Temporal analysis of prevalence and antibiotic-resistance patterns in Stenotrophomonas maltophilia clinical isolates in a 19-year retrospective study.

Stenotrophomonas maltophilia is a nonfermenting gram-negative bacterium associated with multiple nosocomial outbreaks. Antibiotic resistance increases healthcare costs, disease severity, and mortality. Multidrug-resistant infections (such as S. maltophilia infection) are difficult to treat with conventional antimicrobials. This study aimed to investigate the isolation rates, and resistance trends of S. maltophilia infections over the past 19 years, and provide future projections until 2030. In total, 4466 patients with S. maltophilia infection were identified. The adult and main surgical intensive care unit (ICU) had the highest numbers of patients (32.2%), followed by the cardiology department (29.8%), and the paediatric ICU (10%). The prevalence of S. maltophilia isolation increased from 7% [95% confidence interval (CI) 6.3-7.7%] in 2004-2007 to 15% [95% CI 10.7-19.9%] in 2020-2022. Most S. maltophilia isolates were resistant to ceftazidime (72.5%), levofloxacin (56%), and trimethoprim-sulfamethoxazole (14.05%), according to our study. A consistent and significant difference was found between S. maltophilia-positive ICU patients and non-ICU patients (P = 0.0017) during the three-year pandemic of COVID-19 (2019-2021). The prevalence of S. maltophilia isolates is expected to reach 15.08% [95% CI 12.58-17.59%] by 2030. Swift global action is needed to address this growing issue; healthcare authorities must set priorities and monitor infection escalations and treatment shortages.

Characteristics and clinical manifestations of patients, including organ transplant patients, during the surge of JN.1: Insights from Saudi Arabia.

BACKGROUND: Amidst the persistent global health threat posed by the evolving SARS-CoV-2 virus throughout the four-year-long COVID-19 pandemic, the focus has now turned to the Omicron variant and its subvariant, JN.1, which has rapidly disseminated worldwide. This study reports on the characteristics and clinical manifestations of patients during the surge of the JN.1 variant in Saudi Arabia; it also investigates the evolution of SARS-CoV-2 variants in organ transplant patients and identifies patient risk factors. METHODS: A total of 151 nasopharyngeal samples from patients with PCR-confirmed SARS-CoV-2 infection were collected between September 2023 and January 2024. Demographic and clinical data of the patients were obtained from electronic health records. All confirmed positive samples underwent sequencing using Ion GeneStudio and the Ion AmpliSeq™ SARS-CoV-2 panel. RESULTS: During the surge of the JN.1 variant, the average age of the patients was 40 years, ranging from 3 to 93 years, and nearly 50% of the patients were male. Our investigation revealed that the J.N variant predominantly infected patients with comorbidities or organ transplant recipients (57.6%). Moreover, patients with comorbidities or organ transplants exhibited a higher number of mutations. In our organ transplant cohort, an increased total number of spike mutations was associated with a lower risk of developing severe disease (OR = 0.96, 95% CI: 0.93-0.98). CONCLUSIONS: Although JN.1 may not prove to be particularly harmful, it is crucial to recognize the persistent emergence of concerning variants, which create new pathways for the virus to evolve. The ongoing evolution of SARS-CoV-2 is evident in the continuous divergence of these variants from the original strain that marked the onset of the pandemic nearly four years ago.

Genomic Surveillance and Mutation Analysis of SARS-CoV-2 Variants among Patients in Saudi Arabia.

