Direct or DNA Extraction-Free Amplification and Quantification of Foodborne Pathogens.

The use of direct nucleic acid amplification of pathogens from food matrices has the potential to reduce time to results over DNA extraction-based approaches as well as traditional culture-based approaches. Here we describe protocols for assay design and experiments for direct amplification of foodborne pathogens in food sample matrices using loop-mediated isothermal amplification (LAMP) and polymerase chain reaction (PCR). The examples provided include the detection of Escherichia coli in milk samples and Salmonella in pork meat samples. This protocol includes relevant reagents and methods including obtaining target sequences, assay design, sample processing, and amplification. These methods, though used for specific example matrices, could be applied to many other foodborne pathogens and sample types.

Nitric Oxide Synthase 3 Gene Polymorphisms and Their Association with Acute Myocardial Infarction and Chronic Stable Angina: A Case-Control Study from Northern India.

Background: Coronary artery disease (CAD) that encompasses acute myocardial infarction (AMI), chronic stable angina (CSA), and unstable angina (UA) has numerous known risk factors. Genetic predispositions contribute as major risk in the development of CAD and the genes regulating atherosclerosis are important for disease prevention. Nitric oxide synthase 3 (NOS3) gene responsible for nitric oxide (NO) production is of special importance. Aim: To evaluate the role of three NOS3 polymorphisms (-786C/T, 894G/T, and 4a4b) in patients with CAD, particularly in AMI and CSA and their comparison with healthy controls. Materials and Methods: One hundred patients in each AMI and CSA group and 100 controls were included and were typed for three NOS3 polymorphisms (-786C/T, 894G/T, and 4a4b) by polymerase chain reaction-restriction fragment length polymorphism. Plasma NO metabolites (NOx) were also evaluated. Results: A significant association of 894G/T polymorphism with AMI in dominant model (P = 0.052) and with CSA in dominant and codominant models was detected (P = 0.008 and P = 0.006, respectively). Plasma NO levels were found to be significantly higher (P < 0.0001) in healthy controls (43.80 ± 6.28) compared to AMI and CSA patients (37.05 ± 6.75 and 38.67 ± 5.61). No significant association of -786C/T and 4a4b polymorphism with AMI and CSA risk under recessive, dominant, and codominant models was detected. Conclusion: Our study revealed a significant association of 894G/T polymorphism with AMI and independent association of NOx levels with CAD, indicating high risk of CAD in the North Indian population. Our findings will be helpful in identifying the genetic risk factors associated with CAD and better management of the diagnostic as well as therapeutic measures.

Acute Retinal Necrosis Caused by Varicella Zoster Virus and Cytomegalovirus Co-Infection.

PURPOSE: To report the clinical course of two cases of acute retinal necrosis (ARN) caused by varicella zoster virus (VZV) and cytomegalovirus (CMV) co-infection detected by polymerase chain reaction (PCR) on aqueous tap. METHODS: Observational case reports. RESULTS: Two patients presented to our services with unilateral panuveitis suggestive of ARN complicated by hemorrhagic vasculitis and started empirical therapy. Aqueous PCR was performed on the same day and showed double positivity for VZV and CMV, which guided treatment. At follow-up, wide-field color fundus imaging and high-resolution optical coherence tomography showed resolution of active retinitis. CONCLUSION: Our cases suggest that ARN complicated by hemorrhagic vasculitis may be secondary to CMV and VZV co-infection, both in patients with an unremarkable clinical history and in those with immunodeficiency. In our cases, aqueous PCR testing was of paramount importance to determine the aetiology of ARN and to adjust the antiviral therapy accordingly.

Rapid Molecular Diagnostics of Pneumonia Caused by Gram-Negative Bacteria: A Clinician's Review.

