Routine Methods Research Articles

B-165 The EXENT System reliably distinguishes daratumumab from endogenous monoclonal immunoglobulins in multiple myeloma patients

Abstract Background The use of therapeutic monoclonal antibodies (t-mAbs) in the treatment of multiple myeloma (MM) has created challenges in the clinical laboratory. Co-migration of t-mAbs (predominantly IgGK) and the endogenous M-protein during serum protein electrophoresis can result in inability to distinguish between a patient’s M protein and the t-mAb. This can hinder accurate assessment of therapeutic responses or may lead to extensive workup with immunofixation electrophoresis and gel-shift assays for accurate confirmation of a complete response, or to exclude the emergence of a new clone. There is a need for an analytically specific routine method to identify t-mAb interference. MALDI-TOF mass spectrometry is an especially promising tool, as due to its ability to identify M proteins’ molecular mass, it has the potential to recognize any t-mAbs with known amino acid composition. We have used the EXENT® Immunoglobulin Isotypes (GAM) Assay for the EXENT Analyzer (The Binding Site, part of Thermo Fisher Scientific) to assess the reproducibility of daratumumab (anti-CD38, IgGK) detection in an analytical study, and assessed the EXENT System’s ability to detect daratumumab in a cohort of MM patients. Methods The expected mass to charge ratio (m/z) of the +2-charge state of daratumumab’s kappa light chain was calculated based on published information. Daratumumab was spiked into normal human serum at 1.0 g/L, and this material was tested in 5 replicates a day over 5 days. In addition, 109 samples from 34 MM patients receiving daratumumab, carfilzomib, lenalidomide and dexamethasone (Dara-KRd) treatment were also tested. Samples were obtained after 14 or 28 days following daratumumab administration in various treatment cycles, ranging from cycle 4 to end-of-treatment. Results The expected m/z for the +2-charge state of daratumumab’s kappa light chain was calculated as 11,691.01 to 11,693.1. In the reproducibility experiment, the median m/z was 11691.3, within the expected range. The difference between the lowest and highest m/z was 1.3 across the 25 replicates. Daratumumab was detected in 103 out of 109 samples (97.3%): 51/57 (89.5%) in IgGK samples, and in all IgGL (23) and IgA (29) samples. In two IgGK patients, where daratumumab was not detected in 2 out of 4 samples and in 1 out of 3 samples, respectively, the patients’ own M proteins were in the proximity (24.8 and 27.1 m/z) of daratumumab’s m/z. In the other three cases where daratumumab was not modeled by the algorithm, the daratumumab peak overlapped with the endogenous M-protein's adduct peak (estimated m/z 11,688) in one sample; it was masked by the shoulder of a large (8.42 g/L) IgGK M-protein in another sample; and no obvious reason was identified in the third sample. There was no difference in the detection rate between 14 days and 28 days post administration. Conclusions The EXENT Immunoglobulin Isotypes (GAM) Assay has successfully identified the presence of daratumumab in approximately 90% of IgGK patient samples in addition to all other (non-IgGK) MM samples. These results demonstrate the potential of the EXENT Immunoglobulin Isotypes (GAM) Assay for recognizing known t-mAb interference.

A comparative study of aptasensor vs. immunosensor for ultrasensitive detection of aflatoxin B1 using Ag-pSi SERS substrate

Numerous SERS based platforms have been designed to address the emerging need for detecting fungal metabolite contamination in foodstuffs, and specifically the Group 1 carcinogen aflatoxin B1. Herein, 4-aminothiophenol modified silver-coated porous silicon was used as the SERS substrate. Two ratiometric responses were individually assessed upon direct target capture using specific aptamers or antibodies. Under optimized physical features, elevated enhancement factor, wide dynamic range, low detection limits and pronounced recycling capabilities were achieved (7.39 × 107, 0.2–200 ppb, 0.0085 and 0.0110 ppb, 7 and 1 regeneration cycles without impairing the performances for aptasensor and immunosensor, respectively). The accuracy and anti-interference responses in several intricate matrices (maize, peanut, wheat, oats and rice) were compared to a routine HPLC method with equivalent recoveries. Overall, the comparative assessment revealed preferable features of reusability, durability and accuracy of the aptasensor over the immunosensor. Furthermore, the results demonstrate the substantial potential of the proposed SERS substrate for diverse on-site analytical applications using simple and portable monitoring instrumentation.