The genome of severe acute respiratory coronavirus-2 (SARS-CoV-2), the virus responsible for coronavirus disease 2019 (COVID-19), has undergone a rapid evolution, resulting in the emergence of multiple SARS-CoV-2 variants with amino acid changes. This study aimed to sequence the whole genome of SARS-CoV-2 and detect the variants present in specimens from Saudi Arabia. Furthermore, we sought to analyze and characterize the amino acid changes in the various proteins of the identified SARS-CoV-2 variants. A total of 1161 samples from patients diagnosed with COVID-19 in Saudi Arabia, between 1 April 2021 and 31 July 2023, were analyzed. Whole genome sequencing was employed for variant identification and mutation analysis. The statistical analysis was performed using the Statistical Analytical Software SAS, version 9.4, and GraphPad, version 9.0. This study identified twenty-three variants and subvariants of SARS-CoV-2 within the population, with the Omicron BA.1 (21K) variant (37.0%) and the Delta (21J) variant (12%) being the most frequently detected. Notably, the Omicron subvariants exhibited a higher mean mutation rate. Amino acid mutations were observed in twelve proteins. Among these, the spike (S), ORF1a, nucleocapsid (N), and ORF1b proteins showed a higher frequency of amino acid mutations compared to other the viral proteins. The S protein exhibited the highest incidence of amino acid mutations (47.6%). Conversely, the ORF3a, ORF8, ORF7a, ORF6, and ORF7b proteins appeared more conserved, demonstrating the lowest percentage and frequency of amino acid mutations. The investigation of structural protein regions revealed the N-terminal S1 subunit of the S protein to frequently harbor mutations, while the N-terminal domain of the envelope (E) protein displayed the lowest mutation frequency. This study provides insights into the variants and genetic diversity of SARS-CoV-2, underscoring the need for further research to comprehend its genome evolution and the occurrence of mutations. These findings are pertinent to the development of testing approaches, therapeutics, and vaccine strategies.

Diagnosis of Human Cytomegalovirus Drug Resistance Mutations in Solid Organ Transplant Recipients-A Review.

Human cytomegalovirus (HCMV) infection may be asymptomatic in healthy individuals but can cause severe complications in immunocompromised patients, including transplant recipients. Breakthrough and drug-resistant HCMV infections in such patients are major concerns. Clinicians are first challenged to accurately diagnose HCMV infection and then to identify the most effective antiviral drug and determine when to initiate therapy, alter drug dosage, or switch medication. This review critically examines HCMV diagnostics approaches, particularly for immunocompromised patients, and the development of genotypic techniques to rapidly diagnose drug resistance mutations. The current standard method to identify prevalent and well-known resistance mutations involves polymerase chain reaction amplification of UL97, UL54, and UL56 gene regions, followed by Sanger sequencing. This method can confirm clinical suspicion of drug resistance as well as determine the level of drug resistance and range of cross-resistance with other drugs. Despite the effectiveness of this approach, there remains an urgent need for more rapid and point-of-care HCMV diagnosis, allowing for timely lifesaving intervention.

Whole genome sequencing of resistance and virulence genes in multi-drug resistant Pseudomonas aeruginosa.

BACKGROUND: Pseudomonas aeruginosa is an opportunistic bacterium that causes serious hospital-acquired infections. To assess the risk of clinically isolated P. aeruginosa to human health, we analyzed the resistance and virulence mechanisms of a collection of clinical isolates. METHODS: This was a retrospective study in which P. aeruginosa isolates collected from January 1, 2018 to August 31, 2019 were analyzed using phenotypic and whole-genome sequencing (WGS) methods. The analysis included 48 clinical samples. Median patient age was 54.0 (29.5) years, and 58.3% of patients were women. Data from the microbiology laboratory database were reviewed to identify P. aeruginosa isolates. All unique isolates available for further testing were included, and related clinical data were collected. Infections were defined as hospital acquired if the index culture was obtained at least 48 h after hospitalization. RESULTS: High-risk P. aeruginosa clones, including sequence types (STs) ST235 and ST111, were identified, in addition to 12 new STs. The isolates showed varying degrees of biofilm formation ability when evaluated at room temperature, along with reduced metabolic activity, as measured by metabolic staining, suggesting their ability to evade antimicrobial therapy. Most isolates (77.1%) were multidrug resistant (MDR), with the highest resistance and susceptibility rates to beta-lactams and colistimethate sodium, respectively. CONCLUSIONS: The MDR phenotypes of the examined isolates can be explained by the high prevalence of efflux-mediated resistance- and hydrolytic enzyme-encoding genes. These isolates had high cytotoxic potential, as indicated by the detection of toxin production-related genes.

