Detection by PCR of pathogenic protozoa in raw and drinkable water samples in Colombia

Triviño-Valencia, Jessica; Lora, Fabiana; Zuluaga, Juan David; Gómez-Marín, Jorge Enrique

doi:10.1007/s00436-016-4917-5

Cited by 44 publications

(46 citation statements)

References 24 publications

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“…Sequencing of PCR results was also important to discriminate between species for Cryptosporidium and in the present work it was possible to identify C. hominis in stools from children and to detect C. parvum in water and surface. We have reported that C. parvum is detected in tap water more frequently than C. hominis in our city (Triviño‐Valencia et al, ).…”

Section: Discussionmentioning

confidence: 60%

“…For food, live, and surface samples, the DNeasy Blood & Tissue Extraction Kit (Qiagen, Germany) with mechanical lysis and zirconium beads were applied five times for DNA extraction after agitation in a stomacher 400‐W BagMixer (Interscience, France) at 260 rpm for 30 s with glycine 1 M (Biorad, Estados Unidos) and pH of 5.5 and a wash solution (PBS 1X, 100 μl of Tween 80, Merck, Germany, and 10 g of sulfamic acid—Merck, Germany) at pH of 7.5, respectively (Cook et al, ), and a formalin–ether concentration method for water eluates was applied (also for inert and live surface samples), as described (Hernández‐Arango, Pinto, Muñoz‐Sanchez, Lora‐Suarez, & Gómez‐Marín, ; Lora‐Suarez, Rivera, Triviño‐Valencia, & Gomez‐Marin, ; Triviño‐Valencia, Lora, Zuluaga, & Gomez‐Marin, ). For stools, 1 g of sample was diluted in 5 ml of sterile saline solution at 0.9% and filtered through gauze.…”

Section: Methodsmentioning

confidence: 99%

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Food protozoa safety assessment and risk in school restaurants in Armenia, Colombia

Muñoz‐Sánchez

Hernández-Arango

Buitrago‐Lopez

et al. 2019

Journal of Food Safety

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This work assessed the risk of protozoa in 10 school restaurants in Armenia (Quindío, Colombia) by analyzing the presence of Cryptosporidium spp, Giardia duodenalis, Blastocystis, and Cyclospora cayetanensis DNA in the food, water, and living and inert surfaces of school restaurants and in stools of children who ate at these restaurants. Of the 213 food, water, and surface samples, 6.6% were positive using PCR to test DNA for Blastocystis; 3.8% for Cryptosporidium spp; 0.9% for G. duodenalis; and 0% for C. cayetanensis. In 187 stool samples analyzed via microscopy from children who attended the restaurants, 40 (21.4%) were positive for Blastocystis and 21 (11.2%) were positive for Giardia spp. Via PCR, 20 (10.7%) were positive for Cryptosporidium and 0 (0%) for C. cayetanensis. A higher positivity in children's stools for Blastocystis spp was correlated with lower compliance in property conditions and for higher positivity of Giardia spp in children's stool was related to lower knowledge by food manipulators. Inspection scores can identify restaurants with higher risk for protozoa infection.

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Section: Discussionmentioning

confidence: 60%

Section: Methodsmentioning

confidence: 99%

Food protozoa safety assessment and risk in school restaurants in Armenia, Colombia

Muñoz‐Sánchez

Hernández-Arango

Buitrago‐Lopez

et al. 2019

Journal of Food Safety

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“…The supernatant was discarded and 600 lL of DNAzol (Invitrogen, USA), 10 lL of isoamyl alcohol (Fisher Scientific, USA) and 0.3 g of zirconium silicate beads with 0.5 mm diameter (BioSpec, USA) were added. Afterwards, the tubes were shaken five times in a Mini Bead Beater (BioSpec, USA) to maximum speed for 1 minute and 1 minute in ice [41]. Later, a Wizard Genomic DNA extraction kit (Promega, USA) was used for nuclear lysis and purification, following the manufacturer's protocol.…”

Section: Toxoplasma Gondii Dna Extraction Procedures and Pcr Detectionmentioning

confidence: 99%

Detection and genotypes of Toxoplasma gondii DNA in feces of domestic cats in Colombia

et al. 2020

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The high prevalence of Toxoplasma gondii in the human population in Colombia has been linked to the existence of a high density of urban stray cats, exposing the whole population to a high density of oocysts. The goal of this study was to determine the DNA prevalence of T. gondii by conventional PCR and to phylogenetically analyze ROP18 sequences from positive samples in domestic cat (Felis catus) fecal samples in the city of Armenia, Quindío. Fecal samples from 140 cats were collected from 10 districts around the city. Samples were concentrated using Ritchie’s method and analyzed through optical microscopy. Concentrates were used for DNA extraction followed by nested PCR amplification for T. gondii gene B1. PCR for ROP18 was performed on all B1 positive samples; the ROP18 sequences obtained were related to the Archetype I Brazilian and Chinese strains. No oocysts were detected by optical microscopy; however, 17.8% (25/140) B1 and 24% (6/25) ROP18 PCR-positive samples were detected. Phylogenetic analyses showed that isolates clustered into a single group. We assessed whether associations existed between T. gondii positive fecal samples and survey variables such as cat healthcare and socioeconomic characteristics of owners, but no statistically significant associations were found. The presence of T. gondii in cat feces is an important factor contributing to the high prevalence in the human population of this city.

