scholarly journals Human Bocavirus Prevalence In Children With Acute Gastroenteritis From Rural Communities In The Northen Region Of South Africa

2019 ◽  
Author(s):  
Mpumelelo Casper Rikhotso ◽  
Ronewa Khumela ◽  
Jean Pierre Kabue ◽  
Afsatou Ndama Traoré ◽  
Natasha Potgieter
Keyword(s):  
South Africa ◽  
Rural Communities ◽  
Real Time ◽  
Real Time Pcr ◽  
Acute Gastroenteritis ◽  
Enteric Viruses ◽  
Health Care Facilities ◽  
Human Bocavirus ◽  
Stool Samples ◽  
Pcr Targeting

AbstractBACKGROUNDAcute gastroenteritis (AGE) is a leading cause of morbidity and mortality in young children worldwide. Human Bocavirus (HBoV) is an emerging virus globally associated with diarrhea. The aim of this study was to demonstrate the prevalence of HBoV genotypes in children (≤5 years) from rural communities in South Africa (SA) suffering from AGE.MATERIAL AND METHODA total of 141 fecal samples of children ≤5 years with acute gastroenteritis (AGE) were collected from rural Primary Health Care facilities in the Vhembe district of SA between June 2017 and July 2018. Clinical symptoms and demographic data were also recorded. A total of 102 (72%) were outpatients and 39 (28%) were hospitalized patients. Human Bocavirus (HBoV) genotypes were determined using Real-Time Multiplex PCR. DNA extracts of positive samples were confirmed by conventional PCR targeting the NS1 gene. Co-infection with other enteric viruses were determined in HBoV positive samples using Real-Time PCR.RESULTSHBoV was detected in 8 (5.7%) children with AGE. Children were in the age group between 1-24 months. HBoV1 and HBoV3 genotypes were each detected in 3 (37.5%) stool samples and HBoV2 in 2 (25%) stool samples. Co-infection with other enteric viruses included Rotavirus (37.5%); Adenovirus (37.5%); Norovirus (25%) and Astrovirus (12.5%).CONCLUSIONHBoV infections could be seen as a potential emerging diarrheal pathogen in South Africa. Further studies are required to understand the role of HBoV infections in children and adults with acute gastroenteritis.Author summaryAcute gastroenteritis (AGE) is recognized as a major cause for mortality in children ≤5 years of age in Africa and other developing countries. Viruses known to be involved in AGE includes Rotavirus, Norovirus, Astrovirus and Adenovirus and have been reported globally. Recently the Human Bocavirus (HBoV) have been reported in numerous studies globally as a potential cause of diarrhea. In this study, the prevalence and genetic diversity of human Bocavirus in children with AGE from rural communities in Limpopo, South Africa were investigated. In total, 141 stool samples from children ≤ 5 years with AGE were assessed for the presence of HBoV using Real-Time PCR. HBoV were detected in 8 (5.7%) patients and included 3 positive samples for HBoV1 and HBoV3 respectively and 2 positive for HBoV2. No HBoV4 were detected. Among the 8 positive HBoV samples, co-infection with other enteric viruses were found in 7 (87.5%) samples, while mono infection with HBoV alone was detected in 1 (12.5%) patient. HBoV mixed infection with Rotavirus (3/8; 37.5%); Adenovirus (3/8; 37.5%); Norovirus (2/8; 25%) and Astrovirus (1/8; 12.5%) were observed in this study. This study reported for the first time on the prevalence of human Bocavirus in children with AGE from rural communities in South Africa.

scholarly journals Predominance of Human Bocavirus Genotype 1 and 3 in Outpatient Children with Diarrhea from Rural Communities in South Africa, 2017–2018

Pathogens ◽  
2020 ◽  
Vol 9 (4) ◽  
pp. 245
Author(s):  
Mpumelelo Casper Rikhotso ◽  
Ronewa Khumela ◽  
Jean Pierre Kabue ◽  
Afsatou Ndama Traoré-Hoffman ◽  
Natasha Potgieter
Keyword(s):  
South Africa ◽  
Rural Communities ◽  
Real Time ◽  
Acute Gastroenteritis ◽  
Clinical Symptoms ◽  
Demographic Data ◽  
Enteric Viruses ◽  
Health Care Facilities ◽  
Human Bocavirus ◽  
Pcr Targeting

