Clinical, Virologic, and Immunologic Evaluation of Symptomatic Coronavirus Disease 2019 Rebound Following Nirmatrelvir/Ritonavir Treatment.

BACKGROUND: Nirmatrelvir/ritonavir, the first severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) protease inhibitor, reduces the risk of hospitalization and death by coronavirus disease 2019 (COVID-19) but has been associated with symptomatic rebound after therapy completion. METHODS: Six individuals with relapse of COVID-19 symptoms after treatment with nirmatrelvir/ritonavir, 2 individuals with rebound symptoms without prior antiviral therapy and 7 patients with acute Omicron infection (controls) were studied. Soluble biomarkers and serum SARS-CoV-2 nucleocapsid protein were measured. Nasal swabs positive for SARS-CoV-2 underwent viral isolation and targeted viral sequencing. SARS-CoV-2 anti-spike, anti-receptor-binding domain, and anti-nucleocapsid antibodies were measured. Surrogate viral neutralization tests against wild-type and Omicron spike protein, as well as T-cell stimulation assays, were performed. RESULTS: High levels of SARS-CoV-2 anti-spike immunoglobulin G (IgG) antibodies were found in all participants. Anti-nucleocapsid IgG and Omicron-specific neutralizing antibodies increased in patients with rebound. Robust SARS-CoV-2-specific T-cell responses were observed, higher in rebound compared with early acute COVID-19 patients. Inflammatory markers mostly decreased during rebound. Two patients sampled longitudinally demonstrated an increase in activated cytokine-producing CD4+ T cells against viral proteins. No characteristic resistance mutations were identified. SARS-CoV-2 was isolated by culture from 1 of 8 rebound patients; Polybrene addition increased this to 5 of 8. CONCLUSIONS: Nirmatrelvir/ritonavir treatment does not impede adaptive immune responses to SARS-CoV-2. Clinical rebound corresponds to development of a robust antibody and T-cell immune response, arguing against a high risk of disease progression. The presence of infectious virus supports the need for isolation and assessment of longer treatment courses. CLINICAL TRIALS REGISTRATION: NCT04401436.

Ruthenium(II)-Dithiocarbazates as Anticancer Agents: Synthesis, Solution Behavior, and Mitochondria-Targeted Apoptotic Cell Death.

The reaction of the Ru(PPh3 )3 Cl2 with HL1-3 -OH (-OH stands for the oxime hydroxyl group; HL1 -OH=diacetylmonoxime-S-benzyldithiocarbazonate; HL2 -OH=diacetylmonoxime-S-(4-methyl)benzyldithiocarbazonate; and HL3 -OH=diacetylmonoxime-S-(4-chloro)benzyl-dithiocarbazonate) gives three new ruthenium complexes [RuII (L1-3 -H)(PPh3 )2 Cl] (1-3) (-H stands for imine hydrogen) coordinated with dithiocarbazate imine as the final products. All ruthenium(II) complexes (1-3) have been characterized by elemental (CHNS) analyses, IR, UV-vis, NMR (1 H, 13 C, and 31 P) spectroscopy, HR-ESI-MS spectrometry and also, the structure of 1-2 was further confirmed by single crystal X-ray crystallography. The solution/aqueous stability, hydrophobicity, DNA interactions, and cell viability studies of 1-3 against HeLa, HT-29, and NIH-3T3 cell lines were performed. Cell viability results suggested 3 being the most cytotoxic of the series with IC50 6.9±0.2 µM against HeLa cells. Further, an apoptotic mechanism of cell death was confirmed by cell cycle analysis and Annexin V-FITC/PI double staining techniques. In this regard, the live cell confocal microscopy results revealed that compounds primarily target the mitochondria against HeLa, and HT-29 cell lines. Moreover, these ruthenium complexes elevate the ROS level by inducing mitochondria targeting apoptotic cell death.

LDDNet: A Deep Learning Framework for the Diagnosis of Infectious Lung Diseases.

