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15 pages, 6972 KiB

Open AccessArticle

Identification of Intestinal Microbial Community in Gallstone Patients with Metagenomic Next-Generation Sequencing

by Li Ding, Su Wang, Wenrong Jiang, Yingxin Miao, Wenjian Liu, Feng Yang, Jinghao Zhang, Wenjing Chi, Tao Liu, Yue Liu, Shiwen Wang, Yanmei Zhang and Hu Zhao

Diagnostics 2023, 13(16), 2712; https://doi.org/10.3390/diagnostics13162712 - 21 Aug 2023

Cited by 1 | Viewed by 1006

Abstract

Gallstone disease (GD) is one of the most common gastrointestinal diseases worldwide. Nowadays, intestinal microbiota are thought to play important roles in the formation of gallstones. In our study, human fecal samples were extracted for metagenomic next-generation sequencing (mNGS) on the Illumina HiSeq [...] Read more.

Gallstone disease (GD) is one of the most common gastrointestinal diseases worldwide. Nowadays, intestinal microbiota are thought to play important roles in the formation of gallstones. In our study, human fecal samples were extracted for metagenomic next-generation sequencing (mNGS) on the Illumina HiSeq platform, followed by bioinformatics analyses. Our results showed that there was a particular intestinal micro-ecosystem in GD patients. In contrast to healthy people, the sequences of Bacteroidetes, Bacteroides and Thetaiotaomicron were obviously more abundant in GD patients at phylum, genus and species levels, respectively. On the other hand, the glycan metabolism and drug resistance, especially for the β-lactams, were the most profound functions of gut microbes in GD patients compared to those in normal subjects. Furthermore, a correlation analysis drew out that there existed a significant relationship between the serum levels of biochemical indicators and abundances of intestinal microbes in GD patients. Our results illuminate both the composition and functions of intestinal microbiota in GD patients. All in all, our study can broaden the insight into the potential mechanism of how gut microbes affect the progression of gallstones to some extent, which may provide potential targets for the prevention, diagnosis or treatment of GD. Full article

(This article belongs to the Special Issue Laboratory Diagnosis in Microbial Diseases)

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11 pages, 260 KiB

Open AccessArticle

Diagnosis of Latent Tuberculosis Infection in Hemodialysis Patients: TST versus T-SPOT.TB

by Umut Devrim Binay, Ali Veysel Kara, Faruk Karakeçili and Orçun Barkay

Diagnostics 2023, 13(14), 2369; https://doi.org/10.3390/diagnostics13142369 - 14 Jul 2023

Viewed by 894

Abstract

Hemodialysis (HD) patients should be screened for latent tuberculosis (TB) infection. We aimed to determine the frequency of latent TB infection in HD patients and to compare the effectiveness of the tests used. The files of 56 HD patients followed between 1 January [...] Read more.

Hemodialysis (HD) patients should be screened for latent tuberculosis (TB) infection. We aimed to determine the frequency of latent TB infection in HD patients and to compare the effectiveness of the tests used. The files of 56 HD patients followed between 1 January 2021 and 1 October 2022 were retrospectively analyzed. Demographic data, the presence of the Bacillus Calmette-Guerin (BCG) vaccine, whether or not the patients had previously received treatment for TB before, the status of encountering a patient with active TB of patients over 18 years of age, without active tuberculosis and who had a T-SPOT.TB test or a Tuberculin Skin Test (TST) were obtained from the patient files. The presence of previous TB in a posterior–anterior (PA) chest X-ray was obtained by evaluating PA chest X-rays taken routinely. Of the patients, 60.7% (n = 34) were male and their mean age was 60.18 ± 14.85 years. The mean duration of dialysis was 6.43 ± 6.03 years, and 76.8% (n = 43) had 2 BCG scars. The T-SPOT.TB test was positive in 32.1% (n = 18). Only 20 patients (35.7%) had a TST and all had negative results. While the mean age of those with positive T-SPOT.TB results was higher (p = 0.003), the time taken to enter HD was shorter (p = 0.029). T-SPOT.TB test positivity was higher in the group that had encountered active TB patients (p = 0.033). However, no significant difference was found between T-SPOT.TB results according to BCG vaccine, albumin, urea and lymphocyte levels. Although T-SPOT.TB test positivity was higher in patients with a previous TB finding in a PA chest X-ray, there was no statistically significant difference (p = 0.093). The applicability of the TST in the diagnosis of latent TB infection in HD patients is difficult and it is likely to give false-negative results. The T-SPOT.TB test is not affected by the BCG vaccine and immunosuppression. Therefore, using the T-SPOT.TB test would be a more appropriate and practical approach in the diagnosis of latent TB in HD patients. Full article

