High quality haplotype-resolved genome assemblies of Populus tomentosa Carr., a stabilized interspecific hybrid species widespread in Asia.

Populus has a wide ecogeographical range spanning the Northern Hemisphere, and interspecific hybrids are common. Populus tomentosa Carr. is widely distributed and cultivated in the eastern region of Asia, where it plays multiple important roles in forestry, agriculture, conservation, and urban horticulture. Reference genomes are available for several Populus species, however, our goals were to produce a very high quality de novo chromosome-level genome assembly in P. tomentosa genome that could serve as a reference for evolutionary and ecological studies of hybrid speciation throughout the genus. Here, combining long-read sequencing and Hi-C scaffolding, we present a high-quality, haplotype-resolved genome assembly. The genome size was 740.2 Mb, with a contig N50 size of 5.47 Mb and a scaffold N50 size of 46.68 Mb, consisting of 38 chromosomes, as expected with the known diploid chromosome number (2n = 2x = 38). A total of 59,124 protein-coding genes were identified. Phylogenomic analyses revealed that P. tomentosa is comprised of two distinct subgenomes, which we deomonstrate is likely to have resulted from hybridization between Populus adenopoda as the female parent and Populus alba var. pyramidalis as the male parent, with an origin of approximately 3.93 Ma. Although highly colinear, significant structural variation was found between the two subgenomes. Our study provides a valuable resource for ecological genetics and forest biotechnology.

Acute oesophageal necrosis in a patient with recent SARS-CoV-2.

A 57-year-old Hispanic man with diabetes presented with dyspnoea. He had a positive SARS-CoV-2 PCR. He was intubated for severe hypoxia and treated with intermittent pressors, methylprednisolone and supportive care. He was extubated on hospital day (HD) 9 and discharged to a skilled nursing facility (SNF) on HD 18. Approximately 1 month later, he presented with melena. Endoscopy revealed two large 1.5-2 cm wide-based distal oesophageal ulcers without active bleeding. Histology showed ulcerated squamous mucosa with extensive necrosis extending to the muscularis propria and coccoid bacterial colonies with rare fungal forms suggestive of Candida He was treated with fluconazole and pantoprazole and was discharged to a SNF. Approximately 3 weeks later, he was readmitted for complications. Repeat endoscopy demonstrated improvement and histology revealed chronic inflammation with reactive epithelial changes. Incidentally, SARS-CoV-2 PCR was positive during this visit without any respiratory symptoms.

Mental Health Consequences of Lockdown During the COVID-19 Pandemic: A Cross-Sectional Study.

OBJECTIVE: We aimed to provide an overview of the psychological status and behavioral consequences of the lockdown during the COVID-19 pandemic in Libya. METHODS: A cross-sectional study was conducted among the Libyan population through May and June 2020 in more than 20 cities. The survey comprised basic demographic data of the participants and anxiety symptoms measured using the seven-item Generalized Anxiety Disorder scale (GAD-7) with ≥15 as the cut-off score for clinically significant anxiety symptoms. Additionally, a survey regarding the lockdown effect was administered, which consisted of several parts, to measure the lockdown status. RESULTS: A total of 8084 responses were recorded, of which, 5090 (63%) were women and 2994 (37%) were men. The mean age (SD) for study participants was 27.2 (8.9) years. Among the participants, 1145 (14.2%) reached the cut-off score to detect anxiety symptoms; however, of the study variables, only five were predictors of clinically significant anxiety: age, gender, marital status, work status, being a financial supporter for the family, and being infected with COVID-19. Women had 1.19 times higher odds to exhibit anxiety symptoms than men. Increasing age was significantly associated with reduced likelihood of exhibiting anxiety symptoms, whereas being married was significantly associated with higher likelihood of anxiety symptoms, compared to not being married. Being suspended from work was associated with an increase in the likelihood of anxiety symptoms. However, we found that being infected with COVID-19 was associated with a 9.59 times higher risk of exhibiting severe anxiety symptoms. Among the study participants, 1451 (17.9%) reported a physical and/or verbal abuse episode from family members, 958 (11.9%) reported abuse outside the family, and 641 (7.9%) reported abuse from enforcers, during the lockdown. CONCLUSION: Our study provided an overview of the psychological and behavioral status, among those who resided in Libya during the civil war and COVID-19 pandemic. The study demonstrates a concerningly high level of clinically significant anxiety during lockdown among the Libyan population during Libya's lockdown period.

The Sub-Phenotypes of Sickle Cell Disease in Kuwait.