Exposure to Cadmium Telluride Quantum Dots and Gene Expression Profile of Huh-7 Hepatocellular Carcinoma Cell Line.

Nanoparticles have shown promising potential for efficient drug delivery, circumventing biological interferences like immunological and renal clearance and mechanical and enzymatic destruction. However, a handful of research papers have questioned the biomedical use of metal-based nanoparticles like cadmium telluride quantum dots (CdTe-QDs) for their cytotoxic, genotoxic, and carcinogenic potential. Herein, we examined the effects of CdTe-QD NPs on gene expression profile of hepatocellular carcinoma (Huh-7) cell line. Huh-7 cells were treated with CdTe-QD NPs (10 µg/ml for 6, 12, and 24 hours, and 25 µg/ml for 6 and 12 hours), and transcriptomic analysis was performed using microarray to evaluate the global gene expression profile. Differential expressed genes (DEGs) were observed for both the doses (10 and 25 µg/ml) of CdTe-QD NPs at different time points. Gene ontology (GO) analysis revealed that genes involved in molecular function of cell cycle, organizational injury and abnormalities, cell death and survival, gene expression, cancer, organismal survival, and cellular development were differentially expressed. Overall, we have demonstrated differential expression of several genes, involved in maintaining cell survival, metabolism, and genome integrity. These findings were confirmed by RT-qPCR study for some canonical pathway genes signifying possible implication in NP toxicity-mediated cell survival and inhibition of cell death.

Changes in the Fibrinolytic System of Patients Infected with Severe Acute Respiratory Syndrome Coronavirus 2.

INTRODUCTION: In this study, coagulation and fibrinolysis parameters and their association with disease severity were investigated in coronavirus disease (COVID-19) patients. MATERIALS AND METHODS: COVID-19 patients (n = 446) admitted to our institute between 21 February 2021 and 17 March 2022, were recruited. Clinical data and staging were collected from all patients. Blood samples were collected and analyzed for several parameters of fibrinolysis and coagulation, including alpha-2-antiplasmin(α2AP) and plasminogen, thrombin activatable fibrinolysis inhibitor (TAFI), tissue plasminogen activator (tPA), plasminogen activator inhibitor-1 (PAI-1), D-dimer, and fibrinogen levels. RESULTS: The TAFI, fibrinogen, and tPA levels were significantly higher in participants who died compared to that of patients who recovered (p < 0.001). However, PAI-1, tPA, and TAFI were significantly higher in patients admitted to the ICU than those of the healthy controls (p < 0.001 for PAI-1 and tPA; p = 0.0331 for TAFI). Our results showed that stage C and D COVID-19 patients had significantly higher levels of PAI-1 (p = 0.003). Furthermore, stage D COVID-19 patients had significantly higher tPA and TAFI values (p = 0.003). CONCLUSIONS: Hypofibrinolysis was the most prevalent condition among patients with severe COVID-19. In this study, several coagulation markers were elevated, making them suitable prognostic markers for hypofibrinolysis.

Association between interleukin-27 gene polymorphisms and Plasmodium falciparum Malaria.

Malaria is often characterized by a complicated disease course due to multifaceted intrinsic genetic factors of the host and the parasite. This study aimed to investigate the role of interleukin-27 (IL-27) gene polymorphisms in Plasmodium falciparum malaria infection in a Saudi Arabian cohort. This case-control study obtained blood samples from 250 malaria patients with P. falciparum and 200 randomly identified healthy control subjects from the Malaria Center in the Jazan area. Malaria patients were grouped into three cohorts as follow: low (<500 parasites/µl of blood), moderate (500-1000 parasites/µl of blood), and high (>1000 parasites/µl of blood) parasitemia. The results show that the IL-27 variant rs181209 was significantly associated with malaria patients (P = 0.026). Similarly, the homozygous GG genotype of rs26528 was also associated with risk of developing P. falciparum malaria (P = 0.032). The minor allele C of variant rs181206 exhibited an association with low to moderate parasitemia (P = 0.046). Furthermore, the rs181209 AA genotype was statistically significant in age group 1-5 years (P = 0.049). In conclusion, this study suggests that variant rs181209 and rs26528 could be associated with the risk of malaria infection by P. falciparum in the population studied.

