Identification of differences in the magnitude and specificity of SARS-CoV-2 nucleocapsid antibody responses in naturally infected and vaccinated individuals.

As there are limited data on B-cell epitopes for the nucleocapsid protein in SARS-CoV-2, we sought to identify the immunodominant regions within the N protein, recognized by patients with varying severity of natural infection with the Wuhan strain (WT), delta, omicron, and in those who received the Sinopharm vaccines, which is an inactivated, whole virus vaccine. Using overlapping peptides representing the N protein, with an in-house ELISA, we mapped the immunodominant regions within the N protein, in seronegative (n = 30), WT infected (n = 30), delta infected (n = 30), omicron infected + vaccinated (n = 20) and Sinopharm (BBIBP-CorV) vaccinees (n = 30). We then investigated the sensitivity and specificity of these immunodominant regions and analyzed their conservation with other SARS-CoV-2 variants of concern, seasonal human coronaviruses, and bat Sarbecoviruses. We identified four immunodominant regions aa 29-52, aa 155-178, aa 274-297, and aa 365-388, which were highly conserved within SARS-CoV-2 and the bat coronaviruses. The magnitude of responses to these regions varied based on the infecting SARS-CoV-2 variants, >80% of individuals gave responses above the positive cut-off threshold to many of the four regions, with some differences with individuals who were infected with different VoCs. These regions were found to be 100% specific, as none of the seronegative individuals gave any responses. As these regions were highly specific with high sensitivity, they have a potential to be used to develop diagnostic assays and to be used in development of vaccines.

A comparison of changes in venous lactate and haematocrit during fluid resuscitation of dengue haemorrhagic fever.

BACKGROUND: Judicious fluid resuscitation and stringent monitoring of clinical parameters improve the outcome of dengue haemorrhagic fever (DHF). The usefulness of serum lactate to monitor adequate fluid therapy has not been adequately explored. METHODS: An observational study was conducted in Sri Lanka, recruiting 162 DHF patients within 12 h of diagnosis of the critical phase. Venous lactate level was measured at each time of performing haematocrit (HCT), using a prevalidated handheld lactate analyser. RESULTS: The median venous lactate level was 1.3 (range 0.3-6) mmol/L in the study population and 154 (95.2%) patients had median lactate levels of <2 mmol/L. The HCT values in the study participants ranged from 28 to 62, with a median value of 43. There was no statistically significant correlation between the lactate and HCT values obtained at the same time. A significant reduction in venous lactate was not observed following the administration of fluid boluses. The expected reduction in HCT was seen following the administration of dextran and crystalloid/dextran combination. The maximum recorded lactate level positively correlated with the duration of hospital stay. CONCLUSIONS: This study concludes that venous lactate is not an appropriate parameter with which to monitor the response to fluid therapy in uncomplicated DHF.

Efficacy and safety of Link Natural Sudarshana, an Ayurvedic herbal preparation in COVID-19 patients: A phase II multicenter double-blind randomized placebo-controlled trial.

ETHNOPHARMACOLOGICAL RELEVANCE: In vitro and in vivo studies have shown anti-viral and immunomodulatory actions in components of many traditional medicines. Various constituents of traditional medicines have been found to be effective against coronavirus disease (COVID-19) in several clinical trials and in-silico studies. Sudarshana cúrna, a polyherbal Ayurvedic medicine, has been used over thousands of years for a variety of infectious fevers. AIMS OF THE STUDY: This study aimed to evaluate the efficacy and safety of Link Natural Sudarshana (LNS) tablets, in patients with COVID 19 disease. LNS is a polyherbal preparation comprising 49 medicinal plants included in the Sudarshana cúrna. MATERIALS AND METHODS: A randomized parallel-group double-blind placebo-controlled multi-center phase II clinical trial was conducted in patients with mild to moderate COVID-19 disease. They were randomly allocated to intervention and control groups. The intervention group received LNS tablets whereas the control group received placebo tablets for 10 days or until the patient was discharged from the hospital. All patients received standard symptomatic treatment. The primary outcome, a reduction in mean log viral load was assessed at day 5 of treatment. The secondary outcomes, clinical progression and safety, were assessed by, monitoring changes in symptoms daily on a Likert scale ranging from 1 to 4 and laboratory tests respectively. RESULTS: A total of 171 patients (treatment group 83, control group 88) completed the trial. There were no significant differences between the baseline status of the two groups except that body mass index was significantly higher in the placebo group. The mean log viral load reduction at day 5 was higher in the treatment group (2.20 ± 1.67) compared to the placebo group (1.93 ± 1.80), with a mean difference of -0.278. This difference was not statistically significant at the 5% significant level. Reduction of mean cumulative symptom score, which included 16 symptoms graded according to severity, was higher in the treatment group compared to the placebo group. This difference was not statistically significant. None of the study participants developed hypoxia. Among the 7 lymphopenia patients in the placebo group, 3 continued to have lymphopenia at day 10, whereas 9 lymphopenia patients in the treatment group, reverted to normal counts. C reactive proteins (CRP) showed a greater reduction in the treatment group. None reported adverse effects. No significant changes occurred in hematological and biochemical parameters that assessed safety. CONCLUSIONS: LNS is safe to use in COVID-19 patients and accelerated the decline in viral load, relieved symptoms, reduced CRP levels and reversed lymphopenia earlier, when compared to the placebo.