With approximately half a billion events per year, lower respiratory tract infections (LRTIs) represent a major challenge for the global public health. Among LRTI cases, those caused by Gram-negative bacteria (GNB) are associated with a poorer prognostic. Standard-of-care etiologic diagnostics is lengthy and difficult to establish, with more than half of cases remaining microbiologically undocumented. Recently, syndromic molecular diagnostic panels became available, enabling simultaneous detection of tens of pathogen-related and antimicrobial-resistance genetic markers within a few hours. In this narrative review, we summarize the available data on the performance of molecular diagnostics in GNB pneumonia, highlighting the main strengths and limitations of these assays, as well as the main factors influencing their clinical utility. We searched MEDLINE and Web of Science databases for relevant English-language articles. Molecular assays have higher analytical sensitivity than cultural methods, and show good agreement with standard-of-care diagnostics regarding detection of respiratory pathogens, including GNB, and identification of frequent patterns of resistance to antibiotics. Clinical trials reported encouraging results on the usefulness of molecular assays in antibiotic stewardship. By providing early information on the presence of pathogens and their probable resistance phenotypes, these assays assist in the choice of targeted therapy, in shortening the time from sample collection to appropriate antimicrobial treatment, and in reducing unnecessary antibiotic use.

Mitigation of Diabetes Mellitus Using Euphorbia helioscopia Leaf Ethanolic Extract by Modulating GCK, GLUT4, IGF, and G6P Expressions in Streptozotocin-Induced Diabetic Rats.

Diabetes mellitus is a metabolic disorder. Synthetic antidiabetics are the commonly used treatment options associated with complications. The objective of this study was to explore the antioxidative and antidiabetic potential of Euphorbia helioscopia whole plant ethanolic extract using in vitro and in vivo models. For that purpose, the antioxidative potential was explored by using 2,2-diphenyl-1-picrylhydrazyl analysis. In vitro antidiabetic potential of the extract was evaluated using amylase inhibitory analysis. In vivo antidiabetic activity of the extract was assessed in diabetic rats using streptozotocin/nicotinamide (60 mg/kg/120 mg/kg) as an inducing agent. Metformin was used as standard. The results indicated the presence of significant quantities of phenolic 82.18 ± 1.28 mgg-1 gallic acid equivalent (GAE) and flavonoid 66.55±1.22 mgg-1 quercetin equivalent (QE) contents in the extract. Quantitation of phytoconstituents exhibited the presence of sinapic acid, myricetin, and quercetin using HPLC analysis. The extract inhibited α-amylase by 84.71%, and an antiglycemic potential of 50.34% was assessed in the OGTT assay. Biochemical analysis demonstrated a reduction in urea, creatinine, cholesterol, low-density lipoprotein, and alkaline phosphatase (p < 0.001) as compared to diabetic control rats at the dose of 500 mg/kg. An upregulation in the expressions of glucokinase, glucose transporter 4, peroxisome proliferator-activated receptor γ, and insulin-like growth factor was observed in treated rats in contrast to G6P expression, which was downregulated upon treatment. In conclusion, this study provided evidence of the antioxidative and antidiabetic potential of E. helioscopia whole plant ethanolic extract through in vitro and in vivo analysis and emphasized its promising role as a natural alternative.

Evaluating the clinical efficacy of a long-read sequencing-based approach for carrier screening of spinal muscular atrophy.

Spinal muscular atrophy (SMA) is the second most common fatal genetic disease in infancy. It is caused by deletion or intragenic pathogenic variants of the causative gene SMN1, which degenerates anterior horn motor neurons and leads to progressive myasthenia and muscle atrophy. Early treatment improves motor function and prognosis in patients with SMA, but drugs are expensive and do not cure the disease. Therefore, carrier screening seems to be the most effective way to prevent SMA birth defects. In this study, we genetically analyzed 1400 samples using multiplex ligation-dependent probe amplification (MLPA) and quantitative polymerase chain reaction (qPCR), and compared the consistency of the results. We randomly selected 44 samples with consistent MLPA and qPCR results for comprehensive SMA analysis (CASMA) using a long-read sequencing (LRS)-based approach. CASMA results showed 100% consistency, visually and intuitively explained the inconsistency between exons 7 and 8 copy numbers detected by MLPA in 13 samples. A total of 16 samples showed inconsistent MLPA and qPCR results for SMN1 exon 7. CASMA was performed on all samples and the results were consistent with those of resampling for MLPA and qPCR detection. CASMA also detected an additional intragenic variant c.-39A>G in a sample with two copies of SMN1 (RT02). Finally, we detected 23 SMA carriers, with an estimated carrier rate of 1/61 in this cohort. In addition, CASMA identified the "2 + 0" carrier status of SMN1 and SMN2 in a family by analyzing the genotypes of only three samples (parents and one sibling). CASMA has great advantages over MLPA and qPCR assays, and could become a powerful technical support for large-scale screening of SMA.