Human bocavirus (HBoV) is an emerging virus globally associated with diarrhea in young children. This study aims to investigate the prevalence of HBoV genotypes in children (≤5 years) from rural communities in South Africa (SA) suffering from acute gastroenteritis (AGE). A total of 141 fecal samples of children ≤5 years with acute gastroenteritis (AGE) were collected from rural primary health care facilities in the Vhembe district of SA between June 2017 and July 2018. Clinical symptoms and demographic data were also recorded. A total of 102 (72%) were outpatients, and 39 (28%) were hospitalized patients. Human bocavirus (HBoV) genotypes were determined using real-time multiplex PCR. DNA extracts of positive samples were confirmed by conventional PCR targeting the NS1 gene. Co-infection with other enteric viruses were determined in HBoV-positive samples using real-time PCR. HBoV was detected in eight (5.7%) children with AGE, of which three (37.5%) were HBoV1, three (37.5%) were HBoV3, and two (25%) were HBoV2. The majority of positive cases were identified in outpatients (62%) between the ages of 1 and 24 months. Co-infection in HBoV-positive samples with other enteric viruses included rotavirus (37.5%), adenovirus (37.5%), norovirus (25%), and astrovirus (12.5%). HBoV infections could be seen as a potential emerging diarrheal pathogen in South Africa. However, more studies are needed to understand the role of HBoV infections in children with AGE.

scholarly journals Prevalence and Genetic Characterisation of Human Sapovirus from Children with Diarrhoea in the Rural Areas of Vhembe District, South Africa, 2017–2020

Viruses ◽  
2021 ◽  
Vol 13 (3) ◽  
pp. 393
Author(s):  
Mpho Magwalivha ◽  
Jean-Pierre Kabue Ngandu ◽  
Afsatou Ndama Traore ◽  
Natasha Potgieter
Keyword(s):  
South Africa ◽  
Rural Communities ◽  
Rural Areas ◽  
Rna Extraction ◽  
Diarrhoeal Disease ◽  
Positive Sample ◽  
Prevalent Strain ◽  
Stool Samples ◽  
Vhembe District ◽  
Pcr Targeting

Diarrhoeal disease is considered an important cause of morbidity and mortality in developing areas, and a large contributor to the burden of disease in children younger than five years of age. This study investigated the prevalence and genogroups of human sapovirus (SV) in children ≤5 years of age in rural communities of Vhembe district, South Africa. Between 2017 and 2020, a total of 284 stool samples were collected from children suffering with diarrhoea (n = 228) and from children without diarrhoea (n = 56). RNA extraction using Boom extraction method, and screening for SV using real-time PCR were done in the lab. Positive samples were subjected to conventional RT-PCR targeting the capsid fragment. Positive sample isolates were genotyped using Sanger sequencing. Overall SV were detected in 14.1% (40/284) of the stool samples (16.7% (38/228) of diarrhoeal and 3.6% (2/56) of non-diarrhoeal samples). Significant correlation between SV positive cases and water sources was noted. Genogroup-I was identified as the most prevalent strain comprising 81.3% (13/16), followed by SV-GII 12.5% (2/16) and SV-GIV 6.2% (1/16). This study provides valuable data on prevalence of SV amongst outpatients in rural and underdeveloped communities, and highlights the necessity for further monitoring of SV circulating strains as potential emerging strains.

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 Cryptosporidium spp. during chemotherapy: a cross-sectional study of 94 patients with malignant solid tumor

2021 ◽  
Vol 41 (5) ◽  
pp. 293-298
Author(s):  
Mehmet Karabey ◽  
Hüseyin Can ◽  
Tülay Öncü Öner ◽  
Mert Döşkaya ◽  
Sedef Erkunt Alak ◽  
...  
Keyword(s):  
Sample Size ◽  
Real Time ◽  
Solid Tumors ◽  
Real Time Pcr ◽  
Tertiary Care ◽  
Diagnostic Methods ◽  
Cross Sectional ◽  
Cryptosporidium Spp ◽  
Stool Samples ◽  
Pcr Targeting

