic wellbeing of the countries affected. Lately, substantial gains have been made in reducing malaria morbidity and mortality through intense malaria control initiatives such as use of effective antimalarials, intensive distribution and use of insecticide-treated nets (ITNs), and implementation of massive indoor residual spraying (IRS) campaigns. However, these gains are being threatened by widespread resistance of the parasite to antimalarials, and the vector to insecticides. Over the years the use of vaccines has proven to be the most reliable, cost-effective and efficient method for controlling the burden and spread of many infectious diseases, especially in resource poor settings with limited public health infrastructure. Nonetheless, this had not been the case with malaria until the most promising malaria vaccine candidate, RTS,S/AS01, was approved for pilot implementation programme in three African countries in 2015. This was regarded as the most important breakthrough in the fight against malaria. However, RTS,S/AS01 has been found to have some limitations, the main ones being low efficacy in certain age groups, poor immunogenicity and need for almost three boosters to attain a reasonable efficacy. Thus, the search for a more robust and effective malaria vaccine still continues and a better understanding of naturally acquired immune responses to the various stages, including the transmissible stages of the parasite, could be crucial in rational vaccine design. This review therefore compiles what is currently known about the basic biology of P. falciparum and the natural malaria immune response against malaria and progress made towards vaccine development.
Although many important mediators and critical pathways are found to be involved in host immune responses to Haemonchus contortus infection, the initial responses to infection in the naïve and in the previously exposed s...Although many important mediators and critical pathways are found to be involved in host immune responses to Haemonchus contortus infection, the initial responses to infection in the naïve and in the previously exposed state have not been compared at the transcriptional level. To further understand the development of adaptive immunity to H. contortus infection, we compared the early abomasal gene expression patterns between a primary and a tertiary challenge for four lines of sheep to discover differentially expressed genes (DEGs). The sheep were from the resistant (R) and susceptible (S) lines of two flocks of sheep selected for divergent responses to gastro-intestinal parasites (HSF and TSF). The flocks have separate origins and were initiated using two different strains of Merino sheep. One of the DEGs, mast cell proteinase 1, had significantly lower expression in tertiary compared to primary infections for all four lines of sheep. This gene was not identified in previous studies where resistant and susceptible sheep samples were compared within infection time points. Comparing the differentially expressed genes (DEGs) for the two R lines reveals that responses differed very little between the primary and tertiary challenges for HSFR and only two genes were identified, in contrast to the TSFR where there were 134 genes identified including the two identified using the HSFR animals. Similarly, comparing the primary and tertiary challenges for HSFS identified 15 DEGs, whilst for TSFS there were 128 DEGs identified. It is surprising that so few genes respond similarly between the two challenge regimes across the four lines of sheep, and suggests significant differences in immune mechanisms between the two flocks (across the lines) and also between the lines within flocks. Our results offer a quantitative snapshot comparing the transcriptome in the ovine abomasum between primary and tertiary infections with H. contortus in both genetically resistant and susceptible sheep.
The oncosphere larvae of tapeworms cyclically extend and retract their hooks during the penetration of the intestine of their intermediate hosts. The mechanisms regulating these movements are essentially unknown, in part...The oncosphere larvae of tapeworms cyclically extend and retract their hooks during the penetration of the intestine of their intermediate hosts. The mechanisms regulating these movements are essentially unknown, in part due to the biohazardous nature of oncospheres from human pathogens. In this work, we standardized a method for the analysis of motility of hatched oncospheres (hexacanths) of the model tapeworm Hymenolepis microstoma. We used this assay to explore the relevance of protein kinases C (PKC) and A (PKA) in these processes. Pharmacological inhibition of the PKC pathway resulted in impaired larval motility. On the other hand, the PKA inhibitor H-89 potently blocked larval motility, as well as the motility of other life stages, although other inhibitors of the PKA pathway were not effective. This work represents the first study of the mechanisms that regulate the motility of oncospheres, and provides a path for further exploration.
