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  • 1
    In: Pathogens, MDPI AG, Vol. 10, No. 8 ( 2021-08-19), p. 1053-
    Abstract: Cystoisospora (C.) belli is a coccidian parasite associated with acute or chronic gastroenteritis in immunocompromised patients. Dissatisfactory sensitivity of microscopy as the diagnostic standard approach has been described. Here, we comparatively evaluated two real-time PCRs targeting ribosomal RNA gene sequences of C. belli in stool in a test comparison without a reference standard applying latent class analysis. Therefore, 1000 stool samples from Ghanaian HIV (human immunodeficiency virus) patients (n = 905) as well as military returnees from the tropics (n = 95) were assessed by both assays in parallel. After the exclusion of 33 samples showing PCR inhibition, 29 and 33 positive results were recorded with the 5.8S rRNA gene/ITS-2 sequence PCR and the ITS-2 sequence PCR, respectively, resulting in an accuracy-adjusted prevalence of 3.2%. Nearly perfect agreement between both assays was indicated by Fleiss’ kappa of 0.933 with sensitivity and specificity of 92.8% and 100% as well as 100% and 99.8% for the 5.8S rRNA gene/ITS-2 sequence PCR and the ITS-2 sequence PCR, respectively. Both assays proved to be suitable for the diagnosis of C. belli in human stool samples with slightly better sensitivity of the ITS-2 sequence assay, while the 5.8S rRNA gene/ITS-2 sequence PCR may be considered for confirmatory testing.
    Type of Medium: Online Resource
    ISSN: 2076-0817
    Language: English
    Publisher: MDPI AG
    Publication Date: 2021
    detail.hit.zdb_id: 2695572-6
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  • 2
    In: Microorganisms, MDPI AG, Vol. 11, No. 5 ( 2023-05-17), p. 1313-
    Abstract: Potential etiological relevance for gastroenteric disorders including diarrhea has been assigned to Arcobacter butzleri. However, standard routine diagnostic algorithms for stool samples of patients with diarrhea are rarely adapted to the detection of this pathogen and so, A. butzleri is likely to go undetected unless it is specifically addressed, e.g., by applying pathogen-specific molecular diagnostic approaches. In the study presented here, we compared three real-time PCR assays targeting the genes hsp60, rpoB/C (both hybridization probe assays) and gyrA (fluorescence resonance energy transfer assay) of A. butzleri in a test comparison without a reference standard using a stool sample collection with a high pretest probability from the Ghanaian endemicity setting. Latent class analysis was applied with the PCR results obtained with a collection of 1495 stool samples showing no signs of PCR inhibition to assess the real-time PCR assays’ diagnostic accuracy. Calculated sensitivity and specificity were 93.0% and 96.9% for the hsp60-PCR, 100% and 98.2% for the rpoB/C-PCR, as well as 12.7% and 99.8% for the gyrA-PCR, respectively. The calculated A. butzleri prevalence within the assessed Ghanaian population was 14.7%. As indicated by test results obtained with high-titer spiked samples, cross-reactions of the hsp60-assay and rpoB/C-assay with phylogenetically related species such as A. cryaerophilus can occur but are less likely with phylogenetically more distant species like, e.g., A. lanthieri. In conclusion, the rpoB/C-assay showed the most promising performance characteristics as the only assay with sensitivity 〉 95%, albeit associated with a broad 95%-confidence interval. In addition, this assay showed still-acceptable specificity of 〉 98% in spite of the known cross-reactivity with phylogenetically closely related species such as A. cryaerophilus. If higher certainty is desired, the gyrA-assay with specificity close to 100% can be applied for confirmation testing with samples showing positive rpoB/C-PCR results. However, in case of a negative result in the gyrA-assay, this cannot reliably exclude the detection of A. butzleri in the rpoB/C-assay due to the gyrA-assay’s very low sensitivity.
