Diagnosis of Mucormycosis Using Frozen Section, Histopathology, Culture, and Reverse Transcriptase Polymerase Chain Reaction (RT-PCR) Techniques: A Comparative Study.

Mucormycosis usually occurs in immunocompromised patients or those with uncontrolled diabetes. Along the third wave of SARS-CoV-2, an associated angioinvasive opportunistic infection with Mucor, a life-threatening fungal infection, was rampant and emerging. With an increase in the usage of steroids in the COVID scenario, the rate of mucormycosis did take a rapid and alarming increase in King Edward Memorial Hospital, Pune, India. Any delay in the diagnosis and management of the disease was life-threatening. The most conventional methods to diagnose mucormycosis are microbiological culture and histopathology of the tissue. The microbiological culture method plays an important role in the diagnosis of mucormycosis. However, the technique is labour-intensive, taking seven to eight days. Histopathology leads to false-negative reports if the tissue is not biopsied from representative sites. On the other hand, molecular methods are rapid, reliable, and applicable to different body samples, such as tissue, paraffin-embedded tissue blocks, plasma, and urine. We aimed to use a reverse transcriptase polymerase chain reaction (RT-PCR) method to detect Mucor in plasma samples. Due to a lack of availability of fresh samples, nucleic acid was extracted from the tissue sections of 69 cases diagnosed as Mucor by histopathology. These samples were subjected to RT-PCR using the MucorGenius kit (Pathonostics, Maastricht, Netherlands). A total of 57 tissue samples were sent for culture, and 49% of our cases were positive by culture and equally by RT-PCR. There was 80% sensitivity and 76% specificity between culture and PCR techniques. However, the use of blood/plasma for RT-PCR for early diagnosis of mucormycosis will be the method of choice.

L-Ergothioneine slows the progression of age-related hearing loss in CBA/CaJ mice.

The naturally occurring amino acid, l-ergothioneine (EGT), has immense potential as a therapeutic, having shown promise in the treatment of other disease models, including neurological disorders. EGT is naturally uptaken into cells via its specific receptor, OCTN1, to be utilized by cells as an antioxidant and anti-inflammatory. In our current study, EGT was administered over a period of 6 months to 25-26-month-old CBA/CaJ mice as a possible treatment for age-related hearing loss (ARHL), since presbycusis has been linked to higher levels of cochlear oxidative stress, apoptosis, and chronic inflammation. Results from the current study indicate that EGT can prevent aging declines of some key features of ARHL. However, we found a distinct sex difference for the response to the treatments, for hearing - Auditory Brainstem Responses (ABRs) and Distortion Product Otoacoustic Emissions (DPOAEs). Males exhibited lower threshold declines in both low dose (LD) and high dose (HD) test groups throughout the testing period and did not display some of the characteristic aging declines in hearing seen in Control animals. In contrast, female mice did not show any therapeutic effects with either treatment dose. Further confirming this sex difference, EGT levels in whole blood sampling throughout the testing period showed greater uptake of EGT in males compared to females. Additionally, RT-PCR results from three tissue types of the inner ear confirmed EGT activity in the cochlea in both males and females. Males and females exhibited significant differences in biomarkers related to apoptosis (Cas-3), inflammation (TNF-a), oxidative stress (SOD2), and mitochondrial health (PGC1a).These changes were more prominent in males as compared to females, especially in stria vascularis tissue. Taken together, these findings suggest that EGT has the potential to be a naturally derived therapeutic for slowing down the progression of ARHL, and possibly other neurodegenerative diseases. EGT, while effective in the treatment of some features of presbycusis in aging males, could also be modified into a general prophylaxis for other age-related disorders where treatment protocols would include eating a larger proportion of EGT-rich foods or supplements. Lastly, the sex difference discovered here, needs further investigation to see if therapeutic conditions can be developed where aging females show better responsiveness to EGT.

Virus-like particles derived from bacteriophage MS2 as antigen scaffolds and RNA protective shells.