(This article belongs to the Special Issue Laboratory Diagnosis in Microbial Diseases)

19 pages, 1278 KiB

Open AccessArticle

Performance Evaluation of VIDAS^® Diagnostic Assays Detecting Anti-Chikungunya Virus IgM and IgG Antibodies: An International Study

by Geovana M. Pereira, Erika R. Manuli, Laurie Coulon, Marina F. Côrtes, Mariana S. Ramundo, Loïc Dromenq, Audrey Larue-Triolet, Frédérique Raymond, Carole Tourneur, Carolina dos Santos Lázari, Patricia Brasil, Ana Maria Bispo de Filippis, Glaucia Paranhos-Baccalà, Alice Banz and Ester C. Sabino

Diagnostics 2023, 13(13), 2306; https://doi.org/10.3390/diagnostics13132306 - 07 Jul 2023

Viewed by 1759

Abstract

Chikungunya (CHIK) is a debilitating mosquito-borne disease with an epidemiology and early clinical symptoms similar to those of other arboviruses-triggered diseases such as dengue or Zika. Accurate and rapid diagnosis of CHIK virus (CHIKV) infection is therefore challenging. This international study evaluated the [...] Read more.

Chikungunya (CHIK) is a debilitating mosquito-borne disease with an epidemiology and early clinical symptoms similar to those of other arboviruses-triggered diseases such as dengue or Zika. Accurate and rapid diagnosis of CHIK virus (CHIKV) infection is therefore challenging. This international study evaluated the performance of the automated VIDAS^® anti-CHIKV IgM and IgG assays compared to that of manual competitor IgM and IgG ELISA for the detection of anti-CHIKV IgM and IgG antibodies in 660 patients with suspected CHIKV infection. Positive and negative agreements of the VIDAS^® CHIKV assays with ELISA ranged from 97.5% to 100.0%. The sensitivity of the VIDAS^® CHIKV assays evaluated in patients with a proven CHIKV infection confirmed reported kinetics of anti-CHIKV IgM and IgG response, with a positive detection of 88.2–100.0% for IgM ≥ 5 days post symptom onset and of 100.0% for IgG ≥ 11 days post symptom onset. Our study also demonstrated the superiority of ELISA and VIDAS^® assays over rapid diagnostic IgM/IgG tests. The analytical performance of VIDAS^® anti-CHIKV IgM and IgG assays was excellent, with a high precision (coefficients of variation ≤ 7.4%) and high specificity (cross-reactivity rate ≤ 2.9%). This study demonstrates the suitability of the automated VIDAS^® anti-CHIKV IgM and IgG assays to diagnose CHIKV infections and supports its applicability for epidemiological surveillance and differential diagnosis in regions endemic for CHIKV. Full article

(This article belongs to the Special Issue Laboratory Diagnosis in Microbial Diseases)

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10 pages, 561 KiB

Open AccessArticle

Evaluation of the Performance Characteristics of a New POC Multiplex PCR Assay for the Diagnosis of Viral and Bacterial Neuromeningeal Infections

by Hervé Le Bars, Neil Madany, Claudie Lamoureux, Clémence Beauruelle, Sophie Vallet, Christopher Payan and Léa Pilorgé

Diagnostics 2023, 13(6), 1110; https://doi.org/10.3390/diagnostics13061110 - 15 Mar 2023

Cited by 2 | Viewed by 1882

Abstract

Point-of-care syndromic PCR (POC SPCR) assays are useful tools for the rapid detection of the most common causative agents of community-acquired infections responsible for meningitis and encephalitis infections. We evaluated the performance characteristics of the new QIAstat-Dx^® Meningitis/Encephalitis panel (QS) compared to [...] Read more.

