Molecular typing of Giardia duodenalis isolates from nonhuman primates housed IN a Brazilian zoo.
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SUBMITTER: David EB
PROVIDER: S-EPMC4085826 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:BackgroundGiardia 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 findingsTwo 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/significanceTyping 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.
| S-EPMC8023459 | biostudies-literature
Project description:BackgroundGiardia duodenalis is one of the most prevalent and highly diverse human parasites, encompassing a complex of eight genetically distinct assemblages, each further divided into sub-assemblages. While in recent years, G. duodenalis genotype distribution patterns in humans have been intensely studied, there is still very little information available on the diversity of Giardia genotypes and sub-assemblages infecting people in Romania. In the present study, we investigated the genetic diversity of Giardia duodenalis in asymptomatic patients from Romania.MethodsOver an 11-month period, human feces from 7805 healthy adults were screened by microscopic analysis for G. duodenalis cysts during their obligatory periodic check-ups. DNA extraction was performed from microscopic-positive fecal samples, followed by multilocus sequence typing of four genetic loci of the ITS region, gdh, tpi and bg genes, followed by DNA sequencing and phylogenetic analysis. Statistical analysis was performed using EpiInfo 2000 software.ResultsThe prevalence of giardiasis in the present study was 0.42% (33/7805). Twenty-three samples (76.67%) were successfully genotyped at each locus. The bg and tpi genes had the highest typing success rate (100%). The identified assemblages were assemblage A in 27 cases (subtypes A2 and A3), and B in 3 cases.ConclusionsTo our knowledge, the present study is the first report of multilocus sequence typing of G. duodenalis isolated from humans in Romania. The present results may shed light on G. duodenalis infection in humans at a regional and national level, thus increasing awareness against this parasitic infection.
| S-EPMC7393877 | biostudies-literature
Project description:Only a few studies based on multilocus characterization have been conducted on the molecular epidemiology of Giardia duodenalis in captive nonhuman primates (NHPs). The present article provides the first report on the occurrence of G. duodenalis in the ring-tailed lemur (Lemur catta) and barbary macaque (Macaca sylvanus) kept in the zoo in Košice, Slovakia. All samples were examined by flotation technique, with total prevalence of 17.4% (4/23). The microscopically positive samples were assayed by nested PCR and consecutively sequenced at β-giardin (bg), glutamate dehydrogenase (gdh) and triosephosphate isomerase (tpi) genes. Positive isolates were recognized as assemblage B and sub-assemblage BIV and subtype WB8 were confirmed. The identification and genotyping of this parasite in Slovakia, may help to better understand the epidemiological situation in Europe about the circulation of G. duodenalis zoonotic assemblages in NHPs.
| S-EPMC8254674 | biostudies-literature
Project description:BackgroundThe parasite Giardia duodenalis infects a wide range of vertebrate hosts, including domestic and wild animals as well as humans. Giardia is genotyped into eight assemblages (A-H). Zoonotic assemblages A and B have already been identified in humans and wild and domestic animals (non-human primates and cats) from Brazilian Amazon and in the world. Due to its zoonotic/zooanthroponotic nature, surveillance initiatives and the definition of Giardia assemblages are important in order to characterise the epidemiological scenario and to implement further control measures.ObjectivesDetermine assemblages of G. duodenalis in sloths from the Brazilian Amazon Region.MethodsFaecal parasitological examination of sloths from Amazonas State. Polymerase chain reaction (PCR) targeting the beta giardin (BG), and genes from multilocus sequence typing (MLST) scheme, amplicon sequencing and phylogenetic analysis.FindingsHere, we identified, by microscopy, Giardia in two northern sloths (Bradypus tridactylus). These two samples were submitted to molecular assays and it was revealed that both were infected by G. duodenalis assemblage A. Phylogenetic analysis showed that they belong to assemblage A within sequences from humans and wild and domestic animals.ConclusionTherefore, besides showing, by the first time, the current presence of this parasite in sloths, our findings reveals that this wild animal species would be part of the zoonotic/zooanthroponotic scenario of this parasite in the Brazilian Amazon.