Kuwaiti patients with sickle cell disease generally have a mild phenotype, but exhibit considerable heterogeneity, in spite of high Hb F levels. We have carried out a cross-sectional study of patients with sickle cell disease in the five major hospitals in Kuwait. Details of their hemoglobin (Hb) genotypes, clinical presentations and complications are presented. The study was over a span of 3 years and involved 396 patients, made up of 351 (88.6%) Kuwaitis and 45 (11.4%) expatriates. They were aged <1 to 73 years. Hb SS (ßS/ßS) was the most common (in 246 patients, i.e. 62.1%) followed by Hb S (HBB: c.20A>T)-ß-thalassemia (Hb S-ß-thal) in 138 (34.8%) and 11 (2.8%) Hb S/Hb D-Punjab (HBB: c.364G>C). Hb F ranged from 1.0 to 55.0%, with a mean of 21.2 ± 9.8%. The most common presentation was vaso-occlusive crises (VOCs), with 230 (54.8%) having had at least one prior to the study with 54 (13.2%) and 74 (18.9%) having between 2-3 and >3 VOCs, respectively. Hydroxyurea (HU) was prescribed to 157 (39.6%) patients. The most common complication was gallstones in 131 (33.1%), followed by acute splenic sequestration in 26.8% and avascular necrosis of the femoral head in 21.2% patients, respectively. Stroke, priapism and leg ulcers were rare. Gallstones, splenic sequestration and osteonecrosis were significantly more common in patients aged >16 years. Patients with Hb S-ß-thal were similar to those with Hb SS in their clinical profiles. The phenotypic expression of sickle cell disease in Kuwaitis is unique in many respects. The role(s) of Hb F and other genetic modifiers require further elucidation.

Comparative transcriptome analysis of TUCPs in Gossypium hirsutum Ligon-lintless-1 mutant and their proposed functions in cotton fiber development.

Transcripts of uncertain coding potential (TUCP) are part of the LncRNAs, which encode some polypeptides. However, the abundance of TUCP transcripts and their roles in Ligon-linless-1 (Li-1) cotton mutant during the early termination of fiber development are still not documented. Li-1 mutant is one of the excellent modules for investigating fiber elongation processes due to its unique fiber developmental stages. To examine the function of TUCP in cotton fiber development, it is important to identify TUCPs and their involvement in fiber development. In this study, we found that 11104 TUCP transcripts were removed by coding potential criteria of Pfam domain scan. Additionally, differential expression levels of TUCP transcripts were detected between Li-1 mutant and the wild-type (WT), which imply their possible functions in cotton fiber development. These results further revealed that a great number of differentially expressed TUCP transcripts in cotton were identified at 8 DPA, followed by 0 DPA and stem. However, these might explain an undesirable function in cotton fiber development. The gene ontology and pathway analysis, based on differential expression patterns of TUCP transcripts on targeted genes, identified the transport process, cytoskeleton structure, membrane permeability and fatty acids. These give new insight into significant involvement in early cessation of cotton fiber development and abnormal stem. The RNA-seq and qRT-PCR expression analyses of TUCP transcripts evidently singled out three possible genes, TUCP_010675, TUCP_001475, TUCP_009444 and other targeted mRNAs. The expression pattern of TUCP transcripts and their mRNA targets provided valuable evidence for further investigations on the biological functions of TUCP in cotton fiber development. The study findings may serve as a useful tool for comparative analysis of TUCP transcripts in cotton species and assist in selection of the applicable candidate genes for further functional analyses, genetic improvement and genetic engineering of cotton fiber development.

Clinical and Molecular Characteristics of Non-Transfusion-Dependent Thalassemia in Kuwait.

Although not regularly transfused, patients with non-transfusion-dependent thalassemia (NTDT) are prone to iron overload and its complications. Their molecular, phenotypical and laboratory characteristics vary in different populations and there is a need to document local prevailing patterns. We have reviewed the records of our patients with NTDT in Kuwait and documented their clinical and molecular characteristics in addition to iron status [serum ferritin and liver magnetic resonance imaging (MRI) T2*], management and complications. There were 41 patients, made up of 20 with ß-thalassemia intermedia (ß-TI), 18 with Hb H (ß4) disease and three with Hb E (HBB: c.79G > A)-ß-thalassemia (Hb E-ß-thal); their ages ranged from 3 to 36 years (mean 12.5 ± 7.7). While 18 (43.9%) had been transfused at least once, only three (7.3%) had been transfused on multiple occasions. Three patients had serum ferritin >500 ng/mL; while four of 38 had mild or moderate liver iron overload. Seven (35.0%) of the ß-TI patients were managed with hydroxyurea (HU) with good response. Other complications included five patients with gallstones and one each of hypothyroidism and moyamoya. The most common mutations among the ß-TI patients were IVS-II-1 (G > A) and IVS-I-6 (T > C), while among the Hb H patients, the Saudi α2-globin gene polyadenylation (polyA) (AATAAA > AATAAG) mutation was responsible for all cases either as homozygotes (61.1%) or compound heterozygotes with the α-thal-2 (-α(3.7)) allele (33.3%). Although the pattern of NTDT in Kuwaiti patients is generally mild, there is a need to follow them to adulthood as the complications are cumulative and more prevalent in this group.