Leishmaniasis is one of the fast-growing parasitic diseases worldwide. The treatment of this fatal disease presents a daunting challenge because of its adverse effects, necessity for long-term treatment regime, unavailab...Leishmaniasis is one of the fast-growing parasitic diseases worldwide. The treatment of this fatal disease presents a daunting challenge because of its adverse effects, necessity for long-term treatment regime, unavailability of functional drugs, emergence of drug resistance and the related expenditure. This calls for an urgent need for novel drugs and the evaluation of new targets. Proteins of the fatty acid biosynthetic pathway are validated as drug targets in pathogenic bacteria and certain viruses. Likewise, this pathway has been speculated as a suitable target against parasite infections. Fatty acid synthesis in parasites seems to be very complex and distinct from the counterpart mammalian host due to the presence of unique mechanisms for fatty acid biosynthesis and acquisition. In recent times, there have been few evidences of the existence of this pathway in the bloodstream form of some pathogens. The fatty acid biosynthesis thus presents a viable and attractive target for emerging therapeutics. Understanding the mechanisms underlying fatty acid metabolism is key to identifying a potential drug target. However, investigations in this direction are still limited and this article attempts to outline the existing knowledge, while highlighting the scope and relevance of the fatty acid biosynthetic pathway as a drug target. This review highlights the advances in the treatment of leishmaniasis, the importance of lipids in the pathogen, known facts about the fatty acid biosynthesis in Leishmania and how this pathway can be manipulated to combat leishmaniasis, suggesting novel drug targets.
Neobenedenia melleni, a marine fish ectoparasite, is responsible for considerable losses in the mariculture industry. In maintaining the parasite's homeostasis, sterols are structural and functional lipids that perform v...Neobenedenia melleni, a marine fish ectoparasite, is responsible for considerable losses in the mariculture industry. In maintaining the parasite's homeostasis, sterols are structural and functional lipids that perform vital functions. Thus, understanding the mechanisms of biosynthesis and the uptake of sterols can reveal potential pharmacological targets. The objective of this work was thereby to characterize the N. melleni sterols. The most abundant sterol found was cholesterol in either its free (47.48 ± 15.93 %) or esterified form. However, its precursors, squalene (3.53 ± 0.92 %) and desmosterol (0.25 ± 0.03 %), were also found, suggesting the uptake of these intermediates from hosts or an unusual active pathway of sterol biosynthesis, which can be further explored as pharmacological targets.
Efficient reverse genetics approaches are critical for the study of many organisms. The CRISPR/Cas9 gene editing system has led to a plethora of new tools for geneticists. Here, we successfully established a simplified C...Efficient reverse genetics approaches are critical for the study of many organisms. The CRISPR/Cas9 gene editing system has led to a plethora of new tools for geneticists. Here, we successfully established a simplified CRISPR/Cas9 system for the malaria model parasite Plasmodium berghei. The homologous directed repair (HDR) template is provided as a linear template with homologous arms of 600-700bp while the CRISPR elements sgRNA and Cas9 are encoded from a single plasmid utilizing the Ribozyme-Guide-Ribozyme (RGR) expression strategy. Our approach eliminates the need for negative selection markers since the plasmid cannot be incorporated into the genome. As a test case we inserted the FLAG encoding sequence into the ACT2 locus using this new approach. We showed that the genetic modification of this locus had no adverse effects on the completion of the P. berghei life cycle, including transmission through the mosquito.
