Prevalence and molecular characterization of Entamoeba moshkovskii in diarrheal patients from Eastern India

Background Importance of the amphizoic amoeba Entamoeba moshkovskii is increasing in the study of amoebiasis as a common human pathogen in some settings. Limited studies are found on the genetic and phylogenetic characterization of E. moshkovskii from India; hence remain largely unknown. In this study, we determined the prevalence and characterized the E. moshkovskii isolates in eastern India. Methods A three-year systemic surveillance study among a total of 6051 diarrhoeal patients from ID Hospital and BC Roy Hospital, Kolkata was conducted for E. moshkovskii detection via a nested PCR system targeting 18S rRNA locus. The outer primer set detected the genus Entamoeba and the inner primer pair identified the E. moshkovskii species. The 18S rRNA locus of the positive samples was sequenced. Genetic and phylogenetic structures were determined using DnaSP.v5 and MEGA-X. GraphPad Prism (v.8.4.2), CA, USA was used to analyze the statistical data. Result 4.84% (95%CI = 0.0433–0.0541) samples were positive for Entamoeba spp and 3.12% (95%CI = 0.027–0.036) were infected with E. moshkovskii. E. moshkovskii infection was significantly associated with age groups (X2 = 26.01, P<0.0001) but not with gender (Fisher’s exact test = 0.2548, P<0.05). A unique seasonal pattern was found for E. moshkovskii infection. Additionally, 46.56% (95%CI = 0.396–0.537) were sole E. moshkovskii infections and significantly associated with diarrheal incidence (X2 = 335.5,df = 9; P<0.0001). Sequencing revealed that the local E. moshkovskii strains were 99.59%-100% identical to the prototype (GenBank: KP722605.1). The study found certain SNPs that showed a correlation with clinical features, but it is not necessarily indicative of direct control over pathogenicity. However, SNPs in the 18S rRNA gene could impact the biology of the amoeba and serve as a useful phylogenetic marker for identifying pathogenic E. moshkovskii isolates. Neutrality tests of different coinfected subgroups indicated deviations from neutrality and implied population expansion after a bottleneck event or a selective sweep and/or purifying selection in co-infected subgroups. The majority of FST values of different coinfected subgroups were <0.25, indicating low to moderate genetic differentiation within the subgroups of this geographical area. Conclusion The findings reveal the epidemiological significance of E. moshkovskii infection in Eastern India as the first report in this geographical area and expose this species as a possible emerging enteric pathogen in India. Our findings provide useful knowledge for further research and the development of future control strategies against E. moshkovskii.

[1]  C. Happi,et al.  Genetic diversity and population structure of Plasmodium falciparum in Nigeria: insights from microsatellite loci analysis , 2021, Malaria journal.

[2]  T. Nozaki,et al.  Multilocus sequence typing (MLST) of Entamoeba histolytica identifies kerp2 as a genetic marker associated with disease outcomes. , 2021, Parasitology international.

[3]  L. Jaeger,et al.  Infections with Giardia duodenalis and Entamoeba histolytica/Entamoeba dispar as Hidden and Prevalent Conditions in Periurban Communities in the State of Rio de Janeiro, Brazil , 2020, Journal of tropical medicine.

[4]  R. Abozahra,et al.  Prevalence and Molecular Differentiation of Entamoeba histolytica, Entamoeba dispar, Entamoeba moshkovskii, and Entamoeba hartmanni in Egypt , 2020, Acta Parasitologica.

[5]  Xiaomei Deng,et al.  High Genetic Diversity and Low Differentiation in Michelia shiluensis, an Endangered Magnolia Species in South China , 2020, Forests.

[6]  T. Nozaki,et al.  Prevalence and distribution of Entamoeba species in a rural community in northern South Africa , 2020, Food and waterborne parasitology.

[7]  W. Bulimo,et al.  First report of Entamoeba moshkovskii in human stool samples from symptomatic and asymptomatic participants in Kenya , 2019, Tropical Diseases, Travel Medicine and Vaccines.