Transcranial Doppler and brain MRI in children with sickle cell disease and high hemoglobin F levels.

BACKGROUND/OBJECTIVE: While overt stroke and silent brain infarcts (SBI) are uncommon among Kuwaiti patients with sickle cell disease (SCD), there have been no previous transcranial Doppler (TCD) studies in this population. The main objective of this study is to determine TCD velocities in a group of Kuwaiti children with SCD and correlate same with brain magnetic resonance imaging (MRI) and angiography (MRA) findings. MATERIALS AND METHODS: Forty-three steady-state, pediatric patients with SCD aged 10.1 ± 3.9 years (21 SS, 19 Sß(0)Thal, and 3 SD) were studied. Twenty-six age-matched, normal siblings of the patients served as controls. TCD was performed using a General Electric (GE), Vivid 3 equipment and MRI/MRA with a GE Signa Excite HD 1.5 Tesla magnet. Complete blood count was with an electronic counter and Hb quantitation with cation-exchange high performance liquid chromatography (HPLC). RESULTS: The mean time-averaged mean of the maximum velocity (TAMV) was significantly higher in the SCD group than the controls, but was normal (<170 cm/second) in all. The mean values were comparable among the SS and Sß(0)thal groups. Five (11.1%) patients had SBI and all were between 12 and 16 years of age. There was no significant difference of TAMV in this group compared to those without infarcts. No patient showed evidence of stenosis or any other abnormalities in the circle of Willis vessels. CONCLUSION: The mild phenotype among Kuwaiti patients with SCD is reflected in normal TCD velocities and a low prevalence of SBI in children with the disease.

Hypercoagulable state and methylenetetrahydrofolate reductase (MTHFR) C677T mutation in patients with beta-thalassemia major in Kuwait.

INTRODUCTION: Patients with thalassemia major often present with a hypercoagulable state, the pathogenesis of which is still not understood. MATERIALS AND METHODS: This study evaluates the risk factors for hypercoagulability in 50 beta-thalassemia major patients and 50 healthy controls. Fasting total homocysteine, protein C (PC), protein S (PS), antithrombin (AT), activated protein C resistance (APCR) and lupus anticoagulant (LA) were assessed. MTHFR C677T mutation was determined. RESULTS: Significant reductions in PC, PS and AT were noted in patients. Only 4% of the patients had hyperhomocysteinemia. Thirty-two percent of the patients were heterozygous and 4% were homozygous for MTHFR C677T mutation. CONCLUSION: The natural coagulation inhibitors PC, PS and AT were significantly reduced in patients with beta-thalassemia major and were thus important risk factors for the hypercoagulable state, but hyperhomocysteinemia and MTHFR mutation do not seem to be significant risk factors for thromboembolic events.

Effects of cigarette smoking on hematological parameters and von Willebrand factor functional activity levels in asymptomatic male and female Arab smokers.

OBJECTIVES: This study aimed at determining the effects of cigarette smoking based on gender, on several hematological parameters and von Willebrand factor protein in the asymptomatic Arab population of Kuwait. SUBJECTS AND METHODS: Ninety-two subjects participated in this study: 55 males (31 smokers and 24 nonsmokers) and 37 females (18 smokers and 19 nonsmokers). Complete blood count results were obtained using Beckman Coulter Hematology Analyzer. Von Willebrand factor functional activity was determined using an enzyme-linked immunoassay-based test in which anti-von Willebrand factor IgG monoclonal antibody was used that recognizes a functional epitope of the protein. The coagulation profile was obtained using ACL 9000 coagulation analyzer. RESULTS: Male smokers had significantly higher levels of white blood cell count (p = 0.03) and von Willebrand factor protein levels (p = 0.029), and a significantly shorter thrombin time (p = 0.019) than nonsmokers. Smoking did not appear to affect any of the parameters analyzed in females as no significant difference was found between smokers and nonsmokers (p > 0.05). CONCLUSION: Our results showed that smoking affected white blood cell count and von Willebrand factor levels in males and not in females, and as such could be potential markers for smoking-induced endothelial damage in asymptomatic Arab male smokers.