Association of SARS-CoV-2 Nucleocapsid Protein Mutations with Patient Demographic and Clinical Characteristics during the Delta and Omicron Waves.

SARS-CoV-2 genomic mutations outside the spike protein that may increase transmissibility and disease severity have not been well characterized. This study identified mutations in the nucleocapsid protein and their possible association with patient characteristics. We analyzed 695 samples from patients with confirmed COVID-19 in Saudi Arabia between 1 April 2021, and 30 April 2022. Nucleocapsid protein mutations were identified through whole genome sequencing. 𝜒2 tests and t tests assessed associations between mutations and patient characteristics. Logistic regression estimated the risk of intensive care unit (ICU) admission or death. Of the 60 mutations identified, R203K was the most common, followed by G204R, P13L, E31del, R32del, and S33del. These mutations were associated with reduced risk of ICU admission. P13L, E31del, R32del, and S33del were also associated with reduced risk of death. By contrast, D63G, R203M, and D377Y were associated with increased risk of ICU admission. Most mutations were detected in the SR-rich region, which was associated with low risk of death. The C-tail and central linker regions were associated with increased risk of ICU admission, whereas the N-arm region was associated with reduced ICU admission risk. Consequently, mutations in the N protein must be observed, as they may exacerbate viral infection and disease severity. Additional research is needed to validate the mutations' associations with clinical outcomes.

SARS-CoV-2 spike gene Sanger sequencing methodology to identify variants of concern.

The global demand for rapid identification of circulating SARS-CoV-2 variants of concern has led to a shortage of commercial kits. Therefore, this study aimed to develop and validate a rapid, cost-efficient genome sequencing protocol to identify circulating SARS-CoV-2 (variants of concern). Sets of primers flanking the SARS-CoV-2 spike gene were designed, verified and then validated using 282 nasopharyngeal positive samples for SARS-CoV-2. Protocol specificity was confirmed by comparing these results with SARS-CoV-2 whole-genome sequencing of the same samples. Out of 282 samples, 123 contained the alpha variant, 78 beta and 13 delta, which were indicted using in-house primers and next-generation sequencing; the numbers of variants found were 100% identical to the reference genome. This protocol is easily adaptable for detection of emerging variants during the pandemic.

Molecular Epidemiology of SARS-CoV-2 and Clinical Manifestations among Organ Transplant Recipients with COVID-19.

RNA viruses, including SARS-CoV-2, rely on genetic mutation as a major evolutionary mechanism, leading to the emergence of variants. Organ transplant recipients (OTRs) may be particularly vulnerable to such mutations, making it crucial to monitor the spread and evolution of SARS-CoV-2 in this population. This cohort study investigated the molecular epidemiology of SARS-CoV-2 by comparing the SARS-CoV-2 whole genome, demographic characteristics, clinical conditions, and outcomes of COVID-19 illness among OTRs (n = 19) and non-OTRs with (n = 38) or without (n = 30) comorbid conditions. Most patients without comorbidities were female, whereas most OTRs were male. Age varied significantly among the three groups: patients with comorbidities were the oldest, and patients without comorbidities were the youngest. Whole-genome sequencing revealed that OTRs with mild disease had higher numbers of unusual mutations than patients in the other two groups. Additionally, OTRs who died had similar spike monoclonal antibody resistance mutations and 3CLpro mutations, which may confer resistance to nirmatrelvir, ensitrelvir, and GC37 therapy. The presence of those unusual mutations may impact the severity of COVID-19 illness in OTRs by affecting the virus's ability to evade the immune system or respond to treatment. The higher mutation rate in OTRs may also increase the risk of the emergence of new virus variants. These findings highlight the importance of monitoring the genetic makeup of SARS-CoV-2 in all immunocompromised populations and patients with comorbidity.