Adipokine levels and their association with clinical disease severity in patients with dengue.

Adipokines have not been studied in acute dengue, despite their emerging role in inducing and regulating inflammation. Therefore, we sought to identify adipokine levels in patients with varying severities of acute dengue to understand their role in disease pathogenesis. We determined the levels of leptin, resistin, omentin, adiponectin, as well as IFNß, and NS1 using quantitative ELISA in patients with dengue fever (DF = 49) and dengue haemorrhagic fever (DHF = 22) at admission (febrile phase) and at the time of discharge (recovery phase). The viral loads and serotypes of all samples were quantified using quantitative real-time RT-PCR. Resistin levels (p = 0.04) and omentin (p = 0.006) levels were significantly higher in patients who developed DHF. Omentin levels in the febrile phase also correlated with the AST (Spearman's r = 0.38, p = 0.001) and ALT levels (Spearman's r = 0.24, p = 0.04); as well as serum leptin levels with both AST (Spearman's r = 0.27, p = 0.02) and ALT (Spearman's r = 0.28, p = 0.02). Serum adiponectin levels in the febrile phase did not correlate with any of the other adipokines or with liver enzymes, but inversely correlated with CRP levels (Spearman's r = -0.31, p = 0.008). Although not significant (p = 0.14) serum IFNß levels were lower in the febrile phase in those who progressed to develop DHF (median 0, IQR 0 to 39.4 pg/ml), compared to those who had DF (median 37.1, IQR 0 to 65.6 pg.ml). The data suggest that adipokines are likely to play a role in the pathogenesis of dengue, which should be further explored for the potential to be used as prognostic markers and as therapeutic targets.

Hirudin versus citrate as an anticoagulant for ROTEM platelet whole blood impedance aggregometry in thrombocytopenic patients.

Citrate is widely used as an anticoagulant for platelet function tests (PFTs). Due to an intrinsic inhibitory effect of citrate on platelet function, hirudin is used as an alternative. However, studies comparing the effect of these anticoagulants on rotational thromboelastometry (ROTEM) platelet whole blood impedance aggregometry in thrombocytopenic patients are scant. Cross-sectional study was done in 105 patients who entered the critical phase of Dengue hemorrhagic fever with plasma leakage and severe thrombocytopenia (<100 × 109/L). Samples were collected on two consecutive days and considered as a combined data set for analysis, out of which 200 have been included in the data analysis. Platelet count was used from routine full blood count. ROTEM platelet used TRAPTEM assay, which was performed with 3.2% sodium citrate and 525 ATU/ml hirudin anticoagulated blood. Means of all the TRAPTEM parameters were significantly higher in hirudin, compared to citrate samples (p < .05). Significantly higher overall platelet aggregation was observed in hirudinized samples with a significant mean difference (p < .05) compared to citrate in each quartile of platelet count. Higher platelet aggregation was observed with hirudin compared to citrate in ROTEM platelet whole blood impedance aggregometry in thrombocytopenic patients elaborating the importance of using hirudin anticoagulation in PFTs, particularly in patients with severe thrombocytopenia.