Activation of innate immune responses by IL-2-expressing Salmonella typhimurium is independent of Toll-like receptor 4.

We previously demonstrated that an attenuated strain of Salmonella enterica serovar Typhimurium, engineered to express IL-2 (strain GIDIL2), is cleared more rapidly than its parental, non-cytokine-expressing, strain (designated BRD509) from the reticuloendothelial system of susceptible BALB/c mice. This early clearance correlated with the induction of a strong innate immune response within a few hours of administration of GIDIL2 organisms. In the present study, we wished to assess the contribution of LPS recognition to GIDIL2-induced immune responses using Toll-like receptor 4 (TLR4) mutant C3H/HeJ mice. In contrast to LPS responder mice, both BRD509 and GIDIL2 strains persisted at higher levels in LPS non-responder animals. However, the GIDIL2 bacterial loads in the peritoneal cavity and spleen, recovered over a period of 21 days post infection, were consistently lower than the corresponding CFUs of the BRD509 strain. Direct evidence for the induction of innate immunity was shown by demonstrating increased NK cell cytotoxicity, NOS2 gene expression, and nitric oxide synthesis by peritoneal cells obtained as early as 2h after infection with GIDIL2, but not BRD509, organisms. Unlike BALB/c mice, however, these responses failed to afford any protection against virulent challenge in C3H/HeJ mice. Taken together, our data demonstrate that despite the induction of innate immune responses by IL2-expressing organisms, this was not sufficient to induce protection in TLR4 mutant mice.

Alpha tocopherol protects against immunosuppressive and immunotoxic effects of lead.

Chronic exposure to lead (Pb) is associated with multiorgan toxicity. The precise mechanism(s) involved, however, remains incompletely defined. The present study was undertaken to analyze the effect of Pb on the immune system and determine the ability of alpha tocopherol (AT) to reverse Pb-induced immunotoxicity. Groups of TO Mice (6 per group) were treated ip for 2 weeks with saline alone, Pb acetate alone, Pb plus AT, or with AT alone. Spleens were then analyzed for (i) cellular composition by flow cytometry, (ii) cellular response to B and T cell mitogens and (iii) production of nitric oxide (NO). Pb treatment resulted in a significant state of splenomegaly associated mainly with an influx of CD11b+ myeloid cells. Surprisingly, however, these cells exhibited no upregulation in expression of activation markers and did not produce NO. The lymphocyte mitogenic responses were inhibited by > or = 70% in Pb-treated group. Concurrent treatment with Pb and AT resulted in almost a complete reversal of Pb-induced splenic cellular influx. Despite this, however, mitogenic responses in Pb + AT treated group were approximately 50% of those observed in normal (saline-treated) controls. We conclude that (1) chronic treatment with Pb acetate induces a state of splenomegaly and decreased proliferation in response to mitogenic stimuli and (2) co-treatment with AT largely reversed the cellular influx but this was associated with only a partial improvement of the mitogenic responses. These results highlight the role of AT as a potentially effective antioxidant in the immune system.

Induction of innate immunity by IL-2-expressing Salmonella confers protection against lethal infection.

Previously, we demonstrated that an attenuated Salmonella strain expressing IL-2 (designated GIDIL2) is cleared from the circulation at a much faster rate than the non-cytokine-expressing parental strain (designated BRD509). These findings suggested that IL-2 expression led to the rapid induction of innate immune responses that, in turn, accounted for the accelerated rate of bacterial clearance. In the present study, the mechanism by which this early antibacterial response is mediated was investigated. We demonstrate that as early as 2 h after infection with GIDIL2, but not BRD509, peritoneal excudate cells exhibited enhanced NK cytotoxic activity and upregulated NOS2 mRNA and NO production. This early response coincided with an enhancement of GIDIL2 clearance from the peritoneal cavity, first evidenced 22 h post-infection. Moreover, it conferred a high level of protection against virulent Salmonella challenge given within 16-20 h of GIDIL2 administration. These findings highlight the importance of innate immunity in the control of early bacterial proliferation and demonstrate the rapidity by which these responses are induced following bacterial entry.

