Recent Publications by Panagiotis Karanis, Professor of Anatomy

Javanmard E, Rahimi HM, Niyyati M, Aghdaei HA, Sharifdini M, Mirjalali H, Zali MR, Karanis P. Molecular analysis of Blastocystis sp. and its subtypes from treated wastewater routinely used for irrigation of vegetable farmlands in Iran. Journal of Water and Health, 2019 Oct; 17 (5): 837-844. doi: 10.2166/wh.2019.045.

Abstract

Treated wastewater samples were collected, filtered using sterile 47-mm cellulose nitrate membrane and DNA extracted from the filtered materials. The presence of Blastocystis sp. was confirmed via polymerase chain reaction (PCR) targeting the SSU rRNA gene of Blastocystis sp. in 5/12 of samples. Based on the subtype analysis after sequencing, 2, 2 and 1 of ST2, ST6 and ST8 were detected among the isolates, respectively. Furthermore, both ST6s were allele 139, alleles 11 and 138 were identified in ST2 and the only ST8 was allele 95. The phylogenetic tree showed that one of ST2 was clustered together with those ST2 that were already reported from humans and animals. The presence of Blastocystis sp. in treated wastewater can indicate the potential role of this type of water for irrigation in the transmission of pathogenic microorganisms to downstream farmlands.

Lass A, Ma L, Kontogeorgos I, Xueyong Z, Li X, Karanis P. Contamination of wastewater with Echinococcus multilocularis – possible implications for drinking water resources in the QTP China. Water Research 170: 115334.

Abstract

Echinococcus multilocularis is a parasite that causes a dangerous zoonosis, alveolar echinococcosis (AE). Its presence in water sources, however, has scarcely been studied heretofore. Accordingly, 222 samples of different origin including wastewater from wastewater treatment plants (WWTPs) (n = 137), slaughterhouse (n = 49) as well as water from rivers (n = 26) and a cattle farm (n = 10) were collected from Xining City and a rural area in Qinghai-Tibet Plateau (QTP), an endemic area. Material obtained after processing of 10 L volume samples was subsequently analysed using three molecular detection methods: nested PCR, real-time PCR and LAMP. E. multilocularis DNA was found in 13 (5.85%) water samples; including 8 (5.8%), 3 (6%), 2 (20%) and 0 positive samples found in WWTPs, a slaughterhouse, a cattle farm and rivers, respectively. All three (LAMP, PCR, RT-PCR) molecular tools displayed high agreement and effectiveness in their ability of detecting the parasite’s DNA in environmental material. This is the first investigation describing E. multilocularis detection in wastewater samples, using three sensitive molecular diagnostic tools. Results indicate the role of wastewater in dissemination of E. multilocularis and the risk of contamination of water sources.

Masangkay FR, Milanez GD, Tsiami A, Somsak V, Kotepui M, Tangpong J, Karanis P. First report of Cryptosporidium hominis in a freshwater sponge. Sci Total Environ. 700:134447. doi: 10.1016/j.scitotenv.2019.134447. Epub 2019 Oct 4.

 Abstract

Identification of Cryptosporidium oocyst is essential in ensuring water quality fit for human use, consumption, and recreation. This communication proposes the supplemental analysis of substrate-associated biofilms, in particular, freshwater sponges in improving case finding of waterborne-protozoan pathogens (WBPP) in environmental aquatic samples. In this study, a small portion of a mature freshwater sponge under the Genus Spongilla was subjected to microscopic and molecular analysis to identify the presence of Cryptosporidium. Microscopic screening with modified Kinyoun’s staining (MK) and microscopic confirmation using direct antibody fluorescent testing (IFT) returned with Cryptosporidium spp. positive findings. Molecular investigation resulted in the confirmation of Cryptosporidium hominis upon sequencing of PCR products and phylogenetic analysis. This is the first report of a pathogenic protozoan, C. hominis isolated from a freshwater sponge. The results of this study provide evidence of the value of expanding water quality assessment strategies to the analysis of substrate-associated biofilms and sponges in improving case finding of WBPP in natural aquatic environments.

Masangkay Frederick, Milanez Giovanni, Tsiami Amalia,Hapan FreidaSomsak, VoravuthKotepui, Manas Kotepui, Tangpong Jitbanjong and Karanis, Panagiotis. Waterborne protozoan pathogens in environmental aquatic biofilms: implications for water quality assessment strategies. Environmental Pollution, 259. ISSN 0269-7491.