Citrate is the most commonly used anticoagulant for coagulation studies including rotational thromboelastometry (ROTEM).Hirudin is an alternative option to be used as an anticoagulant for PFTs because of the inhibitory effect of citrate on platelet function.One study (Nissen et al. (2020)) reported higher precision and platelet aggregation with hirudinized blood of healthy individuals, over citrate using ROTEM platelet.However, none of the studies were performed in patients in actual clinical context.We evaluated the potential benefit of using hirudin anticoagulated blood over citrate in thrombocytopenic patients due to Dengue hemorrhagic fever using ROTEM platelet.We observed higher platelet aggregation with hirudin compared to citrate suggesting the importance of using hirudin anticoagulation in PFTs, particularly in patients with severe thrombocytopenia.

Identification of differences in the magnitude and specificity of SARS-CoV-2 nucleocapsid antibody responses in naturally infected and vaccinated individuals.

Background: As there are limited data on B cell epitopes for the nucleocapsid protein in SARS-CoV-2, we sought to identify the immunodominant regions within the N protein, recognized by patients with varying severity of natural infection with the Wuhan strain (WT), delta, omicron and in those who received the Sinopharm vaccines, which is an inactivated, whole virus vaccine. Methods: Using overlapping peptides representing the N protein, with an in-house ELISA, we mapped the immunodominant regions within the N protein, in seronegative (n=30), WT infected (n=30), delta infected (n=30), omicron infected+vaccinated (n=20) and Sinopharm (BBIBP-CorV) vaccinees (n=30). We then investigated the sensitivity and specificity of these immunodominant regions and analysed their conservation with other SARS-CoV-2 variants of concern, seasonal human coronaviruses and bat Sarbecoviruses. We then investigated the kinetics of responses to these regions in those with varying severity of acute COVID-19. Results: We identified four immunodominant regions aa 29-52, aa 155-178, aa 274 to 297 and aa 365 to 388, were highly conserved within SARS-CoV-2 and the bat coronaviruses. The magnitude of responses to these regions varied based on the infecting SARS-CoV-2 variants, with WT infected individuals predominantly recognizing aa155 to 178 regions, delta infected individuals and vaccinated+omicron infected individuals predominantly recognizing regions aa 29 to 52 and aa 274 to 294 regions. Sinopharm vaccinees recognized all four regions, with the magnitude of responses significantly lower than other groups. >80% of individuals gave responses above the positive cut-off threshold to many of the four regions, with some differences with individuals who were infected with different VoCs. These regions were found to be 100% specific, as none of the seronegative individuals gave any responses. Conclusions: N-protein specific responses appear to be detectable in over 90% of those who were naturally infected or vaccinated with a whole virus inactivated vaccine, with responses mainly directed against four regions of the protein, which were highly conserved. As these regions were highly specific with high sensitivity, they have a potential to be used to develop diagnostic assays and to be used in development of vaccines.

Adipokine levels and their association with clinical disease severity in patients with dengue.

Adipokines have not been studied in acute dengue, despite their emerging role in inducing and regulating inflammation. Therefore, we sought to identify adipokine levels in patients with varying severities of acute dengue to understand their role in disease pathogenesis. We determined the levels of leptin, resistin, omentin, adiponectin, as well as IFNß, and NS1 using quantitative ELISA in patients with dengue fever (DF=49) and dengue haemorrhagic fever (DHF=22) at admission (febrile phase) and at the time of discharge (recovery phase). The viral loads and serotypes of all samples were quantified using quantitative real-time RT-PCR. Resistin levels (p =0.04) and omentin (p=0.006) levels were significantly higher in patients who developed DHF. Omentin levels in the febrile phase also correlated with the AST (Spearman's r=0.38, p=0.001) and ALT levels (Spearman's r=0.24, p=0.04); as well as serum leptin levels with both AST (Spearman's r=0.27, p=0.02) and ALT (Spearman's r=0.28, p=0.02). Serum adiponectin levels in the febrile phase did not correlate with any of the other adipokines or with liver enzymes, but inversely correlated with CRP levels (Spearman's r=-0.31, p=0.008). Although not significant (p=0.14) serum IFNß levels were lower in the febrile phase in those who progressed to develop DHF (median 0, IQR 0 to 39.4 pg/ml), compared to those who had DF (median 37.1, IQR 0 to 65.6 pg.ml). The data suggest that adipokines are likely to play a role in the pathogenesis of dengue, which should be further explored for the potential to be used as prognostic markers and as therapeutic targets.

Response and Survival Estimates of Patients With Plasma Cell Myeloma in a Resource-Constrained Setting Using Protocols From High-Income Countries: A Single-Center Experience From Sri Lanka.