Identification methods as a factor affecting the performance of clinical microbiology laboratories participating in an external quality assessment program: a cross-sectional, retrospective analysis.

Introduction. Laboratory participation in external quality assessment (EQA) programmes including proficiency testing (PT) is a requirement of clinical laboratory conformance to ISO 15189:2022 Medical laboratories - Requirements for quality and competence. PT is one EQA method whereby laboratories are sent blinded samples for characterization by routine laboratory diagnostic methods. Importantly, PT enables a laboratory's performance to be evaluated in comparison to the standard reference methods and to the performance of other peer laboratories using similar diagnostic methods.Gap statement. The desired outcome of participating in PT is to help laboratories identify possible sources of error in each step of the total testing process and particularly in their test methods during the analytical phase.Aim. This cross-sectional study investigated the impact of using matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) compared to conventional phenotypic biochemical testing on laboratory performance in a clinical bacteriology PT scheme.Methodology. During a 6-year period from 2017-2022, the Canadian Microbiology Proficiency Testing implemented 112 PT challenges comprising 22 different sample types and included 61 different bacterial species. This was translated into 5883 graded test events for analysis. Multiple logistic regression techniques were employed to explore the association between the test method employed and laboratory performance. The sample type and aerobic classification of challenge organisms were included as confounding variables.Results. Laboratories using MALDI-TOF MS performed significantly better in characterizing microorganisms than laboratories using phenotypic biochemical testing alone [odds ratio OR = 5.68, confidence interval (CI): 3.92, 8.22] regardless of the sample type and aerobic classification. Notably, our analysis identified a significant association between anaerobic organisms and laboratory performance (OR: 0.24, CI: 0.17-0.35), suggesting that culturing and identifying fastidious organisms remains a significant obstacle for many clinical microbiology laboratories.Conclusions. Although no method is infallible and its performance will depend on the validation and quality assurance procedures, this finding may help the management in the decision for implementing MALDI-TOF MS in the microbiology laboratory. This study highlights the important role PT providers play in the objective assessment of laboratory performance and how it can provide evidence for quality improvement.

Microenvironmental alkalization promotes the therapeutic effects of MSLN-CAR-T cells

Triple-negative breast cancer (TNBC) is characterized by high invasion, prone metastasis, frequent recurrence and poor prognosis. Unfortunately, the curative effects of current clinical therapies, including surgery, radiotherapy, chemotherapy and immunotherapy,...

Clinical, Morphological and Multiplex Analysis of a Case of COVID-19-Associated Placental and Fetal Organ Damage at 16 Weeks Gestation

Abstract. COVID-19-associated damage to the placenta and fetal organs is possible among infected pregnant women. This phenomenon is associated with the presence of the virus in the placental tissue itself and transplacental transmission of SARS-CoV-2 from mother to fetus and occurs in case of severe course of the disease in the mother. However, reliable and clinically significant morphological changes in COVID-19-associated placental damage are still a subject of controversy and speculation. The aim of this article is to present a clinical case analysis and evaluation of the morphological features and results of multiplex analysis of COVID-19- associated fetal and placental tissue damage. Materials and methods. Placental and fetal tissue samples were examined using routine histological methods and immunohistochemical studies of the expression of antibodies to CD15 (MMA), CD3 (MRQ-39), CD68 (KP-1), CD138 (DB-A38). The spectroscopic study was performed using the InSpectr M spectrometer with an excitation wavelength of 532 nm. Results The study revealed an inflammatory infiltrate in the fetal membranes, in the umbilical vein wall, in the stroma of the villi, basal plate and intervillous space, forming thrombi of the vessels of the villi, chorionic plate, intervillous space, trophoblast necrosis, abundant deposition of perivillous fibrin. Spectroscopically, signs of changes in the quantitative and qualitative composition of the amino acids proline, phenylalanine, tyrosine and tryptophan, as well as the appearance of non-standard amino acids (hydroxyproline) and protein isoforms, signs of instability of the acid-base balance were revealed. Conclusion. Among the identified changes, both those characteristic of COVID-19-associated placental damage and non-specific ones associated with the body's inflammatory/cytokine response to COVID-19 and persistent placental infection are determined. The results of the multiplex analysis identified a promising direction in understanding the biological basis and essence of this pathology.