This paper proposes a new deep learning (DL) framework for the analysis of lung diseases, including COVID-19 and pneumonia, from chest CT scans and X-ray (CXR) images. This framework is termed optimized DenseNet201 for lung diseases (LDDNet). The proposed LDDNet was developed using additional layers of 2D global average pooling, dense and dropout layers, and batch normalization to the base DenseNet201 model. There are 1024 Relu-activated dense layers and 256 dense layers using the sigmoid activation method. The hyper-parameters of the model, including the learning rate, batch size, epochs, and dropout rate, were tuned for the model. Next, three datasets of lung diseases were formed from separate open-access sources. One was a CT scan dataset containing 1043 images. Two X-ray datasets comprising images of COVID-19-affected lungs, pneumonia-affected lungs, and healthy lungs exist, with one being an imbalanced dataset with 5935 images and the other being a balanced dataset with 5002 images. The performance of each model was analyzed using the Adam, Nadam, and SGD optimizers. The best results have been obtained for both the CT scan and CXR datasets using the Nadam optimizer. For the CT scan images, LDDNet showed a COVID-19-positive classification accuracy of 99.36%, a 100% precision recall of 98%, and an F1 score of 99%. For the X-ray dataset of 5935 images, LDDNet provides a 99.55% accuracy, 73% recall, 100% precision, and 85% F1 score using the Nadam optimizer in detecting COVID-19-affected patients. For the balanced X-ray dataset, LDDNet provides a 97.07% classification accuracy. For a given set of parameters, the performance results of LDDNet are better than the existing algorithms of ResNet152V2 and XceptionNet.

Yearlong COVID-19 Infection Reveals Within-Host Evolution of SARS-CoV-2 in a Patient With B-Cell Depletion.

B-cell-depleting therapies may lead to prolonged disease and viral shedding in individuals infected with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and this viral persistence raises concern for viral evolution. We report sequencing of early and late samples from a 335-day infection in an immunocompromised patient. The virus accumulated a unique deletion in the amino-terminal domain of the spike protein, and complete deletion of ORF7b and ORF8, the first report of its kind in an immunocompromised patient. Unique viral mutations found in this study highlight the importance of analyzing viral evolution in protracted SARS-CoV-2 infection, especially in immunosuppressed hosts.

Success and Challenges Associated with Large-Scale Collaborative Surveillance for Carbapenemase Genes in Gram-Negative Bacteria.

The emergence and spread of antimicrobial resistance, especially in Gram-negative bacteria, has led to significant morbidity and increased cost of health care. Large surveillance studies such as the one performed by the Antibiotic Resistance Laboratory Network are immensely valuable in understanding the scope of resistance mechanisms, especially among carbapenemase-producing Gram-negative bacteria. However, the routine laboratory detection of carbapenemases in these bacteria remains challenging and requires further optimization.

What's New in Antibiograms? Updating CLSI M39 Guidance with Current Trends.

A vast amount of antimicrobial susceptibility test (AST) data is generated from routine testing in diagnostic laboratories for the primary purpose of guiding clinicians in antimicrobial therapy decisions for their patients. However, there is additional value for these data when they are compiled at the local, regional, national, and global levels. Cumulative AST data can be used to prepare antibiograms at the individual health care facility level. These reports can be used to gain insight into appropriate empirical therapy options prior to the availability of AST results on an individual patient's isolate. Different types of cumulative AST data reports can also be compiled at the regional, national, and global levels to estimate susceptibility rates in geographic regions, document trends in evolving microbial populations, and recognize the appearance and spread of emerging antimicrobial resistance threats. The first CLSI M39 Guideline for Analysis and Presentation of Cumulative AST Data was published in 2000. Since that time, there have been changes to AST and reporting recommendations as well as the introduction of advanced informatics technologies to analyze and present data. The 5th edition of M39 has taken into consideration these changes to assist those who analyze, present, and utilize routine antibiograms and other types of cumulative AST data reports as well as those who design information systems for the capturing and analyzing of AST data. Furthermore, antimicrobial stewardship programs (ASPs) have expanded considerably, and uses of the antibiogram by ASPs have been addressed. This minireview will remind users of the basic recommendations for analysis and presentation of antibiograms and provide new suggestions to enhance these reports.

Pooled Saliva Specimens for SARS-CoV-2 Testing.