Trichomoniasis is the most common non-viral sexually transmitted infection, caused by the protozoan parasite Trichomonas vaginalis, affecting millions of people worldwide. The main treatment against trichomoniasis is met...Trichomoniasis is the most common non-viral sexually transmitted infection, caused by the protozoan parasite Trichomonas vaginalis, affecting millions of people worldwide. The main treatment against trichomoniasis is metronidazole and other nitroimidazole derivatives, but up to twenty percent of clinical cases of trichomoniasis are resistant to these drugs. In this study, we used high-performance virtual screening to search for molecules that specifically bind to the protein, triosephosphate isomerase from T. vaginalis (TvTIM). By in silico molecular docking analysis, we selected six compounds from a chemical library of almost 500,000 compounds. While none of the six inhibited the enzymatic activity of recombinant triosephosphate isomerase isoforms, one compound (A4; 3,3'-{[4-(4-morpholinyl)phenyl]methylene}bis(4- hydroxy-2H-chromen-2-one) altered their fluorescence emission spectra, suggesting that this chemical might interfere in an important non-glycolytic function of TvTIM. In vitro assays demonstrate that A4 is not cytotoxic but does have trichomonacidal impact on T. vaginalis cultures. With these results, we propose this compound as a potential drug with a new therapeutic target against Trichomonas vaginalis.
Mol Biochem Parasitol
· 2021 Sep · PMID 34492240
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During schistosomiasis, the paired Schistosoma mansoni female produces about 300 eggs each day. These eggs are responsible for the clinical picture and the transmission of the disease. During female development and egg p...During schistosomiasis, the paired Schistosoma mansoni female produces about 300 eggs each day. These eggs are responsible for the clinical picture and the transmission of the disease. During female development and egg production, fs800 is expressed only in female vitelline cells. Blast search of fs800 did not show similarities with any published sequences by NCBI. We hypothesize that the product of this gene plays a role in S. mansoni egg production. By using RNA interference to knockdown fs800 and quantitative PCR to measure the gene expression in the female schistosomes, we were able to demonstrate that fs800 product is crucial for viable egg production, it has no effect on worm health or male-female pairing. Our data suggest fs800 inhibition as a potential target to prevent transmission and pathology of schistosomiasis.
Mol Biochem Parasitol
· 2021 Sep · PMID 34492239
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Toxoplasma gondii is a protozoan parasite that causes opportunistic infection in immunocompromised individuals. The parasite forms latent tissue cysts that are refractory to current treatments and give rise to life-threa...Toxoplasma gondii is a protozoan parasite that causes opportunistic infection in immunocompromised individuals. The parasite forms latent tissue cysts that are refractory to current treatments and give rise to life-threatening reactivated infection following immune suppression. Previously, we showed that guanabenz sharply reduces brain cyst count in BALB/c mice harboring latent toxoplasmosis; however, whether cyst count would change once drug treatment stopped was not addressed. In the present study, we observed a rebound in brain cysts following the discontinuation of guanabenz or a guanabenz-pyrimethamine combination therapy. The re-expansion of brain cysts was not accompanied by symptoms of acute toxoplasmosis. We also tested whether the rebound in cyst counts could be ameliorated by administering pyrimethamine during or after guanabenz treatment.
The phenomenon of non-uniform usage of the synonymous codons, where some codons are given more preference to others, is known as codon usage bias (CUB). CUB is known to be determined by two major evolutionary forces i.e....The phenomenon of non-uniform usage of the synonymous codons, where some codons are given more preference to others, is known as codon usage bias (CUB). CUB is known to be determined by two major evolutionary forces i.e. mutation pressure and selection. We used various approaches to understand the codon usage pattern in mitochondrial CO (MT-CO) genes involved in complex IV of the respiratory chain (RC) as no work was reported yet. Our present study revealed that CUB was relatively high and the coding sequences were rich in A and T. Correspondence analysis further indicated that A/T compositional properties under mutational pressure might be affecting the codon usage pattern and was different in different classes for MT-CO gene. A highly significant correlation between A% and A3%, T% and T3%, G% and G3%, C% and C3%, GC% and GC3% in all the classes indicated that compositional constraints under mutational pressure and natural selection might affect the CUB. Neutrality plot indicated that both natural selection and mutational bias affected the CUB, where, natural selection played the major role as compared to mutational pressure.