 Abstract

Biofilms containing pathogenic organisms from the water supply are a potential source of protozoan parasite outbreaks and a general public health concern. The aim of the present study was to demonstrate the simultaneous and multispatial occurrence of waterborne protozoan pathogens (WBPP) in substrate-associated biofilms (SAB) and compare it to surface water (SW) and sediments with bottom water (BW) counterparts using manual filtration and elution from low-volume samples. For scenario purposes, simulated environmental biofilm contamination was created from in-situ grown one-month-old SAB (OM-SAB) that were spiked with Cryptosporidium parvum oocysts. Samples were collected from the largest freshwater reservoirs in Luzon, Philippines and a University Lake in Thailand. A total of 69 samples (23 SAB, 23 SW, and 23 BW) were evaluated using traditional staining techniques for Cryptosporidium, and immunofluorescence staining for the simultaneous detection of Cryptosporidium and Giardia. In the present study, WBPP was found in 43% SAB, 39% SW, and 39% BW samples tested with SAB results reflecting SW and BW results. Further, the potential and advantages of using low-volume sampling for the detection of parasite (oo)cysts in aquatic matrices were also demonstrated. Scanning electron microscopy of OM-SAB revealed a naturally-associated testate amoeba shell, while Cryptosporidium oocysts spiked samples provided a visual profile of what can be expected from naturally contaminated biofilms. This study provides the first evidence for the simultaneous and multi-spatial occurrence of waterborne protozoan pathogens in low-volume environmental aquatic matrices and warrants SAB testing along with SW and BW matrices for improved water quality assessment strategies (iWQAS).

Link:https://www.sciencedirect.com/science/article/pii/S0269749119368915?via%3Dihub

R. Mahmoudi, F. Mahdavi, K. Ashrafi, K. Forghanparast, B. Rahmati, A. Mirzaei, Z. Atrkar Roshan and P. Karanis  Report of Giardia assemblages and giardiasis in residents of Guilan province – Iran. Parasitology Research, 21 Jan 2020.

Abstract

Giardia duodenalis is considered a highly diverse organism that infects a variety of mammalian hosts. Giardiasis is a significant public health problem in Iran. The purpose of this study was to investigate the occurrence of Giardia duodenalis (G. lamblia, G. intestinalis) infections in humans residing in the Guilan province of Iran. Stool samples were collected during 12 months from 8356 individuals that had been referred to certain hospitals in the capital city of Rasht in the Guilan province, of which 4126 were males and 4230 were females. The samples were separated into three groups according to patient age: group A 1–9 years old (n = 483); group B 10–19 years old (n = 491); and group C greater than 20 years old (n = 7382). The wet mount technique was performed directly on 8356 fecal samples for microscopy. Samples were examined using a saline and iodine direct smear technique in order to confirm the presence of G. duodenalis. The results indicated that 2.5% (206/8356) of the samples were identified as positive for G. duodenalis. A total of 30% of the infected patients (n = 62) had no symptoms. In symptomatic cases, the most common symptoms (46%, n = 95) were abdominal cramps and bloating. Twenty-four percent of patients (n = 50) had cramps, bloating, nausea, and diarrhea. Sixty positive samples were sent for G. duodenalis genotyping based on the amplification of the gdh gene. Forty-one PCR products were successfully selected and sequenced, where 38 (92.6%) samples were identified as genotype A/subgenotype II and in three samples (7.4%) genotype B/subgenotype IV. Genotype A-II had a dominant prevalence as compared to the genotype B-IV samples that were identified in the study. Based on the samples provided by the regional teaching hospitals and subsequent sample analysis, the authors concluded that assemblage A-II is most likely the most common Giardia subgroup infection in the Guilan region. Assemblages have been reported in both humans and animals; however, further studies need to investigate the role of domestic animals and water reservoirs as potential sources of Giardia infection in the Guilan region.

Link: https://link.springer.com/article/10.1007%2Fs00436-019-06595-1

Ilknur Koyuna, Zeynep Kolorena, Ulku Karaman, Amalia Tsiami, Panagiotis Karanis. Acanthamoeba in river water samples of Black Sea, Turkey. Journal of Water and Health (in press, 18 November 2020).

Abstract

The present study aims to investigate the occurrence of free-living amoeba (FLA) in water resources (rivers and tap water) in Samsun in Black Sea. The presence of Acanthamoeba spp. was confirmed in 98 of 192 water samples collected from 32 sites of Samsun province (Samsun center, Terme, Carsamba, Tekkekoy, Bafra) by PCR. Acanthamoeba spp. was found in 15/36 river samples from Samsun, in 58/90 from Terme, in 12/30 from Carsamba, in 7/18 from Tekkekoy, and in 6/18 from Bafra. No Acanthamoeba species were detected in tap water samples. The highest rate in river waters contaminated with Acanthamoeba species was in Terme followed by Samsun center (41.7%), Carsamba (40%), Tekkekoy (38.9%) and Bafra districts (33.3%), respectively. The result of the subsequent sequence analysis showed Haplotype I (A. triangularis) was 5%, Haplotype II (A. polyphaga) 29.6%, Haplotype III (Acanthamoeba spp.) in 62%, and Haplotype IV (A. lenticulata) in 3%. The most common genotype was Acanthamoeba T4 (Acanthamoeba spp., A. polyphaga, A. triangularis) and T5 genotype was also found in 3%. The T4 genotype is the most common genotype worldwide associated with Acanthamoeba keratitis (AK); therefore humans and animals living in the area area risk after contact with such waters.