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“…To date, this parasitic occurrence has been reported in different geographical location of river [5]. In fact, 11.5% of Malaysian river water samples were reported to get contaminated with Cryptosporidium oocysts [6]. Recently, two cases have been reported on Cryptosporidium occurrence in Kuantan, Pahang [7,8].…”

Section: Introductionmentioning

confidence: 99%

CHEMICAL COMPONENTS OF POLYMERASE CHAIN REACTION IN 18S rRNA FOR DETECTION OF Cryptosporidium FROM RIVER WATER SAMPLES

2019

MJAS

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The gene of 18S ribosomal RNA or 18S rRNA is the universal gene function as a general genetic marker for species identification of microorganisms including parasites. Cryptosporidium has distinct 18S rRNA genes along different species within the same genus. In this study, polymerase chain reaction or PCR was used to study chemical components of PCR setup in amplification of 18S rRNA gene of this parasite. Cryptosporidium was collected from river water samples and its presence was confirmed using specific immunofluorescence detection of this parasite. Isolated water containing Cryptosporidium was then subjected to genomic DNA extraction before PCR step. The chemical components of PCR consisting of MgCI 2 , deoxynucleotide triphosphate (DNTPs), Polymerases, free DNase-water, universal primers and PCR buffer were studied in different volume and concentration. Each chemical component of PCR was optimized differently in yielding the same final volume of 20 µL per each reaction. The value range of chemical components of PCR consisted of MgCI 2 (0.1 µM-0.5 µM), dNTPs (50-250 mM), free DNase water (5-10 µL), polymerases (0.2-0.5 U) and universal primers (2-20 µM). The result indicated that 0.2 µM of MgCI 2 , 100 mM of dNTPs, less than 10 µL of free DNase water, 0.5 U of polymerases and 10 mM of universal primers were the best combination to get better result for molecular identification of 18S rRNA Cryptosporidium. As a conclusion, accurate and proper concentration or volume of each PCR chemical components is essential for molecular identification of 18S rRNA Cryptosporidium gene. In future studies, study on gradient temperature parameters of PCR run can be included to study the chemical nature of amplified genes either in denaturation, annealing or extension steps. AbstrakGen 18S ribosomal RNA adalah gen universal yang berfungsi sebagai penanda genetik umum untuk pengenalpastian spesies mikroorganisma termasuk parasit. Cryptosporidium mempunyai gen 18S rRNA yang berlainan daripada spesies berbeza dalam genus yang sama. Dalam kajian ini, tindak balas berantai polymerase atau PCR digunakan untuk mengkaji komponen kimia susun atur PCR dalam amplifikasi gen 18S rRNA bagi parasit ini. Cryptosporidium telah diambil dari sampel air sungai dan disahkan kehadirannya menggunakan pengesanan immunopendaflour terhadap parasit ini. Sampel air yang diambil mengandungi Cryptosporidium yang kemudiannya diteruskan untuk pengekstrakan genomik DNA sebelum peringkat PCR. Komponen kimia PCR yang terdiri daripada MgCI 2 , deoksinukleotida trifosfat (dNTPs), polimerase, air yang bebas DNase, primer umum dan larutan penimbal PCR telah dikaji dalam isipadu dan kepekatan yang berbeza. Setiap komponen kimia PCR dioptimakan secara ISSN 1394 -2506 Mohd Aiman et al: CHEMICAL COMPONENTS OF POLYMERASE CHAIN REACTION IN 18S rRNA FOR DETECTION OF Cryptosporidium FROM RIVER WATER SAMPLES 402 berbeza dalam menghasilkan isipadu akhir 20 uL bagi setiap tindak balas. Nilai julat komponen kimia PCR terdiri daripada MgCI 2 (0.1 µM-0.5 µM), dNTPs (50-250 mM), ai...

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Detection by PCR of pathogenic protozoa in raw and drinkable water samples in Colombia

Cited by 44 publications

References 24 publications

Food protozoa safety assessment and risk in school restaurants in Armenia, Colombia

Food protozoa safety assessment and risk in school restaurants in Armenia, Colombia

Detection and genotypes of Toxoplasma gondii DNA in feces of domestic cats in Colombia

CHEMICAL COMPONENTS OF POLYMERASE CHAIN REACTION IN 18S rRNA FOR DETECTION OF Cryptosporidium FROM RIVER WATER SAMPLES

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