[8]  A. Schiller,et al.  Entamoeba Histolytica: Updates in Clinical Manifestation, Pathogenesis, and Vaccine Development , 2018, Canadian journal of gastroenterology & hepatology.

[9]  Sudhir Kumar,et al.  MEGA X: Molecular Evolutionary Genetics Analysis across Computing Platforms. , 2018, Molecular biology and evolution.

[10]  Longxian Zhang,et al.  Prevalence, molecular epidemiology, and zoonotic potential of Entamoeba spp. in nonhuman primates in China. , 2017, Infection, genetics and evolution : journal of molecular epidemiology and evolutionary genetics in infectious diseases.

[11]  H. Al-Mekhlafi,et al.  First molecular epidemiology of Entamoeba histolytica, E. dispar and E. moshkovskii infections in Yemen: different species‐specific associated risk factors , 2017, Tropical medicine & international health : TM & IH.

[12]  J. Saugar,et al.  Molecular prevalence of Entamoeba histolytica/dispar infection among patients attending four health centres in north-west Ethiopia , 2017, Tropical doctor.

[13]  Surinder Kumar,et al.  Prevalence of Entamoeba histolytica and Giardia lamblia infection in a Rural Area of Haryana, India , 2016 .

[14]  M. López,et al.  Identification of Entamoeba moshkovskii in Treated Waste Water Used for Agriculture , 2016, EcoHealth.

[15]  S. Ghosh,et al.  Molecular Epidemiology of Amoebiasis: A Cross-Sectional Study among North East Indian Population , 2015, PLoS neglected tropical diseases.

[16]  Hui Liu,et al.  Genetic diversity of Trichomonas vaginalis clinical isolates from Henan province in central China , 2015, Pathogens and global health.

[17]  J. M. Peralta,et al.  Differential Diagnosis of Entamoeba spp. in Clinical Stool Samples Using SYBR Green Real-Time Polymerase Chain Reaction , 2014, TheScientificWorldJournal.

[18]  Anthony K. Waruru,et al.  Intestinal parasitic infections in children presenting with diarrhoea in outpatient and inpatient settings in an informal settlement of Nairobi, Kenya , 2013, BMC Infectious Diseases.

[19]  R. Mahmud,et al.  Differentiating Entamoeba histolytica, Entamoeba dispar and Entamoeba moshkovskii using nested polymerase chain reaction (PCR) in rural communities in Malaysia , 2012, Parasites & Vectors.

[20]  T. Anuar,et al.  First molecular identification of Entamoeba moshkovskii in Malaysia , 2012, Parasitology.

[21]  R. Haque,et al.  Entamoeba bangladeshi nov. sp., Bangladesh , 2012, Emerging infectious diseases.

[22]  M. López,et al.  Entamoeba moshkovskii perspectives of a new agent to be considered in the diagnosis of amebiasis. , 2012, Acta tropica.

[23]  Shihab U. Sobuz,et al.  Entamoeba moshkovskii is associated with diarrhea in infants and causes diarrhea and colitis in mice. , 2012, The Journal of infectious diseases.

[24]  M. Norhayati,et al.  Public health and clinical importance of amoebiasis in Malaysia: a review. , 2011, Tropical biomedicine.

[25]  C. Clark,et al.  Increased sampling reveals novel lineages of Entamoeba: consequences of genetic diversity and host specificity for taxonomy and molecular detection. , 2011, Protist.

[26]  T. Nozaki,et al.  Trend of Entamoeba histolytica infestation in Kolkata , 2010, Gut Pathogens.

[27]  K. Rajendran,et al.  Hospital-based surveillance of enteric parasites in Kolkata , 2009, BMC Research Notes.

[28]  S. Rasti,et al.  Entamoeba dispar: genetic diversity of Iranian isolates based on serine-rich Entamoeba dispar protein gene. , 2008, Pakistan journal of biological sciences : PJBS.

