A deep learning-based model for detecting Leishmania amastigotes in microscopic slides: a new approach to telemedicine.

BACKGROUND: Leishmaniasis, an illness caused by protozoa, accounts for a substantial number of human fatalities globally, thereby emerging as one of the most fatal parasitic diseases. The conventional methods employed for detecting the Leishmania parasite through microscopy are not only time-consuming but also susceptible to errors. Therefore, the main objective of this study is to develop a model based on deep learning, a subfield of artificial intelligence, that could facilitate automated diagnosis of leishmaniasis. METHODS: In this research, we introduce LeishFuNet, a deep learning framework designed for detecting Leishmania parasites in microscopic images. To enhance the performance of our model through same-domain transfer learning, we initially train four distinct models: VGG19, ResNet50, MobileNetV2, and DenseNet 169 on a dataset related to another infectious disease, COVID-19. These trained models are then utilized as new pre-trained models and fine-tuned on a set of 292 self-collected high-resolution microscopic images, consisting of 138 positive cases and 154 negative cases. The final prediction is generated through the fusion of information analyzed by these pre-trained models. Grad-CAM, an explainable artificial intelligence technique, is implemented to demonstrate the model's interpretability. RESULTS: The final results of utilizing our model for detecting amastigotes in microscopic images are as follows: accuracy of 98.95 1.4%, specificity of 98 2.67%, sensitivity of 100%, precision of 97.91 2.77%, F1-score of 98.92 1.43%, and Area Under Receiver Operating Characteristic Curve of 99 1.33. CONCLUSION: The newly devised system is precise, swift, user-friendly, and economical, thus indicating the potential of deep learning as a substitute for the prevailing leishmanial diagnostic techniques.

Imaging patterns of Lophomonas blattarum infection in the respiratory tract: a registry-based analysis.

BACKGROUND: Lophomonas blattarum is an emerging protozoan that mostly infects the lower respiratory tract and causes pulmonary lophomoniasis. Radiologic findings in patients with pulmonary lophomoniasis have yet to be studied. Thus, we conducted a registry-based clinical investigation to evaluate the radiologic findings of lophomoniasis. METHODS: In this cross-sectional study, 34 Lophomonas positive patients were enrolled. Demographic data, relevant characteristics, and radiologic findings of the patients were recorded and analyzed. RESULTS: Thirty-four (male = 18, female = 16) patients with an average age of 52.21 ± 20.48 years old were examined. Radiological findings such as Alveolar consolidation (26.5%), Ground glass opacity (5.9%), Centrilobular nodules (23.5%), Tree -in- bud (38.2%), Cavitation (23.5%), Pleural effusion (23.5%), Interstitial opacity (8.8%), Lymphadenopathy (23.5%), Bronchocele (5.9%), Bronchiectasis (29.4%), Nodules (8.8%) and Mass (11.8%) were obtained, that the frequency of all radiological findings was less than 50%. CONCLUSION: In this study, the most common radiological findings in patients with lophomoniasis were tree-in-bud nodules, alveolar consolidation, bronchiectasis, and centrilobular nodules which were mostly seen in the right lung and its middle and lower lobes. Given that the radiologic findings of this disease are unknown, it can be considered in differential diagnosis.

Detection of circulatory E. granulosus-derived cell-free DNA in the plasma and urine of human cystic echinococcosis using an in-house PCR: a potential promising diagnostic biomarker.

BACKGROUND: The diagnostic tool for identifying cystic echinococcosis (CE) patients at an early stage is currently lacking. However, circulatory cell-free DNA (cfDNA) has shown potential as a biomarker for parasitic infections and could be used for diagnosing CE. RESEARCH DESIGN AND METHODS: The plasma and urine samples were collected from 39 patients with confirmed CE through imaging and histopathological techniques. All plasma samples were tested for anti-echinococcal antibodies using a commercial ELISA test. Total plasma and urine cfDNA were extracted and an in-house PCR assay was developed to detect E. granulosus specific cfDNA in the samples of CE patients. RESULTS: Out of the 39 patients, 30 tested positive for E. granulosus using serology, with a sensitivity of 76.9%. Moreover, the detection rates for the cfDNA were 79.5% in plasma samples and 58.97% in urine samples using the 80 bp COX1 gene. The plasma-based PCR and serology test showed the highest agreement (Kappa = 0.53). CONCLUSIONS: Plasma-based PCR has been found to be a reliable diagnostic tool for identifying CE patients at different cyst stages. It offers validity, speed, and sufficient sensitivity, making it an alternative to serology in diagnosing CE in endemic areas.

Isolation, characterization, and pathogenicity assay of Acanthamoeba and its endosymbionts in respiratory disorders and COVID-19 hospitalized patients, northern Iran.