    Type of Medium: Online Resource
    ISSN: 2076-2607
    Language: English
    Publisher: MDPI AG
    Publication Date: 2023
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  • 3
    In: Pathogens, MDPI AG, Vol. 11, No. 7 ( 2022-07-02), p. 760-
    Abstract: Both Schistosoma spp. (species) and Leishmania spp. are prevalent in Ghana in West Africa. However, little is known about their local occurrence in immunocompromised individuals. In the study presented here, the real-time PCR-(polymerase chain reaction-)based screening for repetitive DNA (deoxyribonucleotide acid) sequences from the genomes of Leishmania (L.) spp. and Schistosoma (S.) spp. was performed in the serum of HIV-(human immunodeficiency virus-)infected Ghanaian patients. In 1083 assessed serum samples from HIV-positive and HIV-negative Ghanian patients, Leishmania spp.-specific DNA was not detected, while the diagnostic accuracy-adjusted prevalence estimation suggested a 3.6% prevalence of the S. mansoni complex and a 0.5% prevalence of the S. haematobium complex. Associations of schistosomiasis with younger age, as well as with the male sex, could be shown but not with an HIV status. Weakly significant signals for the associations of schistosomiasis with an increased viral load, reduced CD4+ (CD = cluster of differentiation) T cell count, and a reduced CD4+/CD8+ ratio could be observed but was inconsistently lost in the case of the stratification on the species complex level. So, it is concluded that factors other than HIV status are more likely to have influenced the occurrence of Schistosoma spp. infections in the assessed Ghanaian patients. Potential associations between HIV infection-associated factors, such as the viral load and the immune status of the patients, for which weak signals were observed in this hypothesis-forming retrospective assessment, should be confirmed by prospective, sufficiently powered investigations.
    Type of Medium: Online Resource
    ISSN: 2076-0817
    Language: English
    Publisher: MDPI AG
    Publication Date: 2022
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  • 4
    In: Pathogens, MDPI AG, Vol. 12, No. 8 ( 2023-07-30), p. 999-
    Abstract: Medical complications during pregnancy have been frequently reported from Western Africa with a particular importance of infectious complications. Placental tissue can either become the target of infectious agents itself, such as, e.g., in the case of urogenital schistosomiasis, or be subjected to contamination with colonizing or infection-associated microorganisms of the cervix or the vagina during vaginal delivery. In the retrospective cross-sectional assessment presented here, the quantitative dimension of infection or colonization with selected resistant or pathogenic bacteria and parasites was regionally assessed. To do so, 274 collected placental tissues from Ivory Coastal and Ghanaian women were subjected to selective growth of resistant bacteria, as well as to molecular screening for beta-lactamase genes, Schistosoma spp. and selected bacterial causative agents of sexually transmitted infections (STI). Panton–Valentine-negative methicillin-resistant Staphylococcus aureus (MRSA) was grown from 1.8% of the tissue samples, comprising the spa types t008 and t688, as well as the newly detected ones, t12101 (n = 2) and t12102. While the culture-based recovery of resistant Enterobacterales and nonfermentative rod-shaped Gram-negative bacteria failed, molecular assessments confirmed beta-lactamase genes in 31.0% of the samples with multiple detections of up to four resistance genes per sample and blaCTX-M, blaIMP, blaGES, blaVIM, blaOXA-58-like, blaNDM, blaOXA-23-like, blaOXA-48-like and blaKPC occurring in descending order of frequency. The beta-lactamase genes blaOXA-40/24-like, blaNMC_A/IMI, blaBIC, blaSME, blaGIM and blaDIM were not detected. DNA of the urogenital schistosomiasis-associated Schistosoma haematobium complex was recorded in 18.6% of the samples, but only a single positive signal for S. mansoni with a high cycle-threshold value in real-time PCR was found. Of note, higher rates of schistosomiasis were observed in Ghana (54.9% vs. 10.3% in Ivory Coast) and Cesarean section was much more frequent in schistosomiasis patients (61.9% vs. 14.8% in women without Schistosoma spp. DNA in the placenta). Nucleic acid sequences of nonlymphogranuloma-venereum-associated Chlamydia trachomatis and of Neisseria gonorrhoeae were recorded in 1.1% and 1.9% of the samples, respectively, while molecular attempts to diagnose Treponema pallidum and Mycoplasma genitalium did not lead to positive results. Molecular detection of Schistosoma spp. or STI-associated pathogens was only exceptionally associated with multiple resistance gene detections in the same sample, suggesting epidemiological distinctness. In conclusion, the assessment confirmed considerable prevalence of urogenital schistosomiasis and resistant bacterial colonization, as well as a regionally expected abundance of STI-associated pathogens. Continuous screening offers seem advisable to minimize the risks for the pregnant women and their newborns.