Rapid Diagnostic PCR Assay Method for Species Identification of Mantidis Ootheca (Sangpiaoxiao) Based on Cytochrom C Oxidase I (COI) Barcode Analysis.

Mantidis Ootheca (sangpiaoxiao), the egg case of the mantis, is a type of insect-derived traditional medicine widely used in East Asia. However, species identification based on egg morphology is challenging, leading to the distribution of counterfeit and adulterated products. The use of inauthentic ingredients can pose serious health risks to consumers. This study aimed to develop PCR markers that can rapidly and accurately differentiate between authentic and counterfeit Mantidis Ootheca. The mitochondrial cytochrome c oxidase I (COI) region was sequenced in thirteen samples from four mantis species: Tenodera angustipennis, Statilia maculata, Hierodula patellifera, and T. sinensis. Four sets of SCAR primers were designed based on species-specific nucleotide polymorphisms, and a multiplex SCAR assay was developed by combining all sets of the primers. The sequence-characterized amplified region (SCAR) primers successfully produced amplicons for each target species, even with low-DNA templates or templates containing DNA from multiple samples. No amplification was observed for nontarget species. This study presents a novel approach for identifying authentic Mantidis Ootheca species using DNA-based diagnostic marker assays, which enable rapid and precise species identification. The SCAR assays developed in this study will aid in maintaining quality control and promoting the standardization of commercial Mantidis Ootheca products.

Assessment of High-Resolution Melting Curve Analysis for Leishmania spp. Detection in Different Clinical Manifestations of Leishmaniasis in India.

The accurate diagnosis and identification of Leishmania species are crucial for the therapeutic selection and effective treatment of leishmaniasis. This study aims to develop and evaluate the use of high-resolution melting curve analysis (HRM)-PCR for Leishmania species identification causing visceral leishmaniasis (VL), post-kala-azar dermal leishmaniasis (PKDL) and cutaneous leishmaniasis (CL) in the Indian subcontinent. Two multi-copy targets (ITS-1 and 7SL-RNA genes) were selected, and an HRM-PCR assay was established using L. donovani, L. major, and L. tropica standard strain DNA. The assay was applied on 93 clinical samples with confirmed Leishmania infection, including VL (n = 30), PKDL (n = 50), and CL (n = 13) cases. The ITS-1 HRM-PCR assay detected as little as 0.01 pg of template DNA for L. major and up to 0.1 pg for L. donovani and L. tropica. The detection limit for the 7SL-RNA HRM-PCR was 1 pg for L. major and 10 pg for L. donovani and L. tropica. The ITS-1 HRM-PCR identified 68 out of 93 (73.11%) leishmaniasis cases, whereas 7SL-RNA HRM-PCR could only detect 18 out of 93 (19.35%) cases. A significant correlation was observed between the kDNA-based low Ct values and ITS-1 HRM-PCR positivity in the VL (p = 0.007), PKDL (p = 0.0002), and CL (p = 0.03) samples. The ITS-1 HRM-PCR assay could identify Leishmania spp. causing different clinical forms of leishmaniasis in the Indian subcontinent, providing rapid and accurate results that can guide clinical management and treatment decisions.

Relationship Between IL-10 Single Nucleotide Polymorphisms (rs1800871, rs1800872, and rs1800896) and the Severity of COVID-19.