Acanthamoeba spp., are common free-living amoebae found in nature that can serve as reservoirs for certain microorganisms. The SARS-CoV-2 virus is a newly emerged respiratory infection, and the investigation of parasitic infections remains an area of limited research. Given that Acanthamoeba can act as a host for various endosymbiotic microbial pathogens and its pathogenicity assay is not fully understood, this study aimed to identify Acanthamoeba and its bacterial and fungal endosymbionts in patients with chronic respiratory disorders and hospitalized COVID-19 patients in northern Iran. Additionally, a pathogenicity assay was conducted on Acanthamoeba isolates. Urine, nasopharyngeal swab, and respiratory specimens were collected from two groups, and each sample was cultured on 1.5% non-nutrient agar medium. The cultures were then incubated at room temperature and monitored daily for a period of two weeks. Eight Acanthamoeba isolates were identified, and PCR was performed to confirm the presence of amoebae and identify their endosymbionts. Four isolates were found to have bacterial endosymbionts, including Stenotrophomonas maltophilia and Achromobacter sp., while two isolates harbored fungal endosymbionts, including an uncultured fungus and Gloeotinia sp. In the pathogenicity assay, five isolates exhibited a higher degree of pathogenicity compared to the other three. This study provides significant insights into the comorbidity of acanthamoebiasis and COVID-19 on a global scale, and presents the first evidence of Gloeotinia sp. as a fungal endosymbiont. Nevertheless, further research is required to fully comprehend the symbiotic patterns and establish effective treatment protocols.

Phylogenetic analysis and antimony resistance of Leishmania major isolated from humans and rodents.

Cutaneous Leishmaniasis (CL) is one of the world's neglected diseases which is caused by Leishmania spp. The aim of this study was to assess molecular profile and antimony resistance of Leishmania isolated from human and rodent hosts. Samples were collected from suspected CL patients referred to health centres and wild rodent's traps in Gonbad-e-Qabus region, north-eastern Iran. Smears were subjected to PCR-RFLP to identify Leishmania species. In addition, ITS1-PCR products were sequenced for phylogenetic analysis. Clinical isolates and rodent samples were subjected to MTT assay to determine IC50 values and in vitro susceptibilities. Expression levels of antimony resistance-related genes were determined in CL isolates. Out of 1,949 suspected patients with CL and 148 rodents, 1,704 (87.4%) and 6 (4.05%) were positive with direct smear, respectively. Digestion patterns of BusRI (HaeIII) endonuclease enzyme were similar to what expected for Leishmania major. Phylogenetic analysis revealed that the highest interspecies similarity was found between current L. major sequences with L. major obtained from Russia and Uzbekistan. Out of 20 L. major samples tested, 13 (65%) were resistant to meglumine antimoniate (MA) treatment, with an activity index (AI) exceeding 4. The remaining 7 samples (35%) responded to MA treatment and were classified as sensitive isolates, with a confirmed sensitive phenotype based on their AI values. The comparison expression analysis of three major antimony resistance-associated genes in unresponsive clinical isolates demonstrated significant fold changes for TDR1 (4.78-fold), AQP1 (1.3-fold), and γ-GCS (1.17-fold) genes (P < 0.05). Herein, we demonstrate genetic diversity and antimony resistance of L. major isolated from human and reservoir hosts in north-eastern Iran, which could be the basis for planning future control strategies.

Potential Association Between Latent Toxoplasma gondii Infection and Suicide Attempts: A Case-Control Registry-Based Study.

Toxoplasma gondii is a ubiquitous parasitic protozoan that may be an important cause of neurological and psychiatric diseases. The purpose of this case-control registry-based study was to evaluate the prevalence of T. gondii infection and related risk factors among subjects who attempted suicide by drug use and a control group at the Iranian National Registry Center for Toxoplasmosis in Mazandaran Province, northern Iran. Baseline data were collected from participants using a questionnaire, and a blood sample was taken from each individual. The plasma was prepared for serological analysis, whereas the buffy coat was used for molecular analysis. Out of 282 individuals (147 cases with suicide attempters [SA] and 135 controls), 42.9% of patients and 16.3% of control subjects were positive for anti-Toxoplasma immunoglobin G (IgG), but all participants were negative for T. gondii DNA and anti-Toxoplasma immunoglobin M. Based on multiple logistic regressions, IgG seropositivity in SA in the age group of 20-30 years was 3.22 times higher than that in the control group (p < 0.001). These findings suggest that latent T. gondii infection among SA is significantly higher than that in healthy individuals, indicating a potential association between latent toxoplasmosis and SA at least in the studied area. Further research is needed to shed light on the potential association between T. gondii and suicide among different populations and areas of the world.

Anti-leishmanial effects of resveratrol and resveratrol nanoemulsion on Leishmania major.