We evaluated saliva (SAL) specimens for SARS-CoV-2 reverse transcriptase PCR (RT-PCR) testing by comparison of 459 prospectively paired nasopharyngeal (NP) or midturbinate (MT) swabs from 449 individuals with the aim of using saliva for asymptomatic screening. Samples were collected in a drive-through car line for symptomatic individuals (n = 380) and in the emergency department (ED) (n = 69). The percentages of positive and negative agreement of saliva compared to nasopharyngeal swab were 81.1% (95% confidence interval [CI], 65.8% to 90.5%) and 99.8% (95% CI, 98.7% to 100%), respectively. The percent positive agreement increased to 90.0% (95% CI, 74.4% to 96.5%) when considering only samples with moderate to high viral load (cycle threshold [CT ] for the NP, ≤34). Pools of five saliva specimens were also evaluated on three platforms, bioMérieux NucliSENS easyMAG with ABI 7500Fast (CDC assay), Hologic Panther Fusion, and Roche Cobas 6800. The average loss of signal upon pooling was 2 to 3 CT values across the platforms. The sensitivities of detecting a positive specimen in a pool compared with testing individually were 94%, 90%, and 94% for the CDC 2019-nCoV real-time RT-PCR, Panther Fusion SARS-CoV-2 assay, and Cobas SARS-CoV-2 test, respectively, with decreased sample detection trending with lower viral load. We conclude that although pooled saliva testing, as collected in this study, is not quite as sensitive as NP/MT testing, saliva testing is adequate to detect individuals with higher viral loads in an asymptomatic screening program, does not require swabs or viral transport medium for collection, and may help to improve voluntary screening compliance for those individuals averse to various forms of nasal collections.

Sensitivity in Detection of Antibodies to Nucleocapsid and Spike Proteins of Severe Acute Respiratory Syndrome Coronavirus 2 in Patients With Coronavirus Disease 2019.

BACKGROUND: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the cause of coronavirus disease 2019 (COVID-19), is associated with respiratory-related disease and death. Assays to detect virus-specific antibodies are important to understand the prevalence of infection and the course of the immune response. METHODS: Quantitative measurements of plasma or serum antibodies to the nucleocapsid and spike proteins were analyzed using luciferase immunoprecipitation system assays in 100 cross-sectional or longitudinal samples from patients with SARS-CoV-2 infection. A subset of samples was tested both with and without heat inactivation. RESULTS: At >14 days after symptom onset, antibodies against SARS-CoV-2 nucleocapsid protein showed 100% sensitivity and 100% specificity, whereas antibodies to spike protein were detected with 91% sensitivity and 100% specificity. Neither antibody levels nor the rate of seropositivity were significantly reduced by heat inactivation of samples. Analysis of daily samples from 6 patients with COVID-19 showed anti-nucleocapsid and spike protein antibodies appearing between days 8 and 14 after initial symptoms. Immunocompromised patients generally had a delayed antibody response to SARS-CoV-2, compared with immunocompetent patients. CONCLUSIONS: Antibody to the nucleocapsid protein of SARS-CoV-2 is more sensitive than spike protein antibody for detecting early infection. Analyzing heat-inactivated samples with a luciferase immunoprecipitation system assay is a safe and sensitive method for detecting SARS-CoV-2 antibodies.

Clinical Impact of Rapid Point-of-Care PCR Influenza Testing in an Urgent Care Setting: a Single-Center Study.