Glycosylphosphatidylinositol-phospholipase C (GPI-PLC) is an enzyme that has been implicated in GPI-dependent protein trafficking and phosphoinositide metabolism in the bloodstream stage of African trypanosomes. However,...Glycosylphosphatidylinositol-phospholipase C (GPI-PLC) is an enzyme that has been implicated in GPI-dependent protein trafficking and phosphoinositide metabolism in the bloodstream stage of African trypanosomes. However, despite the fact that it is associated with the cytoplasmic face of internal organellar compartments, its role in general membrane trafficking has not been investigated. Using a GPI-PLC null cell line, we determine the effect of GPI-PLC deficiency on these processes. Biosynthetic trafficking of lysosomal cargo, soluble cathepsin L and membrane bound p67, are unaffected. Likewise, secretory transport, recycling and ultimate lysosomal turnover of the GPI-anchored and transmembrane glycoproteins, transferrin receptor and invariant surface glycoprotein 65, respectively, were unaffected. A significant decrease in the endocytic uptake of transferrin was observed, confirming a prior report, but ultimate delivery to the lysosome was unimpacted. These results contribute to our understanding of the roles of this enigmatic enzyme in trypanosome cell biology.
Cryptosporidium parvum (C. parvum) is an intracellular parasite of the intestinal cells. It causes cryptosporidiosis that can be fatal in immunosuppressed individuals. Autophagy is a process to eliminate intracellular mi...Cryptosporidium parvum (C. parvum) is an intracellular parasite of the intestinal cells. It causes cryptosporidiosis that can be fatal in immunosuppressed individuals. Autophagy is a process to eliminate intracellular microbes. The autophagy-related 16 - like 1 (ATG16L1) gene encodes proteins involved in the autophagy pathway. Single nucleotide polymorphism (SNP) in this gene increases the invasion and survival of the intracellular microbes. This study aimed to assess whether SNP in the ATG16L1 gene influences the risk and severity of cryptosporidiosis. Group I: cases with C. parvum infection (C. parvum, n = 40) and group II: healthy control (HC, n = 120) were included. Genotyping of the ATG16L1 gene was done for all participants to determine the polymorphism status as AA, GG, or AG genotype. A significant association between C. parvum infection and ATG16L1 genotypes was detected. C. parvum group had a significantly higher frequency of GG genotype and G allele when compared to HC group. The genotypes (AG + GG) and G allele had 2.428 and 2.13 folds risk of C. parvum infection when compared to the AA genotype and the A allele. Patients with the AG + GG genotype had statistically significant higher Cryptosporidium oocyst counts in stool, higher infection intensity, more frequency of vomiting and dehydration, longer disease duration, and more recurrence. The GG or AG genotypes were independent risk factors in the disease severity (p- value = 0.013). In conclusion, ATG16L1 SNP increased the risk and severity of cryptosporidiosis. Thus, individuals with such SNP can benefit from autophagy up-regulating approaches in decreasing the risk and controlling C. parvum infection.
Thioredoxin peroxidase (TPx) plays an important role in protecting parasites against oxidative damage. However, studies on the role of TPxs in Echinococcus multilocularis are limited. In this study, one tpx gene of E. mu...Thioredoxin peroxidase (TPx) plays an important role in protecting parasites against oxidative damage. However, studies on the role of TPxs in Echinococcus multilocularis are limited. In this study, one tpx gene of E. multilocularis, named as emtpx-1, was identified. EmTPx-1 shares two positionally conserved cysteine residues (Cys48 and Cys169) with orthologs from other platyhelminths. EmTPx-1 is highly expressed in the germinal layer and present in exosome-like vesicles secreted by E. multilocularis metacestodes. EmTPx-1 displays peroxidase activity, which removes hydrogen peroxide in the presence of dithiothreitol. Furthermore, EmTPx-1 could protect DNA from oxidative damages, and EmTPx-1-expressing E. coli cells had an enhanced resistance to oxidative stress. In addition, EmTPx-1 enhanced the expression of arg1, ym1, and il-10, but suppressed inos, tnf-α, and il-1β expression in LPS-stimulated macrophages. Our data suggest a critical role for EmTPx-1 in oxidative stresses and M2 macrophage polarization.
