X-linked recessive ichthyosis (XLI) is clinically characterized by dark brown, widespread dryness with polygonal scales. We describe the identification of STS and PUDP deletions using targeted panel sequencing combined with copy-number variation (CNV) analysis in XLI. A 9-month-old infant was admitted for genetic counseling. Since the second day after birth, the infant\'s skin tended to be dry and polygonal scales had accumulated over the abdomen and upper extremities. The infant\'s maternal uncle and brother (who had also exhibited similar skin symptoms from birth) presented with polygonal scales on their trunks. CNV analysis revealed a hemizygous deletion spanning 719.3 Kb on chromosome Xp22 (chrX:7,108,996-7,828,312), which included a segment of the STS gene and exhibited a Z ratio of -2 in the proband. Multiplex ligation-dependent probe amplification (MLPA) confirmed this interstitial Xp22.31 deletion. Our report underscores the importance of implementing CNV screening techniques, including sequencing data analysis and gene dosage assays such as MLPA, to detect substantial deletions that encompass the STS gene region of Xq22 in individuals suspected of having XLI.

Digital PCR (dPCR) is a nucleic acid quantification method that is widely used in genetic analysis. One of the most significant advantages of dPCR over other methods is the possibility of absolute quantitative determination of genetic material without construction of calibration curves, which allows one to detect even single molecules of nucleic acids, and, hence, provides early diagnosis of diseases. One specific characteristic of dPCR is the detection of the analyzed biological object in each microreaction, followed by the presentation of the analysis results in a binary system, thereby giving the method its name. The key aspects of developing the dPCR method, i.e., from the first devices based on microfluidic chip technology to modern systems capable of measuring a target at a concentration of up to 1 in 100000 copies are shown in the current work. We analyzed the data on the detection of various pathogens using dPCR, as well as summarizing various study results demonstrating the innovativeness of this method. Both the possibilities of multiplex dPCR analysis and its potential in clinical practice are presented. This review also addresses the issue of the role of dPCR in the development of noninvasive methods for analysis of oncological diseases. Possible ways of developing dPCR technology were emphasized, including its use as a \"point-of-care\" system.

Introduction. The FilmArray blood culture identification panel (BCID) panel is a multiplex PCR assay with high sensitivity and specificity to identify the most common pathogens in bloodstream infections (BSIs).Hypothesis. We hypothesize that the BCID panel has good diagnostic performance for BSIs and can be popularized in clinical application.Aim: To provide summarized evidence for the diagnostic accuracy of the BCID panel for the identification of positive blood cultures.Methodology. We searched the MEDLINE, EMBASE and Cochrane databases through March 2021 and assessed the efficacy of the diagnostic test of the BCID panel. We performed a meta-analysis and calculated the summary sensitivity and specificity of the BCID panel. Systematic review protocols were registered in the International Prospective Register of Systematic Reviews (PROSPERO) (registration number CRD42021239176).Results. A total of 16 full-text articles were eligible for analysis. The overall sensitivities of the BCID panel on Gram-positive bacteria, Gram-negative bacteria and fungi were 97 % (95 % CI, 0.96-0.98), 100 % (95 % CI, 0.98-01.00) and 99 % (95 % CI, 0.87-1.00), respectively. The pooled diagnostic specificities were 99 % (95 % CI, 0.97-1.00), 100 % (95 % CI, 1.00-1.00) and 100 % (95 % CI, 1.00-1.00) for Gram-positive bacteria, Gram-negative bacteria and fungi, respectively.Conclusions. The BCID panel has high rule-in value for the early detection of BSI patients. The BCID panel can still provide valuable information for ruling out bacteremia or fungemia in populations with low pretest probability.

BACKGROUND: Accuracy and timing of antibiotic therapy remain a challenge for lower respiratory tract infections. New molecular techniques using Multiplex Polymerase Chain Reaction, including the FilmArray® Pneumonia Plus Panel [FAPP], have been developed to address this. The aim of this study is to evaluate the FAPP diagnostic performance for the detection of the 15 typical bacteria of the panel from respiratory samples in a meta-analysis from a systematic review.
METHODS: We searched PubMed and EMBASE from January 1, 2010, to December 31, 2022, and selected any study on the FAPP diagnostic performance on respiratory samples compared to the reference standard, bacterial culture. The main outcome was the overall diagnostic accuracy with sensitivity and specificity. We calculated the log Diagnostic Odds Ratio and analyzed performance for separate bacteria, antimicrobial resistance genes, and according to the sample type. We also reported the FAPP turnaround time and the out-of-panel bacteria number and species. This study is registered with PROSPERO (CRD42021226280).
RESULTS: From 10 317 records, we identified 30 studies including 8 968 samples. Twenty-one were related to intensive care. The overall sensitivity and specificity were 94% [95% Confidence Interval (CI) 91-95] and 98% [95%CI 97-98], respectively. The log Diagnostic Odds Ratio was 6.35 [95%CI 6.05-6.65]. 9.3% [95%CI 9.2-9.5] of bacteria detected in culture were not included in the FAPP panel.
CONCLUSIONS: This systematic review reporting the FAPP evaluation revealed a high accuracy. This test may represent an adjunct tool for pulmonary bacterial infection diagnostic and antimicrobial stewardship. Further evidence is needed to assess the impact on clinical outcome.

