scholarly journals Suitability of current typing procedures to identify epidemiologically linked human Giardia duodenalis isolates

2021 ◽  
Vol 15 (3) ◽  
pp. e0009277
Author(s):  
Andreas Woschke ◽  
Mirko Faber ◽  
Klaus Stark ◽  
Martha Holtfreter ◽  
Frank Mockenhaupt ◽  
...  
Keyword(s):  
Primary Care ◽  
High Frequency ◽  
Giardia Duodenalis ◽  
Multiple Infections ◽  
Mixed Infections ◽  
Single Strain ◽  
Allelic Sequence ◽  
Mlst Type ◽  
Result Interpretation ◽  
Assemblage B

Background Giardia duodenalis is a leading cause of gastroenteritis worldwide. Humans are mainly infected by two different subtypes, i.e., assemblage A and B. Genotyping is hampered by allelic sequence heterozygosity (ASH) mainly in assemblage B, and by occurrence of mixed infections. Here we assessed the suitability of current genotyping protocols of G. duodenalis for epidemiological applications such as molecular tracing of transmission chains. Methodology/Principal findings Two G. duodenalis isolate collections, from an outpatient tropical medicine clinic and from several primary care laboratories, were characterized by assemblage-specific qPCR (TIF, CATH gene loci) and a common multi locus sequence typing (MLST; TPI, BG, GDH gene loci). Assemblage A isolates were further typed at additional loci (HCMP22547, CID1, RHP26, HCMP6372, DIS3, NEK15411). Of 175/202 (86.6%) patients the G. duodenalis assemblage could be identified: Assemblages A 25/175 (14.3%), B 115/175 (65.7%) and A+B mixed 35/175 (20.0%). By incorporating allelic sequence heterozygosity in the analysis, the three marker MLST correctly identified 6/ 9 (66,7%) and 4/5 (80.0%) consecutive samples from chronic assemblage B infections in the two collections, respectively, and identified a cluster of five independent patients carrying assemblage B parasites of identical MLST type. Extended MLST for assemblage A altogether identified 5/6 (83,3%) consecutive samples from chronic assemblage A infections and 15 novel genotypes. Based on the observed A+B mixed infections it is estimated that only 75% and 50% of assemblage A or B only cases represent single strain infections, respectively. We demonstrate that typing results are consistent with this prediction. Conclusions/Significance Typing of assemblage A and B isolates with resolution for epidemiological applications is possible but requires separate genotyping protocols. The high frequency of multiple infections and their impact on typing results are findings with immediate consequences for result interpretation in this field.

scholarly journals Cryptosporidium in fish: alternative sequencing approaches and analyses at multiple loci to resolve mixed infections

Parasitology ◽  
2017 ◽  
Vol 144 (13) ◽  
pp. 1811-1820 ◽  
Author(s):  
ANDREA PAPARINI ◽  
RONGCHANG YANG ◽  
LINDA CHEN ◽  
KAISING TONG ◽  
SUSAN GIBSON-KUEH ◽  
...  
Keyword(s):  
Genetic Diversity ◽  
High Frequency ◽  
Sanger Sequencing ◽  
Multiple Infections ◽  
Mixed Infections ◽  
Multiple Loci ◽  
The Rich ◽  
Next Generation Sequencing Ngs ◽  
Molecular Parasitology ◽  
Generation Sequencing

SUMMARYCurrently, the systematics, biology and epidemiology of piscine Cryptosporidium species are poorly understood. Here, we compared Sanger ‒ and next-generation ‒ sequencing (NGS), of piscine Cryptosporidium, at the 18S rRNA and actin genes. The hosts comprised 11 ornamental fish species, spanning four orders and eight families. The objectives were: to (i) confirm the rich genetic diversity of the parasite and the high frequency of mixed infections; and (ii) explore the potential of NGS in the presence of complex genetic mixtures. By Sanger sequencing, four main genotypes were obtained at the actin locus, while for the 18S locus, seven genotypes were identified. At both loci, NGS revealed frequent mixed infections, consisting of one highly dominant variant plus substantially rarer genotypes. Both sequencing methods detected novel Cryptosporidium genotypes at both loci, including a novel and highly abundant actin genotype that was identified by both Sanger sequencing and NGS. Importantly, this genotype accounted for 68·9% of all NGS reads from all samples (249 585/362 372). The present study confirms that aquarium fish can harbour a large and unexplored Cryptosporidium genetic diversity. Although commonly used in molecular parasitology studies, nested PCR prevents quantitative comparisons and thwarts the advantages of NGS, when this latter approach is used to investigate multiple infections.

