Genetic diversity of Cryptosporidium isolates from patients in North India

SummaryBackgroundCryptosporidiosis is a significant cause of diarrheal illness in both immunocompetent and immunocompromised populations. Cryptosporidium species infect a wide range of hosts including humans. Different species are morphologically indistinguishable, and molecular techniques have become the key to detection and source tracking. The present study was designed to study the genetic diversity of human Cryptosporidium isolates in North India.MethodsCryptosporidium oocysts were detected in stool samples by special staining of fecal smears. DNA was extracted with a Qiagen kit and all samples were genotyped by small subunit ribosomal ribonucleic acid (SSU rRNA)-based nested PCR-restriction fragment length polymorphism (RFLP) tool using enzymes SspI and VspI. Cryptosporidium hominis and Cryptosporidium parvum isolates were subtyped by sequence analysis of the nested PCR amplified gp60 gene.ResultsFifty-three fecal samples were found to be positive for Cryptosporidium oocysts. RFLP analysis revealed 39 isolates as C. hominis and 13 isolates of C. parvum; one sample failed amplification. gp60-based sequencing of C. hominis and C. parvum divided them into eight subgenotype families and 17 subtypes. gp60-based sequencing identified seven cases of mixed infection with C. hominis and C. parvum/Cryptosporidium meleagridis and showed the presence of C. meleagridis in six HIV-positive patients that were indistinguishable in RFLP.ConclusionsCryptosporidium isolates obtained in the present study from patients in North India belonged to three species, eight subgenotype families, and 17 subtypes. The existence of many Cryptosporidium species, subgenotypes, and subtypes along with mixed infections reveals the complexity of Cryptosporidium transmission; this heterogeneity indicates stable cryptosporidiosis transmission in North India. The results may have further implications in understanding the epidemiology and control of this infection

Serum immunoglobulin G, M and A response to Cryptosporidium parvum in Cryptosporidium-HIV co-infected patients

<p>Abstract</p> <p>Background</p> <p><it>Cryptosporidium parvum</it>, the protozoan parasite, causes a significant enteric disease in immunocompromised hosts such as HIV patients. The present study was aimed to compare serum IgG, IgM and IgA responses to crude soluble antigen of <it>C. parvum </it>in HIV seropositive and seronegative patients co-infected with <it>Cryptosporidium </it>and to correlate the responses with symptomatology.</p> <p>Methods</p> <p><it>Cryptosporidium parvum </it>specific serum antibody (IgG, IgM and IgA) responses were assessed by ELISA in 11 HIV seropositive <it>Cryptosporidium </it>positive (Group I), 20 HIV seropositive <it>Cryptosporidium </it>negative (Group II), 10 HIV seronegative <it>Cryptosporidium </it>positive (Group III), 20 HIV seronegative <it>Cryptosporidium </it>negative healthy individuals (Group IV) and 25 patients with other parasitic diseases (Group V).</p> <p>Results</p> <p>A positive IgG and IgA antibody response was observed in significantly higher number of <it>Cryptosporidium </it>infected individuals (Gp I and III) compared to <it>Cryptosporidium </it>un-infected individuals (Gp II, IV and V) irrespective of HIV/immune status. Sensitivity of IgG ELISA in our study was found to be higher as compared to IgM and IgA ELISA. The number of patients with positive IgG, IgM and IgA response was not significantly different in HIV seropositive <it>Cryptosporidium </it>positive patients with diarrhoea when compared to patients without diarrhoea and in patients with CD4 counts <200 when compared to patients with CD4 counts >200 cells/μl.</p> <p>Conclusion</p> <p>The study showed specific serum IgG and IgA production in patients infected with <it>Cryptosporidium</it>, both HIV seropositive and seronegative as compared to uninfected subjects suggesting induction of <it>Cryptosporidium </it>specific humoral immune response in infected subjects. However, there was no difference in number of patients with positive response in HIV seropositive or seronegative groups indicating that HIV status may not be playing significant role in modulation of <it>Cryptosporidium </it>specific antibody responses. The number of patients with positive IgG, IgM and IgA response was not significantly different in patients with or without history of diarrhoea thereby indicating that <it>Cryptosporidium </it>specific antibody responses may not be necessarily associated with protection from symptomatology.</p

An Insight into the Genome of Pathogenic and Non-Pathogenic Acanthamoeba

Background: Acanthamoeba are amphizoic amoeba majorly responsible for causing Acanthamoeba keratitis (AK) and Granulomatous amoebic encephalitis (GAE). Despite its ubiquitous nature, the frequency of infections is not high, probably due to the existence of non-pathogenic isolates. The whole-genome sequencing and an annotated genome assembly can unravel the biological functions and help in identifying probable genes related to pathogenicity. Methods: Illumina and Nanopore sequencing were performed for keratitis, encephalitis, and non-pathogenic environmental isolates. Hybrid assembly was prepared for the AK and GAE isolates, while only the Illumina reads were utilized for a non-pathogenic environmental isolate. Protein coding genes were identified using the GeneMark-ES program and BLASTx module of Diamond used for gene prediction. Additionally, the Kyoto Encyclopedia of Genes and Genomes annotation and cluster of orthologous group&rsquo;s annotation using RPS-blast against the CDD database was performed. The subsequent data analysis and validation will help identify probable pathogenic genes. Results: The genome assemblies of 9.67, 8.34, and 8.89 GBs were reported for GAE, AK, and non-pathogenic isolate, respectively. KEGG reported 22,946 in GAE, 24,231 in keratitis, and 9367 genes in the environmental isolate. The COG annotation revealed 3232 in GAE, 3403 in keratitis, and 1314 genes in the non-pathogenic isolate. Conclusion: The present study has attempted to generate de novo hybrid genome assemblies of Acanthamoeba that would help decode the genome of free-living amoeba and will provide genomic data for a better understanding of virulence-related factors

Dermatoscopy in diagnosis of cutaneous myiasis arising in pemphigus vulgaris lesions

Myiasis, infestation of live human and vertebrate animals by larvae, can complicate ulcers and open wounds. Although myiasis occurs in neglected erosions of pemphigus, such a complication is not documented in the literature. Herein, we report a case of myiasis complicating pemphigus vulgaris and describe its dermatoscopic features