The clinical impact of rapid sample-to-answer \"syndromic\" multiplex polymerase chain reaction (PCR) testing for respiratory viruses is not clearly established. We performed a systematic literature review and meta-analysis to evaluate this impact for patients with possible acute respiratory tract infection in the hospital setting.
We searched EMBASE, MEDLINE, and Cochrane databases from 2012 to present and conference proceedings from 2021 for studies comparing clinical impact outcomes between multiplex PCR testing and standard testing.
Twenty-seven studies with 17,321 patient encounters were included in this review. Rapid multiplex PCR testing was associated with a reduction of - 24.22 h (95% CI -28.70 to -19.74 h) in the time to results. Hospital length of stay was decreased by -0.82 days (95% CI -1.52 to -0.11 days). Among influenza positive patients, antivirals were more likely to be given (RR 1.25, 95% CI 1.06-1.48) and appropriate infection control facility use was more common with rapid multiplex PCR testing (RR 1.55, 95% CI 1.16-2.07).
Our systematic review and meta-analysis demonstrates a reduction in time to results and length of stay for patients overall along with improvements in appropriate antiviral and infection control management among influenza-positive patients. This evidence supports the routine use of rapid sample-to-answer multiplex PCR testing for respiratory viruses in the hospital setting.

BACKGROUND: The traditional diagnosis model has great challenges for the etiological diagnosis of the central nervous system (CNS) diseases with similar clinical manifestations, especially for the diagnosis of rare pathogens. It is very important to make rapid and accurate identification of pathogens for guiding clinical choices in administering countermeasures.
METHODS: On August 22, 2020, a 49 years old Chinese male patient had a headache for two days, and then the computed tomography (CT) scan of the brain showed subarachnoid hemorrhage. Subsequently, he underwent twice craniotomy and about 3 weeks of hospitalization. Since September 20, the patient was in the local rehabilitation hospital for hyperbaric oxygen therapy for about three weeks. Then the patient developed acute purulent meningoencephalitis. In the absence of diagnosis of specific pathogenic bacteria, vancomycin (1 g every 12 hours), ceftazidime (2 g every 8 hours), mannitol dehydration (125 mL, every 8 hours), and sodium valproate (0.4 g tid) was used timely according to cerebrospinal fluid (CSF) examination and clinical manifestations. CSF smear and routine culture test were negative during hospitalization. We used the metagenomic next-generation sequencing (mNGS) analysis of CSF for quick and accurate diagnosis, which identified human herpesvirus type 4 (EBV), Corynebacterium corynebacterium, Achromobacter xylose oxidation, and Acinetobacter baumannii, But the mapping degree was not high. Then, we used the modified method-multiplex PCR-based targeted gene sequencing platform (ptNGS) to detect CSF samples and found that the sequences detected were Acinetobacter pittii (A. pittii) and Staphylococcus epidermidis. S. epidermidis might come from skin colonization during lumbar puncture, so it was excluded from the etiological diagnosis. Therefore, we highly suspected that A. pittii was the pathogen in this case. After about three weeks of hospitalization treatment, the patient\'s symptoms were relieved.
CONCLUSIONS: In conclusion, empirical medication before the identification of pathogens is very important. The ptNGS may be an effective method for the diagnosis of pathogens.

The primary aim is to provide a summary of evidence for the diagnostic accuracies of multiplex PCR gastrointestinal (GI) panels-BioFire FilmArray and Luminex xTAG on the detection of gastroenteritis pathogens. The secondary aim is to compare the performance of these GI panels head to head.
A comprehensive search up to 1 December 2019 was conducted on PubMed, Embase, Ovid Medline and Web of Science for studies that used FilmArray or Luminex xTAG Gastrointestinal Pathogen Panel (GPP) for diagnosis of acute gastroenteritis. A summary of diagnostic accuracies for the 16 pathogens were calculated by comparing the GI panels to the current gold standards (conventional standard microbiology techniques such as culture or PCR for bacteria, PCR or enzyme immunoassay (EIA) for viruses, microscopy or EIA for parasite). Hierarchical summary receiver operating characteristic (HSROC) curve analysis, pretest and post-test probabilities were used for estimating the pathogen detection performance.
A total of 11 studies with 7085 stool samples were eligible for analysis. Multiplex PCRs demonstrated high diagnostic accuracy, with specificity ≧0.98 and area under the ROC curve (AUROC) ≧0.97 for all the pathogens except for Yersinia enterocolitica (AUROC 0.91). The FilmArray panel demonstrated a higher sensitivity than xTAG GPP for most of the pathogens with the exception of Rotavirus A (xTAG GPP and FilmArray were both 0.93).
This is the first meta-analysis that is a head-to-head comparison examining the performance of the novel multiplex PCR-based tests Luminex xTAG GPP and FilmArray GI panel in detecting each pathogen. Point estimates calculated from eligible studies showed that both GI panels are highly accurate and may provide important diagnostic information for early identification of gastroenteritis. In addition, although FilmArray has higher sensitivity and post-test probability than xTAG GPP for most of the pathogens, how this will translate to a clinical setting remains unclear.

