T1R2/T1R3 polymorphism affects sweet and fat perception: Correlation between SNP and BMI in the context of obesity development.

Genetic variations in taste receptors are associated with gustatory perception and obesity, which in turn affects dietary preferences. Given the increasing tendency of people with obesity choosing sweet, high-fat meals, the current study assessed the cross-regulation of two polymorphisms of the sweet taste receptor (T1R2/T1R3), rs35874116 and rs307355, on fat sensitivity in Indian adults. We investigated the association between taste sensitivity and BMI in the T1R2, T1R3, and CD36 polymorphic and non-polymorphic groups. The general labelled magnitude scale (gLMS) was used to assess the taste sensitivity of 249 participants in addition to anthropometric data. TaqMan Probe-based RT-PCR was employed to determine the polymorphisms. Additionally, the colorimetric method utilizing 3, 5-dinitro salicylic acid was used to evaluate the participants' salivary amylase activity. The mean detection thresholds for linoleic acid (LA) and sucrose were greater in individuals with obesity (i.e., 0.97 ± 0.08 mM and 0.22 ± 0.02 M, respectively) than in healthy adults (p < 0.0001), indicating lower sensitivity. Moreover, it was found that a greater proportion of persons with obesity fall into the polymorphic groups (i.e., 52% with genotype CD36 AA, 44% with genotype T1R2 CC, and 40% with genotype T1R3 TT). All three single nucleotide polymorphisms support the Hardy-Weinberg equilibrium (p = 0.78). The Pearson correlation analysis between LA and the sucrose detection threshold revealed a significant (p < 0.0001) positive relationship with an r value of 0.5299. Moreover, salivary amylase activity was significantly (p < 0.05) higher in the polymorphic sub-groups. The results of our study imply that genetic variations in T1R2/T1R3 receptors affect perception of both sweetness and fat, which may have an effect on obesity.

CD36 and GPR120 mediated orogustatory perception of dietary lipids and its physiological implication in the pygmy mouse Mus booduga.

Fat taste perception has long been concerned in the regulation of dietary fat intake. Substantial experimental evidence defends fat as a sixth taste modality, but its allied peripheral mechanisms are not yet well established. The present study aimed to analyse the diet-induced changes in fat taste perception and its associated physiological variations in Mus booduga. Four groups of animals were used for the present study and were fed any one of the following diet; normal diet (10% fat), low-fat diet (4% fat), high-fat diet (36% fat), or high-fat diet (HFD) (36% fat) + rapeseed oil (HFRDO) (14%) for 9 weeks. The animals were then subjected to metabolic tolerance, fat preference, and conditioned taste aversion studies. Diet-induced alterations in the expression of genes associated with lipogenesis, inflammation, and fat taste (CD36 and GPR120) were analysed. Capacitative calcium signalling induced by both linoleic acid and grifolic acid in taste bud cells (TBCs) was also analysed. In result, both the HFD and HFDRO groups revealed deterioration in glucose homoeostasis and displayed decreased preference scores for fatty acids, which are associated with lower CD36 expression and increased GPR120 expression in TBCs. Furthermore, change in [Ca2+ ]i induced by LA was also compromised in CD36 positive TBCs along with elevated systemic inflammatory and lipidemic responses in both these obese groups. Overall, for the first time, our results support that chronic HFD feeding alters the CD36 and GPR120 mediated fat taste perception in M. booduga.

Whole-Cell Phenotypic Screening of Medicines for Malaria Venture Pathogen Box Identifies Specific Inhibitors of Plasmodium falciparum Late-Stage Development and Egress.

We report a systematic, cellular phenotype-based antimalarial screening of the Medicines for Malaria Venture Pathogen Box collection, which facilitated the identification of specific blockers of late-stage intraerythrocytic development of Plasmodium falciparum First, from standard growth inhibition assays, we identified 173 molecules with antimalarial activity (50% effective concentration [EC50] ≤ 10 µM), which included 62 additional molecules over previously known antimalarial candidates from the Pathogen Box. We identified 90 molecules with EC50 of ≤1 µM, which had significant effect on the ring-trophozoite transition, while 9 molecules inhibited the trophozoite-schizont transition and 21 molecules inhibited the schizont-ring transition (with ≥50% parasites failing to proceed to the next stage) at 1 µM. We therefore rescreened all 173 molecules and validated hits in microscopy to prioritize 12 hits as selective blockers of the schizont-ring transition. Seven of these molecules inhibited the calcium ionophore-induced egress of Toxoplasma gondii, a related apicomplexan parasite, suggesting that the inhibitors may be acting via a conserved mechanism which could be further exploited for target identification studies. We demonstrate that two molecules, MMV020670 and MMV026356, identified as schizont inhibitors in our screens, induce the fragmentation of DNA in merozoites, thereby impairing their ability to egress and invade. Further mechanistic studies would facilitate the therapeutic exploitation of these molecules as broadly active inhibitors targeting late-stage development and egress of apicomplexan parasites relevant to human health.