Mol Biochem Parasitol
· 2021 Sep · PMID 34324911
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Post-translational modifications (PTMs) allow proteins to regulate their structure, localisation and function in response to cell intrinsic and environmental signals. The diversity and number of modifications on proteins...Post-translational modifications (PTMs) allow proteins to regulate their structure, localisation and function in response to cell intrinsic and environmental signals. The diversity and number of modifications on proteins increase the complexity of cellular proteomes by orders of magnitude. Several proteomic and molecular studies have revealed an abundance of PTMs in malaria parasite proteome, where mediators of PTMs play crucial roles in parasite pathogenesis and transmission. In this article, we discuss recent findings in asexual stages of ten diverse PTMs and investigate whether these proteins are expressed in sexual stages. We discovered 25-50 % of proteins exhibiting post-translational modifications in asexual stages are also expressed in sexual stage gametocytes. Moreover we analyse the function of the modified proteins shared with the gametocyte proteome and try to encourage the scientific community to investigate the roles of diverse PTMs beyond phosphorylation in sexual stages which could not only reveal unique aspects of parasite biology, but also uncover new avenues for transmission blocking.
Rumen flukes in the genus Fischoederius are neglected foodborne parasites of cattle in Asia. Fischoederius elongatus, first described in 1883 from a sample collected in Indonesia is the type-species of the genus and is f...Rumen flukes in the genus Fischoederius are neglected foodborne parasites of cattle in Asia. Fischoederius elongatus, first described in 1883 from a sample collected in Indonesia is the type-species of the genus and is found from South to East Asia. In this study Fischoederius spp were collected from cattle in Thailand. The flukes resembled F. elongatus and images of 48 specimens were taken and their DNA was isolated. The mtDNA sequence of the cytochrome c oxidase subunit I (COX1) gene was amplified by PCR and used for restriction analysis with MseI. Nine restriction patterns (A-I) were observed and the COX1 mtDNA sequence for each pattern was determined. Phylogenetic analysis clustered the nine COX1 sequences into five groups with 4.6-9.6 % sequence differences between the groups. This is beyond intragenic variation observed for the COX1 gene in other organisms and suggested that the analyzed specimens represented several species. A comparative transcriptome analysis of specimens with COX1 MseI patterns A, C, E supported this finding. The observed median base differences, both absolute and relative, in the protein coding sequences of 999 orthologs were similar to those between distinct fruit fly species. It is proposed that the genus Fischoederius contains undescribed species that follow the classic description of F. elongatus.
Kinetoplastid parasites are model eukaryotes with a complex cell cycle that is highly regulated both spatially and temporally. In addition, diseases caused by these parasites continue to have a significant impact on huma...Kinetoplastid parasites are model eukaryotes with a complex cell cycle that is highly regulated both spatially and temporally. In addition, diseases caused by these parasites continue to have a significant impact on human and animal health worldwide. While there have been advancements in chemotherapy for these diseases, there is a continual need for an arsenal of compounds that have robust anti-parasite activity with minimal impact on the human host. While investigating a series of 2,3-diphenyl-2,3-dihydro-4H-1,3-thiaza-4-one heterocycles with potential activity against these parasites, we found a pyridothiazinone that inhibits growth of the monoxenous parasite Crithidia fasciculata and two life cycle stages of Trypanosoma brucei. This inhibition is more pronounced in T. brucei and is associated with an unusual pre-abscission cell cycle arrest. Exploring the mode of action for these and related compounds in kinetoplastids may provide tools with which to explore cell cycle regulation in these important organisms.