Molecular typing ofGiardia duodenalisin humans in Queensland – first report of Assemblage E

Parasitology ◽  
2017 ◽  
Vol 144 (9) ◽  
pp. 1154-1161 ◽  
Author(s):  
ALIREZA ZAHEDI ◽  
DANIEL FIELD ◽  
UNA RYAN
Keyword(s):  
Molecular Typing ◽  
Quantitative Polymerase Chain Reaction ◽  
Giardia Duodenalis ◽  
Protozoan Parasite ◽  
Mixed Infections ◽  
Chain Reaction ◽  
Specific Primers ◽  
First Report ◽  
Polymerase Chain ◽  
Assemblage B

SUMMARYLittle is known about the genetic diversity of the protozoan parasite,Giardia duodenalis,infecting humans in Queensland, Australia. The present study typed 88 microscopicallyGiardia-positive isolates using assemblage-specific primers at the triose phosphate isomerase (tpi) gene and sequenced a subset of isolates at the glutamate dehydrogenase (gdh) gene (n= 30) andtpilocus (n= 27). Using thetpi-assemblage specific primers,G. duodenalisassemblage A and assemblage B were detected in 50% (44/88) and 38·6% (34/88) of samples, respectively. Mixed infections with assemblages A and B were identified in 4·5% (4/88) and assemblage E was identified in 6·8% (6/88) of samples. Sequence analysis at thegdhandtpiloci also confirmed the presence of assemblage E in these isolates. Cyst numbers per gram of feces (g−1) were determined using quantitative polymerase chain reaction and of the isolates that were typed as assemblage E, cyst numbers ranged 13·8–68·3 × 106cysts g−1. This is the first report of assemblage E in humans in Australia, indicating that in certain settings, this assemblage may be zoonotic.

scholarly journals Molecular Characterization of Giardia duodenalis in Children and Adults Sampled in Algeria

2020 ◽  
Vol 9 (1) ◽  
pp. 54
Author(s):  
Salem Belkessa ◽  
Daniel Thomas-Lopez ◽  
Karim Houali ◽  
Farida Ghalmi ◽  
Christen Rune Stensvold
Keyword(s):  
Real Time ◽  
Molecular Characterization ◽  
Real Time Pcr ◽  
Intestinal Parasites ◽  
Giardia Duodenalis ◽  
Specific Pcr ◽  
Stool Samples ◽  
Pcr Targeting ◽  
Assemblage B

The molecular epidemiology of giardiasis in Africa remains unclear. A study was carried out across four hospitals in Algeria. A total of 119 fecal samples from 55 children, 37 adults, and 27 individuals of undetermined age, all scored positive for intestinal parasites by microscopy, and were screened by real-time PCR for Giardia. Molecular characterization of Giardia was performed by assemblage-specific PCR and PCR targeting the triose phosphate isomerase gene (tpi). Of the 119 samples, 80 (67%) were Giardia-positive by real-time PCR. For 48 moderately-highly real-time PCR-positive samples, tpi genotyping assigned 22 samples to Assemblage A and 26 to Assemblage B. Contrary to Assemblage A, Assemblage B exhibited substantial genetic diversity and allelic heterozygosity. Assemblage-specific PCR proved to be specific for discriminating Assemblage A or B but not as sensitive as tpi genotyping. We confirmed that real-time PCR is more sensitive than microscopy for detecting Giardia in stool samples and that robust amplification and sequencing of the tpi gene is feasible when moderate-to-strongly real-time PCR-positive samples are used. This study is one of the few performed in Africa providing genotyping data on Giardia infections in humans. Both assemblages A and B were commonly seen and not associated with specific sociodemographic data.

scholarly journals Longitudinal analysis on parasite diversity in honeybee colonies: new taxa, high frequency of mixed infections and seasonal patterns of variation

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Carolina Bartolomé ◽  
María Buendía-Abad ◽  
María Benito ◽  
Beatriz Sobrino ◽  
Jorge Amigo ◽  
...  
Keyword(s):  
New Taxa ◽  
Longitudinal Analysis ◽  
High Frequency ◽  
Seasonal Patterns ◽  
Mixed Infections ◽  
Parasite Diversity

High frequency of primary hyperaldosteronism among hypertensive patients from a primary care area in Sweden

2006 ◽  
Vol 24 (3) ◽  
pp. 154-159 ◽  
Author(s):  
Christina Westerdahl ◽  
Anders Bergenfelz ◽  
Anders Isaksson ◽  
Anders Wihl ◽  
Christina Nerbrand ◽  
...  
Keyword(s):  
Primary Care ◽  
High Frequency ◽  
Primary Hyperaldosteronism ◽  
Hypertensive Patients

scholarly journals Prevalence of Human Papillomavirus in Women from Mexico City

2012 ◽  
Vol 2012 ◽  
pp. 1-4 ◽  
Author(s):  
María Guadalupe López Rivera ◽  
Maria Olivia Medel Flores ◽  
José D’Artagnan Villalba Magdaleno ◽  
Virginia Sánchez Monroy
Keyword(s):  
Mexico City ◽  
High Frequency ◽  
Multiple Infections ◽  
Mexican Women ◽  
Genotype Distribution ◽  
Common Cancer ◽  
Random Samples ◽  
Hpv Genotypes ◽  
Hpv Types ◽  
Hpv 18