Background: Interleukin-10 (IL-10) is an anti-inflammatory cytokine whose levels are elevated in patients with severe COVID-19. IL-10 polymorphisms may play a role in increasing IL-10 levels and the severity of COVID-19. This study aimed to investigate the relationship between IL-10 single nucleotide polymorphisms (SNPs) (rs1800896 [-1082 C < T], rs1800871 [-819 A > G], and rs1800872 [-592 T > G]) and the severity of COVID-19 in patients from Kermanshah Province, Iran. Methods: A total of 150 patients with mild COVID-19 (84 men and 66 women aged 40.1 ± 12.44 years) and 143 patients with severe COVID-19 (76 men and 67 women aged 61.04 ± 15.65 years) participated in this study. Blood samples were collected from the patients, DNA was extracted, and the genotype of each SNPs was determined using the polymerase chain reaction-restriction fragment length polymorphism method. Result: The results of this study did not show a significant relationship between the genotypes of the three studied SNPs and the severity of COVID-19 (p > 0.05). Conclusion: According to our findings, these SNPs were not associated with COVID-19 severity in patients in Kermanshah.

Minimal residual disease testing for classical Hodgkin lymphoma: A comprehensive review.

Classical Hodgkin lymphoma (cHL) is a common lymphoma that affects young patients. Fortunately, the disease is highly curable as it is susceptible to the currently available treatment modalities. Disease monitoring with Positron Emission Tomography and Computed Tomography (PET/ CT) is an integral part of managing these patients. PET guided protocols are currently used to adjust treatment according to the response. The pivotal idea behind the use of response-adapted approaches is to preserve efficacy while decreasing the toxicity. It also helps to intensify therapy in patients in need because of suboptimal response. However, imaging techniques are limited by their sensitivity and specificity. Minimal Residual Disease (MRD) assessment is a newly emerging concept in many hematologic malignancies. It utilizes various molecular techniques such as polymerase chain reaction (PCR), and next-generation sequencing (NGS) as well as flow cytometry, to detect disease traces. This review looks into MRD detection techniques, its current applications, and the evidence in the literature for its use in cHL.

Evaluating the Impact of Ultrasonic Irrigation on Bacterial Levels and Activity Following Chemomechanical Procedures.

AIM: This single-arm interventional trial aimed to investigate the efficacy of ultrasonic irrigation as a supplementary disinfection approach after chemomechanical procedures using molecular techniques based on ribosomal RNA (rRNA) and rRNA genes (referred to as DNA). METHODOLOGY: Samples were collected from 35 single-rooted teeth with radiographic evidence of apical periodontitis. Samples were taken after gaining root canal access (S1), chemomechanical procedures (CMP, S2), and ultrasonic irrigation (S3). DNA-targeted qPCR using universal primers was used to estimate total bacterial levels, while rRNA-targeted qPCR was used to assess bacterial activity. Ratios between rRNA and DNA levels were calculated to search for active bacteria in the samples (rRNA/ DNA ≥ 1). Wilcoxon matched-pairs signed-rank test was used to compare the differences in DNA levels between samples and DNA and rRNA levels within samples (P <.05). RESULTS: DNA-based methods revealed a significant decrease in bacterial levels from S1 to S2 and S2 to S3 (both P <.05). Notably, 11 out of 35 (31.4%) root canals did not harbor bacterial DNA after CMP, whereas ultrasonic activation increased DNA-negative samples to 17 (48.6%). However, all DNA-positive samples were also positive for rRNA, with significantly higher rRNA than DNA levels (P <.05), indicating bacterial activity at the sampling time. CONCLUSIONS: Ultrasonic irrigation improved the disinfection of root canals after chemomechanical procedures by reducing bacterial levels. However, persisting bacteria remained active in the root canals after CMP and ultrasonic irrigation.

Pure neuritic leprosy: Latest advancements and diagnostic modalities: Diagnosis of Pure Neuritic Leprosy.