We have investigated the main genetic causes for non-syndromic hearing impairment (NSHI) in the hearing impairment individuals from the North-Eastern Romania and proposed a cost-effective diagnosis protocol.
MLPA followed by Sanger Sequencing were used for all 291 patients included in this study.
MLPA revealed abnormal results in 141 cases (48.45%): 57 (40.5%) were c.35delG homozygous, 26 (18.44%) were c.35delG heterozygous, 14 (9.93%) were compound heterozygous and 16 (11.35%) had other types of variants. The entire coding region of GJB2 was sequenced and out of 150 patients with normal results at MLPA, 29.33% had abnormal results: variants in heterozygous state: c.71G>A (28%), c.457G>A (20%), c.269T>C (12%), c.109G>A (12%), c.100A>T (12%), c.551G>C (8%). Out of 26 patients with c.35delG in heterozygous state, 38.46% were in fact compound heterozygous.
We identified two variants: c.109G>A and c.100A>T that have not been reported in any study from Romania. MLPA is an inexpensive, rapid and reliable technique that could be a cost-effective diagnosis method, useful for patients with hearing impairment. It can be adaptable for the mutation spectrum in every population and followed by Sanger sequencing can provide a genetic diagnosis for patients with different degrees of hearing impairment.

Mammalian species identification is one of the important issues in forensic science. Determining the origins of non-human biological material found at crime scenes can increase the possibility of identifying the true culprit by narrowing down the range of suspects. Although many techniques based on mitochondrial DNA (mtDNA) have been developed, challenges remain to cost-effectively identify species from degraded samples containing a mixture of DNA from multiple species and to standardize procedures for mammalian species identification. This review evaluates the reliability and versatility of mtDNA-based techniques to reveal obstacles to the establishment of standard analytical methods, with a particular focus on DNA mixtures. When samples contain a mixture of DNA from multiple species, the interpretation of sequencing analysis results is difficult. Although DNA metabarcoding using next-generation sequencing (NGS) technologies can overcome the DNA mixture problem, DNA metabarcoding is not suitable for the type of small-scale analysis routinely performed by local forensic laboratories, primarily because it is costly and time-consuming. By contrast, fluorescent multiplex PCR analysis enables cost-effective and simultaneous species identification from suboptimal samples, although the number of identifiable species is currently limited in comparison with sequencing techniques. The advantages and limitations of current techniques presented in this review indicate that multiplex PCR analysis will continue to be important for mammalian species identification in forensic casework analysis. Further developments in multiplex PCR analysis that enable the identification of an increased number of species will play a key step for standardization efforts among forensic laboratories.

BACKGROUND: The FilmArray® meningitis/encephalitis (ME) panel is a multiplex PCR assay which can detect the most commonly identified pathogens in central nervous system infections. It significantly decreases the time to diagnosis of ME and data has yielded several positive outcomes. However, in part, reports of both false positive and false negative detections have resulted in concerns about adoption.
OBJECTIVE: The aim was to evaluate the ME panel in a diagnostic test accuracy review.
METHODS: The PubMed and EMBASE databases were systematically searched through May 2019.
METHODS: Eligible studies were those providing sensitivity and specificity data for the ME panel compared with a reference standard. Studies providing details on false positive and false negative results of the panel as well as further investigation (adjudication) of the discordant results between the panel and comparator assays were included and assessed separately.
METHODS: Patients with suspected ME for whom a panel was ordered were included.
METHODS: The ME panel was compared to reference standard methods for diagnosing community-acquired ME. We performed a meta-analysis and calculated the summary sensitivity and specificity of the ME panel. Moreover, we evaluated the false positive and false negative results of the panel.
RESULTS: Thirteen studies (3764 patients) were included in the review and 8 of them (3059 patients) were pooled in a meta-analysis. The summary estimates of sensitivity and specificity with 95% confidence intervals (CI) was 90% (95% CI 86-93%) and 97% (95% CI 94-99%), respectively. When we looked specifically at studies that assessed further the false positive and false negative results, false positive detections were 11.4% and 4% before and after adjudication, respectively. The highest proportion of false positive was observed for Streptococcus pneumoniae followed by Streptococcus agalactiae. False negative isolates were 2.2% and 1.5% before and after adjudication, respectively. Herpes simplex virus 1 and 2, enterovirus and Cryptococcus neoformans/gattii had the highest proportions of false negative determinations. False negative C. neoformans/gattii were mostly patients with positive antigen titres, on treatment or cleared disease.
CONCLUSIONS: The currently available literature suggests that the ME panel has high diagnostic accuracy. However, the decision for implementation should be individualized based on the needs of the patient population, the capabilities of the laboratory, and the knowledge of the healthcare providers that will utilize the test.