RESEARCH & DEVELOPMENT OF P2O5 DETERMINATION METHOD IN PHOSPHATE FERTILIZERS AND ITS VALIDATION

The validation of chemical measurement for Phosphorus determination in fertilizers followed a chain of experimental procedures to ascertain the performance characteristic (PC), connecting Precision, accuracy, reproducibility, recovery, linearity, limit of detection and of quantification. Performance characteristics are accomplished to ensure that analytical method fits the purpose of particular testing. This method was conducted in three stages those were preparation, testing and data processing. The standard test method for determination of P2O5 content in fertilizers used for regulatory purpose is somewhat lengthy and time taking. A modification was done to the standard method to create a more efficient approach that meets validation criteria and is suitable for analyzing advisory fertilizer samples. During the year 2023-24, comprehensive research and development activity were carried out at Soil and Water Testing Laboratory (SWTL) for Research, Faisalabad and analytical data was generated. The obtained results were processed statistically and compared with the acceptance requirements. The coefficient of determination (R2), precision and accuracy/recovery percentage respectively were 0.9996, 0.3-1.10% and 98.01-102.34%, while LOD and LOQ were 0.84 and 2.25%. Based on the results the study concluded that the modified test method applied to determine P2O5was appropriate for its intended use based on the acceptance criteria of performance characteristics set forth in the validation protocol and can be applied as routine method at laboratory.

Raman fiber-optic probe for rapid diagnosis of gastric and esophageal tumors with machine learning analysis or similarity assessments: a comparative study.

Gastric and esophageal cancers, the predominant forms of upper gastrointestinal malignancies, contribute significantly to global cancer mortality. Routine detection methods, including medical imaging, endoscopic examination, and pathological biopsy, often suffer from drawbacks such as low sensitivity and laborious and complex procedures. Raman spectroscopy is a non-invasive and label-free optical technique that provides highly sensitive biomolecular information to facilitate effective tumor identification. In this work, we report the use of fiber-optic Raman spectroscopy for the accurate and rapid diagnosis of gastric and esophageal cancers. Using a database of 14,000 spectra from 140 ex vivo tissue pieces of both tumor and normal tissue samples, we compare the random forest (RF) and our established Euclidean distance Raman spectroscopy (EDRS) model. The RF analysis achieves a sensitivity of 85.23% and an accuracy of 83.05% in diagnosing gastric tumors. The EDRS algorithm with improved diagnostic transparency further increases the sensitivity to 92.86% and accuracy to 89.29%. When these diagnostic protocols are extended to esophageal tumors, the RF and EDRS models achieve accuracies of 71.27% and 93.18%, respectively. Finally, we demonstrate that fewer than 20 spectra are sufficient to achieve good Raman diagnostic accuracy for both tumor tissues. This optimizes the balance between acquisition time and diagnostic performance. Our work, although conducted on ex vivo tissue models, offers valuable insights for in vivo in situ endoscopic Raman diagnosis of gastric and esophageal cancer lesions in the future. Our study provides a robust, rapid, and convenient method as a new paradigm in in vivo endoscopic medical diagnostics that integrates spectroscopic techniques and aRaman probe for detecting upper gastrointestinal malignancies.