Pure neuritic leprosy (PNL) is characterized by exclusive peripheral neuropathy without dermatological alterations. Diagnosis is difficult since skin lesions and acid-fast bacilli (AFB) in slit smears are absent. Presently, the gold standard for diagnosis is the histopathological examination of peripheral nerve biopsy. Even then, the detection of bacteria is difficult, and histological findings may be non-specific. Nerve biopsy is an invasive procedure that is possible only in specialized centers and limited to certain sensory nerves. Therefore, the establishment of serological, immunological, and molecular laboratory tests could be more beneficial for diagnosing pure neuritic leprosy to achieve effective treatment and reduction in its consequent disabilities. This review suggests that the presence of Mycobacterium leprae (M.leprae) in PNL cases can be proven by using non-invasive procedures, viz., multiplex polymerase chain reaction (M-PCR), serological findings, immunological profiling, and improved nerve-imaging. Findings also indicate the necessity for improving the sensitivity of PCR and further research on specificity in ruling out other clinical conditions that may mimic PNL.

Additional reporting of diffuse and homogeneous ROS-1 SP384 immunoreactivity enhances prediction of ROS1 fusion-positive non-small cell lung cancer.

OBJECTIVES: ROS-1 immunohistochemistry (IHC) is a common method for screening ROS1 fusion in the clinical management of non-small cell lung cancer. The interpretation criteria for ROS-1 SP384 IHC, however, remain unestablished. METHODS: Sixty-five non-small cell lung cancer cases underwent AmoyDx ROS1 fusion real-time polymerase chain reaction (PCR) study and ROS-1 SP384 IHC tests, which were retrieved for analysis. ROS-1 IHC tests were interpreted based on the established classifiers as well as the presence of diffuse homogeneous immunoreactivity. The diagnostic accuracies of these ROS-1 IHC interpretation methods were evaluated by comparing them with the ROS1 real-time PCR results. RESULTS: Previous ROS-1 IHC classifiers demonstrated high sensitivity for positive ROS1 real-time PCR results (100%), but they showed low specificities (25%-50%) and overall accuracies (58%-72%). In contrast, the diffuse homogeneous ROS-1 immunoreactivity predicted positive ROS1 real-time PCR results with much higher specificity (94%) and overall accuracy (95%), albeit with a slightly lower sensitivity (97%). Some cases that showed discrepancy between diffuse homogeneous ROS-1 immunoreactivity and real-time PCR results involved rare ROS1::LDLR fusion and suboptimal IHC staining. CONCLUSIONS: A 3-tier reporting system for ROS-1 SP384 IHC testing combining previous interpretation criteria and diffuse and homogeneous immunoreactivity may better predict ROS1 fusion status without decreasing specificity.

Role of WT1 in Measurable Residual Disease Follow-Up in the Post Allogeneic Stem Cell Transplant Setting.

Objectives: The Wilms' tumor gene 1 (WT1) plays a critical role in cell development and the regulation of essential genes involved in cell growth and metabolism. In the context of hematopoietic tumors, including acute myeloid leukemia (AML), WT1 has been identified as a potential marker for measurable residual disease (MRD) assessment. Relapse after allogeneic hematopoietic stem cell transplantation (allo-SCT) remains a significant challenge in AML treatment, highlighting the importance of MRD monitoring for risk stratification and treatment decisions. This study aimed to investigate the clinical significance of WT1 as a molecular marker for MRD and its correlation with chimerism in AML patients post-allo-SCT setting. Methods: We have included 58 patients with WT1-expression-positive acute myeloid leukemia (AML) who received allo-SCT in our center between 2016-2022. The exclusion criteria are as follows: not having WT1 polymerase chain reaction (PCR) measurement at diagnosis, not receiving allo-SCT, and not having a serial measurement of WT1 post-transplant. Pre- and post-transplant assessments were made with flow cytometry, WT1 PCR, and bone marrow morphological evaluations. Statistical analyses were carried out to explore correlations between WT1 levels, MRD markers, and chimerism post-transplantation. Results: We found that WT1 had a significant correlation with flow cytometry and bone marrow morphological evaluation, but not with chimerism. Interestingly, high WT1 expressors exhibited a more robust correlation with chimerism compared to the general cohort. The negative predictive value for post-allo-SCT relapse was 91.8% for the whole WT1 cohort; for high WT1 expressors, it was similar, at 87.5%. The negative predictive value for post-allo-SCT relapse was high for the whole WT1 cohort; for high WT1 expressors, it was similar. The WT1 MRD assay showed a high negative predictive value for post-allo-SCT relapse, consistent across both the entire cohort (91.8%) and high WT1 expressors (87.5%). Conclusions: WT1 expression levels may serve as a valuable ancillary marker in MRD assessment and relapse prediction post-allo-SCT in AML patients, particularly for those lacking specific fusion genes or mutations. However, further large-scale, controlled studies are needed to standardize WT1 MRD assays and establish clear guidelines for their clinical application.