The versatile potential of bacteriophage MS2-derived virus-like particles (VLPs) in medical biotechnology has been extensively studied during the last 30 years. Since the first reports showing that MS2 VLPs can be produced at high yield and relatively easily engineered, numerous applications have been proposed. Particular effort has been spent in developing MS2 VLPs as protective capsules and delivery platforms for diverse molecules, such as chemical compounds, proteins and nucleic acids. Among these, two are particularly noteworthy: as scaffolds displaying heterologous epitopes for vaccine development and as capsids for encapsulation of foreign RNA. In this review, we summarize the progress in developing MS2 VLPs for these two areas.

Hollow, nanosized protein particles have many potential uses. If they can be appropriately engineered, they may for example be able to carry therapeutic cargoes to diseased cells or be used as a vaccine where appropriate antigens are mounted on their external surface. Many viruses offer a ready-made protein particle, the capsid, which can be made hollow by exclusion of the viral genetic material. MS2 is a virus that targets bacteria ­ a bacteriophage ­ which is well characterized and has been developed over many years for a number of applications. It has particular promise for development as a vaccine and for RNA delivery, both of which are reviewed here.

Molecular characterization and functional analysis of the ecdysone receptor isoform (EcR) from the oriental fruit moth Grapholita molesta (Lepidoptera: Tortricidae).

20-Hydroxyecdysone (20E) plays a vital role in a series of biological processes, via the nuclear receptors, EcR/USP by activating the ecdysone regulatory cascade. To clarify the role of EcR during the development of Grapholita molesta, the complementary DNA of ecdysone receptor isoform B1 (GmEcR-B1) was obtained from the transcriptome of G. molesta and verified by PCR. Alignment analysis revealed that the deduced protein sequence of GmEcR-B1 was highly homologous to EcR proteins identified in other lepidopteran species, especially the EcR-B1 isoform in Spodoptera litura. Quantitative real-time PCR showed that GmEcRs was expressed at all test developmental stages, and the expression level of GmEcRs was relatively higher during the period of the 3rd day of fifth instar larvae to 2nd of pupa than those in other stages. Moreover, the messenger RNA of GmEcRs was much more strongly expressed in the Malpighian tubule and epidermis than those in other tissues, which suggests that this gene may function in a tissue-specific manner during larval development. Silencing of GmEcRs could significantly downregulate the transcriptional level of ecdysone-inducible genes and result in increased mortality during metamorphosis and prolonged prepupal duration. Taken together, the present results indicate that GmEcRs may directly or indirectly affect the development of G. molesta.

Effectiveness of a standardized quality control management procedure for COVID-19 RT-PCR testing: a large-scale diagnostic accuracy study in China.

BACKGROUND: The study aimed to construct a standardized quality control management procedure (QCMP) and access its accuracy in the quality control of COVID-19 reverse transcriptase-polymerase chain reaction (RT-PCR). METHODS: Considering the initial RT-PCR results without applying QCMP as the gold standard, a large-scale diagnostic accuracy study including 4,385,925 participants at three COVID-19 RT-PCR testing sites in China, Foshan (as a pilot test), Guangzhou and Shenyang (as validation sites), was conducted from May 21, 2021, to December 15, 2022. RESULTS: In the pilot test, the RT-PCR with QCMP had a high accuracy of 99.18% with 100% specificity, 100% positive predictive value (PPV), and 99.17% negative predictive value (NPV). The rate of retesting was reduced from 1.98% to 1.16%. Its accuracy was then consistently validated in Guangzhou and Shenyang. CONCLUSIONS: The RT-PCR with QCMP showed excellent accuracy in identifying true negative COVID-19 and relieved the labor and time spent on retesting.

Exploration of the genetic influence of MYOT and MB genes on the plumage coloration of Muscovy ducks.

Plumage color, a pivotal attribute delineating diverse Muscovy duck strains, assumes considerable significance within the field of Muscovy duck breeding research. This study extends the existing research by delving into the hereditary aspects of genes associated with plumage coloration in Muscovy ducks. The principal objective is to discern marker genes conducive to targeted breeding strategies based on plumage color, thereby furnishing indispensable technical foundations for the development of novel Muscovy duck varieties. Our investigation focused on scrutinizing the impact of MYOT and MB genes on the genetic expression of plumage color at both the RNA and protein levels in Muscovy ducks. The results elucidate that black Muscovy ducks manifest markedly elevated mRNA and protein expression levels of MYOT and MB genes in comparison to their white counterparts, indicating that both genes may play a constructive regulatory role in the context of plumage coloration in Muscovy ducks. The outcomes of this study delineate a discernible correlation between MYOT and MB genes and the plumage coloration in Muscovy ducks. Employing gene expression analysis, we successfully identified candidate genes that may be intricately linked to the determination of plumage color in these ducks.