PURPOSE: There is a significant disparity in global cancer care and outcome between countries. Progress in the treatment of symptomatic plasma cell myeloma (PCM) in high-income countries is not seen in low- and middle-income countries. MATERIALS AND METHODS: This is was a retrospective cohort study of all patients diagnosed with PCM between May 1, 2013, and September 30, 2021, at the first hemato-oncology center in Sri Lanka. We aimed to provide data on clinicopathologic characteristics, response, and survival estimates. RESULTS: A total of 79 patients with PCM received first-line therapy during the study period. The median age was 64 years, and approximately one third (33%) of patients were older than 70 years. There were 42 (53%) males and 37 females. Hypercalcemia, renal impairment, anemia, and bone disease were detected in 36.7%, 38%, 72.1%, and 81%, respectively. Thirty-nine, 34, and six patients received a combination of cyclophosphamide, thalidomide, and dexamethasone; bortezomib, thalidomide, and dexamethasone; and other treatments, respectively. The overall response rate (≥ partial response) was approximately 97% for both cyclophosphamide, thalidomide, and dexamethasone and bortezomib, thalidomide, and dexamethasone. Twenty-three (29%) of these patients died during the study period, but only 14 (18%) died due to PCM or associated sepsis. After a median follow-up of 40.6 months (range, 35.2-59.07 months), the median overall survival was 84.2 months (95% CI, 60.87 to not available). The 5-year estimated overall survival was 65%. CONCLUSION: To our knowledge, this is the only well-characterized study on long-term survival of patients with PCM in Sri Lanka. We have shown that it is possible to successfully apply Western treatment and supportive care protocols to the local population. These published data will help to benchmark and improve the treatment and develop blood cancer care in the local setting.

Efficacy of rupatadine in reducing the incidence of dengue haemorrhagic fever in patients with acute dengue: A randomised, double blind, placebo-controlled trial.

BACKGROUND: Rupatadine was previously shown to reduce endothelial dysfunction in vitro, reduced vascular leak in dengue mouse models and to reduce the extent of pleural effusions and thrombocytopenia in patients with acute dengue. Therefore, we sought to determine the efficacy of rupatadine in reducing the incidence of dengue haemorrhagic fever (DHF) in patients with acute dengue. METHODS AND FINDINGS: A phase 2, randomised, double blind, placebo controlled clinical trial was carried out in patients with acute dengue in Sri Lanka in an outpatient setting. Patients with ≤3 days since the onset of illness were either recruited to the treatment arm of oral rupatadine 40mg for 5 days (n = 123) or the placebo arm (n = 126). Clinical and laboratory features were measured daily to assess development of DHF and other complications. 12 (9.7%) patients developed DHF in the treatment arm compared to 22 (17.5%) who were on the placebo although this was not significant (p = 0.09, relative risk 0.68, 95% CI 0.41 to 1.08). Rupatadine also significantly reduced (p = 0.01) the proportion of patients with platelet counts <50,000 cells/mm3 and significantly reduced (p = 0.04) persisting vomiting, headache and hepatic tenderness (p<0.0001) in patients. There was a significant difference in the duration of illness (p = 0.0002) although the proportion of individuals who required hospital admission in both treatment arms. Only 2 patients on rupatadine and 3 patients on the placebo developed shock, while bleeding manifestations were seen in 6 patients on rupatadine and 7 patients on the placebo. CONCLUSIONS: Rupatadine appeared to be safe and well tolerated and showed a trend towards a reducing proportion of patients with acute dengue who developed DHF. Its usefulness when used in combination with other treatment modalities should be explored. TRIAL REGISTRATION: International Clinical Trials Registration Platform: SLCTR/2017/024.

Kinetics of immune responses to SARS-CoV-2 proteins in individuals with varying severity of infection and following a single dose of the AZD1222.