Point-of-care syndromic PCR (POC SPCR) assays are useful tools for the rapid detection of the most common causative agents of community-acquired infections responsible for meningitis and encephalitis infections. We evaluated the performance characteristics of the new QIAstat-Dx^® Meningitis/Encephalitis panel (QS) compared to the laboratory reference methods and the POC SPCR Biofire^® FilmArray^® Meningitis Encephalitis Panel (FA). Viral (Enterovirus, Parechovirus, HSV-1, HSV-2, HHV-6, VZV) and bacterial (E. coli K1, H. influenzae, L. monocytogenes, encapsulated N. meningitidis, M. pneumoniae, S. agalactiae, S. pneumoniae, S. pyogenes) pathogens were suspended at low concentrations and tested with the POC SPCR systems. The reproducibility, analytical specificity, carryover contamination, interferences and clinical samples were evaluated. All samples tested positive with both QS and FA except for those containing the lowest concentrations of Enterovirus-D68-B3, Echovirus-30 and S. agalactiae which were only detected by FA. In terms of analytical specificity, we observed 3 false positive results out of 48 QS tests versus 1 out of 37 FA tests. For the other studied criteria, both QS and FA performed as expected. Our results suggest that the performance characteristics of QS are close to those of FA. A prospective multicenter study would be useful to complete the performances evaluation of QS. Full article

(This article belongs to the Special Issue Laboratory Diagnosis in Microbial Diseases)

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9 pages, 3263 KiB

Open AccessArticle

Prevalence of Schistosoma haematobium and Intestinal Helminth Infections among Nigerian School Children

by Tolulope Alade, Thuy-Huong Ta-Tang, Sulaiman Adebayo Nassar, Akeem Abiodun Akindele, Raquel Capote-Morales, Tosin Blessing Omobami and Pedro Berzosa

Diagnostics 2023, 13(4), 759; https://doi.org/10.3390/diagnostics13040759 - 16 Feb 2023

Cited by 2 | Viewed by 2658

Abstract

Schistosomiasis and soil-transmitted helminthiases (STH) are two parasitic diseases mainly affecting school children. The purpose of this study was to estimate the current prevalence and infection intensity, in addition to the associations of these infections with age and sex, in children aged 4–17 [...] Read more.

Schistosomiasis and soil-transmitted helminthiases (STH) are two parasitic diseases mainly affecting school children. The purpose of this study was to estimate the current prevalence and infection intensity, in addition to the associations of these infections with age and sex, in children aged 4–17 years living in Osun State, Nigeria. From each participant (250 children), one urine and one stool sample were taken for the study, for the microscopic detection of eggs or larvae in faeces by means of the Kato–Katz method and eggs in filtrated urine. The overall prevalence of urinary schistosomiasis was 15.20%, with light infection. The intestinal helminthic species identified (and their prevalence) were S. stercoralis (10.80%), S. mansoni (8%), A. lumbricoides (7.20%), hookworm (1.20%), and T. trichiura (0.4%), all of them being classified as light infections. Single infections (67.95%) are more frequent than multiple infections (32.05%). With this study, schistosomiasis and STH are still endemic in Osun State, but with a light to moderate prevalence and light infection intensity. Urinary infection was the most prevalent, with higher prevalence in children over 10 years. The >10 years age group had the highest prevalence for all of the intestinal helminths. There were no statistically significant associations between gender and age and urogenital or intestinal parasites. Full article

(This article belongs to the Special Issue Laboratory Diagnosis in Microbial Diseases)

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11 pages, 738 KiB

Open AccessArticle

Gram Stain and Culture of Sputum Samples Detect Only Few Pathogens in Community-Acquired Lower Respiratory Tract Infections: Secondary Analysis of a Randomized Controlled Trial

by Mariana B. Cartuliares, Helene Skjøt-Arkil, Christian B. Mogensen, Thor A. Skovsted, Steen L. Andersen, Andreas K. Pedersen and Flemming S. Rosenvinge

Diagnostics 2023, 13(4), 628; https://doi.org/10.3390/diagnostics13040628 - 08 Feb 2023

Viewed by 3481

Abstract

Identification of the bacterial etiology of lower respiratory tract infections (LRTI) is crucial to ensure a narrow-spectrum, targeted antibiotic treatment. However, Gram stain and culture results are often difficult to interpret as they depend strongly on sputum sample quality. We aimed to investigate [...] Read more.