Molecular Characterization of Leishmania Species among Patients with Cutaneous Leishmaniasis in Asir Province, Saudi Arabia.

Anthroponotic cutaneous leishmaniais (ACL) and zoonotic cutaneous leishmaniasis (ZCL) caused by Leishmania tropica and Leishmania major, respectively, are endemic vector-borne diseases in southern Saudi Arabia. In 2021, an outbreak of cutaneous leishmaniasis occurred in the province of Asir. The main objective of our investigation was to analyze the epidemiological features of CL in southern Saudi Arabia. The ministry of health recorded 194 CL patients between January and December 2021 from the Asir province. Our findings showed that the majority of CL patients (87.1%) originated from the governorates of Khamis-Mushait and Abha. Most of the patients were males (62.3%). While CL affected all age groups, those under 13 years old were the most affected (38.1%). For both genders, CL patients were mostly Saudi citizens (90.7%) compared to non-Saudi expatriates. The majority of CL patients (75.2%) suffered from a single lesion, and the majority of lesions (61.3%) were located on the face. The seasonal prevalence of CL showed two peaks, a small one in July-August and a larger one in March. Of a total of 194 Giemsa slides samples, 188 showed positive amplification of Leishmania ITS1 gene. Based on PCR-RFLP and PCR-HMR, 183 patients showed positive amplification of L. tropica and five patients showed positive amplification of L. major. Phylogenetic analysis revealed a clear distinct separation between L. major and L. tropica sequences. Our results provided strong evidence of the pre-domination of L. tropica, the main etiological agent of ACL in Asir province. We reported for the first time the presence of L. major, an etiological agent of ZCL in the study areas. The co-circulation of ACL and ZCL highlighted the complexity of the epidemiology of CL in southern Saudi Arabia, and subsequently, further studies to identify competent vectors and reservoir hosts for the establishment of control strategies are needed.

SARS-CoV-2 modulates inflammatory responses of alveolar epithelial type II cells via PI3K/AKT pathway.

Background: SARS-CoV-2 infects through the respiratory route and triggers inflammatory response by affecting multiple cell types including type II alveolar epithelial cells. SARS-CoV-2 triggers signals via its Spike (S) protein, which have been shown to participate in the pathogenesis of COVID19. Aim: Aim of the present study was to investigate the effect of SARS-CoV2 on type II alveolar epithelial cells, focusing on signals initiated by its S protein and their impact on the expression of inflammatory mediators. Results: For this purpose A549 alveolar type II epithelial cells were exposed to SARS CoV2 S recombinant protein and the expression of inflammatory mediators was measured. The results showed that SARS-CoV-2 S protein decreased the expression and secretion of IL8, IL6 and TNFα, 6 hours following stimulation, while it had no effect on IFNα, CXCL5 and PAI-1 expression. We further examined whether SARS-CoV-2 S protein, when combined with TLR2 signals, which are also triggered by SARS-CoV2 and its envelope protein, exerts a different effect in type II alveolar epithelial cells. Simultaneous treatment of A549 cells with SARS-CoV-2 S protein and the TLR2 ligand PAM3csk4 decreased secretion of IL8, IL6 and TNFα, while it significantly increased IFNα, CXCL5 and PAI-1 mRNA expression. To investigate the molecular pathway through which SARS-CoV-2 S protein exerted this immunomodulatory action in alveolar epithelial cells, we measured the induction of MAPK/ERK and PI3K/AKT pathways and found that SARS-CoV-2 S protein induced the activation of the serine threonine kinase AKT. Treatment with the Akt inhibitor MK-2206, abolished the inhibitory effect of SARS-CoV-2 S protein on IL8, IL6 and TNFα expression, suggesting that SARS-CoV-2 S protein mediated its action via AKT kinases. Conclusion: The findings of our study, showed that SARS-CoV-2 S protein suppressed inflammatory responses in alveolar epithelial type II cells at early stages of infection through activation of the PI3K/AKT pathway. Thus, our results suggest that at early stages SARS-CoV-2 S protein signals inhibit immune responses to the virus allowing it to propagate the infection while in combination with TLR2 signals enhances PAI-1 expression, potentially affecting the local coagulation cascade.