BACKGROUND: Leishmaniasis is a vector-borne disease that is endemic in the tropical and sub-tropical areas of the world. Low efficacy and high cytotoxicity of the current treatment regimens for leishmaniasis is one of the most important health problems. In this experimental study, anti-leishmanial effects of different concentrations of resveratrol and resveratrol nano-emulsion (RNE) were assessed. METHODS: RNE was prepared using the probe ultra-sonication method. The cytotoxicity was evaluated using the MTT technique on the L929 cell line. The anti-leishmanial activities on promastigotes of leishmania were assessed using vital staining and infected BALB/c mice were used to assess the in vivo anti-leishmanial effects. RESULTS: In vitro and in vivo assays revealed that all concentrations of resveratrol and RNE had valuable inhibitory effects against Leishmania major in comparison to the control group (P < 0.05). The half maximal inhibitory concentration (IC50) values were calculated as 16.23 and 35.71 µg/mL for resveratrol and RNE, respectively. Resveratrol and RNE showed no cytotoxicity against the L929 cell line. CONCLUSIONS: According to the potent in vitro and in vivo anti-leishmanial activity of RNE at low concentration against L. major, we suggest that it could be a promising anti-leishmanial therapeutic against L. major in the future.

Leishmania RNA virus 2 (LRV2) exacerbates dermal lesions caused by Leishmania major and comparatively unresponsive to meglumine antimoniate treatment.

PURPOSE: The present study investigated the possible role of Leishmania RNA virus 2 (LRV2) in the severity of dermal lesions and treatment failure due to Leishmania major. METHODS: The drug susceptibility of 14 clinical isolates of L.major, including resistant (n = 7) and sensitive (n = 7) isolates, was checked in the J774A.1 macrophage cell line. The presence of LRV2 among isolates was investigated by the RdRp gene and semi-nested PCR. Moreover, 1 × 106 sensitive L. major LRV2+ and LRV2- promastigotes were inoculated subcutaneously into the base tails of the 40 BALB/c mice divided into 4 groups (n = 10 in each group), including clinical LRV2+, clinical LRV2-, positive control LRV2+ and negative control LRV2-. The groups were infected with a unique isolate. The lesion size and parasite burden were evaluated. RESULTS: Sensitive and resistant isolates were determined by the drug susceptibility method. A higher presence of LRV2 was observed among MA-resistant isolates (6/7) compared with susceptible isolates (4/7), which was not statistically significant (P = 0.237). On the other hand, a comparison of the lesion sizes between the LRV2+ and LRV2- BALB/c mice groups revealed that the mean size of the lesion in the LRV2+ groups was significantly higher than the LRV2- (P = 0.034). In the same direction, there was an increased parasite burden in mice inoculated with LRV2+ groups compared with the LRV2- BALB/c mice groups (P = 0.002). CONCLUSIONS: Our findings showed that the presence of LRV2 could be one of the factors contributing to exacerbating CL. Although we found a higher presence of LRV2 in the resistant isolates, it seems that further investigations are recommended to determine the detailed association between lesions' aggravation and being comparatively unresponsive to treatment.

Molecular identification and genotyping of Acanthamoeba spp., in bronchoalveolar lavage fluid from immunocompetent patients with chronic respiratory disorders (CRD).

This study aimed to investigate the presence and genotyping of Acanthamoeba spp., in the bronchoalveolar lavage fluid (BALF) of immunocompetent patients with chronic respiratory disorders (CRD). In this study, 211 BALF samples were collected from patients with CRD during the COVID-19 pandemic who were candidates for fiberoptic bronchoscopy (FOB) at Imam Khomeini Hospital, Sari, Mazandaran Province, northern Iran and investigated for Acanthamoeba spp., by PCR. A total of 211 FBAL samples were examined; 5 (5/211; 2.36%) were positive by using the PCR test for Acanthamoeba spp. According to sequence analysis, three strains belonged to the T4 genotype and one strain to the T2 genotype. Our data demonstrate that the presence of Acanthamoeba (T4 and T2) in BALF specimens of patients with respiratory infections. However, it is important to note that these findings may be merely accidental. Our findings suggest further investigation to fully understand the role of Acanthamoeba spp. in the pathogenesis of lung infections.

Comparative evaluation of amniotic fluid as an alternative to fetal bovine serum in the maintenance of Leishmania major and Toxoplasma gondii.

This study aimed to evaluate the efficacy of amniotic fluid (AF) as an alternative to fetal bovine serum (FBS) in the maintenance of Leishmania major promastigotes and Toxoplasma gondii tachyzoites. AF was collected by an obstetrician using sterile syringes during a cesarean section. The culture medium was supplemented with 5 different concentrations of FBS or AF including 2, 5, 10, 20, and 30%. These concentrations were used to maintain both mentioned parasites. L. major was maintained at temperatures 4 and 24 °C and examined once a week for 4 weeks, while T. gondii was maintained at temperatures 4, 24, and 37 °C and examined at hours 24, 48, 72, and 96. For L. major, at both 4 and 24 °C, we observed no significant difference between FBS and AF on day 7. However, on days 14, 21, and 28, the difference between FBS and AF was significant at both temperatures. For T. gondii, no significant difference was observed between FBS and AF at hour 24 and all temperatures. However, this difference was significant at hours 48, 72, and 96 and all temperatures. According to our results, although FBS had a greater efficacy than AF in the growth of L. major and the survival of T. gondii, the number of promastigotes increased over time in AF-containing medium and the number of tachyzoites reduced slowly with a mild slop. Therefore, AF can be a potential alternative to FBS.