To characterize the IgG and IgA responses to different SARS-CoV-2 proteins, we investigated the antibody responses to SARS-CoV-2 following natural infection and following a single dose of AZD1222 (Covishield), in Sri Lankan individuals. The IgG and IgA responses were assessed to S1, S2, RBD, and N proteins in patients at 4 weeks and 12 weeks since the onset of illness or following vaccination. Antibodies to the receptor-binding domain of SARS-CoV-2 wild type (WT), α, ß, and λ and ACE2 (Angiotensin Converting Enzyme 2) receptor blocking antibodies were also assessed in these cohorts. For those with mild illness and in vaccines, the IgG responses to S1, S2, RBD, and N protein increased from 4 weeks to 12 weeks, while it remained unchanged in those with moderate/severe illness. In the vaccines, IgG antibodies to the S2 subunit had the highest significant rise (P < 0.0001). Vaccines had several-fold lower IgA antibodies to all the SARS-CoV-2 proteins tested than those with natural infection. At 12 weeks, the haemagglutination test (HAT) titres were significantly lower to the α in vaccines and significantly lower in those with mild illness and in vaccines to ß and for λ. No such difference was seen in those with moderate/severe illness. Vaccines had significantly less IgA to SARS-CoV-2, but comparable IgG responses those with natural infection. However, following a single dose vaccines had reduced antibody levels to the VOCs, which further declined with time, suggesting the need to reduce the gap between the two doses, in countries experiencing outbreaks due to VOCs.

Rotational thromboelastometry in critical phase of dengue infection: Association with bleeding.

Background: The critical phase of dengue carries a high risk of bleeding. Associations of coagulation test parameters and the risk of bleeding in the critical phase is unclear. This study examines the association of rotational thromboelastometry (ROTEM delta and ROTEM platelet) with bleeding risk of patients with dengue in the critical phase. Methods: A total of 105 patients with confirmed dengue in the critical phase were recruited, with two subsequent prospective time point analyses of ROTEM parameters and platelet count within 24 and 48 hours from the onset of the critical phase. Conventional coagulation tests were performed only at the initial time point. Results: Twenty of 105 patients developed bleeding after onset of the critical phase. Within the first 24 hours of critical-phase onset, platelet count, coagulation tests, and ROTEM delta were unable to differentiate patients with bleeding manifestations from those without (P < .05). Area under the curve of thrombin receptor activating peptide-6 assay of ROTEM platelet (TRAPTEM) discriminated patients with bleeding manifestations from those without, at a cutoff value of <12.5 Ω*min at a sensitivity and specificity of 73.7%, and 60.2%. In patients who developed bleeding, the maximum lysis of extrinsic pathway of ROTEM was significantly lower in patients with severe bleeding compared to those with mild to moderate bleeding. (4.3 ± 3.4% vs 9.4 ± 7.5%; P = .01). Conclusion: An association with bleeding manifestations and TRAPTEM suggest a potential role for defective platelet aggregation in the pathogenesis of bleeding in the critical phase of dengue.

Transcriptomics of Acute DENV-Specific CD8+ T Cells Does Not Support Qualitative Differences as Drivers of Disease Severity.

While several lines of evidence suggest a protective role of T cells against disease associated with Dengue virus (DENV) infection, their potential contribution to immunopathology in the acute phase of DENV infection remains controversial, and it has been hypothesized that the more severe form of the disease (dengue hemorrhagic fever, DHF) is associated with altered T cell responses. To address this question, we determined the transcriptomic profiles of DENV-specific CD8+ T cells in a cohort of 40 hospitalized dengue patients with either a milder form of the disease (dengue fever, DF) or a more severe disease form (dengue hemorrhagic fever, DHF). We found multiple transcriptomic signatures, one associated with DENV-specific interferon-gamma responding cells and two other gene signatures, one specifically associated with the acute phase and the other with the early convalescent phase. Additionally, we found no differences in quantity and quality of DENV-specific CD8+ T cells based on disease severity. Taken together with previous findings that did not detect altered DENV-specific CD4 T cell responses, the current analysis argues against alteration in DENV-specific T cell responses as being a correlate of immunopathology.

A Population of CD4+CD8+ Double-Positive T Cells Associated with Risk of Plasma Leakage in Dengue Viral Infection.

According to the WHO 2009 classification, dengue with warning signs is at the risk of developing severe form of dengue disease. One of the most important warning signs is plasma leakage, which can be a serious complication associated with higher morbidity and mortality. We report that the frequency of CD4+CD8+ double-positive (DP) T cells is significantly increased in patients at risk of developing plasma leakage. Transcriptomic analysis demonstrated that CD4+CD8+ DP cells were distinct from CD4+ Single Positive (SP) T cells but co-clustered with CD8+ SP cells, indicating a largely similar transcriptional profile. Twenty significant differentially expressed (DE) genes were identified between CD4+CD8+ DP and CD8+ SP cells. These genes encode OX40 and CCR4 proteins as well as other molecules associated with cell signaling on the cell surface (NT5E, MXRA8, and PTPRK). While comparing the profile of gene expression in CD4+CD8+ DP cells from patients with and without warning signs of plasma leakage, similar expression profile was observed, implying a role of CD4+CD8+ DP cells in plasma leakage through a quantitative increase rather than functional alteration. This study provided novel insight into the host immune response during the acute febrile phase of DENV infection and the role of CD4+CD8+ DP T cells in the pathogenesis of plasma leakage.