Significance of the Wnt signaling pathway in coronary artery atherosclerosis.

Introduction: The progression of coronary atherosclerosis is an active and regulated process. The Wnt signaling pathway is thought to play an active role in the pathogenesis of several cardiovascular diseases; however, a better understanding of this system in atherosclerosis is yet to be unraveled. Methods: In this study, real-time quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) and Western blotting were used to quantify the expression of Wnt3a, Wnt5a, and Wnt5b in the human coronary plaque, and immunohistochemistry was used to identify sites of local expression. To determine the pathologic significance of increased Wnt, human vascular smooth muscle cells (vSMCs) were treated with Wnt3a, Wnt5a, and Wnt5b recombinant proteins and assessed for changes in cell differentiation and function. Results: RT-PCR and Western blotting showed a significant increase in the expression of Wnt3a, Wnt5a, Wnt5b, and their receptors in diseased coronary arteries compared with that in non-diseased coronary arteries. Immunohistochemistry revealed an abundant expression of Wnt3a and Wnt5b in diseased coronary arteries, which contrasted with little or no signals in normal coronary arteries. Immunostaining of Wnt3a and Wnt5b was found largely in inflammatory cells and myointimal cells. The treatment of vSMCs with Wnt3a, Wnt5a, and Wnt5b resulted in increased vSMC differentiation, migration, calcification, oxidative stress, and impaired cholesterol handling. Conclusions: This study demonstrates the upregulation of three important members of canonical and non-canonical Wnt signaling pathways and their receptors in coronary atherosclerosis and shows an important role for these molecules in plaque development through increased cellular remodeling and impaired cholesterol handling.

Biological and molecular characterization of citrus bent leaf viroid.

Background and aim: Citrus bent leaf viroid (CBLVd) is one of the emerging and widely distributed viroids in citrus-growing areas of the world, including Pakistan. Previously, CBLVd has been reported in Pakistan for the first time in 2009. Therefore, characterization of CBLVd is required to monitor the viroid status in the citrus orchards concerning citrus decline. Methods: Biological and molecular characterization of CBLVd was studied through biological indexing and confirmation through RT-PCR, followed by phylogenetic analysis of selected CBLVd isolates. Among four citrus cultivars viz., Kinnow (Citrus nobilis × Citrus deliciosa), Mosambi (C. sinensis), Futrell's Early (C. reticulata) and Lemon (C. medica) used as indicator plants for two transmission trials viz., graft inoculation and mechanical inoculation. Graft inoculation was more efficient than mechanical inoculation. Results: Symptoms such as mild mosaic, slight backward leaf bending, and leaf curling were observed after eight months' post-inoculation. Citrus nobilis × Citrus deliciosa, C. reticulata and C. sinensis were more sensitive to CBLVd as compared to C. medica. Inoculated plants were reconfirmed through RT-PCR amplicons of 233 bp. The phylogenetic tree of submitted sequences showed more than 90% relevance of CBLVd in Pakistan compared to the rest of the world. Conclusions: There was slight genetic variability, but more than 90% relevance was found among the submitted and already reported CBLVd isolate from Pakistan. Scanty literature is available regarding the biological and molecular studies of CBLVd in Pakistan. Therefore, the transmission and molecular characterization of CBLVd in Pakistan were studied for the first time.

Profiling of Groundnut bud necrosis orthotospovirus-responsive microRNA and their targets in tomato based on deep sequencing.

Tomato, an extensively cultivated vegetable crop produces miRNAs in response to infection with Groundnut bud necrosis orthotospovirus, a viral pathogen causing significant economic losses. High-throughput miRNA sequencing was performed on tomato leaves inoculated with GBNV and mock-inoculated leaves as controls. Analysis revealed 73 known miRNAs belonging to 24 miRNA families, with variable expression levels. Interestingly, 39 miRNAs were upregulated, and 34 were downregulated in response to GBNV infection. Stem-loop quantitative reverse transcription PCR validated the differential expression of selected miRNAs. Additionally, 30 miRNA encoded proteins were identified to be involved in disease resistance and susceptibility. The miRNA-target interactions were found to play significant roles in cellular and metabolic activities, as well as modulating signaling pathways during the plant-virus interaction. The findings shed light on the intricate regulatory network of miRNAs in tomato response to viral infection and may contribute to developing strategies for improving crop protection against viral diseases.