Synthesis and in vitro evaluation of hydrazinyl phthalazines against malaria parasite, Plasmodium falciparum.

In this report, we describe the synthesis of 1-(Phthalazin-4-yl)-hydrazine using bronsted acidic ionic liquids and demonstrate their ability to inhibit asexual stage development of human malaria parasite, Plasmodium falciparum. Through computational studies, we short-listed chemical scaffolds with potential binding affinity to an essential parasite protein, dihydroorotate dehydrogenase (DHODH). Further, these compounds were synthesized in the lab and tested against P. falciparum. Several compounds from our library showed inhibitory activity at low micro-molar concentrations with minimal cytotoxic effects. These results indicate the potential of hydralazine derivatives as reference scaffolds to develop novel antimalarials.

The gustin gene variation at rs2274333 and PROP taster status affect dietary fat perception: a stepwise multiple regression model study.

Gustin, a trophic factor for taste bud development, and its polymorphism at rs2274333 influence taste perception of 6-n-propylthiouracil (PROP) and fungiform papillae (FP) density. The PROP taster status affects dietary fat sensing and body composition. However, there is a paucity of research on the gustin genotype with dietary fat perception, PROP tasting ability, and body mass index (BMI). Thus, taste sensitivity to fat and bitterness was evaluated in 178 healthy individuals. The general labeled magnitude scale was used to determine suprathreshold taste intensity ratings, whereas the alternative forced choice approach was used to estimate the taste-sensing ability. The FP density was assessed by applying blue-colored food dye over the anterior region of the tongue. Restriction fragment length polymorphism was used to detect the genetic polymorphism (rs2274333) in the carbonic anhydrase VI (CA-VI) gene. Fisher's chi-square analysis showed that the CA-VI genotype and allelic frequencies significantly correlated (p<0.001) with the PROP taster status and BMI. Healthy individuals with AA genotypes of the CA-VI polymorphism and PROP super-tasters demonstrated stronger gustatory sensitivity for linoleic acid (LA) with greater FP density in comparison to individuals with AG/GG genotypes and other PROP taster groups. Stepwise forward multiple regression analysis indicates that BMI and PROP taster status significantly influence the LA sensing ability. The suprathreshold intensity rating for LA was also significantly impacted by PROP taster status and CA-VI genotypes, with a variation of 73.3%. Overall, our findings show a relationship between the taste papillae environment and the CA-VI genetic mutation at rs2274333, which influenced the gustatory preference for dietary fat and bitter taste.

Breaking barriers: Innovative approaches for skin delivery of RNA therapeutics.

RNA therapeutics represent a rapidly expanding platform with game-changing prospects in personalized medicine. The disruptive potential of this technology will overhaul the standard of care with reference to both primary and specialty care. To date, RNA therapeutics have mostly been delivered parenterally via injection, but topical administration followed by intradermal or transdermal delivery represents an attractive method that is convenient to patients and minimally invasive. The skin barrier, particularly the lipid-rich stratum corneum, presents a significant hurdle to the uptake of large, charged oligonucleotide drugs. Therapeutic oligonucleotides need to be engineered for stability and specificity and formulated with state-of-the-art delivery strategies for efficient uptake. This review will cover various passive and active strategies deployed to enhance permeation through the stratum corneum and achieve effective delivery of RNA therapeutics to treat both local skin disorders and systemic diseases. Some strategies to achieve selectivity between local and systemic administration will also be discussed.

MicroRNAs: a symphony orchestrating evolution and disease dynamics.