BACKGROUND: Cryptosporidium spp . is a protozoan parasite that infects many vertebrate animals, including humans. Since Cryptosporidium spp . can cause chronic life-threatening diarrhea and severe malabsorption in immunocompromised patients, we investigated the prevalence of this parasite among patients undergoing chemotherapy for malignant solid tumors. OBJECTIVE: Investigate the prevalence of Cryptosporidium spp . in stool samples. DESIGN: Cross-sectional. SETTING: Tertiary care. PATIENTS AND METHODS: Stool samples were collected from adult patients with malignant solid tumors receiving chemotherapy and diarrhea. Cryptosporidium spp . prevalence was determined using Ziehl–Neelsen staining, ELISA, and real-time PCR targeting of the COWP gene. MAIN OUTCOME MEASURE: The prevalence of Cryptosporidium spp . in patients undergoing chemotherapy for malignant solid tumors. SAMPLE SIZE: 94 RESULTS: The prevalence was 2.1% (2/94), 5.3% (5/94), and 5.3% (5/94) as detected by Ziehl–Neelsen staining, real-time PCR and ELISA, respectively. The prevalence reached 8.5% (8/94) using all results obtained from the three methods. Among eight positive stool samples, four were positive by at least two different methods (Ziehl–Neelsen staining-ELISA or ELISA-real-time PCR) whereas the remaining four were positive by either ELISA or real-time PCR. CONCLUSION: These findings show the risk of cryptosporidiosis in cancer patients and the necessity to use at least two diagnostic methods during the diagnosis of cryptosporidiosis to reach more accurate and trustworthy results. LIMITATIONS: Further studies with a larger sample size are recommended. CONFLICT OF INTEREST: None.

Isolation and detection of Shiga toxin-producing Escherichia coli in clinical stool samples using conventional and molecular methods

2009 ◽  
Vol 58 (7) ◽  
pp. 905-911 ◽  
Author(s):  
Matthew W. Gilmour ◽  
Linda Chui ◽  
Theodore Chiu ◽  
Dobryan M. Tracz ◽  
Kathryn Hagedorn ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Real Time ◽  
Shiga Toxin ◽  
Real Time Pcr ◽  
Molecular Methods ◽  
Genetic Profile ◽  
Pcr Assay ◽  
Suspension Array ◽  
Stool Samples ◽  
Pcr Targeting

The isolation of Shiga toxin-producing Escherichia coli (STEC) other than serogroup O157 from clinical stool samples is problematic due to the lack of differential phenotypic characteristics from non-pathogenic E. coli. The development of molecular reagents capable of identifying both toxin and serogroup-specific genetic determinants holds promise for a more comprehensive characterization of stool samples and isolation of STEC strains. In this study, 876 stool samples from paediatric patients with gastroenteritis were screened for STEC using a cytotoxicity assay, commercial immunoassay and a conventional PCR targeting Shiga-toxin determinants. In addition, routine culture methods for isolating O157 STEC were also performed. The screening assays identified 45 stools presumptively containing STEC, and using non-differential culture techniques a total of 20 O157 and 22 non-O157 strains were isolated. These included STEC serotypes O157 : H7, O26 : H11, O121 : H19, O26 : NM, O103 : H2, O111 : NM, O115 : H18, O121 : NM, O145 : NM, O177 : NM and O5 : NM. Notably, multiple STEC serotypes were isolated from two clinical stool samples (yielding O157 : H7 and O26 : H11, or O157 : H7 and O103 : H2 isolates). These data were compared to molecular serogroup profiles determined directly from the stool enrichment cultures using a LUX real-time PCR assay targeting the O157 fimbrial gene lpfA, a microsphere suspension array targeting allelic variants of espZ and a gnd-based molecular O-antigen serogrouping method. The genetic profile of individual stool cultures indicated that the espZ microsphere array and lpfA real-time PCR assay could accurately predict the presence and provide preliminary typing for the STEC strains present in clinical samples. The gnd-based molecular serogrouping method provided additional corroborative evidence of serogroup identities. This toolbox of molecular methods provided robust detection capabilities for STEC in clinical stool samples, including co-infection of multiple serogroups.

scholarly journals 1777. Metagenomic Approach for the Detection of Viruses in Stool Samples from Infants and Children with Acute Gastroenteritis in Kuwait

2019 ◽  
Vol 6 (Supplement_2) ◽  
pp. S654-S655
Author(s):  
Hawraa Adel ◽  
Nada Madi ◽  
Widad Al-Nakib
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Acute Gastroenteritis ◽  
Human Adenovirus ◽  
Infants And Children ◽  
Kappa Statistics ◽  
Next Generation Sequencing Technology ◽  
Stool Samples ◽  
Combined Infection ◽  
Metagenomic Approach

