scholarly journals High Frequency of Cryptosporidium hominis Infecting Infants Points to A Potential Anthroponotic Transmission in Maputo, Mozambique

Pathogens ◽  
2021 ◽  
Vol 10 (3) ◽  
pp. 293
Author(s):  
Idalécia Cossa-Moiane ◽  
Hermínio Cossa ◽  
Adilson Fernando Loforte Bauhofer ◽  
Jorfélia Chilaúle ◽  
Esperança Lourenço Guimarães ◽  
...  
Keyword(s):  
Public Hospitals ◽  
Diagnostic Methods ◽  
Molecular Diagnostic ◽  
P Value ◽  
Cryptosporidium Hominis ◽  
Cryptosporidium Spp ◽  
Pcr Rflp ◽  
Maputo City ◽  
Stool Samples ◽  
Polymerase Chain

Cryptosporidium is one of the most important causes of diarrhea in children less than 2 years of age. In this study, we report the frequency, risk factors and species of Cryptosporidium detected by molecular diagnostic methods in children admitted to two public hospitals in Maputo City, Mozambique. We studied 319 patients under the age of five years who were admitted due to diarrhea between April 2015 and February 2016. Single stool samples were examined for the presence of Cryptosporidium spp. oocysts, microscopically by using a Modified Ziehl–Neelsen (mZN) staining method and by using Polymerase Chain Reaction and Restriction Fragment Length Polymorphism (PCR-RFLP) technique using 18S ribosomal RNA gene as a target. Overall, 57.7% (184/319) were males, the median age (Interquartile range, IQR) was 11.0 (7–15) months. Cryptosporidium spp. oocysts were detected in 11.0% (35/319) by microscopy and in 35.4% (68/192) using PCR-RFLP. The most affected age group were children older than two years, [adjusted odds ratio (aOR): 5.861; 95% confidence interval (CI): 1.532–22.417; p-value < 0.05]. Children with illiterate caregivers had higher risk of infection (aOR: 1.688; 95% CI: 1.001–2.845; p-value < 0.05). An anthroponotic species C. hominis was found in 93.0% (27/29) of samples. Our findings demonstrated that cryptosporidiosis in children with diarrhea might be caused by anthroponomic transmission.

Unusual cryptosporidiosis cases in Swedish patients: extended molecular characterization ofCryptosporidium viatorumandCryptosporidiumchipmunk genotype I

Parasitology ◽  
2013 ◽  
Vol 140 (14) ◽  
pp. 1735-1740 ◽  
Author(s):  
MARIANNE LEBBAD ◽  
JESSICA BESER ◽  
MONA INSULANDER ◽  
LILLEMOR KARLSSON ◽  
JENS G. MATTSSON ◽  
...  
Keyword(s):  
Molecular Characterization ◽  
Cryptosporidium Parvum ◽  
Geographical Variation ◽  
Indian Subcontinent ◽  
Common Species ◽  
Diagnostic Methods ◽  
Molecular Diagnostic ◽  
Cryptosporidium Hominis ◽  
Genotype I ◽  
Protein Locus

SUMMARYMost human cases of cryptosporidiosis are caused byCryptosporidium parvumorCryptosporidium hominis, but the use of molecular diagnostic methods has revealed that several other less common species or genotypes can also be involved. Here, we describe two unusual causes of cryptosporidiosis, one being the recently described speciesCryptosporidium viatorumand the otherCryptosporidiumchipmunk genotype I. Two Swedish patients who were infected withC. viatorumhad travelled to Kenya and Guatemala, respectively, and two others had been infected withCryptosporidiumchipmunk genotype I in Sweden. None of these four patients were immunocompromised, and all four showed classical symptoms of cryptosporidiosis. We performed extensive molecular characterization, including analysis of four loci. The twoC. viatorumisolates were found to differ slightly at the 70-kDa heat shock protein locus, which may indicate a local geographical variation in this species that has previously been described exclusively on the Indian subcontinent.

scholarly journals Detection of LytA Genes in Streptococcus pneumoniae Isolated from sputum pneumonia patients

Biomedika ◽  
2020 ◽  
Vol 13 (1) ◽  
pp. 23-30
Author(s):  
Mustika Sari Hutabarat ◽  
Firdaus Hamid ◽  
Irawaty Djaharuddin ◽  
Alfian Zainuddin ◽  
Rossana Agus ◽  
...  
Keyword(s):  
Streptococcus Pneumoniae ◽  
False Negative ◽  
Pneumococcal Infection ◽  
Diagnostic Methods ◽  
Molecular Diagnostic ◽  
Clinical Samples ◽  
Biochemical Tests ◽  
Negative Results ◽  
False Negative Results ◽  
Polymerase Chain