Identification of the bacterial etiology of lower respiratory tract infections (LRTI) is crucial to ensure a narrow-spectrum, targeted antibiotic treatment. However, Gram stain and culture results are often difficult to interpret as they depend strongly on sputum sample quality. We aimed to investigate the diagnostic yield of Gram stain and culture from respiratory samples collected by tracheal suction and expiratory technique from adults admitted with suspected community-acquired LRTI (CA-LRTI). In this secondary analysis of a randomized controlled trial, 177 (62%) samples were collected by tracheal suction, and 108 (38%) by expiratory technique. We detected few pathogenic microorganisms, and regardless of sputum quality, there were no significant differences between the sample types. Common pathogens of CA-LRTI were identified by culture in 19 (7%) samples, with a significant difference between patients with or without prior antibiotic treatment (p = 0.007). The clinical value of sputum Gram stain and culture in CA-LRTI is therefore questionable, especially in patients treated with antibiotics. Full article

(This article belongs to the Special Issue Laboratory Diagnosis in Microbial Diseases)

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10 pages, 966 KiB

Open AccessArticle

Effect of the Hematocrit and Storage Temperature of Dried Blood Samples in the Serological Study of Mumps, Measles and Rubella

by Mariano Rodríguez-Mateos, Javier Jaso, Paula Martínez de Aguirre, Silvia Carlos, Leire Fernández-Ciriza, África Holguín and Gabriel Reina

Diagnostics 2023, 13(3), 349; https://doi.org/10.3390/diagnostics13030349 - 18 Jan 2023

Cited by 2 | Viewed by 1772

Abstract

Dried blood spots (DBSs) are an economical and convenient alternative to serum/plasma, which allow for the serological and molecular study of different pathogens. Sixty-four blood samples were collected by venipuncture and spotted onto Whatman™ 903 cards to evaluate the utility of DBSs and [...] Read more.

Dried blood spots (DBSs) are an economical and convenient alternative to serum/plasma, which allow for the serological and molecular study of different pathogens. Sixty-four blood samples were collected by venipuncture and spotted onto Whatman™ 903 cards to evaluate the utility of DBSs and the effect of the storage temperature for 120 days after sample collection to carry out serological diagnosis. Mumps, measles and rubella IgG were investigated from DBSs and plasma using an automated chemiluminescent immunoassay. Using a calculated optimal cut-off value, the serological evaluation of mumps, measles and rubella using DBSs achieved high sensitivity (100%, 100% and 82.5%, respectively) and specificity (100%, 87.5% and 100%, respectively). The correlation observed between the plasma and the DBSs processed after sample collection was high (0.914–0.953) for all antibodies studied, both considering hematocrit before sample elution or not. For the different storage conditions, the correlation with plasma was high at 4 °C (0.889–0.925) and at −20 °C (0.878–0.951) but lower at room temperature (0.762–0.872). Measles IgG results were more affected than other markers when DBSs were stored at any temperature for 120 days. To summarize, hematocrit does not affect the processing of DBSs in the study of serological markers of mumps, measles and rubella. DBS stability for serological diagnosis of mumps and rubella is adequate when samples are stored at −20 °C or 4 °C, but not at room temperature, for a period of 4 months. Full article

(This article belongs to the Special Issue Laboratory Diagnosis in Microbial Diseases)

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15 pages, 1970 KiB

Open AccessArticle

A Comparative Study on Visual Detection of Mycobacterium tuberculosis by Closed Tube Loop-Mediated Isothermal Amplification: Shedding Light on the Use of Eriochrome Black T

by Alireza Neshani, Hosna Zare, Hamid Sadeghian, Hadi Safdari, Bamdad Riahi-Zanjani and Ehsan Aryan

Diagnostics 2023, 13(1), 155; https://doi.org/10.3390/diagnostics13010155 - 02 Jan 2023

Cited by 6 | Viewed by 2869

Abstract

Loop-mediated isothermal amplification is a promising candidate for the rapid detection of Mycobacterium tuberculosis. However, the high potential for carry-over contamination is the main obstacle to its routine use. Here, a closed tube LAMP was intended for the visual detection of Mtb [...] Read more.