The genesis of human disease lies in our evolutionary past. Evolution has featured a general trend towards increased morphological complexity, partly conferred by expansion in gene regulatory capacity via microRNA (miRNA) innovation. Many human diseases are directly related to the evolved roles of these miRNAs, and miRNA-based therapies are emerging as an appealing strategy for precision medicine. We focus on three categories of human disease - cancer, inflammation-linked pathologies, and neurological disorders - which are highly prevalent and are associated with substantial disease burden worldwide. In each category we discuss the pathogenic roles of miRNAs in the context of their evolved functions, as well as current and potential advances in targeting these miRNAs for disease therapy.

Tongue papillae density and fat taster status- a cardinal role on sweet and bitter taste perception among Indian population.

Obesity is a complex nutritional disorder that may be influenced by calorie intake and eating behaviours. Aside from many studies, the influence of papillae count on obesity is still debated. Despite the multiple variables connected to weight gain and altered taste perception, determining the association between papillae count and taste sensitivity to fat, sweet and bitter tastes, in particular, has recently become a focus of attention. This study aimed to rule out the relationship between the number of papillae on different areas of the tongue and taste sensitivity in people (n = 150) among the various groups depending on their body mass index (BMI) and fat taste sensitivity. The general labelled magnitude scale (gLMS) was used for the taste sensitivity analysis, and participants were asked to rate the intensity of each concentration of the different tastants. Using a digital camera to obtain a picture of the tongue, the density of the papillae on the tongue was counted manually by three different operators. The study reveals that the total papillae density and BMI had a direct negative correlation (r = -0.43), with papillae density (PD) decreasing as BMI increased. Concurrently, persons with higher BMIs had lower papillae distributions (32.38 ± 1.85 PD/cm2) and significantly lower perceptions of the intensity of fat taste. Further examining papillae density in the anterior front part of the tongue, the front-right section, showed significantly higher papillae distribution (74.04 ± 2.11 PD/cm2) than the front-left section. When considering the sensitivity in the tip of the tongue, middle tongue, and whole mouth, high-sensitivity individuals for fat are more sensitive to both sweet and bitter tastes. Overall, the results of this study demonstrated a strong relationship between taste sensitivity in the Indian population, BMI, and tongue papillae density in various regions of the tongue.

Exercise modifies fatty acid perception and metabolism.

AIM: Obesity is a major public health issue, which is associated with several chronic diseases. In rodents, voluntary wheel running (VWR) is a type of exercise that influences ingestive behavior. This study aims to investigate the possible function of VWR activity in the perception of fat taste and if it mitigates the immediate effects of fatty acid (FA) ingestion. METHODS: Male C57BL/6 mice were arbitrarily assigned to either a sedentary (SED) lifestyle or free access to a running wheel after 5 weeks of dietary regimen. Later these mice groups were used in the investigations on fat preference, metabolic tolerance, and electrophysiology. Diet-induced alterations in CD36 and GPR120 expression that are related to fat perception and the capacitative calcium signaling caused by FA in taste bud cells (TBCs) were also examined. RESULTS: In obese groups, VWR temporarily reduced body weight, demonstrated improvement in preference scores for FA, and recovered from a deterioration in glucose homeostasis. In CD36-positive TBCs, electrophysiological investigations showed alterations in [Ca2+ ]i caused by FA. Further, in the TBCs of circumvallate papillae, there are differences in the expression of the genes CD36 and GPR120 between the active and SED controls. Obese mice also show lower incentive salience for long-chain fatty acids (LCFA) and adapted to the reward system of VWR which may lead to improved incentive salience accredited to wheel running. CONCLUSION: In conclusion, this study provides the first evidence that VWR causes orosensory adaptations to fat and appears to alter taste preference for LCFAs.

CD36 genetic polymorphism and salivary cues are associated with oleic acid sensitivity and dietary fat intake.