Comparison of the effect of surfactant administration during nasal continuous positive airway pressure with that of nasal continuous positive airway pressure alone on respiratory distress syndrome

Background. The American Academy of Pediatrics states that prophylactic surfactant is no longer advised when continuous positive airway pressure (CPAP) is frequently used to stabilize preterm newborns and where the rate of prenatal corticosteroid is high. On the other hand, the European Consensus Guidelines on the Management of Respiratory Distress Syndrome (RDS) urge giving prophylactic surfactants to infants with RDS as early as possible. In this study, we aim to investigate the appropriate guidelines to be adopted in our neonatal intensive care unit about selective or elective INSURE (intubation-surfactant-extubation) method. Materials and methods. This is a clinical trial in which a total number of 120 neonate patients were diagnosed with RDS and had gestational age of 30–34 weeks. Two neonatal intensive care units have been included in the study: group A (n = 87) was treated with CPAP plus INSURE and group B (n = 33) was treated with CPAP only. The primary outcome we searched for is the fate of the patients. The secondary outcomes were length of hospital stay, CPAP management, oxygen therapy, and RDS complications. Results. Group B show a shorter length of hospital stay and CPAP treatment; however, this difference was not statistically significant. On the other hand, oxygen therapy duration was significantly shorter in group B than in group A. Both groups show very close rates of RDS complications (32 vs 33 %). Regarding the primary outcomes, group A had a better survival rate in comparison to group B (88.5 vs 78.8 %); however, this difference was not significant, with P-value of 0.173. Conclusions. In neonates with RDS, the routine INSURE method followed by CPAP gives no more superiority in outcomes (both primary, i.e. survival rate, and secondary, i.e. complications) than CPAP alone.

Effects of a single oral dose of azithromycin in laboring women on antimicrobial resistance (AMR) and the microbiome: a protocol for the Antimicrobial Resistance Sub-Study of the A-PLUS trial

Background The Azithromycin Prevention in Labor Use Study (A-PLUS), a large, multi-national randomized trial, was performed to evaluate the improvement of maternal and newborn outcomes following a single dose of azithromycin (AZ) given during labor. However, the immediate and long-term impact of this single dose on the microbiome and the development or prevalence of antimicrobial resistance are unknown. We designed a sub-study to assess AZ resistance of bacterial isolates from clinical infections and of three target bacteria (Staphylococcus aureus, Streptococcus pneumoniae, and Escherichia coli) from the serial surveillance of the nasopharynx and rectum. Additionally, the serially surveilled samples will be evaluated for changes to the microbiome and the resistome of the nasopharynx and rectum. Methods and Analysis As part of the large, randomized APLUS trial, maternal and neonatal clinical infections were monitored for up to 42 days postpartum, and samples collected for culture using site-specific routine methods. For this sub-study, cultured bacterial isolates will be assessed for AZ resistance using an antibiotic susceptibility method. Additionally, a random subset of maternal-neonatal dyads from the main trial will be selected for serial surveillance with aseptic swab collections of the nasopharynx and the rectum at baseline, and subsequently at 1-week, 6-weeks, 3-months, 6-months, and 12-months postpartum. The serial samples will undergo selective culturing of sentinel bacterial species and screened for AZ resistance. An additional set of serial samples will be stored for future microbiome and resistome analyses. Ethics and Dissemination The study protocol was reviewed, and ethics approval obtained from all the relevant ethical review boards at each research site. All participants will provide informed written consent prior to their enrollment. Results of the trial will be disseminated through peer-reviewed publications, presentations at relevant conferences, and at dissemination meetings with the ministries of health and local stakeholders at each research site. Trial Registration NCT03871491.

SIALOLITHIASIS: REVIEW

Objectives. Sialolithiasis is the most common obstructive salivary gland disease, responsible for approximately 50% of salivary glands pathology cases. The aim of this study is to provide the literature review to present the incident rate, etiology, diagnostic and treatment trends of sialolithiasis Materials and methods: Data from the Medline database, PubMed.gov and supplementary sources were used to conduct a systematic literature search. Results: The 116 studies were analyzed. The relevant data were exported, summarized and presented. Conclusion: The most of available studies regarding the incidence rate of sialolithiasis have been based on selected hospital data and there are a few multicenter analyses. The etiology of salivary stones formation remains not completely clear and various hypotheses have been put forward, thus research into etiologic factors is still continues. Despite the introduction of new technologies in the diagnostics of salivary gland, the routine methods like radiographs and ultrasound examination are still relevant and widely used. Gland preserving techniques in the treatment of sialolithiasis have continuously replaced radical surgery

ESKAPE pathogens in pediatric cardiac surgery patients: 5-year microbiological monitoring inatertiary hospital in Kazakhstan.