Genetic Diversity and Population Genetic Analysis of Plasmodium falciparum Thrombospondin Related Anonymous Protein (TRAP) in Clinical Samples from Saudi Arabia.

The thrombospondin related anonymous protein (TRAP) is considered one of the most important pre-erythrocytic vaccine targets. Earlier population genetic studies revealed the TRAP gene to be under strong balancing natural selection. This study is the first attempt to analyze genetic diversity, natural selection, phylogeography and population structure in 199 clinical samples from Saudi Arabia using the full-length PfTRAP gene. We found the rate of nonsynonymous substitutions to be significantly higher than that of synonymous substitutions in the clinical samples, indicating a strong positive or diversifying selection for the full-length gene and the Von Willebrand factor (VWF). The nucleotide diversity was found to be π~0.00789 for the full-length gene; however, higher nucleotide diversity was observed for the VWF compared to the thrombospondin repeat region (TSP). Deduction of the amino acid sequence alignment of the PNP repeat region in the Saudi samples revealed six genotypes characterized by tripeptide repeat motifs (PNP, ANP, ENP and SNP). Haplotype network, population structure and population differentiation analyses indicated four distinct sub-populations in spite of the low geographical distance between the sampling sites. Our results suggest the likeliness of independent parasite evolution, creating opportunities for further adaptation, including host transition, and making malaria control even more challenging.

Comparison of HIT Tests in Patients with COVID-19 and Thrombocytopenia.

Thrombosis and coagulopathy have been found to be the most prevalent complications in patients with COVID-19. Thromboprophylaxis to prevent thromboembolic events is recommended for hospitalized COVID-19 patients. Heparin-induced thrombocytopenia (HIT) is a known complication of heparin use. This study aimed to determine the incidence of HIT among admitted patients with confirmed COVID-19 by PCR. In this study, two different HIT assays, rapid immunoassay (STic Expert HIT, Stago) and H-PF4 specific enzyme-linked immunosorbent assay (Asserachrom® HPIA - IgG), were performed. Of 200 patients with confirmed COVID-19, we identified 49 patients who met the possibility of HIT (low platelet count and high D-Dimer level). Only five (10.2%) had a positive HIT rapid test. However, none of the tested samples tested positive by ELISA. Thrombosis was reported in two of five (40%) patients. Further extensive studies are required to determine the prevalence and clinical significance of a positive HIT test among patients with COVID-19.

Sociodemographic predictors of confirmed COVID-19 mortality and hospitalization among patients in Saudi Arabia: Analyzing a national COVID-19 database.