Streptococcus pneumoniae (pneumococcus) is a Gram-positive facultative anaerobic bacterium that is a major cause of morbidity and mortality worldwide. But the lack of reporting of disease by this bacterium in Indonesia, one of the causes is because the diagnosis of pneumococcal infection is often clinically not typical and conventional methods which are still the standard gold method often give false-negative results. So the purpose of this study was to evaluate the performance of culture and molecular diagnostic methods using the Polymerase Chain Reaction (PCR) technique in detecting Streptococcus pneumoniae in sputum clinical samples using the Autolysin (LytA) gene which is a virulence factor of this bacterium. 57 isolates from 60 samples were confirmed as Streptococcus sp through microscopic identification, culture, and biochemical tests. Then the sensitivity test with an optochin test of 9 (9%) compared the results descriptively with the PCR technique using the Autolysin A (LytA) gene which was obtained more sensitive by 15 (25%).

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.

scholarly journals Comparison of multi-parallel qPCR and Kato-Katz for detection of soil-transmitted helminth infection among children in rural Bangladesh

2019 ◽  
Author(s):  
Jade Benjamin-Chung ◽  
Nils Pilotte ◽  
Ayse Ercumen ◽  
Jessica R. Grant ◽  
Jacqueline R.M.A. Maasch ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Sensitivity And Specificity ◽  
Diagnostic Method ◽  
Quantitative Polymerase Chain Reaction ◽  
Infection Intensity ◽  
Molecular Diagnostic ◽  
Chain Reaction ◽  
Rural Bangladesh ◽  
Stool Samples ◽  
Polymerase Chain

AbstractAn active area of research investigates whether soil-transmitted helminths (STH) can be locally eliminated in endemic settings. In such settings, highly sensitive diagnostics are needed to detect STH infection. We compared double-slide Kato-Katz, the most commonly used copromicroscopic detection method, to multi-parallel quantitative polymerase chain reaction (qPCR) in 2,800 stool samples from children 2-12 years in rural Bangladesh. We estimated the sensitivity and specificity of each diagnostic using Bayesian latent class analysis. Compared to Kato-Katz, STH prevalence using qPCR was almost 3-fold higher for hookworm species and nearly 2-fold higher forTrichuris trichiura.Ascaris lumbricoidesprevalence was lower using qPCR, and 26% of samples classified asA. lumbricoidespositive by Kato-Katz were negative by qPCR. Amplicon sequencing of the 18S rDNA from 10 samples confirmed thatA. lumbricoideswas absent in samples classified as positive by Kato-Katz and negative by qPCR. The sensitivity of Kato-Katz was 49% forA. lumbricoides, 32% for hookworm, and 52% forT. trichiura; the sensitivity of qPCR was 79% forA. lumbricoides, 93% for hookworm, and 90% forT. trichiura. Specificity was ≥ 97% for both tests for all STH except for Kato-Katz forA. lumbricoides(specificity = 68%). There were moderate negative, monotonic correlations between qPCR cycle quantification values and eggs per gram quantified by Kato-Katz. While it is widely assumed that Kato-Katz has few false positives, our results indicate otherwise. Our findings suggest that qPCR is more appropriate than Kato-Katz in low intensity infection settings because of its higher sensitivity and specificity.Author summarySoil-transmitted helminth infections (STH) (e.g.,Ascaris, hookworm,Trichuris) contribute to a large burden of disease among children in low- and middle-income countries. There is increasing interest in implementing large-scale deworming programs to eliminate STH in certain settings. Efforts to monitor whether local elimination has occurred require sensitive diagnostic tests that will not miss positive cases. Kato-Katz, a microscopy-based diagnostic test, has commonly been used to identify STH eggs in stool, but in settings where infection intensity is low, this method frequently misses positive samples because it requires visual identification of small numbers of eggs, and eggs may degrade prior to visualization. Quantitative polymerase chain reaction (qPCR) is a molecular diagnostic method that may miss fewer infections because it identifies STH DNA in stool, which can be detected in very small quantities and is less likely to degrade. This study compared the performance of Kato-Katz and qPCR using 2,800 stool samples from children aged 2-12 years in rural Bangladesh. qPCR detected substantially more hookworm andTrichurisinfections than Kato-Katz. 26% of samples were classified asAscarispositive by Kato-Katz and negative by qPCR. We conclude that qPCR is a more appropriate diagnostic method than Kato-Katz in low infection intensity settings.

scholarly journals Molecular diagnosis of Anaplasma marginale in cattle: quantitative evaluation of a real-time PCR (Polymerase Chain Reaction) based on msp5 gene