Abstract Background Metagenomics techniques are target-independent tools that enable the identification of uncommon disease etiologies and genomic characterization of all microorganisms present in a sample in less time and at a lower cost than previous sequencing techniques. In this study, we developed a metagenomic approach using next-generation sequencing technology to identify known and unknown viruses in stool samples from infants and children with acute gastroenteritis in Kuwait. Methods We have investigated 84 stool samples from infants and children aged one month to 10 years old with signs and symptoms of gastroenteritis who attended Mubarak Al-Kabeer and Al-Amiri hospitals in Kuwait from January to December 2017 using both multiplex real-time PCR and metagenomics sequencing (Illumina Miseq instrument) methods. Results The metagenomics analysis of viral sequences verified that human adenovirus was the leading cause of gastroenteritis among infants and children in Kuwait, and was detected in 23% of the samples, rotavirus A was detected in 16% of the samples, and the combined infection of human adenovirus and rotavirus was detected in 7% of the samples. Also, newly discovered viruses known to cause gastroenteritis were identified; astrovirus MLB2 and primate bocaparvovirus-1 were detected in 5% of the samples. Also, each of the following new viruses was detected in 2% of the samples; aichivirus A, cardiovirus, parechovirus A, astrovirus VA4, cosavirus F, and bufavirus-3. On the other hand, multiplex real-time PCR showed that the combined infection of human adenovirus and rotavirus was the leading cause of gastroenteritis among infants and children in Kuwait, which was detected in 27% of the samples. However, the rotavirus was the second most common cause of diarrhea, which was detected in 20% of the samples. And the human adenovirus alone was detected in 18% of the samples. Our results showed a 69% agreement between both methods. By applying the Cohen’s Kappa statistics for a measure of agreement, the result gave fair agreement between the two methods (k = 0.388, P = 0.0). Conclusion Our findings revealed the capability of a metagenomic approach to detect many viruses causing gastroenteritis in stool samples from infants and children in Kuwait. Disclosures All authors: No reported disclosures.

scholarly journals Novel Human Astroviruses: Prevalence and Association with Common Enteric Viruses in Undiagnosed Gastroenteritis Cases in Spain

Viruses ◽  
2019 ◽  
Vol 11 (7) ◽  
pp. 585 ◽  
Author(s):  
Diem-Lan Vu ◽  
Aurora Sabrià ◽  
Nuria Aregall ◽  
Kristina Michl ◽  
Virginia Rodriguez Garrido ◽  
...  
Keyword(s):  
Real Time ◽  
Acute Gastroenteritis ◽  
Enteric Viruses ◽  
Epidemiological Studies ◽  
Diagnostic Methods ◽  
Rt Pcr ◽  
Routine Diagnostics ◽  
Human Astroviruses ◽  
Stool Samples ◽  
Pcr Assays

A remarkable percentage of acute gastroenteritis cases remain etiologically undiagnosed. The aim of the study was to determine the prevalence of common and emerging enteric viruses, such as novel human astroviruses, among undiagnosed samples from children with acute gastroenteritis. Epidemiological studies for novel human astroviruses are still scarce. Stool samples collected over two consecutive winter seasons (2016–2017) from children with gastroenteritis in Spain, which were negative for bacteria, rotavirus, and adenovirus by routine diagnostics were screened by real-time RT-PCR assays for the presence of classical and novel astrovirus, rotavirus, norovirus GI and GII, sapovirus, and adenovirus. Overall, 220/384 stool samples (57.3%) were positive for at least one virus. Co-infections were identified in 21% of cases. Among a total of 315 viruses identified, adenovirus was the most prevalent (n = 103), followed by rotavirus (n = 51), sapovirus (n = 50), classical astrovirus (n = 43), novel astroviruses (n = 42), and norovirus (n = 26). Novel astroviruses were present in 13.3% of virus-positive cases. Most novel astroviruses were found in children <2-year-old (30/39 children, 77%, p = 0.01) and were found in co-infection (66%). Only classical astroviruses demonstrated significant differences in the Cq values during mono-infections compared to co-infections. In conclusion, common enteric viruses may be frequently found in children with undiagnosed gastroenteritis, indicating the need to implement more sensitive diagnostic methods. Novel astroviruses circulate in the community and could be the cause of gastroenteritis among young children.

scholarly journals Association of Ct Values from Real-Time PCR with Culture in Microbiological Clearance Samples for Shiga Toxin-Producing Escherichia coli (STEC)

2020 ◽  
Vol 8 (11) ◽  
pp. 1801
Author(s):  
Michael Bording-Jorgensen ◽  
Brendon D. Parsons ◽  
Gillian A.M. Tarr ◽  
Binal Shah-Gandhi ◽  
Colin Lloyd ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Real Time ◽  
Shiga Toxin ◽  
Real Time Pcr ◽  
Gene Detection ◽  
Culture Positivity ◽  
Hands On ◽  
Culture Independent ◽  
Chromogenic Agar ◽  
Stool Samples