| S-EPMC10644951 | biostudies-literature
Project description:Giardiasis is considered the most common intestinal parasitic disease in humans worldwide. In Cuba, this infection has particularly a strong clinical impact on the child population. Giardia duodenalis is a highly diverse protozoan, which comprises a complex of eight morphologically identical genetic assemblages, further divided into sub-assemblages. The present study used triose phosphate isomerase (tpi) and small-subunit ribosomal RNA (SSU rRNA) genes as genetic markers for the identification of G. duodenalis assemblages and sub-assemblages in correlation with clinical and epidemiological data in children attended at the Paediatric Hospital "William Soler" and at Pedro Kouri Institute, between 2015 and 2016. A prevalence of 8% of G. duodenalis infection was recorded in stool samples after concentration techniques from 68 children out of 847 analysed. A 100% detection of Giardia DNA was achieved by a SSU-rRNA PCR, whereas DNA from 63 of 68 (92.6%) was successfully amplified by tpi-PCR. By this assemblage-specific tpi-PCR 32 (50.8%) assemblage B, 17 (27.0%) assemblage A and 14 (22.2%) mixed infection (A + B) were identified. Assemblage B was significantly (P < 0.02) more frequently found in children with diarrhoea. Sequence analysis of the tpi gene of Giardia isolates from symptomatic children showed that assemblage A belonged to the sub-assemblage AII, and 4 sub assemblages BIV and 1 sub assemblage BIII were also recorded. Only 2 discordant genotyping results were observed by phylogenetic comparison of SSU-rRNA and tpi sequences. Further studies with novel molecular tools for a better discrimination at the sub-assemblage level are needed to identify the dynamics of spread of giardiasis and to verify possible correlations between Giardia genetic diversity and clinical manifestation.
| S-EPMC5952668 | biostudies-literature
Project description:Giardia duodenalis is a cryptic protozoan, which has eight assemblages (A-H). Assemblages A and B are the main genotypes reported from humans with probable anthroponotic and zoonotic transmission. The current study aimed to characterize G. duodenalis assemblages in tuberculosis (TB) patients and healthy subjects using multilocus genotyping (MLG). Thirty Giardia-positive stool samples, which were obtained from TB patients and healthy subjects were included in the study. After total DNA extraction, three β-giardin (bg), triosephosphate isomerase (tpi), glutamate dehydrogenase (gdh) genes were amplified and sequenced. Obtained sequences were compared to the GenBank database to characterize assemblages. Phylogenetic analysis using Maximum Likelihood (ML) and Tamura 3-parameter was performed for each gene. From 30 Giardia-positive subjects, 17 (57%) and 13 (43%) were from healthy and TB-infected subjects, respectively. There was no significant co-existence of Giardia and tuberculosis (P-value = 0.051). In addition, 14 (46.7%) and 16 (53.3%) of Giardia isolates were from asymptomatic and symptomatic subjects, respectively. PCR amplification was successful in 25 single samples (83.3%) consisted of 20 for tpi, 15 for bg, and 13 for gdh genes. Accordingly, 13/25 (52%) and 8/25 (32%) belonged to assemblage A and assemblages B, respectively, whereas 4/25 (16%) were either assemblage A or B with different genes at the same time. Significant correlation between assemblages and TB, age, and symptoms was not seen. The phylogenetic analyses represented no separation based on TB and gastrointestinal symptoms. Assemblage A was the predominant genotype in samples. The high frequency of assemblage AII indicated importance of anthroponotic transmission of Giardia in both healthy and TB patients. In addition, considering the exclusive reports of sub-assemblage AIII in wild ruminants, the presence of AIII in the current study have to be carefully interpreted. The inconsistency between the assemblage results of either bg or gdh loci with tpi gene signifies the insufficiency of single gene analysis and the necessity for MLG in molecular epidemiology of G. duodenalis.