2014 ◽  
Vol 34 (1) ◽  
pp. 29-33 ◽  
Author(s):  
Gisele M. Bacanelli ◽  
Carlos A. N. Ramos ◽  
Flábio R. Araújo
Keyword(s):  
Polymerase Chain Reaction ◽  
Real Time ◽  
Real Time Pcr ◽  
Anaplasma Marginale ◽  
Diagnostic Methods ◽  
Molecular Diagnostic ◽  
Chain Reaction ◽  
The Real ◽  
End Point ◽  
Polymerase Chain

The rickettsia Anaplasma marginale is considered the main agent of bovine anaplasmosis. Due the nonspecific clinical signs of the anaplasmosis, the diagnosis of infection depends of laboratory confirmation. In recent years, molecular diagnostic methods have been used to detect A. marginale in cattle. However, the existence of a large number of assays of different sensitivity and cost makes the choice of an appropriate test difficult. In the present study, a real-time Polymerase Chain Reaction (PCR) based on the msp5 target gene was quantitatively assessed and compared to an end point PCR. Both reactions were subjected to sensitivity and specificity evaluation using plasmid DNA and samples from cattle experimentally infected with A. marginale. A comparative field trial of the tests was carried out using samples of cattle from a stable enzootic area for A. marginale. The real-time PCR showed a higher sensitivity than the end point PCR. This reaction (i.e. real-time PCR) was able to detect one copy of the msp5 gene in 100 ηg of plasmidial DNA, and more than 80% of its results were positive among experimentally infected animals seven days after infection. In addition, based on in silico analysis, the real-time PCR evaluated in the present study appears to be useful for the detection of A. ovis.

scholarly journals Nucleic Acid Testing of SARS-CoV-2

2021 ◽  
Vol 22 (11) ◽  
pp. 6150
Author(s):  
Hee-Min Yoo ◽  
Il-Hwan Kim ◽  
Seil Kim
Keyword(s):  
Polymerase Chain Reaction ◽  
Nucleic Acid ◽  
Reverse Transcription ◽  
Diagnostic Methods ◽  
Detection Methods ◽  
Molecular Diagnostic ◽  
Nucleic Acid Testing ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Polymerase Chain

The coronavirus disease 2019 (COVID-19) has caused a large global outbreak. It is accordingly important to develop accurate and rapid diagnostic methods. The polymerase chain reaction (PCR)-based method including reverse transcription-polymerase chain reaction (RT-PCR) is the most widely used assay for the detection of SARS-CoV-2 RNA. Along with the RT-PCR method, digital PCR has emerged as a powerful tool to quantify nucleic acid of the virus with high accuracy and sensitivity. Non-PCR based techniques such as reverse transcription loop-mediated isothermal amplification (RT-LAMP) and reverse transcription recombinase polymerase amplification (RT-RPA) are considered to be rapid and simple nucleic acid detection methods and were reviewed in this paper. Non-conventional molecular diagnostic methods including next-generation sequencing (NGS), CRISPR-based assays and nanotechnology are improving the accuracy and sensitivity of COVID-19 diagnosis. In this review, we also focus on standardization of SARS-CoV-2 nucleic acid testing and the activity of the National Metrology Institutes (NMIs) and highlight resources such as reference materials (RM) that provide the values of specified properties. Finally, we summarize the useful resources for convenient COVID-19 molecular diagnostics.

scholarly journals Potential Application of PCR Based Molecular Methods in Fish Pathogen Identification: A Review

2021 ◽  
Vol 22 (1) ◽  
Author(s):  
Md. Ali Arman Ador ◽  
Md. Shameul Haque ◽  
Sulav Indra Paul ◽  
Jui Chakma ◽  
Rakib Ehsan ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Molecular Techniques ◽  
Diagnostic Methods ◽  
Molecular Diagnostic ◽  
Resistant Bacteria ◽  
Fish Disease ◽  
Fish Pathogen ◽  
Pathogen Identification ◽  
Chain Reaction ◽  
Polymerase Chain

Molecular biology developments have led to fast growth in new methods for fish disease diagnosis. Molecular diagnostic methods are rapid and more specific, more sensitive than the culture of pathogens, serology, histology, and biochemical methods which are traditionally utilized to identify causative agent fish disease. Molecular diagnostic methods are valuable for detecting specific pathogens that are difficult to culture in vitro or require a long cultivation period and it significantly more rapid in providing results compared to culture. It enables earlier informed decision-making and rapid diagnosis of bacteremia, particularly for low levels of bacteria in specimens. Molecular techniques which have the major significance are mainly PCR-based molecular diagnostic methods including Polymerase Chain Reaction (PCR), Real-Time Polymerase Chain Reaction (RT-PCR), Multiplex Polymerase Chain Reaction (multiplexPCR), and Random Amplified Polymorphic DNA (RAPD). These have been increasingly utilized to diagnose fish disease for the last recent years. Molecular diagnostic methods can detect pathogens from asymptomatic fish, so disease outbreaks could be prevented. As a consequence, antibiotic treatment can be reduced and the development of antibiotic-resistant bacteria can be eliminated. In this review paper, we attempt to summarize the potentiality of PCR-based molecular diagnostic methods and their application in fish pathogen identification.