Loop-mediated isothermal amplification is a promising candidate for the rapid detection of Mycobacterium tuberculosis. However, the high potential for carry-over contamination is the main obstacle to its routine use. Here, a closed tube LAMP was intended for the visual detection of Mtb to compare turbidimetric and two more favorable colorimetric methods using calcein and hydroxy naphthol blue (HNB). Additionally, a less studied dye (i.e., eriochrome black T (EBT)) was optimized in detail in the reaction for the first time. Mtb purified DNA and 30 clinical specimens were used to respectively determine the analytical and diagnostic sensitivities of each method. The turbidimetric method resulted in the best analytical sensitivity (100 fg DNA/reaction), diagnostic sensitivity and specificity (100%), and time-to-positivity of the test (15 min). However, this method is highly prone to subjective error in reading the results. Moreover, HNB-, calcein-, and EBT-LAMP could respectively detect 100 fg, 1 pg, and 1 pg DNA/reaction (the analytical sensitivities) in 30, 15, and 30 min, while the diagnostic sensitivity and specificity were respectively 93.3% and 100% for them all. Interestingly, EBT-LAMP showed the lowest potential for subjective error in reading the results. This report helps judiciously choose the most appropriate visual method, taking a step forward toward the field applicability of LAMP for the detection of Mtb, particularly in resource-limited settings. Full article

(This article belongs to the Special Issue Laboratory Diagnosis in Microbial Diseases)

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15 pages, 4347 KiB

Open AccessArticle

Performance Evaluation of Developed Bangasure™ Multiplex rRT-PCR Assay for SARS-CoV-2 Detection in Bangladesh: A Blinded Observational Study at Two Different Sites

by Mamudul Hasan Razu, Zabed Bin Ahmed, Md. Iqbal Hossain, Mohammad Fazle Alam Rabbi, Maksudur Rahman Nayem, Md. Akibul Hassan, Gobindo Kumar Paul, Md. Robin Khan, Md. Moniruzzaman, Pranab Karmaker and Mala Khan

Diagnostics 2022, 12(11), 2617; https://doi.org/10.3390/diagnostics12112617 - 28 Oct 2022

Viewed by 1698

Abstract

In this study, we evaluated the performance of the in-house developed rRT-PCR assay for SARS-CoV-2 RNA targeting the envelope (E) and nucleocapsid (N) genes with internal control as human RNase P. A total of 50 positive samples and [...] Read more.

In this study, we evaluated the performance of the in-house developed rRT-PCR assay for SARS-CoV-2 RNA targeting the envelope (E) and nucleocapsid (N) genes with internal control as human RNase P. A total of 50 positive samples and 50 negative samples of SARS-CoV-2 were tested by a reference kit at site 1 and a subset (30 positives and 16 negatives) of these samples are tested blindly at site 2. The limit of detection (LoD) was calculated by using a replication-deficient complete SARS-CoV-2 genome and known copy numbers, where Pseudo-virus samples were used to evaluate accuracy. On site 1, among the 50 SARS-CoV-2 positive samples 24, 18, and eight samples showed high (Ct < 26), moderate (26 < Ct ≤ 32), and low (32 < Ct ≤ 38) viral load, respectively, whereas in site 2, out of 30 SARS-CoV-2 positive samples, high, moderate, and low viral loads were found in each of the 10 samples. However, SARS-CoV-2 was not detected in the negative sample. So, in-house assays at both sites showed 100% sensitivity and specificity with no difference observed between RT PCR machines. The Ct values of the in-house kit had a very good correlation with the reference kits. LoD was determined as 100 copies/mL. It also displayed 100% accuracy in mutant and wild-type SARS-CoV-2 virus. This Bangasure™ RT-PCR kit shows excellent performance in detecting SARS-CoV-2 viral RNA compared to commercially imported CE-IVD marked FDA authorized kits. Full article