Seasonal influenza virus causes significant morbidity and mortality each year. Point-of-care (POC) testing using rapid influenza diagnostic tests (RIDTs), immunoassays that detect viral antigens, are often used for diagnosis by physician offices and urgent care centers. These tests are rapid but lack sensitivity, which is estimated to be 50 to 70%. Testing by PCR is highly sensitive and specific, but historically these assays have been performed in centralized clinical laboratories necessitating specimen transport and increasing the time to result. Recently, Clinical Laboratory Improvement Amendments (CLIA)-waived, POC PCR influenza assays have been developed with >95% sensitivity and specificity compared to centralized PCR assays. To determine the clinical impact of a POC PCR test for influenza, we compared antimicrobial prescribing patterns of one urgent care location using the Cobas LIAT Influenza A/B assay (LIAT assay; Roche Diagnostics, Indianapolis, IN) to other urgent care centers in our health system using traditional RIDT, with negative specimens being reflexed to PCR. Antiviral prescribing was lower in patients with a negative LIAT PCR result (2.3%) than in patients with a negative RIDT result (25.3%; P < 0.005). Antivirals were prescribed more often in patients that tested positive by LIAT PCR (82.4%) than in those testing positive by either RIDT or reflex PCR (69.9%; P < 0.05). Antibacterial prescriptions for patients testing negative by LIAT PCR were higher (44.5%) than for those testing negative by RIDT (37.7%), although the difference was not statistically significant. In conclusion, having results from a PCR POC test during the clinic visit improved antiviral prescribing practices compared to having rapid results from an RIDT.

Functional Involvement of Leishmania donovani Tryparedoxin Peroxidases during Infection and Drug Treatment.

The parasite Leishmania donovani causes visceral leishmaniasis, a potentially fatal disease. The parasites survive within mammalian macrophages and express a unique set of enzymes, the tryparedoxin peroxidases, for their defense against oxidative stress generated by the host. In this study, we demonstrate different roles of two distinct enzymes, the mitochondrial tryparedoxin peroxidase (mTXNPx) and the cytosolic tryparedoxin peroxidase (cTXNPx), in defending the parasites against mitochondrial and exogenous oxidative stress during infection and drug treatment. Our findings indicate a greater increase in cTXNPx expression in response to exogenous oxidative stress and a higher elevation of mTXNPx expression in response to mitochondrial or endogenous stress created by respiratory chain complex inhibitors. Overexpression of cTXNPx in Leishmania showed improved protection against exogenous stress and enhanced protection against mitochondrial stress in parasites overexpressing mTXNPx. Further, parasites overexpressing cTXNPx infected host cells with increased efficiency at early times of infection compared to control parasites or parasites overexpressing mTXNPx. The mTXNPx-overexpressing parasites maintained higher infection at later times. Higher mTXNPx expression occurred in wild-type parasites on exposure to miltefosine, while treatment with antimony elevated cTXNPx expression. Parasites resistant to miltefosine or antimony demonstrated increased expression of mTXNPx, as well as cTXNPx. In summary, this study provides evidence of distinct roles of the two enzymes defined by virtue of their localization during infection and drug treatment.

Total Laboratory Automation and Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry Improve Turnaround Times in the Clinical Microbiology Laboratory: a Retrospective Analysis.

Technological advances have changed the practice of clinical microbiology. We implemented Bruker matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) and BD Kiestra total laboratory automation (TLA) 4 and 3 years ago, respectively. To assess the impact of these new technologies, we compared turnaround times (TATs) for positive and negative urine cultures before and after implementation. In comparison I, TATs for 61,157 urine cultures were extracted for two periods corresponding to pre-TLA and post-TLA, both using MALDI-TOF MS for organism identification. In comparison II, time to organism identification (ID) and antimicrobial susceptibility (AST) reports were calculated for 5,402 positive culture reports representing four different periods: (i) manual plating and conventional biochemical identification (CONV), (ii) manual plating and MALDI-TOF MS identification (MALDI), (iii) MALDI-TOF MS identification and early phase implementation of TLA (TLA1), and (iv) MALDI-TOF MS identification and late phase implementation of TLA (TLA2). By the comparison I results, median pre- and post-TLA TATs to organism IDs (18.5 to 16.9 h), AST results (41.8 to 40.8 h), and preliminary results for negative cultures (17.7 to 13.6 h), including interquartile ranges for all comparisons, were significantly decreased post-TLA (P < 0.001). By the comparison II results, MALDI significantly improved TAT to organism ID compared to CONV (21.3 to 18 h). TLA further improved overall TAT to ID (18 to 16.5 h) and AST (42.3 to 40.7 h) results compared to MALDI (P < 0.001). In summary, TLA significantly improved TAT to organism ID, AST report, and preliminary negative results. MALDI-TOF MS significantly improved TAT for organism ID. Use of MALDI-TOF MS and TLA individually and together results in significant decreases in microbiology report TATs.