Shiga toxin-producing Escherichia coli (STEC) are associated with acute gastroenteritis worldwide, which induces a high economic burden on both healthcare and individuals. Culture-independent diagnostic tests (CIDT) in frontline microbiology laboratories have been implemented in Alberta since 2019. The objectives of this study were to determine the association between gene detection and culture positivity over time using STEC microbiological clearance samples and also to establish the frequency of specimen submission. Both stx genes’ amplification by real-time PCR was performed with DNA extracted from stool samples using the easyMAG system. Stools were inoculated onto chromogenic agar for culture. An association between gene detection and culture positivity was found to be independent of which stx gene was present. CIDT can provide rapid reporting with less hands-on time and technical expertise. However, culture is still important for surveillance and early cluster detection. In addition, stool submissions could be reduced from daily to every 3–5 days until a sample is negative by culture.

scholarly journals Differing Effects of Standard and Harsh Nucleic Acid Extraction Procedures on Diagnostic Helminth Real-Time PCRs Applied to Human Stool Samples

Pathogens ◽  
2021 ◽  
Vol 10 (2) ◽  
pp. 188
Author(s):  
Tanja Hoffmann ◽  
Andreas Hahn ◽  
Jaco J. Verweij ◽  
Gérard Leboulle ◽  
Olfert Landt ◽  
...  
Keyword(s):  
Nucleic Acid ◽  
Real Time ◽  
Real Time Pcr ◽  
Acid Extraction ◽  
Nucleic Acid Extraction ◽  
Pcr Assay ◽  
Bead Beating ◽  
Stool Samples ◽  
Pcr Assays ◽  
Human Stool

This study aimed to assess standard and harsher nucleic acid extraction schemes for diagnostic helminth real-time PCR approaches from stool samples. A standard procedure for nucleic acid extraction from stool and a procedure including bead-beating as well as proteinase K digestion were compared with group-, genus-, and species-specific real-time PCR assays targeting helminths and nonhelminth pathogens in human stool samples. From 25 different in-house and commercial helminth real-time PCR assays applied to 77 stool samples comprising 67 historic samples and 10 external quality assessment scheme samples positively tested for helminths, higher numbers of positive test results were observed after bead-beating-based nucleic acid extraction for 5/25 (20%) real-time PCR assays irrespective of specificity issues. Lower cycle threshold values were observed for one real-time PCR assay after the standard extraction scheme, and for four assays after the bead-beating-based scheme. Agreement between real-time PCR results after both nucleic acid extraction strategies according to Cohen’s kappa ranged from poor to almost perfect for the different assays. Varying agreement was observed in eight nonhelminth real-time PCR assays applied to 67 historic stool samples. The study indicates highly variable effects of harsh nucleic acid extraction approaches depending on the real-time PCR assay used.

scholarly journals Comparative Assessment of In-House Real-Time PCRs Targeting Enteric Disease-Associated Microsporidia in Human Stool Samples

Pathogens ◽  
2021 ◽  
Vol 10 (6) ◽  
pp. 656
Author(s):  
Konstantin Tanida ◽  
Andreas Hahn ◽  
Kirsten Alexandra Eberhardt ◽  
Egbert Tannich ◽  
Olfert Landt ◽  
...  
Keyword(s):  
Real Time ◽  
Indigenous People ◽  
Real Time Pcr ◽  
Latent Class ◽  
Enterocytozoon Bieneusi ◽  
Enteric Disease ◽  
Immunosuppressed Patients ◽  
Study Population ◽  
Stool Samples ◽  
Pcr Assays

Microsporidiosis is an infection predominantly occurring in immunosuppressed patients and infrequently also in travelers. This study was performed to comparatively evaluate the diagnostic accuracy of real-time PCR assays targeting microsporidia with etiological relevance in the stool of human patients in a latent class analysis-based test comparison without a reference standard with perfect accuracy. Thereby, two one-tube real-time PCR assays and two two-tube real-time PCR assays targeting Enterocytozoon bieneusi and Encephalocytozoon spp. were included in the assessment with reference stool material (20), stool samples from Ghanaian HIV-positive patients (903), and from travelers, migrants and Colombian indigenous people (416). Sensitivity of the assays ranged from 60.4% to 97.4% and specificity from 99.1% to 100% with substantial agreement according to Cohen’s kappa of 79.6%. Microsporidia DNA was detected in the reference material and the stool of the HIV patients but not in the stool of the travelers, migrants, and the Colombian indigenous people. Accuracy-adjusted prevalence was 5.8% (n = 78) for the study population as a whole. In conclusion, reliable detection of enteric disease-associated microsporidia in stool samples by real-time PCR could be demonstrated, but sensitivity between the compared microsporidia-specific real-time PCR assays varied.

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