    Type of Medium: Online Resource
    ISSN: 2076-0817
    Language: English
    Publisher: MDPI AG
    Publication Date: 2023
    detail.hit.zdb_id: 2695572-6
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  • 5
    In: Diagnostics, MDPI AG, Vol. 12, No. 4 ( 2022-04-16), p. 1007-
    Abstract: Prior to the implementation of new diagnostic techniques, a thorough evaluation is mandatory in order to ensure diagnostic reliability. If positive samples are scarcely available, however, such evaluations can be difficult to perform. Here, we evaluated four SeeGene Allplex real-time PCR assays amplifying a total of 28 bacteria, microsporidal and parasitic nucleic acid sequence targets in human stool samples in a multicentric approach. In the assessments with strongly positive samples, sensitivity values ranging between 13% and 100% were recorded for bacteria, between 0% and 100% for protozoa and between 7% and 100% for helminths and microsporidia; for the weakly positive samples, the recorded sensitivity values for bacteria ranged from 0% to 100%; for protozoa, from 0% to 40%; and for helminths and microsporidia, from 0% to 53%. For bacteria, the recorded specificity was in the range between 87% and 100%, while a specificity of 100% was recorded for all assessed PCRs targeting parasites and microsporidia. The intra- and inter-assay variations were generally low. Specifically for some helminth species, the sensitivity could be drastically increased by applying manual nucleic acid extraction instead of the manufacturer-recommended automatic procedure, while such effects were less obvious for the bacteria and protozoa. In summary, the testing with the chosen positive control samples showed varying degrees of discordance between the evaluated Allplex assays and the applied in-house reference assays associated with higher cycle threshold values in the Allplex assays, suggesting that samples with very low pathogen densities might be missed. As the targeted species can occur as harmless colonizers in the gut of individuals in high-endemicity settings as well, future studies should aim at assessing the clinical relevance of the latter hint.
    Type of Medium: Online Resource
    ISSN: 2075-4418
    Language: English
    Publisher: MDPI AG
    Publication Date: 2022
    detail.hit.zdb_id: 2662336-5
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  • 6
    In: Pathogens, MDPI AG, Vol. 11, No. 2 ( 2022-02-17), p. 256-
    Abstract: In the original publication [...]
    Type of Medium: Online Resource
    ISSN: 2076-0817
    Language: English
    Publisher: MDPI AG
    Publication Date: 2022
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  • 7
    In: Pathogens, MDPI AG, Vol. 10, No. 9 ( 2021-09-02), p. 1131-
    Abstract: As qualified microscopy of enteric parasitoses as defined by high diagnostic accuracy is difficult to maintain in non-endemic areas due to scarce opportunities for practicing with positive sample materials, molecular diagnostic options provide less investigator-dependent alternatives. Here, we compared three molecular targets for the real-time PCR-based detection of Cryptosporidium spp. From a population of 1000 individuals comprising both Ghanaian HIV (human immunodeficiency virus) patients and military returnees after deployment in the tropics, stool samples were assessed for Cryptosporidium spp. by real-time PCR targeting the small subunit ribosomal RNA (SSU rRNA) gene, the Cryptosporidium oocyst wall (COWP) gene, and the DnaJ-like protein gene (DnaJ), respectively. In declining order, sensitivity of 100% for the SSU rRNA gene PCR, 90.0% for the COWP PCR and 88.8% for the DnaJ PCR, respectively, as well as specificity of 99.6% for the COWP PCR and 96.9% for both the SSU rRNA gene PCR and the DnaJ PCR, respectively, were recorded. Substantial agreement (kappa value 0.663) between the three assays was observed. Further, an accuracy-adjusted Cryptosporidium spp. prevalence of 6.0% was calculated for the study population. In conclusion, none of the assessed real-time PCR assays were associated with perfect test accuracy. However, a combination of highly sensitive SSU rRNA gene PCR for screening purposes and more specific COWP PCR for confirmatory testing should allow reliable diagnosis of Cryptosporidium spp. in stool samples even in low prevalence settings.