Multicenter Evaluation of the Modified Carbapenem Inactivation Method and the Carba NP for Detection of Carbapenemase-Producing Pseudomonas aeruginosa and Acinetobacter baumannii.

The purpose of this study was to develop the modified carbapenem inactivation method (mCIM) for the detection of carbapenemase-producing Pseudomonas aeruginosa (CP-PA) and carbapenemase-producing Acinetobacter baumannii (CP-AB) and perform a multicenter evaluation of the mCIM and Carba NP tests for these nonfermenters. Thirty P. aeruginosa and 30 A. baumannii isolates previously characterized by whole-genome sequencing from the CDC-FDA Antibiotic Resistance Isolate Bank were evaluated, including CP isolates (Ambler class A, B, and D), non-carbapenemase-producing (non-CP) carbapenem-resistant isolates, and carbapenem-susceptible isolates. Initial comparison of a 1-µl versus 10-µl loop inoculum for the mCIM was performed by two testing sites and showed that 10 µl was required for reliable detection of carbapenemase production among P. aeruginosa and A. baumannii Ten testing sites then evaluated the mCIM using a 10-µl loop inoculum. Overall, the mean sensitivity and specificity of the mCIM for detection of CP-PA across all 10 sites were 98.0% (95% confidence interval [CI], 94.3 to 99.6; range, 86.7 to 100) and 95% (95% CI, 89.8 to 97.7; range, 93.3 to 100), whereas the mean sensitivity and specificity among CP-AB were 79.8% (95% CI, 74.0 to 84.9; range, 36.3 to 95.7) and 52.9% (95% CI, 40.6 to 64.9; range, 28.6 to 100), respectively. At three sites that evaluated the performance of the Carba NP test using the same set of isolates, the mean sensitivity and specificity of the Carba NP test were 97.8% (95% CI, 88.2 to 99.9; range, 93.3 to 100) and 97.8% (95% CI, 88.2 to 99.9; range, 93.3 to 100) for P. aeruginosa and 18.8% (95% CI, 10.4 to 30.1; range, 8.7 to 26.1) and 100% (95% CI, 83.9 to 100; range, 100) for A. baumannii Overall, we found both the mCIM and the Carba NP test to be accurate for detection of carbapenemase production among P. aeruginosa isolates and less reliable for use with A. baumannii isolates.

Molecular, biochemical characterization and assessment of immunogenic potential of cofactor-independent phosphoglycerate mutase against Leishmania donovani: a step towards exploring novel vaccine candidate.

Despite immense efforts, vaccine against visceral leishmaniasis has yet not been developed. Earlier our proteomic study revealed a novel protein, cofactor-independent phoshoglycerate mutase (LdiPGAM), an important enzyme in glucose metabolism, in T helper cells type 1 (Th1) stimulatory region of soluble Leishmania donovani antigen. In this study, LdiPGAM was biochemically and molecularly characterized and evaluated for its immunogenicity and prophylactic efficacy against L. donovani. Immunogenicity of recombinant LdiPGAM (rLdiPGAM) was initially assessed in naïve hamsters immunized with it by analysing mRNA expression of inducible nitric oxide (NO) synthase (iNOS) and other Th1/T helper cells type 2 cytokines, which revealed an upregulation of Th1 cytokines along with iNOS. Immunogenicity of rLdiPGAM was further evaluated in lymphocytes of treated Leishmania-infected hamsters and peripheral blood mononuclear cells of Leishmania patients in clinical remission by various parameters, viz. lymphoproliferation assay and NO production (hamsters and patients) and levels of various cytokines (patients). rLdiPGAM induced remarkable Lymphoproliferative response and NO production in treated Leishmania-infected hamsters as well as in patients and increase in interferon gamma (IFN-γ), interleukin-12 (IL-12p40) responses in Leishmania patients in clinical remission. Vaccination with rLdiPGAM exerted considerable prophylactic efficacy (73%) supported by increase in mRNA expression of iNOS, IFN-γ and IL-12p40 with decrease in transforming growth factor beta and interleukin-10. Above results indicate the importance of rLdiPGAM protein as a potential vaccine candidate against visceral leishmaniasis.