Advances in Malaria Diagnostic Methods in Resource-Limited Settings: A Systematic Review.

Malaria continues to pose a health challenge globally, and its elimination has remained a major topic of public health discussions. A key factor in eliminating malaria is the early and accurate detection of the parasite, especially in asymptomatic individuals, and so the importance of enhanced diagnostic methods cannot be overemphasized. This paper reviewed the advances in malaria diagnostic tools and detection methods over recent years. The use of these advanced diagnostics in lower and lower-middle-income countries as compared to advanced economies has been highlighted. Scientific databases such as Google Scholar, PUBMED, and Multidisciplinary Digital Publishing Institute (MDPI), among others, were reviewed. The findings suggest important advancements in malaria detection, ranging from the use of rapid diagnostic tests (RDTs) and molecular-based technologies to advanced non-invasive detection methods and computerized technologies. Molecular tests, RDTs, and computerized tests were also seen to be in use in resource-limited settings. In all, only twenty-one out of a total of eighty (26%) low and lower-middle-income countries showed evidence of the use of modern malaria diagnostic methods. It is imperative for governments and other agencies to direct efforts toward malaria research to upscale progress towards malaria elimination globally, especially in endemic regions, which usually happen to be resource-limited regions.

S. aureus bacteremia with acute transverse myelitis case report: utility of molecular techniques.

We report a rare case of bacteremia with concomitant acute transverse myelitis (ATM) without evidence of a primary infectious focus or secondary localization due to Staphylococcus aureus in a 60-year-old man admitted for hyperpyrexia, quadriplegia, and respiratory failure. Bacterial ATM is a rare clinical entity with confusing clinical presentation and challenging diagnosis; isolated bacterial infections of the spinal cord without secondary localization or contiguous foci are exceptionally rare, as is S. aureus as the cause of infection. In this case, a rapid etiologic diagnosis was made possible by close collaboration between clinicians, infectious disease specialists and clinical microbiologists combined with extended molecular testing on CSF guided by incoming results.

Conventional PCR-based versus next-generation sequencing-based approach for T-cell receptor γ gene clonality assessment in mature T-cell lymphomas: A phase 3 diagnostic accuracy study.