Determination of anti-cancer effects of Nigella sativa seed oil on MCF7 breast and AGS gastric cancer cells.

BACKGROUND: This study aimed to investigate the cytotoxic, apoptotic, invasion, metastasis, and heat shock proteins (HSPs) effects of N. sativa oil on breast and gastric cancer cells. METHODS: We assessed the cytotoxic and apoptotic effects of various concentrations of N. sativa oil (10-50-100-200 µg/mL) on MCF7 breast cancer and AGS, an adenocarcinoma of the gastric cell line, at 24, 48 and 72 h using the MTT test. Additionally, the expression of the Caspase-3, BCL2/Bax, MMP2-9 and HSP60-70 gene was examined using RT-PCR in cell lines treating with N. sativa. RESULTS: The MTT experiments demonstrate that N. sativa has a time and dose-dependent inhibitory effect on the proliferation of MCF7 and AGS cancer cells. The vitality rates of MCF7 and AGS cells treated with N. sativa were 77.04-67.50% at 24 h, 65.28-39.14% at 48 h, and 48.95-32.31% at 72 h. The doses of 100 and 200 µg/mL were shown to be the most effective on both cancer cells. RT-PCR analysis revealed that N. sativa oil extract increased caspase-3 levels in both cell lines at higher concentrations and suppressed BCL2/Bax levels. Exposure of MCF7 and AGS cell lines to N. sativa caused a significant decrease in the expression of MMP2-9 and HSP60-70 genes over time, particularly at a dosage of 200 µg/mL compared to the control group (p < 0.05). CONCLUSIONS: Our findings indicate that N. sativa oil has a dose-dependent effect on cytotoxicity and the expression of apoptotic, heat shock proteins, and matrix metalloproteinases genes in breast and gastric cancer.

The CoLab score is associated with SARS-CoV-2 viral load during admission in individuals admitted to the intensive care unit: the CoLaIC cohort study.

OBJECTIVES: The present study examines the temporal association between the changes in SARS-CoV-2 viral load during infection and whether the CoLab-score can facilitate de-isolation. METHODS: Nasal swabs and blood samples were collected from ICU-admitted SARS-CoV-2 positive patients at Maastricht UMC+ from March 25, 2020 to October 1, 2021. The CoLab-score was calculated based on 10 blood parameters and age and can range from -43 to 6. Three mixed effects analyses compared patient categories based on initial PCR Ct values (low; Ct≤20, mid; 20>Ct≤30, high; Ct>30), serial PCR Ct values to CoLab-scores over time, and the association between within-patient delta Ct values and CoLab-scores. RESULTS: In 324 patients, the median Ct was 33, and the median CoLab-score was -1.78. Mid (n=110) and low (n=41) Ct-categories had higher CoLab-scores over time (+0.60 points, 95 % CI; 0.04-1.17, and +0.28 points, 95 % CI -0.49 to 1.04) compared to the high Ct (n=87) category. Over time, higher serial Ct values were associated with lower serial CoLab-scores, decreasing by -0.07 points (95 % CI; -0.11 to -0.02) per day. Increasing delta Ct values were associated with a decreasing delta CoLab-score of -0.12 (95 % CI; -0.23; -0.01). CONCLUSIONS: The study found an association between lower viral load on admission and reduced CoLab-score. Additionally, a decrease in viral load over time was associated with a decrease in CoLab-score. Therefore, the CoLab-score may make patient de-isolation an option based on the CoLab-score.

Association Between Multiple Single Nucleotide Polymorphisms in Folate Metabolism Pathway and Breast Cancer Risk in Georgian Women: A Case-Control Study.