Comparison of two rapid test kits with real time polymerase chain reaction for early diagnosis of dengue in Sri Lanka.

Dengue is among the deadliest insect-borne diseases circulating in Sri Lanka. Most of the infections that are diagnosed early are manageable. However, delays in diagnosis may cause fatalities. We evaluated the dengue NS1 antigen card and NS1 SD kit for early diagnosis of dengue using samples from 116 RT-PCR-positive patients admitted within 5 days of the fever onset. RT-PCR tests were performed as standard tests. IgM and IgG ELISA tests were carried out to identify primary and secondary infections. Of the 116 patients who tested positive for dengue using PCR, 48 were positive using NS1 antigen card and 45 were positive using NS1 SD. Patients with 100 copies or higher viral load showed a higher sensitivity in both antigen card and NS1 SD. Of 34 primary infections evaluated, 23 were positive by NS1 antigen card, while the positivity was 21 by NS1 SD. Of the 30 secondary infections evaluated, 15 were positive by NS1 antigen card while 14 by NS1 SD. Our findings showed that while the rapid tests are convenient and much easier to use than PCR, they are less sensitive and need improvement. Until then, clinical diagnosis should have more emphasis on the early diagnosis of dengue.

Genomic and Epidemiological Analysis of SARS-CoV-2 Viruses in Sri Lanka.

Background: In order to understand the molecular epidemiology of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) in Sri Lanka, since March 2020, we carried out genomic sequencing overlaid on available epidemiological data until April 2021. Methods: Whole genome sequencing was carried out on diagnostic sputum or nasopharyngeal swabs from 373 patients with COVID-19. Molecular clock phylogenetic analysis was undertaken to further explore dominant lineages. Results: The B.1.411 lineage was most prevalent, which was established in Sri Lanka and caused outbreaks throughout the country until March 2021. The estimated time of the most recent common ancestor (tMRCA) of this lineage was June 1, 2020 (with 95% lower and upper bounds March 30 to July 27) suggesting cryptic transmission may have occurred, prior to a large epidemic starting in October 2020. Returning travellers were identified with infections caused by lineage B.1.258, as well as the more transmissible B.1.1.7 lineage, which has replaced B.1.411 to fuel the ongoing large outbreak in the country. Conclusions: The large outbreak that started in early October, is due to spread of a single virus lineage, B.1.411 until the end of March 2021, when B.1.1.7 emerged and became the dominant lineage.

Applicability of protocols from high-income countries in a resource limited setting; real world data of histopathology, clinical features and long-term outcome of Hodgkin Lymphoma in Sri Lanka.

Background:: There is a significant disparity in global cancer care and out-come between countries. We aimed to provide data on characteristics, average cost of treatment and survival estimates in patients with Hodgkin Lymphoma in Sri Lanka. Methods: All patients diagnosed with Hodgkin Lymphoma between 01.05.2013 and 01.10.2020 were included in the analysis. Findings: Classical Hodgkin Lymphoma(cHL) diagnosed in 85%; 68% presented with B symptoms and 61% had advanced stage of disease. Treatment was discontinued by 23% either before or just after starting treatment of whom 72% percent were females. The complete response (CR) rate of patients who continued treatment was 86% while the estimated five-year survival rate is 92%. Seventeen percent of these patients died but only two percent due to Hodgkin Lymphoma or associated treatment in the group which continued treatment compared to 45% in the group who defaulted treatment (p-value 0.0002). Five-year survival rate of patients who defaulted treatment was 50% while patients who continued treatment have an estimated five-year survival rate of 90%. Average cost of first line treatment was between US$ 2280 and US$ 7642. First treatment failure may incur substantially higher health care costs. Interpretation: This is the only well characterized study on long-term survival of patients with Hodgkin Lymphoma in Sri Lanka. We have shown that it is possible to successfully apply western treatment and supportive care protocols to the local population. This published data will help to bench mark and improve the treatment and develop blood cancer care in the local setting.