Modified Carbapenem Inactivation Method for Phenotypic Detection of Carbapenemase Production among Enterobacteriaceae.

The ability of clinical microbiology laboratories to reliably detect carbapenemase-producing carbapenem-resistant Enterobacteriaceae (CP-CRE) is an important element of the effort to prevent and contain the spread of these pathogens and an integral part of antimicrobial stewardship. All existing methods have limitations. A new, straightforward, inexpensive, and specific phenotypic method for the detection of carbapenemase production, the carbapenem inactivation method (CIM), was recently described. Here we describe a two-stage evaluation of a modified carbapenem inactivation method (mCIM), in which tryptic soy broth was substituted for water during the inactivation step and the length of this incubation was extended. A validation study was performed in a single clinical laboratory to determine the accuracy of the mCIM, followed by a nine-laboratory study to verify the reproducibility of these results and define the zone size cutoff that best discriminated between CP-CRE and members of the family Enterobacteriaceae that do not produce carbapenemases. Bacterial isolates previously characterized through whole-genome sequencing or targeted PCR as to the presence or absence of carbapenemase genes were tested for carbapenemase production using the mCIM; isolates with Ambler class A, B, and D carbapenemases, non-CP-CRE isolates, and carbapenem-susceptible isolates were included. The sensitivity of the mCIM observed in the validation study was 99% (95% confidence interval [95% CI], 93% to 100%), and the specificity was 100% (95% CI, 82% to 100%). In the second stage of the study, the range of sensitivities observed across nine laboratories was 93% to 100%, with a mean of 97%; the range of specificities was 97% to 100%, with a mean of 99%. The mCIM was easy to perform and interpret for Enterobacteriaceae, with results in less than 24 h and excellent reproducibility across laboratories.

Plasmodium falciparum K76T pfcrt Gene Mutations and Parasite Population Structure, Haiti, 2006-2009.

Hispaniola is the only Caribbean island to which Plasmodium falciparum malaria remains endemic. Resistance to the antimalarial drug chloroquine has rarely been reported in Haiti, which is located on Hispaniola, but the K76T pfcrt (P. falciparum chloroquine resistance transporter) gene mutation that confers chloroquine resistance has been detected intermittently. We analyzed 901 patient samples collected during 2006-2009 and found 2 samples showed possible mixed parasite infections of genetically chloroquine-resistant and -sensitive parasites. Direct sequencing of the pfcrt resistance locus and single-nucleotide polymorphism barcoding did not definitively identify a resistant population, suggesting that sustained propagation of chloroquine-resistant parasites was not occurring in Haiti during the study period. Comparison of parasites from Haiti with those from Colombia, Panama, and Venezuela reveals a geographically distinct population with highly related parasites. Our findings indicate low genetic diversity in the parasite population and low levels of chloroquine resistance in Haiti, raising the possibility that reported cases may be of exogenous origin.

Characterization of the proliferating cell nuclear antigen of Leishmania donovani clinical isolates and its association with antimony resistance.