Background: The folate metabolism pathway plays an integral part in DNA synthesis, methylation, and repair. Methylenetetrahydrofolate reductase (MTHFR) and methylenetetrahydrofolate dehydrogenase (MTHFD1) are both enzymes that are involved in this pathway, and the single nucleotide polymorphisms (SNPs) in genes coding for them have modulatory effects on DNA expression. This study aimed to investigate the relationship between MTHFR C677T (rs1801133) and MTHFD1 G1958A (rs2236225) polymorphisms and the risk of developing breast cancer in Georgian women. Methods: A case-control study was performed examining the MTHFR C677T and MTHFD1 G1958A SNP in breast cancer-confirmed cases and healthy matched controls. Real time-polymerase chain reaction (PCR) was used to genotype SNPs. The case individuals' pathology reports were obtained following surgeries for cancer characteristic data. Statistical analysis was performed to investigate the significance of the acquired data. Results: Statistical analysis of MTHFR C677T SNP revealed that the CT genotype increased the risk of breast cancer by 2.17 folds in the over-dominant model. Statistical analysis of MTHFD1 G1958A SNP showed that the GA genotype increased the risk of breast cancer by 4.12 folds in the codominant model and 2.41 folds in the over-dominant model. No statistically significant link was found between genotypes and lymph node status, however, patients with the CT genotype had higher percentages of proliferative activity. Conclusions: Breast cancer seems to have a statistically significant association with the CT genotype in MTHFR C677T and the GA genotype in MTHFD1 G1958A in Georgian women.

Zingiber officinale extract maximizes the efficacy of simvastatin as a hypolipidemic drug in obese male rats.

Obesity became a serious public health problem with enormous socioeconomic implications among the Egyptian population. The present investigation aimed to explore the efficacy of Zingiber officinale extract as a hypolipidemic agent combined with the commercially well-known anti-obesity drug simvastatin in obese rats. Thirty-five male Wister rats were randomly divided into five groups as follows: group I received a standard balanced diet for ten weeks; high-fat diet was orally administered to rats in groups II-V for ten weeks. From the fifth week to the tenth week, group III orally received simvastatin (40 mg/kg B.W.), group IV orally received Z. officinale root extract (400 mg/kg B.W.), and group V orally received simvastatin (20 mg/kg B.W.) plus Z. officinale extract (200 mg/kg B.W.) separately. Liver and kidney function tests, lipid profiles, serum glucose, insulin, and leptin were determined. Quantitative RT-PCR analysis of PPAR-γ, iNOS, HMG-CoA reductase, and GLUT-4 genes was carried out. Caspase 3 was estimated in liver and kidney tissues immunohistochemically. Liver and kidney tissues were examined histologically. The administration of Z. officinale extract plus simvastatin to high-fat diet-fed rats caused a significant reduction in the expression of HMG-coA reductase and iNOS by 41.81% and 88.05%, respectively, compared to highfat diet (HFD)-fed rats that received simvastatin only. Otherwise, a significant increase was noticed in the expression of PPAR-γ and GLUT-4 by 33.3% and 138.81%, respectively, compared to those that received simvastatin only. Immunohistochemistry emphasized that a combination of Z. officinale extract plus simvastatin significantly suppressed caspase 3 in the hepatic tissue of high-fat diet-fed rats. Moreover, the best results of lipid profile indices and hormonal indicators were obtained when rats received Z. officinale extract plus simvastatin. Z. officinale extract enhanced the efficiency of simvastatin as a hypolipidemic drug in obese rats due to the high contents of flavonoid and phenolic ingredients.

Detection of Avian Orthoavulavirus-1 genotypes VI.2.1 and VII.1.1 with neuro-viscerotropic tropism in some backyard pigeons (Columbidae) in Eastern Saudi Arabia.