| S-EPMC10035859 | biostudies-literature
Project description:BackgroundThe flagellate protozoan Giardia duodenalis is an enteric parasite causing human giardiasis, a major gastrointestinal disease of global distribution affecting both developing and industrialised countries. In Spain, sporadic cases of giardiasis have been regularly identified, particularly in pediatric and immigrant populations. However, there is limited information on the genetic variability of circulating G. duodenalis isolates in the country.MethodsIn this longitudinal molecular epidemiological study we report the diversity and frequency of the G. duodenalis assemblages and sub-assemblages identified in 199 stool samples collected from 184 individual with symptoms compatible with giardiasis presenting to two major public hospitals in Madrid for the period December 2013-January 2015. G. duodenalis cysts were initially detected by conventional microscopy and/or immunochomatography on stool samples. Confirmation of the infection was performed by direct immunofluorescence and real-time PCR methods. G. duodenalis assemblages and sub-assemblages were determined by multi-locus genotyping of the glutamate dehydrogenase (GDH) and β-giardin (BG) genes of the parasite. Sociodemographic and clinical features of patients infected with G. duodenalis were also analysed.Principal findingsOf 188 confirmed positive samples from 178 giardiasis cases a total of 124 G. duodenalis isolates were successfully typed at the GDH and/or the BG loci, revealing the presence of sub-assemblages BIV (62.1%), AII (15.3%), BIII (4.0%), AI (0.8%), and AIII (0.8%). Additionally, 6.5% of the isolates were only characterised at the assemblage level, being all of them assigned to assemblage B. Discordant genotype results AII/AIII or BIII/BIV were also observed in 10.5% of DNA isolates. A large number of multi-locus genotypes were identified in G. duodenalis assemblage B, but not assemblage A, isolates at both the GDH and BG loci, confirming the high degree of genetic variability observed in other molecular surveys. BIV was the most prevalent genetic variant of G. duodenalis found in individuals with symptomatic giardiasis in the population under study.ConclusionsHuman giardiasis is an ongoing public health problem in Spain affecting primarily young children under four years of age but also individuals of all age groups. Our typing and sub-typing results demonstrate that assemblage B is the most prevalent G. duodenalis assemblage circulating in patients with clinical giardiasis in Central Spain. Our analyses also revealed a large genetic variability in assemblage B (but not assemblage A) isolates of the parasite, corroborating the information obtained in similar studies in other geographical regions. We believe that molecular data presented here provide epidemiological evidence at the population level in support of the existence of genetic exchange within assemblages of G. duodenalis.
| S-EPMC4671680 | biostudies-literature
Project description:BackgroundGiardia duodenalis is an important intestinal protozoan in humans worldwide with high infection rates occurring in densely populated and low resource settings. The parasite has been recorded to cause diarrhea in children. This study was carried out to identify G. duodenalis assemblages and sub-assemblages in children presenting with diarrhea in Kenya.MethodsA total of 2112 faecal samples were collected from children aged ≤ 5 years and screened for the presence of Giardia cysts using microscopy. A total of 96 (4.5%) samples were identified as Giardia positive samples and were genotyped using glutamate dehydrogenase (gdh), triose phosphate isomerase (tpi) and β-giardin loci.ResultsThe three markers successfully genotyped 72 isolates and grouped 2 (1.4) isolates as Assemblage A, 64 (88.9) as Assemblage B and 7 (9.7%) consisted of mixed infections with assemblage A and B. A further analysis of 50 isolates using GDH Polymerase Chain Reaction and Restriction Fragment Length Polymorphism (PCR-RFLP) categorized 2 assemblage A isolates as sub-assemblage AII while 6 and 14 assemblage B isolates were categorized into sub-assemblage BIII and BIV respectively. A mixed infection with sub-assemblage BIII and BIV was recorded in 28 isolates. Over half (55.6%) of Giardia infections were recorded among the children between 13 to 48 months old.ConclusionThis paper reports the first data on the assemblages and sub-assemblages of Giardia duodenalis in children representing with diarrhea in Kenya.