Hospital acquired infections greatly affect recovery and survival in pediatric surgical patients. We evaluated prevalence and antimicrobial resistance of ESKAPE pathogens in neonates and infants subjected to cardiac surgery in a tertiary hospital in Central Kazakhstan between 2019 and 2023 (2,278 patients) using routine methods of microbiological detection. ESKAPE pathogens were found in 1,899 out of 2,957 samples (Staphylococcus aureus - 35.3%, Klebsiella pneumoniae - 27.8%, Acinetobacter baumannii - 14.5%, Pseudomonas aeruginosa - 12.4%, Enterobacter sp. - 8.8%, Enterococcus faecium - 1.2%). The total prevalence of ESKAPE increased significantly from 45.1 to 76.9% (P = 0.005) during the study period. The resistance significantly increased in methicillin-resistant S. aureus (MRSA, from13.7 to 41.9%, P = 0.041) but decreased in carbapenem-resistant P. aeruginosa (from 64.3 to 37.7%, P = 0.037) and carbapenem-resistant A. baumannii (from 48.5 to 19.1%, P = 0.039). Gradual but non-significant changes were shown in third-generation cephalosporin resistant K. pneumoniae (from 63.6 to 45.2%) and carbapenem-resistant K. pneumoniae (from 0 to 8.3%). The relative prevalence of ESKAPE pathogens steadily increased in our pediatric cardiac surgery patients in 2019-2023. The most frequent were S. aureus, K. pneumoniae, and A. baumannii, with dramatically increasing tendencies for MRSA. Our results highlight the necessity for a well-designed infection control strategy and constant microbiological monitoring in pediatric cardiac surgery departments.

OCCURRENCE OF LAMPROGLENA PULCHELLA (NORDMANN, 1832) (COPEPODA: LERNAEIDAE) IN SOME CYPRINID FISH FROM OHRID LAKE (MACEDONIA)

The representatives of the class Copepoda are of great importance in fish pathology. Lamproglena pulchella (Nordmann, 1832) (Copepoda: Lernaeidae) is an ectoparasite of many cyprinid fish, widespread in Europe up to the Aral Sea. During the long-term investigations of 12 species of cyprinid fish from the Lake Ohrid, a total of 7 specimens of Lamproglena pulchella was found in 5 fish from 4 fish species: Rutilus ohridanus, Squalius squalus, Scardinius knezevici and Chondrostoma ohridanus. Only fresh fish were subjected to routine parasitological examinations, dissection, and observation methods. Parasite identification was performed morphologically, based on the character of their maxilla, maxilliped, antennae, legs and uropods, using a referent key for determination. The 3 parasite specimens were found in two samples of Rutilus ohridanus, followed by 2 parasite specimens in Squalius squalus and one specimen in Scardinius knezevici and Chondrostoma ohridanus, each. These parasites attach to gills and general body surfaces by their maxilla and maxilliped, causing wounds that become the spots of secondary infection by microbes. The damage caused by this parasite is more pronounced in aquaculture systems. We did not find many parasites, but considering the potential danger that Lamproglena pulchella poses to the fish health, we point out the need to monitor infestations with this parasite and take preventive measures so that the parasite does not spread massively among fish in Lake Ohrid and to prevent its introduction into other waters in Macedonia. The record of Lamproglena pulchella in all four cyprinid fish in the present study is considered as the first from the Ohrid Lake. At the same time, Rutilus ohridanus, Scardinius knezevici and Chondrostoma ohridanus represent new hosts for this parasite, worldwide.

Ability assessment of the craniofacial structures radiological anatomy on panoramic radiography among dental student during pandemic of Covid-19. Ability assessment on OPG among dental student during pandemic of Covid-19.