Cytokine expression by attenuated intracellular bacteria regulates the immune response to infection: the Salmonella model.

Attenuated Salmonella strains have shown excellent efficacy as mucosal vaccine delivery systems. In the present report, several recombinant strains of Salmonella enterica serovar Typhimurium, engineered to express defined murine cytokines, were used to study their potential immunoregulatory capacity in the mouse model of typhoid fever. Specifically, recombinant strains expressing IL-2 (known as GIDIL2) or TNF-alpha (GIDTNF) were compared with the parental, non-cytokine-secreting, strain (BRD509) for their ability to induce a variety of immune responses in susceptible BALB/c mice. Our findings indicate that bacterially-expressed cytokines are functional in vivo and do induce a unique pattern of responses, quite distinct from that induced by BRD509 organisms. Both the type and magnitude of specific immune parameters were affected. These included the capacity to induce an inflammatory response resulting in a state of profound splenomegaly and hepatomegaly, activation of individual immune cells (particularly macrophages and other myeloid lineage cells), and the induction of nitric oxide (NO) secretion. Furthermore, a structural analysis using light as well as electron microscopy was undertaken to examine the host cellular response to infection with the different bacterial strains. The results indicate that cytokine expression by the invading pathogen can dramatically influence host immunity from a very early stage following infection. These findings may well have important consequences for the potential utilization of bacterial vector-encoded cytokines in immunoregulation in different disease settings.

Cross-reactivity between six Enterobacteriaceae complete lipopolysaccharide core chemotypes.

To gain insight into the value of lipopolysaccharide (LPS) core determinants for cross-protective immunisation the serological relationships between six complete (LPS) core types from Enterobacteriaceae were investigated. Hyperimmune sera were raised in mice by repeated immunisation with heat-killed strains of Salmonella choleraesuis (Ra core type) or Escherichia coli (core types R1, R2, R3, R4 and K12) and characterised for reactivity with complete and incomplete core chemotypes by ELISA and immunoblotting. Three sera (anti-Ra, anti-R2 and anti-R3) reacted strongly with 3-5 different complete core types whereas the other three (anti-R1, anti-R4 and anti-K12) reacted strongly only with their homologous core types in these assays. Two approaches were used to examine further the structural bases for cross-reactivity between these cores. By the first approach the anti-complete-core sera were tested for cross-reactivity with truncated forms of the Salmonella species core (incomplete cores) derived from core-defective mutants. By the second approach, antisera raised against some core-defective mutants were tested for cross-reactivity with complete cores. The results of these investigations revealed that several pair-wise combinations of core types can be used as immunogens to elicit immune responses that recognise all six core types and that the major determinants which mediate cross-reactivity between complete cores are localised in the outer core region.

Serodiagnosis of tuberculosis and leprosy by enzyme immunoassay.

OBJECTIVE: To evaluate the use of serodiagnosis for tuberculosis and leprosy using mycobacterial antigen 38 kDa, with kits from Omega laboratories, to detect IgG by enzyme immunoassay (EIA). METHODS: The study population consisted of 58 patients with evidence of tuberculous infection (culture of Mycobacterium tuberculosis complex or microscopic evidence), of whom 23 had pulmonary and 35 had extrapulmonary disease. There were six subjects who had recently been treated for tuberculosis, 11 patients on treatment for leprosy and 137 patients suspected of having tuberculosis on clinical or radiologic grounds (without laboratory evidence). A control group comprised 35 healthy individuals or patients suffering from diseases other than tuberculosis. RESULTS: The tests showed that there was a significant difference in antibody levels between the patients with active pulmonary disease, extrapulmonary tuberculosis and leprosy in comparison with the control group (p<0.001). The sensitivities of the two tests together for proven pulmonary tuberculosis were 100% and 95.7% at 1.0--1.5 and >1.6 EIA cut-off points respectively, while the specificities were 88.5% and 100% at the same cut-off points. The sensitivities for extrapulmonary tuberculosis were 71.4% and only 51.4% at 1.0--1.5 and >1.6 EIA cut-off points. The test was positive in 30 (21.9%) of the 137 suspected patients, while 43 (31.4%) had an equivocal result and the remaining 64 (47.7%) suspects were definitely negative. There was again a significant difference in positivity rates between suspects and the control group. CONCLUSIONS: Omega IgG test is useful in the serodiagnosis of active pulmonary tuberculosis and leprosy, but less sensitive in extrapulmonary disease, particularly in children. Equivocal results may only add to the evidence of tuberculosis in early or minimal disease.