There is a lack of research on the combined effects of genetic variations (specifically CD36 SNPs-rs1761667 and rs1527483), dietary food habits (vegetarian or not), and the salivary environment on obesity and taste sensitivity, especially in the Indian population. The current study aims to better understand the relationship between impaired taste perception, fat consumption, higher BMI and obesity development by examining the combined association between CD36 SNPs, oleic acid (OA) detection threshold, and food habits among Indian participants. Furthermore, the relationship between oral fatty acid (FAs) sensitivity and taste physiology factors linked to inflammation and salivary proteins was considered. Participants with the minor allele (AA/AG) of CD36 (in both rs1527483 and rs1761667) consumed more fat, particularly saturated FAs (p = 0.0351). Salivary lipopolysaccharide, which causes inflammation, was significantly greater in non-vegetarians with a higher BMI (p < 0.05), and it exhibited a negative correlation (r = -0.232 and p < 0.05) with Ki67 gene expression, a marker for taste progenitor cells. A positive correlation (r = 0.474, p = 0.04) between TLR4 mRNA levels and the OA detection threshold was also observed. Participants with BMI > 25 kg/m2 had substantially higher TNF-α and IL-6 receptor mRNA expression levels, but there were no significant differences between the vegetarian and non-vegetarian groups. However, salivary CA-VI, which has a buffering capability on the oral environment, was lower in non-vegetarian adults with BMI >25. Thus, it was shown that non-vegetarians with overweight and obesity in India were in at-risk groups for the CD36 SNP (AA/AG at rs1761667 and rs1527483) and had higher levels of inflammatory markers, which exacerbated alterations in food behaviour and physiological changes, indicating their relevance in the development of obesity.

Genetic variation in sweet taste receptors and a mechanistic perspective on sweet and fat taste sensation in the context of obesity.

Taste sensation enables humans to make nutritionally important decisions such as food preference and consumption. It functions as deterministic factors for unpropitious eating behavior, leading to overweight and obesity. The hedonistic feeling on consumption of fat and sugar-rich meals, in particular, has a negative influence on health. In addition, impairment in the taste receptors alters the downstream signaling of taste transduction pathway. Hence, genetic polymorphism in typical taste receptors is a predictor of taste sensitivity variance across individuals. The present review summarizes the effect of a single nucleotide polymorphism (SNP) in sweet taste receptors (T1R2/T1R3) on taste perception among individuals of various body mass index (BMI). Furthermore, in the context of obesity, we discussed the possibility of crosstalk between fat and sweet receptors as well as taste dysfunction in diseased individuals. In overall, a greater understanding of the physiological relationship between taste receptors, altered taste sensitivity, and genetic polymorphisms should lead to more effective obesity prevention approaches.

Repurposing Belinostat for Alleviation of Atopic Dermatitis.

Atopic dermatitis (AD) is a highly prevalent chronic inflammatory skin disease that is characterized by intense pruritus, seriously affecting patients' quality of life. Its pathophysiology, which involves both the adaptive and innate immune responses as well as skin barrier defects, is still poorly understood. We recently identified a microRNA, miR-335, as a key driver of keratinocyte differentiation and cornification, which is essential for the establishment of a healthy skin barrier. However, expression of miR-335 is lost in AD, leading to barrier defect. We further demonstrated how belinostat, a histone deacetylase inhibitor, can effectively restore miR-335 and resolve the barrier defect in a dry skin model. Here, in this commentary, we highlight the role of belinostat in the treatment of AD and discuss the need for more research into crosstalk between epigenetic and non-coding RNA-based regulation, as well as possible therapeutic strategies targeting the epigenome.

Measurements of the swimming speeds of motile microorganisms using object tracking and their correlation with water pollution and rheology levels.

Self-propelled microscopic organisms are ubiquitous in water. Such organisms' motility depends on hydrodynamic and physical factors related to the rheology of the surrounding media and biological factors depending on the organisms' state and well-being. Here we demonstrate that the swimming speed of Paramecium aurelia, a unicellular protozoan, globally found in fresh, brackish, and salt waters, can be used as a measurable frugal indicator of the presence of pollutants in water. This study establishes a significant and consistent relationship between Paramecia's swimming speed and the presence of five different organic and inorganic contaminants at varying concentrations centered around drinking water thresholds. The large size and ubiquity of the targeted microorganism, the avoidance of reagents or specialized tools for the measurement, and the simple data collection based on an object tracking algorithm enable the automatization of the assessment and real-time results using globally available technology.

Anti-Obesity Effect of T. Chebula Fruit Extract on High Fat Diet Induced Obese Mice: A Possible Alternative Therapy.