Previously, through a proteomic analysis, proliferating cell nuclear antigen (PCNA) was found to be overexpressed in the sodium antimony gluconate (SAG)-resistant clinical isolate compared to that in the SAG-sensitive clinical isolate of Leishmania donovani. The present study was designed to explore the potential role of the PCNA protein in SAG resistance in L. donovani. For this purpose, the protein was cloned, overexpressed, purified, and modeled. Western blot (WB) and real-time PCR (RT-PCR) analyses confirmed that PCNA was overexpressed by ≥ 3-fold in the log phase, stationary phase, and peanut agglutinin isolated procyclic and metacyclic stages of the promastigote form and by ~5-fold in the amastigote form of the SAG-resistant isolate compared to that in the SAG-sensitive isolate. L. donovani PCNA (LdPCNA) was overexpressed as a green fluorescent protein (GFP) fusion protein in a SAG-sensitive clinical isolate of L. donovani, and modulation of the sensitivities of the transfectants to pentavalent antimonial (Sb(V)) and trivalent antimonial (Sb(III)) drugs was assessed in vitro against promastigotes and intracellular (J774A.1 cell line) amastigotes, respectively. Overexpression of LdPCNA in the SAG-sensitive isolate resulted in an increase in the 50% inhibitory concentrations (IC50) of Sb(V) (from 41.2 ± 0.6 µg/ml to 66.5 ± 3.9 µg/ml) and Sb(III) (from 24.0 ± 0.3 µg/ml to 43.4 ± 1.8 µg/ml). Moreover, PCNA-overexpressing promastigote transfectants exhibited less DNA fragmentation compared to that of wild-type SAG-sensitive parasites upon Sb(III) treatment. In addition, SAG-induced nitric oxide (NO) production was found to be significantly inhibited in the macrophages infected with the transfectants compared with that in wild-type SAG-sensitive parasites. Consequently, we infer that LdPCNA has a significant role in SAG resistance in L. donovani clinical isolates, which warrants detailed investigations regarding its mechanism.

Impaired immune responses in the airways are associated with poor outcome in critically ill COVID-19 patients.

Introduction: Mounting evidence indicates that an individual's humoral adaptive immune response plays a critical role in the setting of SARS-CoV-2 infection, and that the efficiency of the response correlates with disease severity. The relationship between the adaptive immune dynamics in the lower airways with those in the systemic circulation, and how these relate to an individual's clinical response to SARS-CoV-2 infection, are less understood and are the focus of this study. Material and methods: We investigated the adaptive immune response to SARS-CoV-2 in paired samples from the lower airways and blood from 27 critically ill patients during the first wave of the pandemic (median time from symptom onset to intubation 11 days). Measurements included clinical outcomes (mortality), bronchoalveolar lavage fluid (BALF) and blood specimen antibody levels, and BALF viral load. Results: While there was heterogeneity in the levels of the SARS-CoV-2-specific antibodies, we unexpectedly found that some BALF specimens displayed higher levels than the paired concurrent plasma samples, despite the known dilutional effects common in BALF samples. We found that survivors had higher levels of anti-spike, anti-spike-N-terminal domain and anti-spike-receptor-binding domain IgG antibodies in their BALF (p<0.05), while there was no such association with antibody levels in the systemic circulation. Discussion: Our data highlight the critical role of local adaptive immunity in the airways as a key defence mechanism against primary SARS-CoV-2 infection.

Nasal Carriage of Epidemic Methicillin-Resistant Staphylococcus aureus 15 (EMRSA-15) Clone Observed in Three Chicago-Area Long-Term Care Facilities.

The spread of pandemic methicillin-resistant Staphylococcus aureus (MRSA) clones such as USA300 and EMRSA-15 is a global health concern. As a part of a surveillance study of three long-term care facilities in the Greater Chicago area, phenotypic and molecular characterization of nasal MRSA isolates was performed. We report a cluster of pandemic EMRSA-15, an MRSA clone rarely reported from the United States, detected during this study.

Recent Advances in Mitochondria-Localized Luminescent Ruthenium(II) Metallodrugs as Anticancer Agents.

Presently, the most effective way to transport drugs specifically to mitochondria inside the cells is of pharmacophoric interest, as mitochondria are recognized as one of the most important targets for new drug design in cancer diagnosis. To date, there are many reviews covering the photophysical, photochemical, and anticancer properties of ruthenium(II) based metallodrugs owing to their high interest in biological applications. There are, however, no reviews specifically covering the mitochondria-localized luminescent Ru(II) complexes and their subsequent mitochondria-mediated anticancer activities. Therefore, this review describes the physicochemical basis for the mitochondrial accumulation of ruthenium complexes, their synthetic strategies to localize and monitor the mitochondria in living cells, and their related underlying anticancer results. Finally, we review the related areas from previous works describing the mitochondria-localized ruthenium complexes for the treatment of cancer-related diseases. Along with this, we also deliberate the perspectives and future directions for emerging more bifunctional Ru(II) complexes that can target, image, and kill tumors more efficiently in comparison with the existing mitochondria-targeted cancer therapeutics.