Introduction: Avian orthoavulavirus-1 (AOAV1) has a wide host range, including domestic and wild birds. The present study aimed to identify the currently circulating AOAV1 strains from some outbreaks in some backyard pigeons in the eastern region of Saudi Arabia (ERSA). Methods: Tracheal/cloacal swabs and tissue specimens were collected from eight backyards in Al-Ahsa, ERSA, between January 2021 and March 2023. Samples were tested for the presence of AOAV1 using commercial real-time RT-PCR. Part of the fusion gene was also amplified by gel-based RT-PCR, and the obtained amplicons were sequenced. Results and discussion: AOAV1 was detected in samples from the eight flocks. The retrieved sequences from samples of 6/8 pigeon backyards are reported. Phylogenetic analysis based on the obtained sequences from these backyard pigeons showed the segregation of the obtained sequences in AOAV1 genotypes VI.2.1 and VII.1.1. Clinically, nervous manifestations were dominant in pigeons infected with both genotypes. Respiratory manifestations and significantly higher overall mortality rate were induced by genotype VI.2.1. The deduced amino acid sequences of the fusion protein cleavage site (FPCS) showed that all the detected isolates belong to velogenic strains. Differences in clinical profiles induced by the natural infection of pigeons with AOAV1 genotypes VI.2.1 and VII.1.1 were reported. The present findings highlight the potential roles of some backyard pigeons in the long-distance spread and cross-species transmission of the reported AOAVI genotypes. Further research is required to perform biotyping and pathotyping of the reported strains.

Infrared spectroscopy combined with chemometrics in transflectance mode: An alternative approach in the photodiagnosis of COVID-19 using saliva.

The Coronavirus Disease 2019 (COVID-19) pandemic, caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has required the search for sensitive, rapid, specific, and lower-cost diagnostic methods to meet the high demand. The gold standard method of laboratory diagnosis is real-time reverse transcription polymerase chain reaction (RT-PCR). However, this method is costly and results can take time. In the literature, several studies have already described the potential of Fourier transform infrared spectroscopy (FTIR) as a tool in the biomedical field, including the diagnosis of viral infections, while being fast and inexpensive. In view of this, the objective of this study was to develop an FTIR model for the diagnosis of COVID-19. For this analysis, all private clients who had performed a face-to-face collection at the Univates Clinical Analysis Laboratory (LAC Univates) within a period of six months were invited to participate. Data from clients who agreed to participate in the study were collected, as well as nasopharyngeal secretions and a saliva sample. For the development of models, the RT-PCR result of nasopharyngeal secretions was used as a reference method. Absorptions with high discrimination (p < 0.001) between GI (28 patients, RT-PCR test positive to SARS-CoV-2 virus) and GII (173 patients who did not have the virus detected in the test) were most relevant at 3512 cm-1, 3385 cm-1 and 1321 cm-1 after 2nd derivative data transformation. To carry out the diagnostic modeling, chemometrics via FTIR and Discriminant Analysis of Orthogonal Partial Least Squares (OPLS-DA) by salivary transflectance mode with one latent variable and one orthogonal signal correction component were used. The model generated predictions with 100 % sensitivity, specificity and accuracy. With the proposed model, in a single application of an individual's saliva in the FTIR equipment, results related to the detection of SARS-CoV-2 can be obtained in a few minutes of spectral evaluation.

Development and Validation of Three Triplex Real-Time RT-PCR Assays for Typing African Horse Sickness Virus: Utility for Disease Control and Other Laboratory Applications.

The African horse sickness virus (AHSV) belongs to the Genus Orbivirus, family Sedoreoviridae, and nine serotypes of the virus have been described to date. The AHSV genome is composed of ten linear segments of double-stranded (ds) RNA, numbered in decreasing size order (Seg-1 to Seg-10). Genome segment 2 (Seg-2) encodes outer-capsid protein VP2, the most variable AHSV protein and the primary target for neutralizing antibodies. Consequently, Seg-2 determines the identity of the virus serotype. An African horse sickness (AHS) outbreak in an AHS-free status country requires identifying the serotype as soon as possible to implement a serotype-specific vaccination program. Considering that nowadays 'polyvalent live attenuated' is the only commercially available vaccination strategy to control the disease, field and vaccine strains of different serotypes could co-circulate. Additionally, in AHS-endemic countries, more than one serotype is often circulating at the same time. Therefore, a strategy to rapidly determine the virus serotype in an AHS-positive sample is strongly recommended in both epidemiological situations. The main objective of this study is to describe the development and validation of three triplex real-time RT-PCR (rRT-PCR) methods for rapid AHSV serotype detection. Samples from recent AHS outbreaks in Kenia (2015-2017), Thailand (2020), and Nigeria (2023), and from the AHS outbreak in Spain (1987-1990), were included in the study for the validation of these methods.