Background: Clonality assessment is currently the major molecular analysis utilized to support the diagnosis of suspicious lymphoid malignancies. Clonal rearrangements of the V-J segments of T-cell receptor G chain locus (TCRγ or TRG) have been observed in almost all types of T neoplasms, such as T-cell-related non-Hodgkin lymphomas and leukemias. At present, the gold standard for clonality evaluation is multiplex polymerase chain reaction (PCR), plus subsequent capillary electrophoresis/heteroduplex analyses, and/or Sanger sequencing. This approach overcomes the problem with the conventional Southern blot hybridization and is more efficient, simple, fast, and reproducible. In the recent years, the new next-generation sequencing (NGS) technologies provided alternative techniques for the analysis of antigen receptors genes, which presented several advantages, such as increased efficiency, specificity (SP), sensitivity (ST), resolution, and objectivity of the results, leading to a better classification, stratification, and monitoring of lymphoid malignancies. Nonetheless, these technologies are still far from being the new gold standard since further studies are warranted to prove their utility. The present study aimed to assess the diagnostic accuracy of these two methods by comparing a commercial NGS-based assay for the evaluation of TRG locus with the gold standard PCR-based one, to fulfill the requirements of a phase 3 diagnostic accuracy study. Methods: We assessed the TRG gene rearrangements in 72 cases using the conventional and highly-validated PCR-based assay proposed by EuroClonality consortium, an alternative commercial PCR-based assay, namely, IdentiClone® TCR Gamma Gene Rearrangement Assay 2.0, and a commercial NGS-based assay, that is, Invivoscribe LymphoTrack® Dx MiSeq® (both by Invivoscribe Technologies Inc., San Diego, CA, USA), to determine the diagnostic accuracy of the latter, and compare them with reference diagnoses made based on observation of clinical manifestations, cytohistological, and immunohistochemical analyses. Statistical values were calculated using the Oxford CATmaker software package. Results: Using standardized criteria of interpretation, the obtained results showed a diagnostic accuracy of 90.3% (correspondence in 65 out of 72 cases) of the test under investigation, with a ST of 86%, a SP of 95%, a positive predicting value of 94%, and a negative predicting value of 88%, demonstrating that it had high efficiency and reliability in detecting clonal TRG gene rearrangements in T-cell non-Hodgkin lymphomas. Conclusions: This diagnostic accuracy study yielded comparable results using a validated PCR-based approach and a new NGS-based one. Subsequent studies and cost-effectiveness evaluation are needed to put the NGS-based clonality assessment into routine diagnostic practice.

Efficacy and Safety of Valganciclovir in Congenital Cytomegalovirus Infection with Isolated Intrahepatic Cholestasis: A Randomized Controlled Trial.

Purpose: Cytomegalovirus (CMV) infection affects the hepatic, neurologic, hematopoietic, respiratory, gastrointestinal, and other organs, resulting in a high mortality rate and long-term sequelae. It may cause acute or chronic hepatitis, or even lead to hepatic cirrhosis. Valganciclovir (VGCV) is an effective, safe, and well-tolerated treatment for congenital CMV infection, without any serious adverse effects. This study was conducted to evaluate the clinical, biochemical, and virological profiles of infants with CMV with intrahepatic cholestasis and to determine the outcomes with or without treatment with VGCV. Methods: Twenty infants aged <6 months diagnosed with congenital CMV infection with evidence of intrahepatic cholestasis were included in this study. Randomization was used to divide the study participants into 2 groups. The control group (n=10) was treated with only supportive management, and the intervention group (n=10) was treated with oral VGCV at 16 mg/kg/dose 12 hours a day for 6 weeks plus supportive treatments. Physical examinations and biochemical, serological, and virological tests were performed at the time of diagnosis and at the end of 6 weeks and 6 months. Results: The control and intervention groups were compared in terms of clinical and laboratory parameters such as jaundice, dark urine, pale stool, hepatomegaly, total bilirubin, aminotransferases, gamma-glutamyl transferase, alkaline phosphatase, and CMV polymerase chain reaction load, which showed a significant reduction after treatment in the intervention group (p<0.05) with oral VGCV, with very few side effects, whereas the control group showed no significant changes. Conclusion: Oral VGCV can be used to effectively treat CMV infection with intrahepatic cholestasis without notable side effects.

Unveiling the genetic landscape of Streptococcus agalactiae bacteremia: emergence of hypervirulent CC1 strains and new CC283 strains in Tehran, Iran.