Comparison of two assays to detect IgG antibodies to the receptor binding domain of SARS­CoV­2 as a surrogate marker for assessing neutralizing antibodies in COVID-19 patients.

BACKGROUND: Neutralizing antibodies (NAbs) are important for protection against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) reinfection. In this study, two assays that are correlated with NAbs were compared: the haemagglutination test (HAT) and the surrogate virus neutralization test (sVNT). METHODS: The specificity of the HAT was compared with the sVNT, and the sensitivity and persistence of antibodies in patients with varying severity of illness was assessed in a cohort of 71 patients at 4-6 weeks and 13-16 weeks. The kinetics were assessed in the first, second, and third weeks in patients with varying severity of acute illness. RESULTS: The specificity of the HAT was >99%, and sensitivity was similar to the sVNT. The levels of HAT were significantly and positively correlated with those of the sVNT (Spearman's r = 0.78, P < 0.0001). Patients with moderate and severe illness had higher HAT titres when compared to those with mild illness. Six of seven patients with severe illness had a titre of >1:640 during the second week of illness, whereas only five of 31 patients with a mild illness had a titre of >1:160 in the second week of illness. CONCLUSIONS: Since the HAT is a simple and very cheap assay to perform, it would be ideal to use as an indicator of NAbs in resource-poor settings.

Urinary leukotrienes and histamine in patients with varying severity of acute dengue.

BACKGROUND: Vascular leak is a hallmark of severe dengue, and high leukotriene levels have been observed in dengue mouse models, suggesting a role in disease pathogenesis. We sought to explore their role in acute dengue, by assessing levels of urinary LTE4 and urinary histamine in patients with varying severity of acute dengue. METHODS: Urinary LTE4, histamine and creatinine were measured by a quantitative ELISA, in healthy individuals (n = 19), patients with dengue fever (DF = 72) and dengue haemorrhagic fever DHF (n = 48). The kinetics of LTE4 and histamine and diurnal variations were assessed in a subset of patients. RESULTS: Urinary LTE4 levels were significantly higher (p = 0.004) in patients who proceed to develop DHF when compared to patients with DF during early illness (≤ 4 days) and during the critical phase (p = 0.02), which continued to rise in patients who developed DHF during the course of illness. However, LTE4 is unlikely to be a good biomarker as ROCs gave an AUC value of 0.67 (95% CI 0.57 and 0.76), which was nevertheless significant (p = 0.002). Urinary LTE4 levels did not associate with the degree of viraemia, infecting virus serotype and was not different in those with primary vs secondary dengue. Urinary histamine levels were significantly high in patients with acute dengue although no difference was observed between patients with DF and DHF and again did not associate with the viraemia. Interestingly, LTE4, histamine and the viral loads showed a marked diurnal variation in both patients with DF and DHF. CONCLUSIONS: Our data suggest that LTE4 could play a role in disease pathogenesis and since there are safe and effective cysteinyl leukotriene receptor blockers, it would be important to assess their efficacy in reducing dengue disease severity.

SARS-CoV-2 neutralizing antibodies in patients with varying severity of acute COVID-19 illness.

In order to support vaccine development, and to aid convalescent plasma therapy, it would be important to understand the kinetics, timing and persistence of SARS-CoV-2 neutralizing antibodies (NAbs), and their association with clinical disease severity. Therefore, we used a surrogate viral neutralization test to evaluate their levels in patients with varying severity of illness, in those with prolonged shedding and those with mild/asymptomatic illness at various time points. Patients with severe or moderate COVID-19 illness had earlier appearance of NAbs at higher levels compared to those with mild or asymptomatic illness. Furthermore, those who had prolonged shedding of the virus, had NAbs appearing faster and at higher levels than those who cleared the virus earlier. During the first week of illness the NAb levels of those with mild illness was significantly less (p = 0.01), compared to those with moderate and severe illness. At the end of 4 weeks (28 days), although 89% had NAbs, 38/76 (50%) in those with > 90 days had a negative result for the presence of NAbs. The Ab levels significantly declined during convalescence (> 90 days since onset of illness), compared to 4 to 8 weeks since onset of illness. Our data show that high levels of NAbs during early illness associated with clinical disease severity and that these antibodies declined in 50% of individuals after 3 months since onset of illness.