Occurrence of obesity and its associated metabolic disorders continues to escalate. The present study evaluates the anti-obesity effects of ethanolic fruit extract of Terminalia chebula (EETC) on high fat diet induced obese mice. The bioactive compounds present in the EETC is evaluated by Fourier-transform infrared (FT-IR), Gas chromatography-mass spectrometry (GC-MS), and Liquid chromatography-mass spectrometry (LC-MS) analysis. The effects of EETC on energy intake, glucose tolerance, and various biochemical parameters were analyzed using laboratory mice. Relative gene expression of Fatty acid synthase (FAS), Peroxisome proliferator-activated receptors α (PPARα), Carnitine palmitoyltransferase-1 (CPT-1), Tumor necrosis factor alpha (TNF-α) as well as Interleukin 6 (IL-6) were analyzed in liver and adipose tissues. The findings reveal the hypolipidemic and anti-obesity potential of EETC on high fat fed obese mice. EETC exerts its anti-obesity effects by suppressing lipogenesis through reduction in lipogenic enzyme (FAS) expression, increased fatty acid oxidation via PPARα and CPT-1 and by triggering the anti-inflammatory responses. To our knowledge, this is the first report of the effect of EETC on PPARα and CPT-1 in in vivo.

Fat taste signal transduction and its possible negative modulator components.

The incidence of obesity and its associated diseases including diabetes and various cardiovascular disease continues to escalate. Since the energy homeostasis executes a substantial role in fat-rich food intake and body weight regulation, it grows into a prevalent subject of interest for its strong energy density and high palatability. Over the decade, the notion that the dietary fatty acids convey signaling cues to oro-gustatory system embrace profound ability in understanding its function along with its perception of dietary fats. In this review, recent developments in the field of oleogustus and its downstream signaling mechanism in taste bud cells are analyzed. Notably, we made a brief attempt to expose the possible negative modulator components that had the potential to modulate the distinctive fat signal transduction components and its oro-gustatory mechanism. This review is in-sighted to urge the scientific community to work towards that goal to establish the libraries comprising both chemical and natural fat taste modifiers that adhere to fat taste receptors and alters its gustatory sense to proficiently combat obesity-linked complications.

Evaluation of ferrocenyl phosphines as potent antimalarials targeting the digestive vacuole function of Plasmodium falciparum.

Owing to their lipophilic nature and chemical stability, ferrocene and its derivatives have been widely explored as antimicrobial agents, in combination with other active chemical 'war heads'. A prime example is ferroquine, an analogue of chloroquine obtained by covalently bonding ferrocene to 4-aminoquinoline, which possesses superior efficacy against multi-drug resistant malaria parasites. Herein, we explored the possibility of combining the ferrocenyl moiety with a phosphine unit and the subsequent inclusion of gold(i) to derive a molecular framework with demonstrated potential in inhibiting parasitic diseases. A library of 24 compounds consisting of 5 non-functionalized ferrocenyl enones and 19 ferrocenyl phosphine derivatives were synthesized, verified and tested against Plasmodium (P.) falciparum, which allowed us to identify compounds with low micromolar potency against both normal and chloroquine-resistant strains. Through flow cytometry combined with microscopic examination of Giemsa-stained thin smears, we observed that most of the active compounds interfered with trophozoite development as well as schizont maturation. The gold complex, namely G3, derived from the hydrophosphination of the terminal furan bearing an enone substrate showed the highest inhibitory potential. We demonstrate that G3 is affecting the parasite's metabolic processes as evident from the swollen digestive vacuole. Furthermore, G3 significantly affected heme de-toxification as determined through the ß-hematin assay, which caused apparent oxidative stress on parasites leading to death. Collectively, these results point out the potential of gold-conjugated ferrocenyl phosphine derivatives as antimalarials targeting the digestive vacuole function and metabolism of parasites.

A reference document on Permissible Limits for solvents and buffers during in vitro antimalarial screening.

Antimalarial drug discovery expands on targeted and phenotype-based screening of potential inhibitory molecules to ascertain overall efficacy, phenotypic characteristics and toxicity, prior to exploring pharmacological optimizations. Candidate inhibitors may have varying chemical properties, thereby requiring specific reconstitution conditions to ensure solubility, stability or bioavailability. Hence, a variety of solvents, buffers, detergents and stabilizers become part of antimalarial efficacy assays, all of which, above certain threshold could interfere with parasite viability, invasion or red blood cell properties leading to misinterpretation of the results. Despite their routine use across malaria research laboratories, there is no documentation on non-toxic range for common constituents including DMSO, glycerol, ethanol and methanol. We herein constructed a compatibility reference guide for 14 such chemicals and estimated their Permissible Limit against P. falciparum asexual stages at which viability and replication of parasites are not compromised. We also demonstrate that at the estimated Permissible Limit, red blood cells remain healthy and viable for infection by merozoites. Taken together, this dataset provides a valuable reference tool for the acceptable concentration range for common chemicals during in vitro antimalarial tests.