Orthopantomography (OPG) is a routine imaging method in dental practice and an essential di- agnostic tool in dentistry. However, OPGs are challenging to interpret due to many overlapping structures. Graduates of dental schools should be aware of image distortions caused by various factors and be able to distinguish them from typical structures to make an accurate diagnosis. The aim was to determine the correlation between the knowledge regarding the location of craniofacial structures of the 1st through 3rd- year dental students and the ability to recognized them on OPGs. The study was conducted in 2021 on 131 dental students using the Microsoft Teams program. Each participant had to determine the location of 4 anthropometric points on 4 OPGs. Using proprietary software, the authors determined the Articular angle between them. The researchers performed the statistical analysis. The Bioethics Committee approved the research. The results of students from particular years of studies did not show statistical significance. There was no statistically significant difference between males and females. Only the answers from third-year male students regarding the one Articular angle showed statistically significant differences compared to the rest of the participants. Recognizing joint structures on OPG is very important from the clinical point of view, although it is often overlooked in diagnostics. Students in the first years of dental school acquire the ability to assess and analyze TMJ on OPGs properly. OPGs have many advantages, including high availability, low cost and low radiation dose. In conclusion we can say that in the first three years of studies, dental students' image assessment skills in the craniofacial radiology field remain the same.

Optimization of the Diagnosis of Central Nervous System Infections in Vietnamese Hospitals: Results From a Retrospective Multicenter Study.

Central nervous system infections pose significant health challenges, particularly in low- and middle-income countries, because of high morbidity and mortality rates. Rapid and accurate diagnosis is essential for effective treatment to prevent adverse outcomes. Traditional culture-based diagnostics are often slow and lack specificity. This study evaluates the BioFire FilmArray Meningitis/Encephalitis (FAME) Panel against standard diagnostics in Vietnam to assess its clinical impact and suitability for local epidemiology. We conducted a prospective study involving 330 patients with suspected central nervous system infections at 4 hospitals in northern Vietnam from July 2022 to April 2023. Cerebrospinal fluid samples were analyzed using routine culture methods and FAME. We compared pathogen detection rates and assessed the potential clinical impact of FAME results on patient management. Of the 330 cerebrospinal fluid specimens, 64 (19%) were positive by either conventional diagnostics (n = 48) and/or FAME (n = 33). The agreement between FAME and conventional diagnostics was 87%. Key pathogens Mycobacterium tuberculosis (n = 7), Klebsiella pneumoniae (n = 5), Streptococcus suis (n = 5), Epstein-Barr virus (n = 3), Acinetobacter baumannii (n = 1), and Trichosporon asahii (n = 1) were not detected by FAME. Classical meningitis parameter clinical symptoms, altered glucose, protein, and pleocytosis were good predictors of FAME positivity, indicating their utility in optimizing local diagnostic algorithms. FAME complements traditional diagnostics by offering rapid and broad pathogen detection, crucial for timely and appropriate therapy. However, its effectiveness varies with local epidemiology, and it should not replace conventional methods entirely. Tailoring diagnostic panels to regional pathogen prevalence is recommended to enhance diagnostic accuracy and clinical outcomes in low- and middle-income countries.

Comparison of the effect of chewing gum with routine method on ileus after burns: a randomized clinical trial

BackgroundChewing gum is a healthy, cheap, and familiar solution for patients with premature irritation of the stomach and intestines. This study compared the effect of chewing gum and the routine method on ileus after burns.MethodsThis study is a randomized clinical trial conducted in Valiasr Hospital in Arak, Markazi Province, in the center of Iran, from December 2021 to February 2023. After the diagnosis of intestinal ileus in 83 patients hospitalized in the burn department by a general surgeon, with the available sampling method, these patients were evaluated based on the inclusion and exclusion criteria of the study. As a result, 66 patients were selected and divided into groups A (33 patients in the intervention group: routine care + gum chewing) and B (33 patients in the control group: routine care) by random allocation method. For the intervention group, from entering the ward until the beginning of oral feeding, gum (without sugar) was chewed four times a day for 15 min, while control groups received the routine diet of the department. Both groups’ condition of the bowel sounds, time of passing gas, and stool were recorded. The statistical analyses were performed using SPSS version 16. The chi-squared, Fisher’s exact, Independent t-test, and Mann-Whitney U tests were utilized.ResultsThe results showed that the median bowel sound return time, time of the first gas discharge, earliest defecation time, and time to start the diet were significantly shorter in the intervention group than in the control group (P < 0.001).ConclusionThis study showed the effect of chewing gum without sugar in shortening the symptoms of intestinal ileus after burns. However, it was not effective in reducing the hospitalization period of patients. According to these results, it is recommended to add chewing gum to the routine care of people hospitalized due to burns.Trial registrationIranian Registry of Clinical Trials IRCT20180715040478N1, 2021-07-27.