    Type of Medium: Online Resource
    ISSN: 2076-0817
    Language: English
    Publisher: MDPI AG
    Publication Date: 2021
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  • 8
    In: Pathogens, MDPI AG, Vol. 11, No. 2 ( 2022-01-26), p. 165-
    Abstract: Diagnostic real-time PCR for the detection of Cyclospora cayetanensis in human stool samples has been applied for two decades. However, recent comparative assessments between in-house and commercial assays suggested room for improvement regarding the agreement of positive signals of the applied real-time PCRs. In order to assess the effect of the choice of the target sequence, 3 inhouse real time PCR assays targeting the 18S rRNA gene (n = 2, one of them later referred to as SSU rRNA gene assay to avoid confusion) and the hsp70 gene of C. cayetanensis were compared in a head-to-head comparison with 905 samples with high pretest probability for C. cayetanensis infections from Ghanaian HIV patients in a test comparison without a reference standard. Only slight agreement kappa of 0.095 was observed. In the assays targeting the SSU rRNA gene, the 18S rRNA gene, and hsp70, positive signals were recorded in 63, 45, and 0 instances, respectively, with latent class analysis-based estimation of sensitivity of 32.2%, 23.3%, 0% as well as of specificity of 99.7%, 99.9% and 100%, respectively. High cycle threshold values with an average of about 35 indicated low quantities of target DNA in the samples with similar Ct values in concordantly and discordantly positive samples. In conclusion, the study suggested target-gene-specific differences in the diagnostic accuracy of real-time PCR-based diagnosis of C. cayetanensis as well as an ongoing need for further standardization of this diagnostic approach.
    Type of Medium: Online Resource
    ISSN: 2076-0817
    Language: English
    Publisher: MDPI AG
    Publication Date: 2022
    detail.hit.zdb_id: 2695572-6
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  • 9
    In: European Journal of Microbiology and Immunology, Akademiai Kiado Zrt., ( 2023-09-26)
    Abstract: Mansonellosis is a widely neglected helminth disease which is predominantly observed in tropical regions. This study was conducted to assess potential associations of the prevalence of circulating Mansonella perstans -specific cell-free DNA in human serum and HIV infection in Ghanaian individuals. Methods For this purpose, serum samples obtained from Ghanaian HIV-patients ( n = 989) and non-HIV-infected Ghanaian control individuals ( n = 91) were subjected to real-time PCR targeting the ITS-(internal transcribed spacer-)2 sequence of M . perstans and Mansonella sp. Deux. Results Mansonella -specific cell-free DNA was detected in serum samples of only 2 HIV-positive and 0 HIV-negative individuals, making any reliable conclusions on potential associations between HIV and mansonellosis in tropical Ghana unfeasible. Conclusions Future epidemiological studies on hypothetical associations between mansonellosis and HIV infections should focus more specifically on high-endemicity settings for both Mansonella spp.-infections and HIV-infections, include higher case numbers and be based on real-time PCR from whole blood rather than from serum, in which only circulating parasite DNA but no more cell-bound parasite DNA can be detected. However, the study did not show associations of HIV infections in Ghanaian individuals with Mansonella worm loads high enough to detect cell-free Mansonella DNA in serum by PCR.
    Type of Medium: Online Resource
    ISSN: 2062-509X , 2062-8633
    Language: Unknown
    Publisher: Akademiai Kiado Zrt.
    Publication Date: 2023
    detail.hit.zdb_id: 2652327-9
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  • 10
    In: Microorganisms, MDPI AG, Vol. 10, No. 7 ( 2022-07-13), p. 1407-
    Abstract: Background: There is a paucity of information on the contemporary burden, disease patterns, and immunological profile of people living with HIV who are co-infected with C. cayetanensis in the post-antiretroviral therapy era. Methods: For this cross-sectional study, stool samples of 640 HIV-positive and 83 HIV-negative individuals in Ghana were tested for C. cayetanensis. Additionally, sociodemographic parameters, clinical symptoms, medical drug intake, and immunological parameters were assessed. Results: The prevalence of C. cayetanensis was 8.75% (n = 56) in HIV-positive and 1.20% (n = 1) in HIV-negative participants (p = 0.015). Within the group of HIV-positive participants, the prevalence reached 13.6% in patients with CD4+ T cell counts below 200 cells/µl. Frequencies of the clinical manifestations of weight loss and diarrheal disease were significantly higher in patients with C. cayetanensis compared to those without co-infection (36.36% vs. 22.59%, p = 0.034 and 20.00% vs. 4.90%, p 〈 0.001, respectively). The expression of markers of immune activation and exhaustion of T lymphocyte sub-populations was significantly elevated in patients colonized with C. cayetanensis. Conclusions: In the modern post-combined antiretroviral therapy (cART) era, the acquisition of C. cayetanensis among PLWH in Ghana is driven largely by the immunosuppression profile characterized by high expression of markers of immune activation and immune exhaustion.
    Type of Medium: Online Resource
    ISSN: 2076-2607
    Language: English
    Publisher: MDPI AG
    Publication Date: 2022
    detail.hit.zdb_id: 2720891-6
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