BACKGROUND: Even with the widespread availability of vaccines for the COVID-19 disease, there is no sign of decline in the rate of spread of the disease. Based on findings of different studies across the globe, the disease is characterized by poor outcomes in specific sociodemographic categories such as age, gender and presence of symptoms. METHODS: In this study, we carried out a multivariable logistic regression analysis on a national database (HESN+) of confirmed COVID-19 cases in Saudi Arabia to determine predictors of hospitalization and mortality for these patients. RESULTS: Data was extracted for 328,301 confirmed COVID- 19 patients (mean age (SD) = 37.79 (1.68)) with 34.92% females and 65.08% males. Of these, 59.87% were Saudi Arabian citizens and 40.13% were non-Saudi. 68.91% of cases were discovered in Riyadh (n = 67,384), Makkah (n = 72,590) and the Eastern Province (n = 79,666). 72.2% of all cases were diagnosed and treated by the Ministry of Health (MOH). Of all confirmed cases, 95.28% showed one or more symptoms associated with COVID-19. 5.48% of these were hospitalized and 1.11% died. Predictors of mortality and hospitalization, respectively, included age (OR; 1.088 and 1.03), being male (OR; 1.443 and 1.138), nationality (OR; 2.11 and 1.993), presence of symptoms (OR; 1.816 and 4.386), and the health care sector in which patients received treatment (MOH OR; 1.352 and 4.731). CONCLUSION: We found that COVID-19-related hospitalization or mortality was higher among males, older adults, and patients showing one or more symptoms, and mortality likelihood was more than fourfold for patients treated by the MOH. Immigrants were also more likely to be hospitalized or die from COVID-19 infection compared to Saudi nationals.

Epidemiology and Clinical Characteristics of People with Confirmed SARS-CoV-2 Infection during the Early COVID-19 Pandemic in Saudi Arabia.

This study provides epidemiologic and clinical characteristics of 492 consecutive patients diagnosed with SARS-CoV-2 infection at King Faisal Specialist Hospital and Research Centre in Saudi Arabia between March and September 2020. Data were collected from electronic case reports. The cohort was 54% male, with 20.4% aged >60 years, 19.9% aged 31−40 years, and 17% aged 41−50 years. The median incubation period was 16 days, with upper and lower 95% quartiles of 27 and 10 days, respectively. Most patients (79.2%) were symptomatic. Variables significantly different between symptomatic and asymptomatic patients were age, blood oxygen saturation percentage, hemoglobin level, lymphocyte count, neutrophil to lymphocyte (NTL) ratio, and alanine aminotransferase (ALT) and aspartate aminotransferase (AST) level. Asymptomatic patients were mostly younger, with lower body mass index and ALT and AST levels but higher lymphocyte counts, NTL ratio, and CD4, CD8, natural killer cell, IgG, and IgM levels. Factors associated with increased risk of mortality were age (>42 years) and comorbidities, particularly diabetes mellitus and hypertension. Patients who were not given an antiviral regimen were associated with better prognosis than patients who received an antiviral regimen (HR, 0.07; 95% CI, 0.011−0.25). These findings will help clinicians and policymakers adopt best management and treatment options for SARS-CoV-2 infection.

Genetic Diversity, Repeat Motifs, and Natural Selection at the C-Terminal Knob-Associated Histidine Rich Protein (KAHRP) of Plasmodium falciparum Clinical Samples from Saudi Arabia.

Background: Malaria is still a public health problem in Saudi Arabia specifically in the Jazan region. Plasmodium falciparum knob-associated histidine-rich proteins (PfKAHRPs) play an important role in cerebral malaria pathophysiology as well as pathogenesis of P. falciparum infections. The repeat region of PfKAHRP C-terminal interaction domain has been found to bind to the infected red blood cells and the vascular endothelium. Thus, this study aimed to assess the allelic variations, genetic diversity, and natural selection acting at the C-terminal PfKAHRP between parasite isolates from Saudi Arabia. Materials and Methods: The PfKHARP C-terminal interaction domain was successfully PCR-amplified and sequence data from 441 clinical isolates from Saudi Arabia were obtained. The DnaSP v5.10 software was used to determine the genetic diversity, polymorphism, haplotype, and natural selection. Haplotype network analysis was constructed by using the median-joining method in the NETWORK version 5.0.0.1 software. Results: Alignment and analysis of 441 C-terminal PfKAHRP-deduced amino acid sequences identified 5 genotypes (I-V) based on the decapeptide repeat arrangements (TKEASTSKEA, TKEASTSKGA, TKEASTTEGA, and TKEASTSKRA). Among the repeat types, Type I (49.43%, 218/441) was the most abundant in Saudi Arabia, followed by Type II (48.29%, 213/441). Overall, the nucleotide diversity in the PfKHARP C-terminal region was found to be low in Saudi Arabia (π = 0.00142); however, natural selection tests indicated positive selection (dN-dS = 1.64, P < 0.05) which was due to the variations within the repeat motifs. Genealogical relationship haplotype network of PfKAHRP from 4 different countries (i.e., Saudi Arabia, Iran, Burundi, and India) revealed 1 major shared haplotype cluster (H_1) with samples representative from all 4 countries (Saudi Arabia; n = 441, Burundi; n = 4, Iran; n = 13, and India; n = 1). Conclusion: Since this is the first study to report on genetic diversity of C-terminal PfKAHRP interaction domain and the repeat motifs from clinical samples in Saudi Arabia, it will contribute towards the rational design of antiadhesion drug therapies for P. falciparum malaria.