(This article belongs to the Special Issue Laboratory Diagnosis in Microbial Diseases)

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8 pages, 376 KiB

Open AccessArticle

Determination of Diphtheria Toxin in Bacterial Cultures by Enzyme Immunoassay

by Maria A. Simonova, Vyacheslav G. Melnikov, Olga E. Lakhtina, Ravilya L. Komaleva, Anja Berger, Andreas Sing and Sergey K. Zavriev

Diagnostics 2022, 12(9), 2204; https://doi.org/10.3390/diagnostics12092204 - 11 Sep 2022

Cited by 2 | Viewed by 1695

Abstract

Since diphtheria toxin (DT) is the main virulence factor of Corynebacterium diphtheriae and C. ulcerans, the detection of DT in corynebacterial cultures is of utmost importance in the laboratory diagnosis of diphtheria. The need to measure the level of DT production (LTP) [...] Read more.

Since diphtheria toxin (DT) is the main virulence factor of Corynebacterium diphtheriae and C. ulcerans, the detection of DT in corynebacterial cultures is of utmost importance in the laboratory diagnosis of diphtheria. The need to measure the level of DT production (LTP) arises when studying the virulence of a strain for the purpose of diphtheria agent monitoring. To determine the LTP of diphtheria agents, an immunoassay based on monoclonal antibodies (mAbs) has been developed. A pair of mAbs specific to the fragment B of DT was selected, which makes it possible to detect DT in a sandwich ELISA with a detection limit of DT less than 1 ng/mL. Sandwich ELISA was used to analyze 218 liquid culture supernatants of high-, low- and non-toxigenic strains of various corynebacteria. It was shown that the results of ELISA are in good agreement with the results of PCR and the Elek test for the tox gene and DT detection, respectively. The diagnostic sensitivity of the assay was approximately 99%, and specificity was 100%. It has been found that strains of C. ulcerans, on average, produce 10 times less DT than C. diphtheriae. The mAbs used in the ELISA proved to be quite discriminatory and could be further used for the design of the LFIA, a method that can reduce the labor and cost of laboratory diagnosis of diphtheria. Full article

(This article belongs to the Special Issue Laboratory Diagnosis in Microbial Diseases)

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Review

Jump to: Research

29 pages, 1776 KiB

Open AccessReview

Antibiotic Resistance Diagnosis in ESKAPE Pathogens—A Review on Proteomic Perspective

by Sriram Kalpana, Wan-Ying Lin, Yu-Chiang Wang, Yiwen Fu, Amrutha Lakshmi and Hsin-Yao Wang

Diagnostics 2023, 13(6), 1014; https://doi.org/10.3390/diagnostics13061014 - 07 Mar 2023

Cited by 5 | Viewed by 5722

Abstract

Antibiotic resistance has emerged as an imminent pandemic. Rapid diagnostic assays distinguish bacterial infections from other diseases and aid antimicrobial stewardship, therapy optimization, and epidemiological surveillance. Traditional methods typically have longer turn-around times for definitive results. On the other hand, proteomic studies have [...] Read more.

Antibiotic resistance has emerged as an imminent pandemic. Rapid diagnostic assays distinguish bacterial infections from other diseases and aid antimicrobial stewardship, therapy optimization, and epidemiological surveillance. Traditional methods typically have longer turn-around times for definitive results. On the other hand, proteomic studies have progressed constantly and improved both in qualitative and quantitative analysis. With a wide range of data sets made available in the public domain, the ability to interpret the data has considerably reduced the error rates. This review gives an insight on state-of-the-art proteomic techniques in diagnosing antibiotic resistance in ESKAPE pathogens with a future outlook for evading the “imminent pandemic”. Full article

(This article belongs to the Special Issue Laboratory Diagnosis in Microbial Diseases)

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