| S-EPMC4802924 | biostudies-literature
Project description:Almost all human giardiasis infections are caused by Giardia duodenalis assemblages A and B. Differentiation between human infections with these assemblages, as well as between single-assemblage (A or B) and mixed-assemblage (A and B) infections, is therefore needed to better understand the pathological impact of infection with either, or both, assemblages. We assessed the prevalence of G. duodenalis assemblages A and B using 305 fecal samples provided by school-age children situated along the southern shoreline of Lake Malawi. Concurrently, intestinal pathology data were also collected to test for association(s) between assemblage infection status and intestinal health. Prevalence of G. duodenalis infection was 39.3% by real-time polymerase chain reaction. Of all identified infections, 32% were single G. duodenalis assemblage A and 32% were single G. duodenalis assemblage B, whereas 33% were mixed-assemblage infections. Fifteen unique G. duodenalis assemblage A and 13 unique G. duodenalis assemblage B β-giardin haplotypes were identified. There was a positive association between single infection with G. duodenalis assemblage B and both self-reporting of abdominal pain (odds ratio [OR]: 3.05, P = 0.004) and self-reporting of diarrhea (OR: 3.1, P = 0.003). No association between single infection with assemblage A and any form of intestinal pathology was found. Additionally, there was a positive association between mixed-assemblage infections and self-reporting of abdominal pain (OR: 3.1, P = 0.002). Our study highlights the importance G. duodenalis assemblage typing and reaffirms the need for improved access to water, sanitation and hygiene infrastructure in rural areas of low- and middle-income countries.
| S-EPMC10484258 | biostudies-literature
Project description:BackgroundLeptospirosis is caused by pathogenic spirochetes of the genus Leptospira spp. This zoonotic disease is distributed globally and affects domestic animals, including cattle. Leptospira interrogans serogroup Sejroe serovar Hardjo and Leptospira borgpetersenii serogroup Sejroe serovar Hardjo remain important species associated with this reproductive disease in livestock production. Previous studies on Brazilian livestock have reported that L. interrogans serovar Hardjo is the most prevalent leptospiral agent in this country and is related to clinical signs of leptospirosis, which lead to economic losses in production. Here, we described the isolation of three clinical strains (Norma, Lagoa and Bolivia) obtained from leptospirosis outbreaks that occurred in Minas Gerais state in 1994 and 2008.ResultsSerological and molecular typing using housekeeping (secY and 16SrRNA) and rfb locus (ORF22 and ORF36) genes were applied for the identification and comparative analysis of Leptospira spp. Our results identified the three isolates as L. interrogans serogroup Sejroe serovar Hardjo and confirmed the occurrence of this bacterial strain in Brazilian livestock. Genetic analysis using ORF22 and ORF36 grouped the Leptospira into serogroup Sejroe and subtype Hardjoprajitno. Genetic approaches were also applied to compare distinct serovars of L. interrogans strains by verifying the copy numbers of the IS1500 and IS1533 insertion sequences (ISs). The IS1500 copy number varied among the analyzed L. interrogans strains.ConclusionThis study provides evidence that L. interrogans serogroup Sejroe serovar Hardjo subtype Hardjoprajitno causes bovine leptospirosis in Brazilian production. The molecular results suggested that rfb locus (ORF22 and ORF36) could improve epidemiological studies by allowing the identification of Leptospira spp. at the serogroup level. Additionally, the IS1500 and IS1533 IS copy number analysis suggested distinct genomic features among closely related leptospiral strains.
| S-EPMC5471881 | biostudies-literature