Pilocytic Astrocytoma: A Review of General, Clinical, and Molecular Characteristics

2020 ◽  
Vol 35 (12) ◽  
pp. 852-858
Author(s):  
Débora Salles ◽  
Gabriela Laviola ◽  
Andréa Cristina de Moraes Malinverni ◽  
João Norberto Stávale
Keyword(s):  
Tumor Development ◽  
Diagnostic Methods ◽  
Braf V600e ◽  
Molecular Diagnostic ◽  
Molecular Characteristics ◽  
Primary Tumors ◽  
Molecular Alterations ◽  
Pilocytic Astrocytomas ◽  
Polymerase Chain

Pilocytic astrocytomas are the primary tumors most frequently found in children and adolescents, accounting for approximately 15.6% of all brain tumors and 5.4% of all gliomas. They are mostly found in infratentorial structures such as the cerebellum and in midline cerebral structures such as the optic nerve, hypothalamus, and brain stem. The present study aimed to list the main characteristics about this tumor, to better understand the diagnosis and treatment of these patients, and was conducted on search of the published studies available in NCBI, PubMed, MEDLINE, Scielo, and Google Scholar. It was possible to define the main histologic findings observed in these cases, such as mitoses, necrosis, and Rosenthal fibers. We described the locations usually most affected by tumor development, and this was associated with the most frequent clinical features. The comparison between the molecular diagnostic methods showed great use of fluorescent in situ hybridization, polymerase chain reaction (PCR), and reverse transcriptase–PCR, important techniques for the detection of BRAF V600E mutation and BRAF-KIAA1549 fusion, characteristic molecular alterations in pilocytic astrocytomas.

scholarly journals Performance of the Kato-Katz method and real time polymerase chain reaction for the diagnosis of soil-transmitted helminthiasis in the framework of a randomised controlled trial: treatment efficacy and day-to-day variation

2020 ◽  
Vol 13 (1) ◽  
Author(s):  
Ladina Keller ◽  
Chandni Patel ◽  
Sophie Welsche ◽  
Tobias Schindler ◽  
Eveline Hürlimann ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Treatment Efficacy ◽  
Controlled Trial ◽  
Diagnostic Methods ◽  
Diagnostic Tools ◽  
Test Results ◽  
Chain Reaction ◽  
Stool Samples ◽  
Polymerase Chain ◽  
Cure Rates

Abstract Background Accurate, scalable and sensitive diagnostic tools are crucial in determining prevalence of soil-transmitted helminths (STH), assessing infection intensities and monitoring treatment efficacy. However, assessments on treatment efficacy comparing traditional microscopic to newly emerging molecular approaches such as quantitative Polymerase Chain Reaction (qPCR) are scarce and hampered partly by lack of an established diagnostic gold standard. Methods We compared the performance of the copromicroscopic Kato-Katz method to qPCR in the framework of a randomized controlled trial on Pemba Island, Tanzania, evaluating treatment efficacy based on cure rates of albendazole monotherapy versus ivermectin-albendazole against Trichuris trichiura and concomitant STH infections. Day-to-day variability of both diagnostic methods was assessed to elucidate reproducibility of test results by analysing two stool samples before and two stool samples after treatment of 160 T. trichiura Kato-Katz positive participants, partially co-infected with Ascaris lumbricoides and hookworm, per treatment arm (n = 320). As negative controls, two faecal samples of 180 Kato-Katz helminth negative participants were analysed. Results Fair to moderate correlation between microscopic egg count and DNA copy number for the different STH species was observed at baseline and follow-up. Results indicated higher sensitivity of qPCR for all three STH species across all time points; however, we found lower test result reproducibility compared to Kato-Katz. When assessed with two samples from consecutive days by qPCR, cure rates were significantly lower for T. trichiura (23.2 vs 46.8%), A. lumbricoides (75.3 vs 100%) and hookworm (52.4 vs 78.3%) in the ivermectin-albendazole treatment arm, when compared to Kato-Katz. Conclusions qPCR diagnosis showed lower reproducibility of test results compared to Kato-Katz, hence multiple samples per participant should be analysed to achieve a reliable diagnosis of STH infection. Our study confirms that cure rates are overestimated using Kato-Katz alone. Our findings emphasize that standardized and accurate molecular diagnostic tools are urgently needed for future monitoring within STH control and/or elimination programmes.

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