BACKGROUND: The emergence of Streptococcus agalactiae infections in patients with bacteremia is increasing. It is crucial to investigate the epidemiology, molecular characteristics, biofilm status, and virulence analysis of Streptococcus agalactiae in these patients. METHODS: In this cross-sectional study, 61 S. agalactiae isolated from blood infection were subjected to characterization through antimicrobial susceptibility tests, biofilm formation, multilocus sequence typing (MLST), and PCR analysis for detecting resistance (tet and erm family) and virulence genes (alp2/3, alp4, bca, bac, eps, rib, lmb, cylE, and pilus island genes). RESULTS: Overall, 32.7% of the isolates demonstrated an inducible clindamycin resistance phenotype. The results showed that 49.2, 24.6, and 8.2% of confirmed Streptococcus agalactiae strains were classified as strong, intermediate, and weak biofilm-forming strains, respectively. tet(M) (57.1%) was recovered most, followed by tet(M) + tet(L) (14.3%), tet(S) + tet(K) (10.7%), tet(M) + tet(K) (8.9%), tet(M) + tet(K) + tet(O) (5.4%), and tet(S) + tet(L) + tet(O) (3.6%). Three virulence gene profiles of cylE, lmb, bca, rib (24.6%; 15/61), cylE, lmb, rib, alp3 (19.7%; 12/61), and cylE, lmb, bac, rib (16.4%; 10/61) were detected in approximately two-thirds of the isolates. MLST revealed that the 61 isolates belonged to six clonal complexes, including CC1 (49.2%), followed by CC12 (18%), CC19 (13.1%), CC22 (9.8%), CC17 (6.6%), and CC283 (3.3%), and 11 different sequence types (STs), including ST1 (24.6%), ST7 (14.8%), ST918 (13.1%), ST2118 (9.8%), ST19 (9.8%), ST48 (6.6%), ST1372 (4.9%), ST22 (4.9%), ST40 (4.9%), ST734 (3.3%), and ST283 (3.3%). Remarkably, all CC1 and CC12 isolates, three-fourths of CC19, and half of CC22 were confirmed as biofilm producers. Conversely, CC17 and CC28 isolates were found to be nonproducers. The occurrence of strong biofilm formation was limited to specific CCs, namely CC1 (34.4%), CC12 (8.2%), CC19 (3.3%), and CC22 (3.3%). CONCLUSION: The high prevalence of CC1 and CC12 clones among S. agalactiae strains reflects the emergence of these lineages as successful clones in Iran, which is a serious concern and poses a potential threat to patients.

Identification and Validation of an Invasion-Related Disease-Free Survival Prognostic Model for Tongue Squamous Cell Carcinoma.

INTRODUCTION: Tongue squamous cell carcinoma (TSCC) is a common malignant tumour type with aggressive invasion and a poor prognosis. To date, invasion-related gene expression signatures for the prognostic stratification of TSCC patients are unavailable in clinical practice. This study aimed to assess the impact of invasion-related genes on the prognosis of TSCC patients. METHODS: We obtained mRNA profiles and clinical data from The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) databases (TCGA-TSCC and GSE41116, respectively). The TSCC samples from the TCGA-TSCC cohort were randomly divided into TCGA training and TCGA test datasets at a 7:3 ratio. Next, a disease-free survival (DFS) prognostic risk model was established on the basis of univariate and stepwise multivariate Cox regression analyses of the TCGA training cohort. Moreover, prognostic genes were screened. The model was subsequently evaluated and validated using the TCGA test and GSE41116 datasets. In addition, the prognostic genes were validated in the human TSCC cell line UM1 and the human oral keratinocyte (HOK) cell line using quantitative real-time polymerase chain reaction (qRT-PCR) analysis. RESULTS: A total of 70 candidate genes related to invasion were identified in the TCGA-TSCC cohort. DFS data were subsequently constructed, and 6 prognostic genes, HMGN2, MYL12B, ACTB, PPP1CA, PSMB9, and IFITM3, were identified. The TSCC samples were divided into high- and low-risk groups in the TCGA training, TCGA test, and GSE41116 cohorts, respectively. In particular, patients with TSCC in the low-risk group had longer DFS than those in the high-risk group. Furthermore, qRT-PCR analysis confirmed that the expression levels of the 6 prognostic genes were significantly greater in the TSCC cell line UM1 than in the HOK cell line. CONCLUSION: This study identified new invasion-related target genes related to poor prognosis in TSCC patients, providing new insights into the underlying mechanisms of TSCC invasion.