Parvovirus B19 Outbreak in Israel: Retrospective Molecular Analysis from 2010 to 2023.

This study presents an analysis of the epidemiological trends of parvovirus B19 (B19V) in Israel from 2010 to 2023, with particular emphasis on the outbreak in 2023. The analysis utilized molecular diagnostic data from individual patients obtained at the Central Virology Laboratory. Between 2010 and 2022, 8.5% of PCR-tested samples were positive for B19V, whereas in 2023, this percentage surged to 31% of PCR-tested samples. Throughout the study period, annual cycles consistently peaked in early spring/summer, with the most recent prominent outbreak occurring in 2016. Predominantly, diagnoses were made in children and women aged 20-39. Despite the notable surge in 2023, over 80% of positive cases continued to be observed in children and young women, with a decrease in cases during winter months. Furthermore, genotype 1a of the virus remained the predominant strain circulating during the outbreak. In light of these circumstances, consideration should be given to implementing screening measures, particularly among high-risk groups such as pregnant women.

TGFß signaling pathway in salivary gland tumors.

OBJECTIVE: Pleomorphic adenoma (PA), mucoepidermoid carcinoma (MEC), and adenoid cystic carcinoma (ACC) are the most prevalent salivary gland tumors. Their pathogenesis has been recently associated with complex molecular cascades, including the TGFß signaling pathway. The aim of this study was to evaluate the expression of genes associated with the TGFß signaling pathway (TGFB1, ITGB6, SMAD2, SMAD4, FBN1, LTBP1, and c-MYC) to map possible downstream alterations in the TGFß cascade. DESIGN: Thirteen PA, 17 MEC, 13 ACC, and 10 non-neoplastic salivary gland samples were analyzed by real-time RT-PCR. RESULTS: Cases of PA presented increased TGFB1, LTPB1, c-MYC, and FBN1 expressions, whereas SMAD2 expression was decreased when compared to non-neoplastic tissue. MEC patients displayed increased expressions of TGFB1, ITGB6, FBN1, and c-MYC and decreased expressions of SMAD2 and SMAD4. ACC cases exhibited elevated expressions of the investigated genes except TGFB1. The present results suggest that decreased expression of SMAD2 and SMAD4 does not impede the transcriptional regulation of c-MYC, especially in PA and MEC. Increased expressions of ITGB6, TGFB1, LTBP1, and FBN1 appear to be related to the regulation of the TGFß signaling pathway in these tumors. Additionally, we observed a higher expression of SMAD4 in ACC and a raised expression of ITGB6 and lowered expression of SMAD2 in MEC. CONCLUSIONS: Our study demonstrated the differential expression of TGFß cascade members in salivary gland tumors such as SMAD2/SMAD4 and c-MYC as well as the participation of ITGB6, TGFB1, LTBP1, and FBN1, contributing to the understanding of the mechanisms involved in tumor progression.

Pooling of samples to optimise SARS-CoV-2 detection in nasopharyngeal swabs and gargle lavage self-samples for covid-19 diagnostics and surveillance.

BACKGROUND: Testing of pooled samples is an effective strategy for increasing testing capacity while saving resources and time. This study aimed to validate pooled testing and gather real-life data on its use for Covid-19 surveillance with a gargle lavage (GL) self-sampling strategy. METHODS: Two-stage pooled testing with pools of 6 and 12 samples was used for preventive testing of an asymptomatic population and Covid-19 surveillance in Czech schools. Both GL and nasopharyngeal swabs were used for sampling. RESULTS: In total, 61,111 samples were tested. The use of pooled testing for large-scale Covid-19 surveillance reduced consumable costs by almost 75% and increased testing capacity up to 3.8-fold compared to standard methods. RT-PCR experiments revealed a minimal loss of sensitivity (0-2.2%) when using pooled samples, enabling the detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) genes with Ct values >35. The minor loss of sensitivity was counterbalanced by a significantly increased throughput and the ability to substantially increase testing frequencies. CONCLUSIONS: Pooled testing is considerably more cost-effective and less time-consuming than standard testing for large-scale Covid-19 surveillance even when the prevalence of SARS-CoV-2 is fluctuating. Gargle lavage self-sampling is a non-invasive technique suitable for sample collection without a healthcare worker's assistance.