Targeted Phenotypic Screening in Plasmodium falciparum and Toxoplasma gondii Reveals Novel Modes of Action of Medicines for Malaria Venture Malaria Box Molecules.

The Malaria Box collection includes 400 chemically diverse small molecules with documented potency against malaria parasite growth, but the underlying modes of action are largely unknown. Using complementary phenotypic screens against Plasmodium falciparum and Toxoplasma gondii, we report phenotype-specific hits based on inhibition of overall parasite growth, apicoplast segregation, and egress or host invasion, providing hitherto unavailable insights into the possible mechanisms affected. First, the Malaria Box library was screened against tachyzoite stage T. gondii and the half-maximal effective concentrations (EC50s) of molecules showing ≥80% growth inhibition at 10 µM were determined. Comparison of the EC50s for T. gondii and P. falciparum identified a subset of 24 molecules with nanomolar potency against both parasites. Thirty molecules that failed to induce acute growth inhibition in T. gondii tachyzoites in a 2-day assay caused delayed parasite death upon extended exposure, with at least three molecules interfering with apicoplast segregation during daughter cell formation. Using flow cytometry and microscopy-based examinations, we prioritized 26 molecules with the potential to inhibit host cell egress/invasion during asexual developmental stages of P. falciparum. None of the inhibitors affected digestive vacuole integrity, ruling out a mechanism mediated by broadly specific protease inhibitor activity. Interestingly, five of the plasmodial egress inhibitors inhibited ionophore-induced egress of T. gondii tachyzoites. These findings highlight the advantage of comparative and targeted phenotypic screens in related species as a means to identify lead molecules with a conserved mode of action. Further work on target identification and mechanism analysis will facilitate the development of antiparasitic compounds with cross-species efficacy. IMPORTANCE The phylum Apicomplexa includes many human and animal pathogens, such as Plasmodium falciparum (human malaria) and Toxoplasma gondii (human and animal toxoplasmosis). Widespread resistance to current antimalarials and the lack of a commercial vaccine necessitate novel pharmacological interventions with distinct modes of action against malaria. For toxoplasmosis, new drugs to effectively eliminate tissue-dwelling latent cysts of the parasite are needed. The Malaria Box antimalarial collection, managed and distributed by the Medicines for Malaria Venture, includes molecules of novel chemical classes with proven antimalarial efficacy. Using targeted phenotypic assays of P. falciparum and T. gondii, we have identified a subset of the Malaria Box molecules as potent inhibitors of plastid segregation and parasite invasion and egress, thereby providing early insights into their probable mode of action. Five molecules that inhibit the egress of both parasites have been identified for further mechanistic studies. Thus, the approach we have used to identify novel molecules with defined modes of action in multiple parasites can expedite the development of pan-active antiparasitic agents.

A portable image-based cytometer for rapid malaria detection and quantification.

Increasing resistance by malaria parasites to currently used antimalarials across the developing world warrants timely detection and classification so that appropriate drug combinations can be administered before clinical complications arise. However, this is often challenged by low levels of infection (referred to as parasitemia) and presence of predominantly young parasitic forms in the patients' peripheral blood. Herein, we developed a simple, inexpensive and portable image-based cytometer that detects and numerically counts Plasmodium falciparum infected red blood cells (iRBCs) from Giemsa-stained smears derived from infected blood. Our cytometer is able to classify all parasitic subpopulations by quantifying the area occupied by the parasites within iRBCs, with high specificity, sensitivity and negligible false positives (~ 0.0025%). Moreover, we demonstrate the application of our image-based cytometer in testing anti-malarial efficacy against a commercial flow cytometer and demonstrate comparable results between the two methods. Collectively, these results highlight the possibility to use our image-based cytometer as a cheap, rapid and accurate alternative for antimalarial testing without compromising on efficiency and minimal processing time. With appropriate filters applied into the algorithm, to rule out leukocytes and reticulocytes, our cytometer may also be used for field diagnosis of malaria.