Biochemical and Molecular Identification of Azospirillum brasilense Bacteria and Evaluation of Their Efficiency in Producing Hormones, Dissolving Phosphorus, and Fixing Nitrogen

The study aimed to isolate A.brasilense bacteria from the soil of the rhizosphere of different plants and different locations in Al-Diwaniyah Governorate. They were identified in two ways. The first was the routine method, which included studying the microscopic and cultural characteristics and biochemical tests of the isolates. The second method was molecular, using polymerase chain reaction (PCR) technology and using primers. It also included testing the efficiency of these isolates in dissolving tricalcium phosphate (TCP) on Pikovskaya agar medium, fixing nitrogen in the liquid nutrient medium (N.B), and measuring the amount of hormone production using an HPLC device. The results of isolation and regular and molecular identification the presence of ten isolates of bacteria bearing the characteristics of A. brasilense bacteria, out of fifteen local bacterial isolates, took the following symbols and sequences (Az2, Az3, Az5, Az6, Az7, Az9, Az11, Az12, Az13, Az14), as the results showed confirmation of the identification of the bacterial isolates identified by biochemical tests. Using a specialized primer to amplify the 462bp fragment of the 16S ribosomal RNA gene, the results of testing the efficiency of the bacteria in dissolving phosphate (TCP) showed that the isolate (Az13) outperformed the highest value in its effectiveness in dissolving metallic phosphorus through the diameter of the clear zone around the colony, which was effective in dissolving phosphate of up to 3.89 mm. As for the nitrogen fixation efficiency test, the isolate (Az3) excelled in the amount of fixed nitrogen reaching 12.44 mg.L-1. As for the amount of its production of growth regulators for Auxins, Gibberellins, and Cytokinins, the isolate (Az3) recorded the highest value in the amount of Auxins production ( Indol-3-acetic acid) its concentration reached 34.4 µg.ml-1, and as for the production of the hormone Gibberellins (GA3), the isolate (Az3) recorded the highest value amounting to 34.7 µg.ml-1, as for the production of the hormone Cytokinins (CK), The isolate (Az11) recorded the highest value, amounting to 28.8 µg.ml-1.

A Comparative Study of Bacteriological Profile and Susceptibility Pattern by Rapid Direct Manual Identification and Antimicrobial Susceptibility Testing with Routine Blood Culture Method in Neonatal Septicemia.

A Comparative Study of Bacteriological Profile and Susceptibility Pattern by Rapid Direct Manual Identification and Antimicrobial Susceptibility Testing with Routine Blood Culture Method in Neonatal Septicemia.

Clinical decision support system using hierarchical fuzzy diagnosis model for migraine and tension-type headache based on International Classification of Headache Disorders, 3rd edition.