Outcome of SARS-CoV-2 variant breakthrough infection in fully immunized solid organ transplant recipients.

SARS-CoV-2 vaccination in solid organ transplant recipients is associated with suboptimal immune response and risk for breakthrough infection. It is not known whether they are at risk of severe post-vaccine breakthrough infections in the presence of SARSCoV-2 variant of concern. We describe a case series of four fully vaccinated solid organ transplant recipients who developed SARS-CoV-2 variants of concern breakthrough infections. Three patients received BNT162b2 mRNA (Pfizer-BioNTech) and one patient received ChAdOx1 (AZD12220) COVID-19 vaccines. The patients were infected with SARS-CoV-2 variants circulating in Saudi Arabia. Two patients were infected with Alpha variant and had severe pneumonia requiring intensive care admission and ventilatory support and subsequently died. The other two patients recovered; one patient was infected with Beta variant required low supplemental oxygen via nasal flow and the other patient was infected with Delta variant and required high supplemental oxygen nasal flow. Younger patients had a better outcome than older patients. Future large studies are required to confirm our observations and to compare the different vaccine efficacy among solid organ transplants in the era of SARS-CoV-2 variants of concern.

Bio-SCAN: A CRISPR/dCas9-Based Lateral Flow Assay for Rapid, Specific, and Sensitive Detection of SARS-CoV-2.

Simple, rapid, specific, and sensitive point-of-care detection methods are needed to contain the spread of SARS-CoV-2. CRISPR/Cas9-based lateral flow assays are emerging as a powerful alternative for COVID-19 diagnostics. Here, we developed Bio-SCAN (biotin-coupled specific CRISPR-based assay for nucleic acid detection) as an accurate pathogen detection platform that requires no sophisticated equipment or technical expertise. Bio-SCAN detects the SARS-CoV-2 genome in less than 1 h from sample collection to result. In the first step, the target nucleic acid sequence is isothermally amplified in 15 min via recombinase polymerase amplification before being precisely detected by biotin-labeled nuclease-dead SpCas9 (dCas9) on commercially available lateral flow strips. The resulting readout is visible to the naked eye. Compared to other CRISPR-Cas-based pathogen detection assays, Bio-SCAN requires no additional reporters, probes, enhancers, reagents, or sophisticated devices to interpret the results. Bio-SCAN is highly sensitive and successfully detected a clinically relevant level (4 copies/µL) of synthetic SARS-CoV-2 RNA genome. Similarly, Bio-SCAN showed 100% negative and 96% positive predictive agreement with RT-qPCR results when using clinical samples (86 nasopharyngeal swab samples). Furthermore, incorporating variant-specific sgRNAs in the detection reaction allowed Bio-SCAN to efficiently distinguish between the α, ß, and δ SARS-CoV-2 variants. Also, our results confirmed that the Bio-SCAN reagents have a long shelf life and can be assembled locally in nonlaboratory and limited-resource settings. Furthermore, the Bio-SCAN platform is compatible with the nucleic acid quick extraction protocol. Our results highlight the potential of Bio-SCAN as a promising point-of-care diagnostic platform that can facilitate low-cost mass screening for SARS-CoV-2.