[29]  A. Bouratbine,et al.  First molecular identification of Entamoeba moshkovskii in human stool samples in Tunisia. , 2008, The American journal of tropical medicine and hygiene.

[30]  G. Aslan,et al.  Detection of Entamoeba histolytica Antigen in Stool Samples in Mersin, Turkey , 2008, The Journal of parasitology.

[31]  D. Marriott,et al.  Laboratory Diagnostic Techniques for Entamoeba Species , 2007, Clinical Microbiology Reviews.

[32]  N. Beebe,et al.  PCR Detection of Entamoeba histolytica, Entamoeba dispar, and Entamoeba moshkovskii in Stool Samples from Sydney, Australia , 2007, Journal of Clinical Microbiology.

[33]  P. Chavalitshewinkoon-Petmitr,et al.  Differential Detection of Entamoeba histolytica, Entamoeba dispar, and Entamoeba moshkovskii by a Single-Round PCR Assay , 2006, Journal of Clinical Microbiology.

[34]  Krishna Khairnar,et al.  Entamoeba moshkovskii and Entamoeba dispar-associated infections in pondicherry, India. , 2005, Journal of health, population, and nutrition.

[35]  D. Schellenberg,et al.  Etiology of diarrhea in children less than five years of age in Ifakara, Tanzania. , 2004, The American journal of tropical medicine and hygiene.

[36]  W. Petri,et al.  Laboratory Diagnosis of Amebiasis , 2003, Clinical Microbiology Reviews.

[37]  T. Nozaki,et al.  Geographic Diversity among Genotypes of Entamoeba histolytica Field Isolates , 2003, Journal of Clinical Microbiology.

[38]  R. Haque,et al.  Entamoeba moshkovskii Infections in Children in Bangladesh , 2003, Emerging infectious diseases.

[39]  A. Martínez-Palomo,et al.  Pathogenesis of Intestinal Amebiasis: From Molecules to Disease , 2000, Clinical Microbiology Reviews.

[40]  G. Cook Amoebiasis and Giardiasis , 1994 .

[41]  J. Neigel,et al.  Application of a random walk model to geographic distributions of animal mitochondrial DNA variation. , 1993, Genetics.

[42]  M. Nei,et al.  Estimation of the number of nucleotide substitutions in the control region of mitochondrial DNA in humans and chimpanzees. , 1993, Molecular biology and evolution.

[43]  D. Bruckner Amebiasis , 1992, Clinical Microbiology Reviews.

[44]  C. Clark,et al.  The Laredo strain and other 'Entamoeba histolytica-like' amoebae are Entamoeba moshkovskii. , 1991, Molecular and biochemical parasitology.

[45]  J. Felsenstein CONFIDENCE LIMITS ON PHYLOGENIES: AN APPROACH USING THE BOOTSTRAP , 1985, Evolution; international journal of organic evolution.

[46]  L. S. Diamond,et al.  Entamoeba moshkovskii: axenic cultivation. , 1970, Experimental parasitology.

[47]  R. Burrows Morphological differentiation of Entamoeba hartmanni and E. polecki from E. histolytica. , 1959, The American journal of tropical medicine and hygiene.

[48]  Ibne Karim M Ali Intestinal amebae. , 2015, Clinics in laboratory medicine.

[49]  E. Houpt,et al.  High prevalence of Entamoeba moshkovskii in a Tanzanian HIV population. , 2008, Acta tropica.

[50]  M. Scaglia,et al.  Entamoeba moshkovskii (Tshalaia, 1941): morpho-biological characterization of new strains isolated from the environment, and a review of the literature. , 1983, Annales de parasitologie humaine et comparee.

[51]  D. Dreyer Growth of a strain of entamoeba histolytica at room temperature. , 1961, Texas reports on biology and medicine.

[52]  Gov,et al.  Bmc Microbiology the Capsule Polysaccharide Structure and Biogenesis for Non-o1 Vibrio Cholerae Nrt36s: Genes Are Embedded in the Lps Region , 2022 .