To determine whether the diagnostic ability of the newly designed hierarchical fuzzy diagnosis method is consistent with that of headache experts for probable migraine (PM) and probable tension-type headache (PTTH). Clinical decision support systems (CDSS) are computer systems designed to help doctors to make clinician decisions by information technology, and have proven to be effective in improving headache diagnosis by making medical knowledge readily available to users in some studies. However, one serious drawback is that the CDSS lacks the ability to deal with some fuzzy boundaries of the headache features utilized in diagnostic criteria, which might be caused by patients' recall bias and subjective bias. A hybrid mechanism of rule-based reasoning and hierarchical fuzzy diagnosis method based on International Classification of Headache Disorders, 3rd edition (ICHD-3) was designed and then validated by a retrospective study with 325 consecutive patients and a prospective study with 380 patients who were clinically diagnosed with migraine and TTH at the headache clinic of Chinese PLA General Hospital. The results of the diagnostic test in the retrospective study indicated that the fuzzy-based CDSS can be used in the diagnosis of migraine without aura (MO) (sensitivity 97.71%, specificity 100%), TTH (sensitivity 98.57%, specificity 100%), PM (sensitivity 91.25%, specificity 98.75%) and PTTH (sensitivity 90.91%, specificity 99.63%). While in the prospective study, the diagnostic performances were MO (sensitivity 91.62%, specificity 96.52%), TTH (sensitivity 92.17%, specificity 95.47%), PM (sensitivity 85.48%, specificity 98.11%) and PTTH (sensitivity 87.50%, specificity 98.60%). Cohen's kappa values for the consistency test were 0.984 ± 0.018 (MO), 0.991 ± 0.018 (TTH), 0.916 ± 0.051 (PM), 0.932 ± 0.059 (PTTH) in the retrospective study and 0.884 ± 0.047 (MO), 0.870 ± 0.055 (TTH), 0.853 ± 0.073 (PM), 0.827 ± 0.118 (PTTH) in the prospective study, which indicated good consistency with the fuzzy-based CDSS and the gold standard (p < 0.001). We developed a fuzzy-based CDSS performs much more similarly to expert diagnosis and performs better than the routine CDSS method in the diagnosis of migraine and TTH, and it could promote the application of artificial intelligence in the area of headache diagnosis.

A selective, sensitive and fast LC-MS/MS method for cabotegravir quantification in rat plasma and pharmacokinetic investigations.

This study presents a novel liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for quantifying cabotegravir (CAB) in rat plasma. A novel, sensitive, and rapid LC-MS/MS method has been developed and validated. Furthermore, protein precipitation technique allowed us to lowered the limit of quantification (LOQ) to nanogram levels, allowing detection of smaller CAB amounts in plasma samples. A review of scientific literature reveals that this method is superior than published methods in terms of runtime, sensitivity, wide linearity, and cost, using LC-MS/MS to quantify CAB in biological samples. CAB reached its maximum concentration (Cmax) of 78.401 μg/mL in rat plasma at 1.50 h (Tmax). Linearity was evaluated across 0.05-1000 μg/mL for CAB using five calibration curves with at least nine standards each with r2 > 0.9997. The intra- and inter-day precision and accuracy results were below 15% and acceptable as per Food and Drug Administration (FDA) guidelines. Stability of compounds were established in a battery of stability studies, that is, benchtop, autosampler, and long-term storage stability as well as freeze thaw cycles. The validated method can be used as a routine method to support pharmacokinetic studies.

Comparative electron microscopic visualization of the lung alveolar epithelial glycocalyx with different staining and labeling methods.

The alveolar surface of the lung is lined by an epithelium consisting of type I (AECI) and type II alveolar epithelial cells (AECII). This epithelium is covered by a liquid alveolar lining layer (ALL). Besides intra-alveolar surfactant, ALL also contains the alveolar epithelial glycocalyx on the apical side of AECI and AECII. To better understand the alveolar epithelial glycocalyx, its ultrastructural visualization by transmission electron microscopy is required. The aim of this study was to systematically re-evaluate routine cytochemical methods for visualization of the alveolar epithelial glycocalyx and specifically its glycan components. For this purpose, we used chemical fixation by vascular perfusion with aldehydes as a common routine approach in mice. After fixation, staining is needed for glycocalyx visualization. Cytochemical staining agents such as alcian blue, ruthenium red, and lanthanum nitrate were compared. In addition, SNL (Sambucus nigra lectin) and UEA1 (Ulex europaeus agglutinin I) were used for sialic acid and fucose-specific labeling. Alcian blue showed the strongest staining, with cloud-like structures, whereas ruthenium red appeared as thread-like structures. On the other hand, lanthanum nitrate did not stain the alveolar epithelial glycocalyx. For specific sialic acid and fucose labeling, both lectins presented a specific signal. In conclusion, these methods can be used routinely for assessing ultrastructural changes of the alveolar epithelial glycocalyx in experimental invivo models under different physiological and pathological conditions. In addition, cytochemical staining by tissue massage and post-embedding lectin labeling after vascular perfusion support 3R (reduction, refinement, replacement) principles of animal welfare.