Introduction. Cervical cancer is the most common cancer among Mexican women. The goal of the present study was to determine the prevalence and distribution of HPV types in women from Mexico City.Methods. Our study was conducted in the Clinica de Especialidades de la Mujer de la Secretaría de la Defensa Nacional, Mexico. Random samples were taken from 929 healthy women requesting a cervical Papanicolaou examination. Detection and genotyping of HPV were performed by multiplex PCR, with the HPV4A ACE Screening kit (Seegene).Results. 85 of nine hundred twenty-nine women (9.1%) were infected with HPV. Of HPV-positive women, 99% and 1% had high- and low-risk HPV genotypes, respectively. The prevalence of the 16 high-risk (HR) HPV types that were screened was 43% : 42% (18) were HPV positive and 14% (16) were HPV positive, which includes coinfection. Multiple infections with different viral genotypes were detected in 10% of the positive cases. Abnormal cervical cytological results were found in only 15.3% of HPV-positive women, while 84.7% had normal cytological results.Conclusions. We found a similar prevalence of HPV to previous studies in Mexico. The heterogeneity of the HPV genotype distribution in Mexico is evident in this study, which found a high frequency of HPV HR genotypes, the majority of which were HPV 18.

Diagnosing genetically diverse avian malarial infections using mixed-sequence analysis and TA-cloning

Parasitology ◽  
2005 ◽  
Vol 131 (1) ◽  
pp. 15-23 ◽  
Author(s):  
J. PÉREZ-TRIS ◽  
S. BENSCH
Keyword(s):  
Dna Sequences ◽  
Accurate Method ◽  
The Other ◽  
Multiple Infections ◽  
Template Switching ◽  
Mixed Infections ◽  
Nucleotide Substitutions ◽  
Single Nucleotide ◽  
In The Wild ◽  
Host Parasite

Birds harbouring several malarial parasites are common in the wild, and resolving such multiple infections is important for our understanding of host–parasite relationships. We propose a simple and reasonably accurate method for detecting and resolving multiple infections, based on the analysis of parasite cytochrome b DNA sequences: genetically mixed infections are first identified by double nucleotide peaks on sequence electropherograms, and later retrieved by TA-cloning. We applied this method to wild birds, and to experimentally created mixes with varying proportion of two parasites (Plasmodium spp. and Haemoproteus spp.). In general, the method was very efficient in detecting and resolving multiple infections, but some problems were encountered. Several multiple infections were erroneously scored as simple, either because one of the parasite lineages was a better target for the primers used, or because it was much more abundant in the mix. On the other hand, single nucleotide substitutions and template switching during PCR produced artificial sequences in some clones. We discuss the utility of the method, and propose a framework for its use when screening for genetically diverse avian malarial parasites.

SAT0320 High frequency of cardiovascular disease in patients with knee osteoarthritis in a primary care setting:

2013 ◽  
Vol 71 (Suppl 3) ◽  
pp. 580.2-580
Author(s):  
N. Navarro ◽  
C. Orellana ◽  
I. Vázquez ◽  
E. Casado ◽  
J. Gratacόs ◽  
...  
Keyword(s):  
Cardiovascular Disease ◽  
Primary Care ◽  
Knee Osteoarthritis ◽  
High Frequency ◽  
Care Setting ◽  
Primary Care Setting

Detection of potentially human infectious assemblages of Giardia duodenalis in fecal samples from beef and dairy cattle in Scotland

Parasitology ◽  
2018 ◽  
Vol 146 (9) ◽  
pp. 1123-1130 ◽  
Author(s):  
Paul M. Bartley ◽  
Beeke K. Roehe ◽  
Sarah Thomson ◽  
Hannah J. Shaw ◽  
Frederieke Peto ◽  
...  
Keyword(s):  
Sequence Analysis ◽  
Beef Cattle ◽  
Dairy Cattle ◽  
Age Groups ◽  
Giardia Duodenalis ◽  
Host Age ◽  
Fecal Samples ◽  
Different Ages ◽  
Sequence Types ◽  
Assemblage B

AbstractThis study aimed to determine the prevalence and assemblages of Giardia duodenalis present in Scottish beef and dairy cattle at different ages, to try to ascertain if cattle could play a role in the spread of zoonotic assemblages of Giardia. A total of 388 fecal samples (128 beef and 253 dairy, seven of unknown breed) were collected from 19 farms in Scotland. Samples were sub-divided by host age, 1, 2, 3, 4, 5 and 6, 7–24 and ⩾25 weeks. DNA was extracted and tested by PCR to detect G. duodenalis DNA. Of the 388 samples, 126 tested positive, giving an overall prevalence of 32.5%, with positive samples being observed in all age groups tested. The prevalence in dairy cattle was 44.7% (113/235), which was significantly higher (P < 0.001) than the prevalence in beef cattle 10.1% (13/128). Sequence analysis demonstrated the presence of assemblage E (77.2%, sequence types E-S1–E-S5), assemblage B (18.2%) and assemblage A (sub-assemblages AI-AII) (4.6%). These data demonstrate that G. duodenalis is found routinely in both dairy and beef cattle throughout Scotland; the presence of assemblages A and B also indicates that cattle may play a role in the spread of potentially zoonotic assemblages of Giardia.

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